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Lorena LOSI
Professore Associato
Dipartimento di Scienze della Vita sede Policlinico
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Pubblicazioni
2024
- Precision medicine through proteomics studies in drug resistant colorectal cancer
[Abstract in Atti di Convegno]
Costi, Maria Paola; Piazzi, Manuela; Tagliazucchi, Lorenzo; Marverti, Gaetano; D’Arca, Domenico; Moschella, MARIA GAETANA; Losi, Lorena; Bedeschi, Martina; Tesei, Anna; Passardi, Alessandro
abstract
2023
- Can immunohistochemistry improve the pathological diagnosis of placenta accreta spectrum (PAS) disorders?
[Articolo su rivista]
Losi, Lorena; Botticelli, Laura; Mancini, Luciano; Negro, Rosa; Hanspeter, Esther; Dematté, Eva; Grandi, Giovanni; Facchinetti, Fabio; Veneziano, Micaela; Malagoli, Claudia; Masini, Meris; Fabbiani, Luca; Rivasi, Francesco
abstract
Purpose: The term of placenta accreta spectrum (PAS) disorder includes all grades of abnormal placentation. It is crucial for pathologist provide standardized diagnostic assessment to evaluate the outcome of management strategies. Moreover, a correct and safe diagnosis is useful in the medico-legal field when it becomes difficult for the gynecologist to demonstrate the suitability and legitimacy of demolitive treatment. The purposes of our study were: (1) to assess histopathologic features according to the recent guidelines; (2) to determine if immunohistochemistry can be useful to identify extravillous trophoblast (EVT) and to measure the depth of infiltration into the myometrium to improve the diagnosis of PAS. Methods: The retrospective study was conducted on 30 cases of gravid hysterectomy with histopathologic diagnosis of PAS. To identify the depth of EVT, immunohistochemical stainings were performed using anti MNF116 (cytokeratins 5, 6, 8, 17, 19), actin-SM, HPL (Human Placental Lactogen), vimentin and GATA3 antibodies. Results: Our cases were graded based on the degree of invasion of the myometrium. Ten were grade 1 (33.3%), 12 grade 2 (40%) and 8 grade 3A (26.7%). EVT invasion was best seen and evident by double immunostainings with actin-SM and cytokeratins, actin-SM and HPL, actin-SM and GATA3. Conclusion: The role of pathologist is decisive to determine the different grades of PAS. A better understanding of the depth of myometrial invasion can be achieved by the use of immunohistochemistry affording an important tool to obtain reproducible grading of PAS. This purpose is crucial in the setting of postoperative quality reviews and particularly in the forensic medicine field.
2023
- Comparison between Cultivated Oral Mucosa and Ocular Surface Epithelia for COMET Patients Follow-Up
[Articolo su rivista]
Attico, Eustachio; Galaverni, Giulia; Torello, Andrea; Bianchi, Elisa; Bonacorsi, Susanna; Losi, Lorena; Manfredini, Rossella; Lambiase, Alessandro; Rama, Paolo; Pellegrini, Graziella
abstract
Total bilateral Limbal Stem Cell Deficiency is a pathologic condition of the ocular surface due to the loss of corneal stem cells. Cultivated oral mucosa epithelial transplantation (COMET) is the only autologous successful treatment for this pathology in clinical application, although abnormal peripheric corneal vascularization often occurs. Properly characterizing the regenerated ocular surface is needed for a reliable follow-up. So far, the univocal identification of transplanted oral mucosa has been challenging. Previously proposed markers were shown to be co-expressed by different ocular surface epithelia in a homeostatic or perturbated environment. In this study, we compared the transcriptome profile of human oral mucosa, limbal and conjunctival cultured holoclones, identifying Paired Like Homeodomain 2 (PITX2) as a new marker that univocally distinguishes the transplanted oral tissue from the other epithelia. We validated PITX2 at RNA and protein levels to investigate 10-year follow-up corneal samples derived from a COMET-treated aniridic patient. Moreover, we found novel angiogenesis-related factors that were differentially expressed in the three epithelia and instrumental in explaining the neovascularization in COMET-treated patients. These results will support the follow-up analysis of patients transplanted with oral mucosa and provide new tools to understand the regeneration mechanism of transplanted corneas.
2023
- Inhibition of ERK1/2 signaling prevents bone marrow fibrosis by reducing osteopontin plasma levels in a myelofibrosis mouse model
[Articolo su rivista]
Bianchi, Elisa; Rontauroli, Sebastiano; Tavernari, Lara; Mirabile, Margherita; Pedrazzi, Francesca; Genovese, Elena; Sartini, Stefano; Dall'Ora, Massimiliano; Grisendi, Giulia; Fabbiani, Luca; Maccaferri, Monica; Carretta, Chiara; Parenti, Sandra; Fantini, Sebastian; Bartalucci, Niccolò; Calabresi, Laura; Balliu, Manjola; Guglielmelli, Paola; Potenza, Leonardo; Tagliafico, Enrico; Losi, Lorena; Dominici, Massimo; Luppi, Mario; Vannucchi, Alessandro Maria; Manfredini, Rossella
abstract
Clonal myeloproliferation and development of bone marrow (BM) fibrosis are the major pathogenetic events in myelofibrosis (MF). The identification of novel antifibrotic strategies is of utmost importance since the effectiveness of current therapies in reverting BM fibrosis is debated. We previously demonstrated that osteopontin (OPN) has a profibrotic role in MF by promoting mesenchymal stromal cells proliferation and collagen production. Moreover, increased plasma OPN correlated with higher BM fibrosis grade and inferior overall survival in MF patients. To understand whether OPN is a druggable target in MF, we assessed putative inhibitors of OPN expression in vitro and identified ERK1/2 as a major regulator of OPN production. Increased OPN plasma levels were associated with BM fibrosis development in the Romiplostim-induced MF mouse model. Moreover, ERK1/2 inhibition led to a remarkable reduction of OPN production and BM fibrosis in Romiplostim-treated mice. Strikingly, the antifibrotic effect of ERK1/2 inhibition can be mainly ascribed to the reduced OPN production since it could be recapitulated through the administration of anti-OPN neutralizing antibody. Our results demonstrate that OPN is a novel druggable target in MF and pave the way to antifibrotic therapies based on the inhibition of ERK1/2-driven OPN production or the neutralization of OPN activity.
2023
- Serum Mass Spectrometry Proteomics and Protein Set Identification in Response to FOLFOX-4 in Drug-Resistant Ovarian Carcinoma
[Articolo su rivista]
D'Arca, Domenico; Severi, Leda; Ferrari, Stefania; Dozza, Luca; Marverti, Gaetano; Magni, Fulvio; Chinello, Clizia; Pagani, Lisa; Tagliazucchi, Lorenzo; Villani, Marco; D'Addese, Gianluca; Piga, Isabella; Conteduca, Vincenza; Rossi, Lorena; Gurioli, Giorgia; De Giorgi, Ugo; Losi, Lorena; Costi, Maria Paola
abstract
2022
- Destabilizers of the thymidylate synthase homodimer accelerate its proteasomal degradation and inhibit cancer growth
[Articolo su rivista]
Costantino, Luca; Ferrari, Stefania; Santucci, Matteo; MH Salo-Ahen, Outi; Carosati, Emanuele; Franchini, Silvia; Lauriola, Angela; Pozzi, Cecilia; Trande, Matteo; Gozzi, Gaia; Saxena, Puneet; Cannazza, Giuseppe; Losi, Lorena; Cardinale, Daniela; Venturelli, Alberto; Quotadamo, Antonio; Linciano, Pasquale; Tagliazucchi, Lorenzo; Moschella, MARIA GAETANA; Guerrini, Remo; Pacifico, Salvatore; Luciani, Rosaria; Genovese, Filippo; Henrich, Stefan; Alboni, Silvia; Santarem, Nuno; CORDEIRO DA SILVA, Anabela; Giovannetti, Elisa; J Peters, Godefridus; Pinton, Paolo; Rimessi, Alessandro; Cruciani, Gabriele; M Stroud, Robert; C Wade, Rebecca; Mangani, Stefano; Marverti, Gaetano; D'Arca, Domenico; Ponterini, Glauco; Costi, Maria Paola
abstract
2022
- Evidence for mitochondrial Lonp1 expression in the nucleus
[Articolo su rivista]
Gibellini, Lara; Borella, Rebecca; De Gaetano, Anna; Zanini, Giada; Tartaro, Domenico Lo; Carnevale, Gianluca; Beretti, Francesca; Losi, Lorena; De Biasi, Sara; Nasi, Milena; Forcato, Mattia; Cossarizza, Andrea; Pinti, Marcello
abstract
The coordinated communication between the mitochondria and nucleus is essential for cellular activities. Nonetheless, the pathways involved in this crosstalk are scarcely understood. The protease Lonp1 was previously believed to be exclusively located in the mitochondria, with an important role in mitochondrial morphology, mtDNA maintenance, and cellular metabolism, in both normal and neoplastic cells. However, we recently detected Lonp1 in the nuclear, where as much as 22% of all cellular Lonp1 can be found. Nuclear localization is detectable under all conditions, but the amount is dependent on a response to heat shock (HS). Lonp1 in the nucleus interacts with heat shock factor 1 (HSF1) and modulates the HS response. These findings reveal a novel extramitochondrial function for Lonp1 in response to stress.
2022
- Identification of a Quinone Derivative as a YAP/TEAD Activity Modulator from a Repurposing Library
[Articolo su rivista]
Lauriola, A.; Uliassi, E.; Santucci, M.; Bolognesi, M. L.; Mor, M.; Scalvini, L.; Elisi, G. M.; Gozzi, G.; Tagliazucchi, L.; Marverti, G.; Ferrari, S.; Losi, L.; D'Arca, D.; Costi, M. P.
abstract
The transcriptional regulators YAP (Yes-associated protein) and TAZ (transcriptional co-activator with PDZ-binding motif) are the major downstream effectors in the Hippo pathway and are involved in cancer progression through modulation of the activity of TEAD (transcriptional enhanced associate domain) transcription factors. To exploit the advantages of drug repurposing in the search of new drugs, we developed a similar approach for the identification of new hits interfering with TEAD target gene expression. In our study, a 27member in-house library was assembled, characterized, and screened for its cancer cell growth inhibition effect. In a secondary luciferase-based assay, only seven compounds confirmed their specific involvement in TEAD activity. IA5 bearing a p-quinoid structure reduced the cytoplasmic level of phosphorylated YAP and the YAP–TEAD complex transcriptional activity and reduced cancer cell growth. IA5 is a promising hit compound for TEAD activity modulator development.
2022
- LC-MS serum proteomics reveals a panel of proteins prognostic of positive responsiveness to bevacizumab therapy in late-stages ovarian cancer patients
[Abstract in Atti di Convegno]
Tagliazucchi, Lorenzo; Moschella, MARIA GAETANA; D'Addese, Gianluca; Califano, Daniela; Arenare, Laura; Mezzanzanico, Delia; Chinello, Clizia; Magni, Fulvio; Villani, Marco; Losi, Lorena; Marverti, Gaetano; D'Arca, Domenico; Chiodini, Paolo; Pignata, Sandro; Costi, Maria Paola
abstract
2022
- SOX2 Is a Univocal Marker for Human Oral Mucosa Epithelium Useful in Post-COMET Patient Characterization
[Articolo su rivista]
Attico, Eustachio; Galaverni, Giulia; Bianchi, Elisa; Losi, Lorena; Manfredini, Rossella; Lambiase, Alessandro; Rama, Paolo; Pellegrini, Graziella
abstract
Total bilateral Limbal Stem Cells Deficiency is a pathologic condition of the ocular surface due to loss or impairment of corneal stem cell function, altering homeostasis of the corneal epithelium. Cultivated Oral Mucosa Epithelial Transplantation (COMET) is the only autologous treatment for this pathology. During the follow-up, a proper characterization of the transplanted oral mucosa on the ocular surface supports understanding the regenerative process. The previously proposed markers for oral mucosa identification (e.g., keratins 3 and 13) are co-expressed by corneal and conjunctival epithelia. Here, we propose a new specific marker to distinguish human oral mucosa from the epithelia of the ocular surface. We compared the transcriptome of holoclones (stem cells) from the human oral mucosa, limbal and conjunctival cultures by microarray assay. High expression of SOX2 identified the oral mucosa vs. cornea and conjunctiva, while PAX6 was highly expressed in corneal and conjunctival epithelia. The transcripts were validated by qPCR, and immunological methods identified the related proteins. Finally, the proposed markers were used to analyze a 10-year follow-up aniridic patient treated by COMET. These findings will support the follow-up analysis of COMET treated patients and help to shed light on the mechanism of corneal repair and regeneration.
2021
- Dermal Alterations in Clinically Unaffected Skin of Pseudoxanthoma elasticum Patients
[Articolo su rivista]
Boraldi, F; Lofaro, Fd; Losi, L; Quaglino, D
abstract
Background: Pseudoxanthoma elasticum (PXE), due to rare sequence variants in the ABCC6 gene, is characterized by calcification of elastic fibers in several tissues/organs; however, the pathomechanisms have not been completely clarified. Although it is a systemic disorder on a genetic basis, it is not known why not all elastic fibers are calcified in the same patient and even in the same tissue. At present, data on soft connective tissue mineralization derive from studies performed on vascular tissues and/or on clinically affected skin, but there is no information on patients' clinically unaffected skin. Methods: Skin biopsies from clinically unaffected and affected areas of the same PXE patient (n = 6) and from healthy subjects were investigated by electron microscopy. Immunohistochemistry was performed to evaluate p-SMAD 1/5/8 and p-SMAD 2/3 expression and localization. Results: In clinically unaffected skin, fragmented elastic fibers were prevalent, whereas calcified fibers were only rarely observed at the ultrastructural level. p-SMAD1/5/8 and p-SMAD2/3 were activated in both affected and unaffected skin. Conclusion: These findings further support the concept that fragmentation/degradation is necessary but not sufficient to cause calcification of elastic fibers and that additional local factors (e.g., matrix composition, mechanical forces and mesenchymal cells) contribute to create the pro-osteogenic environment.
2021
- Folic Acid-Peptide Conjugates Combine Selective Cancer Cell Internalization with Thymidylate Synthase Dimer Interface Targeting
[Articolo su rivista]
Marverti, Gaetano; Marraccini, Chiara; Martello, Andrea; D'Arca, Domenico; Pacifico, Salvatore; Guerrini, Remo; Spyrakis, Francesca; Gozzi, Gaia; Lauriola, Angela; Santucci, Matteo; Cannazza, Giuseppe; Tagliazucchi, Lorenzo; Cazzato, Addolorata Stefania; Losi, Lorena; Ferrari, Stefania; Ponterini, Glauco; Costi, Maria P
abstract
Drug-target interaction, cellular internalization, and target engagement should be addressed to design a lead with high chances of success in further optimization stages. Accordingly, we have designed conjugates of folic acid with anticancer peptides able to bind human thymidylate synthase (hTS) and enter cancer cells through folate receptor alpha (FRalpha) highly expressed by several cancer cells. Mechanistic analyses and molecular modeling simulations have shown that these conjugates bind the hTS monomer-monomer interface with affinities over 20 times larger than the enzyme active site. When tested on several cancer cell models, these conjugates exhibited FRalpha selectivity at nanomolar concentrations. A similar selectivity was observed when the conjugates were delivered in synergistic or additive combinations with anticancer agents. At variance with 5-fluorouracil and other anticancer drugs that target the hTS catalytic pocket, these conjugates do not induce overexpression of this protein and can thus help combating drug resistance associated with high hTS levels.
2021
- Regenerative Medicine of Epithelia: Lessons From the Past and Future Goals
[Articolo su rivista]
Maurizi, Eleonora; Adamo, Davide; Magrelli, FEDERICA MARIA; Galaverni, Giulia; Attico, Eustachio; Merra, Alessia; Maffezzoni, MARIA BENEDETTA RIZZARDA; Losi, Lorena; Genna, VINCENZO GIUSEPPE; Sceberras, Virginia; Pellegrini, Graziella
abstract
This article explores examples of successful and unsuccessful regenerative medicine on human epithelia. To evaluate the applications of the first regenerated tissues, the analysis of the past successes and failures addresses some pending issues and lay the groundwork for developing new therapies. Research should still be encouraged to fill the gap between pathologies, clinical applications and what regenerative medicine can attain with current knowledge.
2021
- Structural bases for the synergistic inhibition of human thymidylate synthase and ovarian cancer cell growth by drug combinations
[Articolo su rivista]
Pozzi, C.; Santucci, M.; Marverti, G.; D'arca, D.; Tagliazucchi, L.; Ferrari, S.; Gozzi, G.; Losi, L.; Tassone, G.; Mangani, S.; Ponterini, G.; Costi, M. P.
abstract
2021
- TERT promoter methylation and protein expression as predictive biomarkers for recurrence risk in patients with serous borderline ovarian tumours
[Articolo su rivista]
Losi, Lorena; Botticelli, Laura; Garagnani, Lorella; Fabbiani, Luca; Panini, Rossana; Gallo, Graziana; Sabbatini, Roberto; Maiorana, Antonino; Benhattar, Jean
abstract
Epithelial ovarian neoplasms can be divided into three
distinct clinicopathological groups: benign, malignant and
borderline tumours. Borderline tumours are less aggressive
than epithelial carcinomas, with an indolent clinical
course and delayed recurrence. However, a subset of
these cases can progress to malignancy and relapse, and
death from recurrent disease can occasionally occur.
Telomerase activation is a critical element in cellular
immortalisation and cancer. The enzyme telomerase
comprises a catalytic subunit (TERT) expressed in various
types of cancers and regulated by promoter methylation
mainly in epithelial tumours.
The aim of this study was to investigate the promoter
methylation status and the expression of TERT in 50
serous borderline tumours (SBTs) and their correlation
with clinicopathological features and outcome. TERT
methylation was analysed by bisulfite pyrosequencing and
TERTexpression by immunohistochemistry. Methylation of
TERT promoter was only observed in four SBTs. A good
correlation with immunostochemistry was found: nuclear
positivity for TERT expression was observed in the methylated
cases, whereas no expression was detected in
unmethylated tumours. One of these patients had a
recurrence after 7 years and another patient died from the
disease. SBTs with hypomethylated tumours and absence
of TERTexpression showed a good clinical behaviour. Our
study highlights the low presence of TERT methylation in
SBTs, confirming that these tumours have a different
biology than serous carcinomas. Furthermore, the
concordance between TERT promoter methylation and
TERT expression and their association with clinical outcomes
leads to consider TERT alteration as a potential
predictive biomarker for recurrence risk identifying patients
who should undergo a careful and prolonged follow-up.
2021
- Transgenic epidermal cultures for junctional epidermolysis bullosa — 5-year outcomes
[Articolo su rivista]
Kueckelhaus, M.; Rothoeft, T.; de Rosa, L.; Yeni, B.; Ohmann, T.; Maier, C.; Eitner, L.; Metze, D.; Losi, L.; Seconetti, A. S.; de Luca, M.; Hirsch, T.
abstract
Inherited junctional epidermolysis bullosa is a severe genetic skin disease that leads to epidermal loss caused by structural and mechanical fragility of the integuments. There is no established cure for junctional epidermolysis bullosa. We previously reported that genetically corrected autologous epidermal cultures regenerated almost an entire, fully functional epidermis on a child who had a devastating form of junctional epidermolysis bullosa. We now report long-term clinical outcomes in this patient. (Funded by POR FESR 2014–2020 — Regione Emilia-Romagna and others.)
2020
- A Peptidic Thymidylate-Synthase Inhibitor Loaded on Pegylated Liposomes Enhances the Antitumour Effect of Chemotherapy Drugs in Human Ovarian Cancer Cells
[Articolo su rivista]
Marverti, Gaetano; Gozzi, Gaia; Maretti, Eleonora; Lauriola, Angela; Severi, Leda; Sacchetti, Francesca; Losi, Lorena; Pacifico, Salvatore; Ferrari, Stefania; Ponterini, Glauco; Leo, Eliana Grazia; Costi, Maria Paola; D'Arca, Domenico
abstract
There is currently no effective long-term treatment for ovarian cancer (OC) resistant to poly-chemotherapy regimens based on platinum drugs. Preclinical and clinical studies have demonstrated a strong association between development of Pt-drug resistance and increased thymidylate synthase (hTS) expression, and the consequent cross-resistance to the hTS inhibitors 5-fluorouracil (5-FU) and raltitrexed (RTX). In the present work, we propose a new tool to combat drug resistance. We propose to treat OC cell lines, both Pt-sensitive and -resistant, with dual combinations of one of the four chemotherapeutic agents that are widely used in the clinic, and the new peptide, hTS inhibitor, [D-Gln4]LR. This binds hTS allosterically and, unlike classical inhibitors that bind at the catalytic pocket, causes cell growth inhibition without inducing hTS overexpression. The dual drug combinations showed schedule-dependent synergistic antiproliferative and apoptotic effects. We observed that the simultaneous treatment or 24h pre-treatment of OC cells with the peptide followed by either agent produced synergistic effects even in resistant cells. Similar synergistic or antagonistic effects were obtained by delivering the peptide into OC cells either by means of a commercial delivery system (SAINT-PhD) or by pH sensitive PEGylated liposomes. Relative to non-PEGylated liposomes, the latter had been previously characterized and found to allow macrophage escape, thus increasing their chance to reach the tumour tissue. The transition from the SAINT-PhD delivery system to the engineered liposomes represents an advancement towards a more drug-like delivery system and a further step towards the use of peptides for in vivo studies. Overall, the results suggest that the association of standard drugs, such as cDDP and/or 5-FU and/or RTX, with the novel peptidic TS inhibitor encapsulated into PEGylated pH-sensitive liposomes can represent a promising strategy for fighting resistance to cDDP and anti-hTS drugs.
2020
- Cadherins down-regulation: towards a better understanding of their relevance in colorectal cancer
[Articolo su rivista]
Losi, Lorena; ZANOCCO MARANI, Tommaso; Grande, Alexis
abstract
The down-regulation of cadherin expression in colorectal cancer (CRC) has been widely studied. However, existing data on cadherin expression are highly variable and its relevance to CRC development has not been completely established. This review examines published studies on cadherins whose down-regulation has been already demonstrated in CRC, trying to establish a relationship with promoter methylation, the capacity to influence the Wnt / CTNNB1 (catenin beta 1, beta-catenin) signalling pathway and the clinical implications for disease outcome. Moreover, it also analyses factors that may explain data variability and highlights the importance of considering the altered subcellular localization of the examined cadherins. The results of this survey reveal that thirty of one hundred existing cadherins appear to be down-regulated in CRC. Among these, ten are cadherins, sixteen are protocadherins, equally divided between clustered and non clustered, and four are cadherin - related. These findings suggest that, to better define the role played by cadherin down-regulation in CRC pathogenesis, the expression of multiple rather than individual cadherins should be taken into account and further functional studies are necessary to clarify the relative ability of individual cadherins to inhibit CTNNB1 therefore acting as tumor suppressors.
2020
- Preclinical study for treatment of hypospadias by advanced therapy medicinal products
[Articolo su rivista]
Sceberras, V.; Attico, E.; Bianchi, E.; Galaverni, G.; Melonari, M.; Corradini, F.; Fantacci, M.; Ribbene, A.; Losi, L.; Balò, S.; Lazzeri, M.; Trombetta, C.; Rizzo, M.; Manfredini, R.; Barbagli, G.; Pellegrini, G.
abstract
Purpose
This paper explores the feasibility of a new therapy for the treatment of hypospadias patients. Hypospadias is a very common congenital malformation of male genitals, with very high rate of recurrences after surgery. The field of regenerative medicine, which offers innovative solutions for many pathologies, still does not offer reliable solution for this pathology. Here, we propose quality, safety, and clinical feasibility assessment for an oral mucosa advanced therapy medicinal product (ATMP) grown on a biocompatible scaffold for a clinical study on urethral reconstruction of hypospadias patients.
Methods
Urethral and oral mucosal epithelia from donor biopsies were cultivated between two fibrin layers, under clinical-grade conditions for cell and tissue characterization and comparison, aimed at tissue engineering. In addition, single-clone analyses were performed to analyze gene expression profiles of the two epithelia by microarray technology.
Results
Oral mucosa appeared suitable for urethral reconstruction. The resulting ATMP was proven to maintain stem cells and regenerative potency. The preclinical safety studies were performed on human tissues to assess abnormalities and tumorigenicity, and confirmed the safety of the ATMP. Finally, the patient selection and the clinical protocol for the upcoming clinical trial were defined.
Conclusions
Against this backdrop, in this paper, we are proposing a new reproducible and reliable ATMP for the treatment of hypospadias.
2019
- Deep learning techniques for detecting preneoplastic and neoplastic lesions in human colorectal histological images
[Articolo su rivista]
Sena, P.; Fioresi, R.; Faglioni, F.; Losi, L.; Faglioni, G.; Roncucci, L.
abstract
Trained pathologists base colorectal cancer identification on the visual interpretation of microscope images. However, image labeling is not always straightforward and this repetitive task is prone to mistakes due to human distraction. Significant efforts are underway to develop informative tools to assist pathologists and decrease the burden and frequency of errors. The present study proposes a deep learning approach to recognize four different stages of cancerous tissue development, including normal mucosa, early preneoplastic lesion, adenoma and cancer. A dataset of human colon tissue images collected and labeled over a 10-year period by a team of pathologists was partitioned into three sets. These were used to train, validate and test the neural network, comprising several convolutional and a few linear layers. The approach used in the present study is ‘direct’; it labels raw images and bypasses the segmentation step. An overall accuracy of >95% was achieved, with the majority of mislabeling referring to a near category. Tests on an external dataset with a different resolution yielded accuracies >80%. The present study demonstrated that the neural network, when properly trained, can provide fast, accurate and reproducible labeling for colon cancer images, with the potential to significantly improve the quality and speed of medical diagnoses.
2019
- Exome sequencing and bioinformatic approaches reveals rare sequence variants involved in cell signalling and elastic fibre homeostasis: new evidence in the development of ectopic calcification
[Articolo su rivista]
Boraldi, Federica; Lofaro, Francesco Demetrio; Romano, Oriana; Grilli, Andrea; Losi, Lorena; Moscarelli, Pasquale; Bicciato, Silvio; Quaglino, Daniela
abstract
Elastic fibres undergo aberrant mineralization in genetic as well as in acquired pathologic conditions causing severe impairment of tissue mechanical properties. Despite the number of investigations performed so far, the pathogenesis of these alterations is still elusive, due to both the complexity of the elastin network and the involvement of many genes and/or pro-osteogenic signalling pathways. Whole Exome Sequencing (WES) was performed on DNA from three patients affected by beta-thalassemia exhibiting soft connective tissue calcification. WES data were analysed with a bioinformatic approach, allowing to screen and to select genes carrying rare sequence variants. These genes were matched with those present in Extracellular Matrix DB. This approach enables to shed light on the involvement of the extracellular matrix in the occurrence of ectopic calcification. Results revealed a number of rare sequence variants in genes related to elastic fibre assembly and integrity. For instance, the involvement of fibrillins and collagen type VI in the formation of a modified microfibrillar scaffold may lead to elastic fibres less resilient and more prone to hydroxyapatite deposition. Moreover, data reveal that changes in mitochondrial metabolic pathways are sustained by a genetic background and emphasize that a persistent chronic oxidative stress can further influence extracellular matrix homeostasis and cell signalling through the TGFβ-BMP axis. Eventually, the presence of multiple rare sequence variants in the Solute Carrier Family 25 Member 5 (SLC25A5) gene is suggestive of the role of this gene as a key factor linking mitochondria metabolism, ADP/ATP ratio and oxidative stress thus affecting extracellular matrix homeostasis and activation of pro-osteogenic factors.
2019
- Involvement of epigenetic modification of TERT promoter in response to all-trans retinoic acid in ovarian cancer cell lines
[Articolo su rivista]
Losi, Lorena; Lauriola, Angela; Tazzioli, Erica; Gozzi, Gaia; Scurani, Letizia; D'Arca, Domenico; Jean, Benhattar
abstract
Background: All-trans retinoic acid (ATRA) is currently being used to treat hematological malignancies, given the ability to inhibit cell proliferation. This effect seems to be related to epigenetic changes of the TERT (Telomerase Reverse Transcriptase) promoter. When hypomethylated, ATRA-inducible TERT repressors can bind the promoter, repressing transcription of TERT, the rate-limiting component of telomerase. Ovarian carcinomas are heterogeneous tumors characterized by several aberrantly methylated genes among which is TERT. We recently found a hypomethylation of TERT promoter in about one third of serous carcinoma, the most lethal histotype. Our aim was to investigate the potential role of ATRA as an anticancer drug in a sub-group of ovarian carcinoma where the TERT promoter was hypomethylated.
Methods: The potential antiproliferative and cytotoxic effect of ATRA was investigated in seven serous ovarian carcinoma and one teratocarcinoma cell lines and the results were compared to the methylation status of their TERT promoter.
Results: The serous ovarian carcinoma cell line OVCAR3, harboring a hypomethylated TERT promoter, was the best and fastest responder. PA1 and SKOV3, two cell lines with an intermediate methylated promoter, revealed a weaker and delayed response. On the contrary, the other 5 cell lines with a highly methylated promoter did not respond to ATRA, indicative of ATRA-resistant cells.
Conclusions: Our results demonstrate an inverse correlation between the methylation level of TERT promoter and ATRA efficacy in ovarian carcinoma cell lines. Although these results are preliminary, ATRA treatment could become a new powerful, personalized therapy in serous ovarian carcinoma patients, but only in those with tumors harboring a hypomethylated TERT promoter.
2019
- Loss of expression of μ-protocadherin and protocadherin-24 in sporadic and hereditary nonpolyposis colorectal cancers
[Articolo su rivista]
Losi, Lorena; Lancellotti, Cesare; Parenti, Sandra; Scurani, Letizia; Zanocco-Marani, Tommaso; Buffoli, Federico; Grassia, Roberto; Ferrari, Sergio; Grande, Alexis
abstract
Colorectal cancer (CRC) is a neoplastic disease in which normal mucosa undergoes a process of malignant transformation due to the progressive accumulation of molecular alterations affecting proto-oncogenes and oncosuppressor genes. Some of these modifications exert their carcinogenic potential by promoting a constitutive activation of the β-catenin signaling proliferation pathway, and when present, loss of cadherin expression also significantly contributes to the same effect. Using a combined approach of molecular and immunohistochemical analysis, we have previously demonstrated that most sporadic CRCs exhibit a down-regulated expression of a cadherin, named μ-protocadherin, that is generally observed in association with a higher proliferation rate and a worse prognosis. The aim of this report was to perform a comparative immunohistochemical assessment of μ-protocadherin and a similar cadherin, named protocadherin-24, in sporadic CRC and hereditary nonpolyposis colorectal cancer. The data obtained put in evidence that double-negative CRCs, lacking both the analyzed protocadherins, are more represented among sporadic tumors, whereas double-positive CRCs, maintaining their expression, exhibit an opposite trend. As expected, loss of protocadherin expression was accompanied by nuclear localization of β-catenin and increased positivity of the Ki-67 proliferation marker. This finding is consistent with the different clinical evolution of the 2 considered CRC sets according to which patients with hereditary nonpolyposis colorectal cancer experience a better prognosis as compared with those affected by a sporadic CRC.
2019
- Role of evaluating tumor‑infiltrating lymphocytes, programmed death‑1 ligand 1 and mismatch repair proteins expression in malignant mesothelioma
[Articolo su rivista]
Losi, Lorena; Bertolini, Federica; Guaitoli, Giorgia; Fabbiani, Luca; Banchelli, Federico; Ambrosini-Spaltro, Andrea; Botticelli, Laura; Scurani, Letizia; Baldessari, Cinzia; Barbieri, Fausto; Cascinu, Stefano; Maiorana, Antonino
abstract
The tumor immune microenvironment (TME) and immune checkpoints have been reported to serve a role in the pathogenesis of malignant mesothelioma (MM) and treatment outcome. Additionally, mismatch Repair (MMR) deficiency appears to enhance the response to checkpoints blockade in several tumors. The aim of the present study was to analyze programmed death‑1 ligand 1 (PD‑L1) expression in MM and to characterize the TME. This could help to understand the immune response, and evaluate its prognostic and predictive values. We also investigated MMR protein expression. We retrospectively analyzed 55 mesotheliomas to determine PD‑L1, CD4+, CD8+, mutL homolog 1 (MLH1), mutS homolog 2 (MSH2), mutS homolog 6 (MSH6) and PMS1 homolog 2, mismatch repair system component (PMS2) expression. We used an immunoscore (1+, 2+ and 3+) to evaluate tumor‑infiltrating lymphocytes (TILs). TILs were observed in all but two samples (53/55); the majority had an immunoscore 1+ (30/53), while 2+/3+ was reported for 23/53 samples. A predominance of CD8+ was highlighted in 8 cases (15%). PD‑L1 expression of ≥1% on tumor cells was displayed in 40 cases; in 9 of these, ≥50% expression was reported. Of note, alterations in MMR staining was not observed. In addition, survival analysis revealed that epithelioid subtype was associated with better prognosis. We observed a trend towards poorer prognosis for ≥50% PD‑L1 expression on tumor cells, lower immunoscore (1+) and CD8+ TIL predominance. The present study highlighted the importance of exploring the TME and the standardization of PD‑L1 assessment guidelines to apply in the field of immunotherapy.
2018
- Conformational Propensity and Biological Studies of Proline Mutated LR Peptides Inhibiting Human Thymidylate Synthase and Ovarian Cancer Cell Growth
[Articolo su rivista]
Saxena, Puneet; Severi, Leda; Santucci, Matteo; Taddia, Laura; Ferrari, Stefania; Luciani, Rosaria; Marverti, Gaetano; Marraccini, Chiara; Tondi, Donatella; Mor, Marco; Laura, Scalvini; Vitiello, Simone; Losi, Lorena; Fonda, Sergio; Pacifico, Salvatore; Guerrini, Remo; D'Arca, Domenico; Ponterini, Glauco; Costi, Maria Paola
abstract
LR and [D-Gln4]LR peptides bind the mono- mer−monomer interface of human thymidylate synthase and inhibit cancer cell growth. Here, proline-mutated LR peptides were synthesized. Molecular dynamics calculations and circular dichroism spectra have provided a consistent picture of the conformational propensities of the [Pron]-peptides. [Pro3]LR and [Pro4]LR show improved cell growth inhibition and similar intracellular protein modulation compared with LR. These represent a step forward to the identification of more rigid and metabolically stable peptides.
2018
- Distinct DNA Methylation Profiles in Ovarian Tumors: Opportunities for Novel Biomarkers
[Articolo su rivista]
Losi, Lorena; Fonda, Sergio; Saponaro, Sara; Chelbi, Sonia T.; Cesare, Lancellotti; Gozzi, Gaia; Loredana, Alberti; Fabbiani, Luca; Laura, Botticelli; Jean, Benhattar
abstract
Aberrant methylation of multiple promoter CpG islands could be related to the
biology of ovarian tumors and its determination could help to improve treatment strategies.
DNA methylation profiling was performed using the Methylation Ligation-dependent Macroarray
(MLM), an array-based analysis. Promoter regions of 41 genes were analyzed in 102 ovarian tumors
and 17 normal ovarian samples. An average of 29% of hypermethylated promoter genes was
observed in normal ovarian tissues. This percentage increased slightly in serous, endometrioid,
and mucinous carcinomas (32%, 34%, and 45%, respectively), but decreased in germ cell tumors
(20%). Ovarian tumors had methylation profiles that were more heterogeneous than other epithelial
cancers. Unsupervised hierarchical clustering identified four groups that are very close to the
histological subtypes of ovarian tumors. Aberrant methylation of three genes (BRCA1, MGMT,
and MLH1), playing important roles in the different DNA repair mechanisms, were dependent on the
tumor subtype and represent powerful biomarkers for precision therapy. Furthermore, a promising
relationship between hypermethylation of MGMT, OSMR, ESR1, and FOXL2 and overall survival was
observed. Our study of DNA methylation profiling indicates that the different histotypes of ovarian
cancer should be treated as separate diseases both clinically and in research for the development of
targeted therapies.
2018
- Human Thymidylate Synthase Inhibitors Halting Ovarian Cancer Growth
[Articolo su rivista]
Ferrari, Stefania; Severi, Leda; Cecilia, Pozzi; Quotadamo, Antonio; Ponterini, Glauco; Losi, Lorena; Marverti, Gaetano; Costi, Maria Paola
abstract
Human thymidylate synthase (hTS) has an important role in DNA biosynthesis, thus it is
essential for cell survival. TS is involved in the folate pathways, specifically in the de novo
pyrimidine biosynthesis. Structure and functions are intimately correlated, account for
cellular activity and, in a broader view, with in vivo mechanisms. hTS is a target for anticancer
agents, some of which are clinical drugs. The understanding of the detailed
mechanism of TS inhibition by currently used drugs and of the interaction with the
mechanism of action of other anticancer agents can suggest new perspective of TS inhibition
able to improve the anticancer effect and to overcome drug resistance.
TS-targeting drugs in therapy today are inhibitors that bind at the active site and that
mostly resemble the substrates. Nonsubstrate analogs offer an opportunity for allosteric
binding and novel mode of inhibition in the cancer cells. This chapter illustrates
the relationship among the large number of hTS actions at molecular and clinical levels, its role as a target for ovarian cancer therapy, in particular in cases of overexpression
of hTS and other folate proteins such as those induced by platinum drug treatments,
and address the potential combination of TS inhibitors with other suitable anticancer
agents.
2018
- KLF4 mediates the effect of 5-ASA on the b-catenin pathway in colon cancer cells
[Articolo su rivista]
Parenti, Sandra; Montorsi, Lucia; Fantini, Sebastian; Mammoli, Fabiana; Gemelli, Claudia; Atene, Claudio Giacinto; Losi, Lorena; Frassineti, Chiara; Calabretta, Bruno; Tagliafico, Enrico; Ferrari, Sergio; Zanocco-Marani, Tommaso; Grande, Alexis
abstract
Mesalazine (5-ASA) is an aminosalicylate anti-inflammatory drug capable of inducing m-protocadherin, a protein expressed by colorectal epithelial cells that is downregulated upon malignant transformation. Treatment with 5-ASA restores m-protocadherin expression and promotes the sequestration of b-catenin to the plasma membrane. Here, we show that 5-ASA–induced m-protocadherin expression is directly regulated by the KLF4 transcription factor. In addition, we suggest the existence of a dual mechanism whereby 5-ASA–mediated b-catenin inhibition is caused by m-protocadherin–dependent sequestration of b-catenin to the plasma membrane and by the direct binding of KLF4 to b-catenin. In addition, we found that 5-ASA treatment suppresses the expression of miR-130a and miR-135b, which target KLF4 mRNA, raising the possibility that this mechanism is involved in the increased expression of KLF4 induced by 5-ASA.
2018
- LonP1 Differently Modulates Mitochondrial Function and Bioenergetics of Primary Versus Metastatic Colon Cancer Cells
[Articolo su rivista]
Gibellini, L; Losi, L; De Biasi, S; Nasi, M; Lo Tartaro, D; Pecorini, S; Patergnani, S; Pinton, P; De Gaetano, A; Carnevale, G; Pisciotta, A; Mariani, F; Roncucci, L; Iannone, A; Cossarizza, A; Pinti, M.
abstract
Mitochondrial Lon protease (LonP1) is a multi-function enzyme that regulates mitochondrial functions in several human malignancies, including colorectal cancer (CRC). The mechanism(s) by which LonP1 contributes to colorectal carcinogenesis is not fully understood. We found that silencing LonP1 leads to severe mitochondrial impairment and apoptosis in colon cancer cells. Here, we investigate the role of LonP1 in mitochondrial functions, metabolism, and epithelial-mesenchymal transition (EMT) in colon tumor cells and in metastasis. LonP1 was almost absent in normal mucosa, gradually increased from aberrant crypt foci to adenoma, and was most abundant in CRC. Moreover, LonP1 was preferentially upregulated in colorectal samples with mutated p53 or nuclear β-catenin, and its overexpression led to increased levels of β-catenin and decreased levels of E-cadherin, key proteins in EMT, in vitro. LonP1 upregulation also induced opposite changes in oxidative phosphorylation, glycolysis, and pentose pathway in SW480 primary colon tumor cells when compared to SW620 metastatic colon cancer cells. In conclusion, basal LonP1 expression is essential for normal mitochondrial function, and increased LonP1 levels in SW480 and SW620 cells induce a metabolic shift toward glycolysis, leading to EMT.
2018
- Malignant peritoneal mesothelioma in a woman with bilateral ovarian serous borderline tumour: Potential interactions between the two diseases
[Articolo su rivista]
Losi, Lorena; Laura, Botticelli; Gianluca, Taccagni; Ernesto, Longinotti; Cesare, Lancellotti; Scurani, Letizia; Gian Franco Zannoni,
abstract
We report a case of a 59-year-old woman with peritoneal malignant mesothelioma and no previous exposure to
asbestos with a diagnosis of bilateral ovarian serous borderline tumour with peritoneal implants one year before.
We discuss the histopathological and immunohistochemical findings to explain possible and potential interactions
between the two diseases. To our knowledge, the association of both serous borderline ovarian tumour and
malignant peritoneal mesothelioma has never been described before in the same woman and in such a tight
temporal connection. This finding raises numerous issues about the origin of the two tumours and further
biomolecular studies are needed to fully understand the carcinogenetic process. From a clinical point of view,
this case report can be useful to gynaecologists because it leads to recommend a careful examination of the
peritoneal cavity during a surgical resection of borderline serous tumour. Moreover, it may suggest performing a
close follow-up associated with a careful surveillance of the patient, especially in the case of micropapillary
pattern, to oncologists. A complete clinical approach could help to detect sooner possible relapses or other
metachronous malignancies.
2018
- Oxaliplatin plus leucovorin and 5-fluorouracil (FOLFOX-4) as a salvage chemotherapy in heavily-pretreated platinum-resistant ovarian cancer
[Articolo su rivista]
Conteduca, Vincenza; Gurioli, Giorgia; Rossi, Lorena; Scarpi, Emanuela; Lolli, Cristian; Schepisi, Giuseppe; Farolfi, Alberto; De Lisi, Delia; Gallà, Valentina; Luca Burgio, Salvatore; Menna, Cecilia; Amadori, Andrea; Losi, Lorena; Amadori, Dino; Costi, Maria Paola; De Giorgi, Ugo
abstract
Background: The purpose of this study was to evaluate the clinical impact of oxaliplatin, leucovorin, and 5-
fluorouracil (FOLFOX-4) chemotherapy in terms of the response rate, progression-free/overall survival (PFS/OS) and
safety profile in patients with heavily pretreated recurrent epithelial ovarian cancer.
Methods: Clinical data were reviewed in 29 patients who received FOLFOX-4 as more than third-line chemotherapy,
consisting of 85 mg/m2 of oxaliplatin, 200 mg/m2 of leucovorin, and bolus 400 mg/m2 on day 1 of 5-fluorouracil,
followed by a 22-h infusion of 600 mg/m2 of 5-fluorouracil for 2 consecutive days every 3 weeks. We also compared
the efficacy and toxicity of FOLFOX-4 with that of topotecan, a standard treatment, given at a dosage of 1.5 mg/m2
every three weeks in 26 patients.
Results: The median age of enrolled patients was 60 years (range 33 to 85). A median of 4 cycles (range 1–17) of
FOLFOX-4 were administered. Complete response and partial response were observed in one (3.5%) and 5 (17.2.2%)
patients, respectively, while stable disease was reported in 8 (27.6%) patients. Among all patients, grade 3–4 anemia,
neutropenia, and thrombocytopenia were observed in 0 (0%), 5 (17.2%), and 3 (10.3%) cases, respectively. Grade 3–4
fatigue was recorded in one (3.4%) patient and diarrhea in 2 (6.9%). Median PFS and OS were 2.8 months [95%
confidence interval (CI) 1.7–4.9] and 6.2months (95% CI 2.4–14.6), respectively. No significant differences in terms of
efficacy and toxicity were observed between patients receiving FOLFOX-4 and those treated with topotecan.
Conclusions: The FOLFOX-4 regimen would seem to obtain similar survival rates to those of standard therapy with
topotecan in platinum-resistant ovarian cancer. Further randomized trials are warranted to confirm our findings.
2018
- Proteomic and bioinformatic studies for the characterization of response to pemetrexed in platinum drug resistant ovarian cancer
[Articolo su rivista]
Severi, Leda; Losi, Lorena; Fonda, Sergio; Taddia, Laura; Gozzi, Gaia; Marverti, Gaetano; Magni, Fulvio; Chinello, Clizia; Stella, Martina; Sheouli, Jalid; Braicu, Elena I.; Genovese, Filippo; Lauriola, Angela; Marraccini, Chiara; Gualandi, Alessandra; D'Arca, Domenico; Ferrari, Stefania; Costi, Maria P.
abstract
Proteomics and bioinformatics are a useful combined technology for the characterization of protein expression level and modulation associated with the response to a drug and with its mechanism of action. The folate pathway represents an important target in the anticancer drugs therapy. In the present study, a discovery proteomics approach was applied to tissue samples collected from ovarian cancer patients who relapsed after the first-line carboplatin-based chemotherapy and were treated with pemetrexed (PMX), a known folate pathway targeting drug. The aim of the work is to identify the proteomic profile that can be associated to the response to the PMX treatment in pre-treatement tissue. Statistical metrics of the experimental Mass Spectrometry (MS) data were combined with a knowledge-based approach that included bioinformatics and a literature review through ProteinQuest™ tool, to design a protein set of reference (PSR). The PSR provides feedback for the consistency of MS proteomic data because it includes known validated proteins. A panel of 24 proteins with levels that were significantly different in pre-treatment samples of patients who responded to the therapy vs. the non-responder ones, was identified. The differences of the identified proteins were explained for the patients with different outcomes and the known PMX targets were further validated. The protein panel herein identified is ready for further validation in retrospective clinical trials using a targeted proteomic approach. This study may have a general relevant impact on biomarker application for cancer patients therapy selection.
2018
- Repurposing of drugs targeting yap-tead functions
[Articolo su rivista]
Elisi, Gian Marco; Santucci, Matteo; D’Arca, Domenico; Lauriola, Angela; Marverti, Gaetano; Losi, Lorena; Scalvini, Laura; Bolognesi, Maria Laura; Mor, Marco; Costi, Maria Paola
abstract
Drug repurposing is a fast and consolidated approach for the research of new active compounds bypassing the long streamline of the drug discovery process. Several drugs in clinical practice have been reported for modulating the major Hippo pathway’s terminal effectors, namely YAP (Yes1-associated protein), TAZ (transcriptional co-activator with PDZ-binding motif) and TEAD (transcriptional enhanced associate domains), which are directly involved in the regulation of cell growth and tissue homeostasis. Since this pathway is known to have many cross-talking phenomena with cell signaling pathways, many efforts have been made to understand its importance in oncology. Moreover, this could be relevant to obtain new molecular tools and potential therapeutic assets. In this review, we discuss the main mechanisms of action of the best-known compounds, clinically approved or investigational drugs, able to cross-talk and modulate the Hippo pathway, as an attractive strategy for the discovery of new potential lead compounds.
2017
- Desmoplastic melanoma: a challenge for the oncologist
[Articolo su rivista]
Manfredini, Marco; Pellacani, Giovanni; Losi, Lorena; Maccaferri, Monia; Tomasi, Aldo; Ponti, Giovanni
abstract
To evaluate clinical, pathologic and genetic features of desmoplastic melanoma (DM).MATERIALS & METHODS:
Analysis of all DM records from 1991 to 2015.
RESULTS:
The most common location of DMs was the head and neck (69%); median age and follow-up were 60.5 and 7.3 years, respectively. A familial predisposition for DMs and others malignancies was analyzed. Thin Breslow thickness (<4.5 mm) was associated with an intraepidermal component or a previous lentigo maligna, whereas high Breslow thickness (>4.5 mm) was observed in 'pure' DM.
CONCLUSION:
DM could progress from an early phase, characterized by an intraepidermal component, to late phase, characterized by a dermal nodule. This hypothesis correlates with melanoma genetic and NF1 mutation, which could be an early event in the progression of DM.
2017
- Pigment epithelial-derived factor: a new player in dermal elastic fibre calcification?
[Articolo su rivista]
Boraldi, Federica; Losi, Lorena; Quaglino, Daniela
abstract
Pigment epithelium-derived factor (PEDF) is an endogenously produced glycoprotein expressed in several organs during developmental stages and adulthood mainly acting on cell differentiation.(1) In vitro and in vivo studies have demonstrated that PEDF has neurotrophic and antioxidant activities as well as the ability to counteract angiogenesis, tumorigenesis, atherosclerosis, thrombosis and inflammation.(1,2) In addition, PEDF has been also related to bone metabolism, increasing alkaline phosphatase (ALP) expression and promoting osteoblast differentiation.(3) This article is protected by copyright. All rights reserved.
2016
- DNA methylation profiling of esophageal adenocarcinoma using Methylation Ligation-dependent Macroarray (MLM)
[Articolo su rivista]
Guilleret, Isabelle; Losi, Lorena; Chelbi, Sonia T; Fonda, Sergio; Bougel, Stéphanie; Saponaro, Sara; Gozzi, Gaia; Alberti, Loredana; Braunschweig, Richard; Benhattar, Jean
abstract
Most types of cancer cells are characterized by aberrant methylation of promoter genes. In this study, we described a rapid, reproducible, and relatively inexpensive approach allowing the detection of multiple human methylated promoter genes from many tissue samples, without the need of bisulfite conversion.The Methylation Ligation-dependent Macroarray (MLM), an array-based analysis, was designed in order to measure methylation levels of 58 genes previously described as putative biomarkers of cancer. The performance of the design was proven by screening the methylation profile of DNA from esophageal cell lines, as well as microdissected formalin-fixed and paraffin-embedded (FFPE) tissues from esophageal adenocarcinoma (EAC). Using the MLM approach, we identified 32 (55%) hypermethylated promoters in EAC, and not or rarely methylated in normal tissues. Among them, 21promoters were found aberrantly methylated in more than half of tumors. Moreover, seven of them (ADAMTS18, APC, DKK2, FOXL2, GPX3, TIMP3 and WIF1) were found aberrantly methylated in all or almost all the tumor samples, suggesting an important role for these genes in EAC. In addition, dysregulation of the Wnt pathway with hypermethylation of several Wnt antagonist genes was frequently observed. MLM revealed a homogeneous pattern of methylation for a majority of tumors which were associated with an advanced stage at presentation and a poor prognosis. Interestingly, the few tumors presenting less methylation changes had a lower pathological stage. In conclusion, this study demonstrated the feasibility and accuracy of MLMfor DNA methylation profiling of FFPE tissue samples.
2016
- Expression of μ-protocadherin is negatively regulated by the activation of the β-catenin signaling pathway in normal and cancer colorectal enterocytes
[Articolo su rivista]
Montorsi, Lucia; Parenti, Sandra; Losi, Lorena; Ferrarini, F; Gemelli, Claudia; Rossi, A; Manco, Gianrocco; Ferrari, Sergio; Calabretta, Bruno; Tagliafico, Enrico; ZANOCCO MARANI, Tommaso; Grande, Alexis
abstract
Mu-protocadherin (MUCDHL) is an adhesion molecule predominantly expressed by colorectal epithelial cells which is markedly downregulated upon malignant transformation. Notably, treatment of colorectal cancer (CRC) cells with mesalazine lead to increased expression of MUCDHL, and is associated with sequestration of β-catenin on the plasma membrane and inhibition of its transcriptional activity. To better characterize the causal relationship between β-catenin and MUCDHL expression, we performed various experiments in which CRC cell lines and normal colonic organoids were subjected to culture conditions inhibiting (FH535 treatment, transcription factor 7-like 2 siRNA inactivation, Wnt withdrawal) or stimulating (LiCl treatment) β-catenin activity. We show here that expression of MUCDHL is negatively regulated by functional activation of the β-catenin signaling pathway. This finding was observed in cell culture systems representing conditions of physiological stimulation and upon constitutive activation of β-catenin in CRC. The ability of MUCDHL to sequester and inhibit β-catenin appears to provide a positive feedback enforcing the effect of β-catenin inhibitors rather than serving as the primary mechanism responsible for β-catenin inhibition. Moreover, MUCDHL might have a role as biomarker in the development of CRC chemoprevention drugs endowed with β-catenin inhibitory activity.
2016
- Promoter Methylation and Down-regulated Expression of the TBX15 Gene in Ovarian Carcinoma
[Articolo su rivista]
Gozzi, Gaia; Chelbi, Sonia T; Manni, Paola; Alberti, Loredana; Fonda, Sergio; Saponaro, Sara; Fabbiani, Luca; Rivasi, Francesco; Benhattar, Jean; Losi, Lorena
abstract
TBX15 is a gene involved in the development of mesodermal derivatives. As the ovaries and the female reproductive system are of mesodermal origin, the aim of the present study was to determine the methylation status of the TBX15 gene promoter and the expression levels of TBX15 in ovarian carcinoma, which is the most lethal and aggressive type of gynecological tumor, in order to determine the role of TBX15 in the pathogenesis of ovarian carcinoma. This alteration could be used to predict tumor development, progression, recurrence and therapeutic effects. The study was conducted on 80 epithelial ovarian carcinoma and 17 control cases (normal ovarian and tubal tissues). TBX15 promoter methylation was first determined by pyrosequencing following bisulfite modification, then by cloning and sequencing, in order to obtain information about the epigenetic haplotype. Immunohistochemical analysis was performed to evaluate the correlation between the methylation and protein expression levels. Data revealed a statistically significant increase of the TBX15 promoter region methylation in 82% of the tumor samples and in various histological subtypes. Immunohistochemistry showed an inverse correlation between methylation levels and the expression of the TBX15 protein. Furthermore, numerous tumor samples displayed varying degrees of intratumor heterogeneity. Thus, the present study determined that ovarian carcinoma typically expresses low levels of TBX15 protein, predominantly due to an epigenetic mechanism. This may have a role in the pathogenesis of ovarian carcinoma independent of the histological subtype.
2016
- Robbins e Cotran Atlante di Anatomia Patologica
[Traduzione in Volume]
Losi, Lorena
abstract
L'Atlante racchiude le conoscenze di base dei testi di Robbins in un formato essenzialmente visivo, pensato per agevolare lo studio e l'apprendimento. Le immagini macroscopiche, microscopiche e radiologiche utilizzate nell'Atlante servono a rinforzare i contenuti presentati nel volume e nelle altre opere della serie Robbins. L'approccio all'insegnamento della Medicina adottato dall'autore è un approccio integrativo che combina, all'interno dei materiali didattici, elementi attinti dalle scienze di base, da quelle cliniche e dalle scienze comportamentali, promuovendo lo sviluppo delle conoscenze a favore di chi necessita dell'assistenza sanitaria.
2015
- Different Effects of BORIS/CTCFL on Stemness Gene Expression, Sphere Formation and Cell Survival in Epithelial Cancer Stem Cells
[Articolo su rivista]
Alberti, Loredana; Losi, Lorena; Leyvraz, Serge; Benhattar, Jean
abstract
Cancer stem cells are cancer cells characterized by stem cell properties and represent a small population of tumor cells that drives tumor development, progression, metastasis and drug resistance. To date, the molecular mechanisms that generate and regulate cancer stem cells are not well defined. BORIS (Brother of Regulator of Imprinted Sites) or CTCFL (CTCF-like) is a DNA-binding protein that is expressed in normal tissues only in germ cells and is re-activated in tumors. Recent evidences have highlighted the correlation of BORIS/CTCFL expression with poor overall survival of different cancer patients. We have previously shown an association of BORIS-expressing cells with stemness gene expression in embryonic cancer cells. Here, we studied the role of BORIS in epithelial tumor cells. Using BORIS-molecular beacon that was already validated, we were able to show the presence of BORIS mRNA in cancer stem cell-enriched populations (side population and spheres) of cervical, colon and breast tumor cells. BORIS silencing studies showed a decrease of sphere formation capacity in breast and colon tumor cells. Importantly, BORIS-silencing led to down-regulation of hTERT, stem cell (NANOG, OCT4, SOX2 and BMI1) and cancer stem cell markers (ABCG2, CD44 and ALDH1) genes. Conversely, BORIS-induction led to up-regulation of the same genes. These phenotypes were observed in cervical, colon and invasive breast tumor cells. However, a completely different behavior was observed in the non-invasive breast tumor cells (MCF7). Indeed, these cells acquired an epithelial mesenchymal transition phenotype after BORIS silencing. Our results demonstrate that BORIS is associated with cancer stem cell-enriched populations of several epithelial tumor cells and the different phenotypes depend on the origin of tumor cells.
2014
- High Expression of hTERT and stemness genes in BORIS/CTCFL positive cells isolated from embryonic cancer cells
[Articolo su rivista]
Alberti, Loredana; Renaud, Stéphanie; Losi, Lorena; Leyvraz, Serge; Jean, Benhattar
abstract
BORIS/CTCFL is a member of cancer testis antigen family normally expressed in germ cells. In tumors, it is aberrantly expressed although its functions are not completely well-defined. To better understand the functions of BORIS in cancer, we selected the embryonic cancer cells as a model. Using a molecular beacon, which specifically targets BORIS mRNA, we demonstrated that BORIS positive cells are a small subpopulation of tumor cells (3–5% of total). The BORIS-positive cells isolated using BORIS-molecular beacon, expressed higher telomerase hTERT, stem cell (NANOG, OCT4, SOX2) and cancer stem cell marker genes (CD44 and ALDH1) compared to the BORIS-negative tumor cells. In order to define the functional role of BORIS, stable BORIS-depleted embryonic cancer cells were generated. BORIS silencing strongly down-regulated the expression of hTERT, stem cell and cancer stem cell marker genes. Moreover, the BORIS knockdown increased cellular senescence in embryonic cancer cells, revealing a putative role of BORIS in the senescence biological program. Our data indicate an association of BORIS expressing cells subpopulation with the expression of stemness genes, highlighting the critical role played by BORIS in embryonic neoplastic disease.
2014
- Involvement of μ-protocadherin in colorectal carcinogenesis: a promise for clinico-pathological evaluation.
[Articolo su rivista]
Losi, Lorena; Grande, Alexis
abstract
Cell-cell adhesion is a fundamental activity to allow the maintenance of epithelial integrity, and defects impairing this process promote the formation of tumors such as colorectal cancer (CRC). In this regard, a crucial role is played by adhesion molecules, named cadherins, which exert their function through the inhibition of the beta-catenin signaling proliferation pathway, constitutively activated in CRC. A number of reports, published over the last decade, have highlighted the existence of a novel cadherin family member, called μ-protocadherin, to underline the hybrid nature of its extra-cellular region, including both cadherin-like and mucin-like domains. Is has been shown that this protein plays an important role in inter-cellular adhesion processes, inhibits beta-catenin activity in normal colorectal mucosa, undergoes a down-regulated expression in CRC and is up-regulated upon treatment with chemoprevention agents against this tumor.
2014
- Silencing of mitochondrial Lon protease deeply impairs mitochondrial proteome and function in colon cancer cells.
[Articolo su rivista]
Gibellini, Lara; Pinti, Marcello; Boraldi, Federica; Giorgio, Valentina; Bernardi, Paolo; Bartolomeo, Regina; Nasi, Milena; DE BIASI, Sara; Missiroli, Sonia; Carnevale, Gianluca; Losi, Lorena; Tesei, Anna; Pinton, Paolo; Quaglino, Daniela; Cossarizza, Andrea
abstract
Lon is a nuclear-encoded, mitochondrial protease that assists protein folding, degrades oxidized/damaged proteins, and participates in maintaining mtDNA levels. Here we show that Lon is up-regulated in several human cancers and that its silencing in RKO colon cancer cells causes profound alterations of mitochondrial proteome and function, and cell death. We silenced Lon in RKO cells by constitutive or inducible expression of Lon shRNA. Lon-silenced cells displayed altered levels of 39 mitochondrial proteins (26% related to stress response, 14.8% to ribosome assembly, 12.7% to oxidative phosphorylation, 8.5% to Krebs cycle, 6.3% to β-oxidation, and 14.7% to crista integrity, ketone body catabolism, and mtDNA maintenance), low levels of mtDNA transcripts, and reduced levels of oxidative phosphorylation complexes (with >90% reduction of complex I). Oxygen consumption rate decreased 7.5-fold in basal conditions, and ATP synthesis dropped from 0.25 ± 0.04 to 0.03 ± 0.001 nmol/mg proteins, in the presence of 2-deoxy-d-glucose. Hydrogen peroxide and mitochondrial superoxide anion levels increased by 3- and 1.3-fold, respectively. Mitochondria appeared fragmented, heterogeneous in size and shape, with dilated cristae, vacuoles, and electrondense inclusions. The triterpenoid 2-cyano-3,12-dioxooleana-1,9,-dien-28-oic acid, a Lon inhibitor, partially mimics Lon silencing. In summary, Lon is essential for maintaining mitochondrial shape and function, and for survival of RKO cells.-Gibellini, L., Pinti, M., Boraldi, F., Giorgio, V., Bernardi, P., Bartolomeo, R., Nasi, M., De Biasi, S., Missiroli, S., Carnevale, G., Losi, L., Tesei, A., Pinton, P., Quaglino, D., Cossarizza, A. Silencing of mitochondrial Lon protease deeply impairs mitochondrial proteome and function in colon cancer cells.
2013
- Biological parameters determining the clinical outcome of autologous cultures of limbal stem cells
[Articolo su rivista]
Pellegrini, Graziella; Rama, P; Matuska, S; Lambiase, A; Bonini, S; Pocobelli, A; Colabelli, Rg; Spadea, L; Fasciani, R; Balestrazzi, E; Vinciguerra, P; Rosetta, P; Tortori, A; Nardi, M; Gabbriellini, G; Traverso, Ce; Macaluso, C; Losi, Lorena; Percesepe, Antonio; Venturi, Beatrice; Corradini, Francesca; Panaras, Athanasios; DI ROCCO, Antonio; Guatelli, P; DE LUCA, Michele
abstract
Aim: Limbal cultures restore the corneal epithelium in patients with ocular burns. We investigated the
biological parameters instrumental for their clinical success. Methods: We report a long-term multicenter
prospective study on 152 patients carrying corneal destruction due to severe ocular burns, treated with
autologous limbal cells cultured on fibrin and clinical-grade 3T3-J2 feeder cells. Clinical results were statistically
evaluated both by parametric and nonparametric methods. Results: Clinical outcomes were scored as full
success, partial success and failure in 66.05, 19.14 and 14.81% of eyes, respectively. The total number of
clonogenic cells, colony size, growth rate and presence of conjunctival cells could not predict clinical results.
Instead, the clinical data provided conclusive evidence that graft quality and likelihood of a successful outcome
rely on an accurate evaluation of the number of stem cells detected before transplantation as holoclones
expressing high levels of the p63 transcription factor. No adverse effects related to the feeder layer have
been observed and the regenerated epithelium was completely devoid of any 3T3‑J2 contamination.
Conclusion: Cultures of limbal stem cells can be safely used to successfully treat massive destruction of the
human cornea. We emphasize the importance of a discipline for defining the suitability and the quality of
cultured epithelial grafts, which are relevant to the future clinical use of any cultured cell type.
2012
- Brooke-Spiegler syndrome: report of two cases not associated with a mutation in the CYLD and PTCH tumor-suppressor genes.
[Articolo su rivista]
Ponti, Giovanni; Nasti, S; Losi, Lorena; Pastorino, L; Pollio, A; Benassi, L; Giudice, S; Bertazzoni, G; Veratti, E; Azzoni, Paola; Bianchi Scarrà, G; Seidenari, S.
abstract
Brooke-Spiegler syndrome represents an autosomal dominant disease characterized by the occurrence of multiple cylindromas, trichoepitheliomas and (sporadically) spiroadenomas. Patients with Brooke-Spiegler syndrome are also at risk of developing tumors of the major and minor salivary glands. Patients with Brooke-Spiegler syndrome have various mutations in the CYLD gene, a tumor-suppressor gene located on chromosome 16q. To date, 68 unique CYLD mutations have been identified. We describe two families with Brooke-Spiegler syndrome, one with familial cylindromatosis and one with multiple familial trichoepithelioma, which showed wide inter-family phenotypic variability. Analysis of germline mutations of the CYLD and PTCH genes was performed using peripheral blood. In addition, formalin-fixed paraffin-embedded tumor samples were analyzed for PTCH somatic mutations and cylindroma cell cultures were obtained directly from patients for further growth and analysis. Clinically, the major features of Brooke-Spiegler syndrome include the presence of heterogeneous skin tumors and wide inter- and intra-familial phenotypic variability. Histopathologically, both cylindromas and trichoepitheliomas were found in affected individuals. Mutations or loss of heterozygosity was not found in CYLD and PTCH genes. In CYLD and PTCH mutation-negative patients, other genes may be affected and further studies are needed to clarify whether these patients may be affected by de novo germline mutations.
2012
- p16 immunohistochemistry of multiple primary melanomas as screening to identify Familial Melanoma Syndrome
[Articolo su rivista]
Ponti, Giovanni; G., Luppi; Losi, Lorena; A. M., Cesinaro; Sartori, Giuliana; Maiorana, Antonino; Pellacani, Giovanni; Longo, Caterina; E., Boni; P., Pepe; Giannetti, Alberto; Seidenari, Stefania; M. T., Landi
abstract
The Authors describe the use of p16 immunohistochemistry in multiple primary melanomas as screening to identify Familial Melanoma Syndrome.
2011
- Clinico-pathological and biomolecular findings in Italian patients with multiple cutaneous neurofibromas.
[Articolo su rivista]
Ponti, Giovanni; Losi, Lorena; Martorana, D; Priola, M; Boni, E; Pollio, A; Neri, Tm; Seidenari, S.
abstract
BACKGROUND: Neurofibroma occurs as isolated or multiple lesions frequently associated with neurofibromatosis type 1 (NF1), a common autosomal dominant disorder affecting 1 in 3500 individuals. It is caused by mutations in the NF1 gene, which comprises 60 exons and is located on chromosome 17q11.2. NF1 is a fully penetrant gene exhibiting a mutation rate some 10-fold higher compared with most other disease genes. As a consequence, a high number of cases (up to 50%) are sporadic. Mutation detection is complex due to the large size of the NF1 gene, the presence of pseudogenes and the great variety of lesions.METHODS: 110 patients with at least two neurofibroma lesions recorded in the files of the Pathology Department of the University of Modena during the period 1999-2010, were included in this study. Through interviews and examination of clinical charts, pedigrees were drawn for all patients who were affected by at least two neurofibromas. We attempted to delineate the clinical features of NF1 and the mutational spectrum in the cohort of 11 NF1 families identified. For each proband, the whole coding sequence and all splice sites were studied for mutations, either by the protein truncation test (PTT), or, more frequently, by denaturing high performance liquid chromatography (DHPLC). Two GIST tumors of NF1 patients were tested for somatic NF1 mutations.RESULTS: NF1 germline mutations were identified in 7 (68%) patients. A novel mutation, c.3457_3460delCTCA in exon 20, was detected in two unrelated patients and was associated with different clinical features. No NF1 somatic mutations were detected in the GIST tumors. A wide phenotypic and genotypic variability was registered, both in the spectrum of skin lesions and visceral neoplasms, even among members of the same family who had different clinical manifestations. A proclivity to multiple tumors arising in the same subject, and a higher tumor burden per family were the most relevant findings observed in patients affected with the NF1 mutation.CONCLUSIONS: We report a novel NF1 mutation and we contribute data for the refinement of the NF1 genotype-phenotype spectrum.
2011
- Down-regulation of μ-protocadherin expression is a common event in colorectal carcinogenesis
[Articolo su rivista]
Losi, Lorena; Parenti, Sandra; Fabrizio, Ferrarini; Rivasi, Francesco; Margherita, Gavioli; Gianni, Natalini; Ferrari, Sergio; Grande, Alexis
abstract
We have previously reported that treatment of colorectal cancer cells with mesalazine results in the up-regulated expression of a novel member of the cadherin protein superfamily, named μ-protocadherin, which is able to sequester β-catenin on plasmatic membrane of treated cells inhibiting its proliferation signalling pathway. This finding suggests that μ-protocadherin could exert an oncosuppressive effect on colorectal epithelium. The purpose of our study was to assess whether μ-protocadherin expression is down-regulated during colorectal carcinogenesis. This issue was addressed by analyzing the messenger RNA and protein expression of μ-protocadherin in normal and tumor colorectal cell samples using a combination of quantitative real-time polymerase chain reaction, microarray analysis, and immunohistochemical examination. To better contextualize the role played by μ-protocadherin in the pathogenesis of colorectal cancer, this last assay was also extended to β-catenin, E-cadherin, and Ki-67 proteins. The results obtained evidenced that (1) levels of μ-protocadherin transcript were down-regulated in all the analyzed colorectal cancer samples as compared with normal mucosa; (2) expression of μ-protocadherin protein was completely lost in most analyzed colorectal cancer samples (71%); (3) μ-protocadherin retains β-catenin on the plasmatic membrane of normal colon enterocytes, which implies that β-catenin is released from this site and translocated to the nucleus in colorectal cancer cells. Our data consequently suggest that down-regulation of μ-protocadherin expression is a common event in colorectal carcinogenesis and might therefore play an important role in this pathologic process.
2010
- Cardiac arrhythmias after intramyocardial administration of mesenchymal bone marrow stem cells in acute myocardial infarction
[Abstract in Atti di Convegno]
Mattioli, Anna Vittoria; Farinetti, Alberto; Lonardi, Roberto; Enrico, Giuliani; Sonia, Pennella; Losi, Lorena; Barbieri, Alberto
abstract
...
2010
- Cytogenetic abnormalities and clinical features in a patient cohort affected by three or more synchronous or metachronous primitive malignancies.
[Articolo su rivista]
Ponti, Giovanni; Luppi, G; Giacobbi, F; Corradini, G; Temperani, Paola; Losi, Lorena; Ferrara, L; Pagano, M; Seidenari, Stefania; Tagliafico, Enrico; Torelli, Giuseppe; Conte, Pierfranco
abstract
The multiple cancers (MC) phenotype represents an intriguing entity from both the clinical and the biomolecular points of view. Multiple cancers can arise in a patient either synchronously or metachronously and are frequently detected in hereditary disorders. Here we report the clinical and cytogenetic characterization of 48 patients developing at least three malignancies outside the context of a known genetic condition and 30 control individuals. Medical and pathology reports were registered, blood was collected for cytogenetic studies, and the standard G-banding technique was used for chromosomal analysis of the lymphocyte cultures. Chromosomal analysis of the peripheral blood cultures revealed high cytogenetic instability in 83% of patients' karyotypes that displayed structural rearrangements most often involving chromosomes X, 1, 6, and 7. Peculiar telomeric associations and marker chromosomes were detected in patients with a suspected cancer family history. The MC condition can be observed over a wide clinical range, which includes either apparently sporadic cases or families with a strong history of tumors. These findings indicate that Xq, 6p, and 7q are likely to harbor genes of importance in cancer development, and the present cytogenetic mapping may be crucial for further molecular genetic investigations to recognize a predictive cytogenetic signature useful to detect patients with a high risk of multiple malignancies.
2010
- Gastrointestinal stromal tumor and other primary metachronous and synchronous neoplasms as a suspicious criterion for syndromic setting
[Articolo su rivista]
Ponti, Giovanni; Luppi, G; Martorana, D; Rossi, G; Losi, Lorena; Bertolini, Federica; Sartori, G; Pellacani, Giovanni; Seidenari, Stefania; Boni, E; Neri, Tm; Silini, E; Tamburini, E; Maiorana, Antonino; Conte, Pierfranco
abstract
Gastrointestinal stromal tumors (GISTs) may be sporadic or inherited. Although KIT and PDGFRA activating mutations are the oncogenic mechanisms in most sporadic and inherited GISTs, a small subset of GISTs are negative for both. Besides the classical Familial GIST Syndrome, GIST can occur as part of multi-neoplastic disease. The present study was designed to analyze the synchronous and metachronous tumors developed among GIST patients assessed by our institution for GIST Syndrome setting recognition. Patients (n=141) with primary GIST (77 men and 64 women) were recruited between 1988 and 2007 and their clinical and pathological records were reviewed. Mutation analysis of KIT, PDGFRA, NF1 and MMR genes was performed on somatic and peripheral blood DNA. GISTs occurred associated with other primary malignancies in 46 of 141 (32.6%) patients. The most common neoplasms were gastrointestinal and genitourinary. A novel exon 6 germline large deletion of NF1 was identified in the NF1/GIST kindred. The development of GIST associated with other neoplasms is common and diagnosis of peculiar benign associated-neoplasms warrants the search for familial cancer susceptibility. In particular, syndromic or familial settings have to be suspected in the presence of neurofibroma or lung chordoma in C-KIT and PDGFRA negative GIST patients.
2010
- Leser-Trélat syndrome in patients affected by six multiple metachronous primitive cancers.
[Articolo su rivista]
Ponti, Giovanni; Luppi, G.; Losi, L.; Giannetti, A.; Seidenari, S.
abstract
Leser-Trélat syndrome is characterized by the eruptive appearance of multiple seborrheic keratoses in association with underlying malignant disease. Usually, the sign of Leser-Trélat is associated with adenocarcinoma, most frequently of the colon, breast, or stomach, but also of the lung, kidney, liver, and pancreas. Herein, we present a case that we believe is the first report of the sign of Leser-Trélat in association with occult gastric adenocarcinoma and the anamnestic oncologic history of five other multiple primitive cancers. Epidermal growth factor receptor (EGFR) immunohistochemical expression analysis of multiple seborrheic keratoses revealed an intense membranous staining in the basal keratinocytes and in all the upper epidermal layers. Patients with the sign of Leser-Trélat should undergo a diagnostic screening programme for malignant disease along with patients with known Leser-Trélat syndrome who present with a recent acute and florid eruption of their seborrheic keratoses. We propose the importance of combining the molecular features of multiple seborrheic keratoses with EGFR immunohistochemistry analyses to determine the likelihood of Leser-Trélat syndrome and the consequent high risk of underlying multiple visceral malignancies.
2010
- Mesalazine inhibits the beta-catenin signalling pathway acting through the upregulation of mu-protocadherin gene in colo-rectal cancer cells
[Articolo su rivista]
Parenti, Sandra; Ferrarini, F; Zini, Roberta; Montanari, Monica; Losi, Lorena; Canovi, B; Ferrari, Sergio; Grande, Alexis
abstract
BACKGROUND: Several reports indicate that mesalazine (5-aminosalicylic acid, 5-ASA) is a promising candidate for the chemoprevention of colo-rectal cancer because of its ability to reach the purpose avoiding the unwanted side effects usually associated with prolonged administration of nonsteroidal anti-inflammatory drugs. This activity of 5-ASA is probably the consequence of a number of effects determined on colo-rectal cancer cells, consisting of reduced proliferation, increased apoptosis and activation of cell cycle checkpoints and DNA repair processes. A recent observation has suggested that inhibition of beta-catenin signalling could induce these cellular effects. AIM: To characterize better the capacity of 5-ASA to inhibit the beta-catenin signalling pathway. METHODS: Genes belonging to the beta-catenin signalling pathway were analysed in colo-rectal cancer cell lines treated with 5-ASA using a combination of laboratory assays that are able to detect their phenotypic expression and functional activity. RESULTS: The results obtained indicated that 5-ASA induces the expression of a protein called mu-protocadherin that belongs to the cadherin superfamily and is able to sequester beta-catenin on the plasmatic membrane of treated cells hampering its function. CONCLUSION: These findings suggest that mu-protocadherin might be employed as a biological marker to monitor the chemopreventive efficacy of 5-ASA.
2010
- Selection of patients with germline MLH1 mutated Lynch syndrome by determination of MLH1 methylation and BRAF mutation.
[Articolo su rivista]
Bouzourene, H; Hutter, P; Losi, Lorena; Martin, P; Benhattar, J.
abstract
Lynch syndrome is one of the most common hereditary colorectal cancer (CRC) syndrome and is caused by germline mutations of MLH1, MSH2 and more rarely MSH6, PMS2, MLH3 genes. Whereas the absence of MSH2 protein is predictive of Lynch syndrome, it is not the case for the absence of MLH1 protein. The purpose of this study was to develop a sensitive and cost effective algorithm to select Lynch syndrome cases among patients with MLH1 immunohistochemical silencing. Eleven sporadic CRC and 16 Lynch syndrome cases with MLH1 protein abnormalities were selected. The BRAF c.1799T> A mutation (p.Val600Glu) was analyzed by direct sequencing after PCR amplification of exon 15. Methylation of MLH1 promoter was determined by Methylation-Sensitive Single-Strand Conformation Analysis. In patients with Lynch syndrome, there was no BRAF mutation and only one case showed MLH1 methylation (6%). In sporadic CRC, all cases were MLH1 methylated (100%) and 8 out of 11 cases carried the above BRAF mutation (73%) whereas only 3 cases were BRAF wild type (27%). We propose the following algorithm: (1) no further molecular analysis should be performed for CRC exhibiting MLH1 methylation and BRAF mutation, and these cases should be considered as sporadic CRC; (2) CRC with unmethylated MLH1 and negative for BRAF mutation should be considered as Lynch syndrome; and (3) only a small fraction of CRC with MLH1 promoter methylation but negative for BRAF mutation should be true Lynch syndrome patients. These potentially Lynch syndrome patients should be offered genetic counselling before searching for MLH1 gene mutations.
2009
- Influenza di diversi tipi di eparina sulla infiammazione indotta dalla circolazione extracorporea: studio dei potenziali bioelettrici
[Abstract in Rivista]
Mattioli, Anna Vittoria; Corazzari, Tolmino; U., Carletti; D., Meglioli; Losi, Lorena; Farinetti, Alberto
abstract
...
2009
- New incidence and mortality data. 2003-2005
[Articolo su rivista]
Crocetti E, AIRTUM Working G. r. o. u. p.; Buzzoni C., Collaborators:Serraino D; Vicario, G; Angelin, T; Bessega, G; Bidoli, E; Brunetti, D; de Dottori, M; Forgiarini, O; French, S; Stanta, G; Zaina, L; Zanier, L; Zambon, P; Baracco, M; Bovo, E; Dal Cin, A; Fiore, Ar; Greco, A; Guzzinati, S; Monetti, D; Rosano, A; Stocco, Cf; Tognazzo, S; Egarter Vigl, E; Bellù, F; Vittadello, F; Bulatko, A; Lüthy, M; Facchinelli, G; De Valiere, E; Tschugguel, B; Dorfmann, H; Giacomin, A; Vercellino, Pc; Andreone, S; Ferretti, S; Marzola, L; Migliari, E; Carletti, N; Nenci, I; Vitarelli, S; Antonini, S; Federico, Massimo; Artioli, Me; Cirilli, C; Fracca, A; Rashid, I; Valla, K; De Lisi, V; Sgargi, P; Bozzani, F; Donato, A; Iannelli, A; Mari, C; Senatore, G; Zevola, A; Abbamonte, B; Alfano, Ia; Annunziato, L; Barone, S; Ferrante, A; Budroni, M; Cesaraccio, R; Pirino, D; Sechi, O; Piras, D; Sechi, A; Oggiano, M; Piffer, S; Franchini, S; Gentilini, Ma; Battisti, L; Cappelletti, M; Falcini, F; Amadori, D; Balducci, C; Benericetti, E; Bucchi, L; Caprara, L; Colamartini, A; Cordaro, C; Desiderio, F; Fabbri, C; Foca, F; Giorgetti, S; Montanari, E; Naldi, S; Nannini, R; Ravaioli, A; Ravegnani, M; Rinaldi, E; Salvatore, S; Serafini, M; Vattiato, R; Vitali, B; Pannozzo, F; Busco, S; Natali, M; Ramazzotti, V; Macci, L; Bugliarello, E; Bernazza, E; Tamburo, L; Rossi, M; Curatella, S; Sperduti, I; Fusco, M; Bellatalla, C; Fusco, M; Panico, M; Perrotta, C; Vassante, B; Crosignani, P; Tagliabue, G; Contiero, P; Fabiano, S; Maghini, A; Tittarelli, A; Codazzi, T; Frassoldi, E; Costa, E; Nobile, S; Vigano, C; Berrino, F; Mangone, L; Pezzarossi, A; Pellegri, C; Caroli, S; Valentini, M; Cavuto, S; De Felice, E; Vercelli, M; Orengo, Ma; Casella, C; Marani, E; Puppo, A; Celesia, Mv; Cogno, R; Grondona, Am; Giachero, G; Manenti, S; Quaglia, A; Garrone, E; Paci, E; Crocetti, E; Buzzoni, C; Caldarella, A; Corbinelli, A; Dainelli, G; Guadagni, M; Intrieri, T; Manneschi, G; Miccinesi, G; Nemcova, L; Sacchettini, C; Giusti, F; La Rosa, F; Stracci, F; Petrinelli, Am; Costarelli, D; Cassetti, T; Scheibel, M; Romagnoli, C; Mastrandrea, V; Zanetti, R; Rosso, S; Patriarca, S; Vicari, P; Sobrato, I; Gilardi, F; Maglietta, G; Gallesio, L; Tumino, R; Cilia, S; La Rosa, Mg; Cascone, G; Cianciolo, G; Frasca, G; Giurdanella, Mc; Martorana, C; Morana, G; Nicita, C; Rollo, P; Ruggeri, Mg; Sigona, A; Spata, E; Vacirca, S; Bisanti, L; Autelitano, M; Ghilardi, S; Bovini, A; Giubelli, C; Tessandori, R; Ardemagni, G; Traina, A; Candela, P; Contrino, Ml; Tisano, F; Madeddu, A; Ponz de Leon, M; di Gregorio, C; Roncucci, Luca; Benfatti, P; Losi, Lorena; Ponti, Giovanni; Pedroni, Monica; Rossi, G; Roncari, B; Maffei, S; Menigatti, M; Rossi, F; Pecone, L; Domati, F; Pastore, G; Magnani, C; Terracini, B; Alessi, D; Dal masso, P; Dama, E; Macerata, V; Maule, M; Mosso, Ml; Nonnato, M; Zuccolo, L; Merletti, F; Pannelli, F; Pascucci, C; Gennaro, V; Benfatto, L; Bianchelli, M; Lazzarotto, A; Viarengo, P.
abstract
In Italy cancer incidence and mortality rates are similar to those in northern European countries and in USA among males, but they are still lower than women.
2009
- Ventricular Arrhythmias After Intramyocardial Administration of Mesenchymal Bone Marrow Stem Cells in Acute Myocardial Infarction: An Animal Model
[Abstract in Rivista]
Mattioli, Anna Vittoria; Farinetti, Alberto; Lonardi, Roberto; Losi, Lorena; Barbieri, Alberto; Enrico, Giuliani; Mattioli, Giorgio
abstract
Background. Mesenchymal bone marrow derived cells (MSCs) have been shown to home to injured tissue and produce growth factors that aid angiogenesis. We aim to evaluate ventricular premature contractions (VPCs) and mechanical properties of cardiomyocytes after MSC transplantation using the micro-electrical mechanical systems (MEMS) Technology; these tiny microchips implanted in the heart may be used to monitor heart physiology. Methods. An acute myocardial infarction was induced by ligation of anterior coronary artery in New Zealand rabbits, weighting >3 kg (10 min of ischemia followed by reperfusion). MSCs were isolated, cultured and re-suspended in saline for injection. Injections were done at the peri-infarcted areas, in 5 rabbits we injected MSC whereas in 5 control rabbits we injected saline (to evaluate pro-angiogenic effect of intramyocardial injections i.m.). An arrhythmias recorder was located on the animal and the ECG was recorded for 7 days. Animals were sacrificed and histological and morphological analysis was performed.
2008
- Malignant melanoma in patients with hereditary nonpolyposis colorectal cancer
[Articolo su rivista]
Ponti, Giovanni; Losi, Lorena; Pellacani, Giovanni; L., Wannesson; A. M., Cesinaro; T., Venesio; C., Petti; Seidenari, Stefania
abstract
Background Malignant melanoma (MM) is the most aggressive skin cancer. Most MMs are sporadic, and in this setting an association with mismatch repair (MMR) gene mutations, typical of hereditary nonpolyposis colorectal cancer (HNPCC) tumours, has been proposed. Objectives To characterize clinically and/or by molecular biology the patients with MM belonging to a cohort of 60 kindreds with HNPCC. Methods Patients with HNPCC with a diagnosis of MM were studied by immunohistochemistry (IHC) on tumour tissue using antibodies to MLH1, MSH2, p16, beta-catenin and E-cadherin, and by direct sequencing of MMR genes on germline DNA, and BRAF and NRAS on somatic DNA extracted from MM. Results Nine cutaneous MMs were detected in the tumour spectrum of eight families with HNPCC. The median age at diagnosis was 46 years. In one HNPCC family the diagnosis of MM was made in two first-degree relatives fitting the clinical definition of familial melanoma. IHC and sequencing analysis showed an MSH2 mutation in one patient with MM. Conclusions Dermatological surveillance should be recommended to families in which MM is diagnosed in at least one member, especially at a young age. The combination of MMR gene mutations and abnormalities of p16 or other molecular pathways is needed to induce melanocytic carcinogenesis in a familial setting as well as in sporadic MM.
2008
- Wnt pathway, angiogenetic and hormonal markers in sporadic and familial adenomatous polyposis-associated juvenile nasopharyngeal angiofibromas (JNA)
[Articolo su rivista]
Ponti, Giovanni; Losi, Lorena; Pellacani, Giovanni; G. B., Rossi; Presutti, Livio; Mattioli, Francesco; Villari, Domenico; L., Wannesson; ALICANDRI CIUFELLI, Matteo; P., Izzo; M., De Rosa; P., Marone; Seidenari, Stefania
abstract
Juvenile nasopharyngeal angiofibroma (JNA) is a rare, invasive, and locally destructive tumor of the nasopharynx. The Writ pathway, angiogenctic and hormonal factors are involved in the pathophysiology of JNA; it can result in an extracolonic manifestation of familial adenomatous polyposis (FAP) or in a sporadic tumor. All patients who underwent resection of JNA between 1991 and 2006 at the University of Modena and Reggio Emilia were studied to identify immunohistochemical markers of associated FAP syndrome. Paraffin-embedded JNA samples were analyzed immunohistochemically for the expression of adenomatous polyposis coli (APC), beta-catenin, E-cadherin, androgen receptor, and vascular endothelial growth factors receptor (VEGFR(2)). In one out of the 4 (25%) young patients affected by JNA the diagnosis of FAP syndrome linked to APC mutation was made. All of the sporadic and familial JNA tumors showed nuclear staining of beta-catenin, whereas altered APC expression was seen only in FAP-associated JNA. All cases were stained with VEGFR(2). A combined clinical, immunohistochemical, and biomolecular screening may be useful for the identification of FAP among patients with a diagnosis of JNA. The Writ pathway can be involved in the JNA pathogenesis either by somatic mutations of beta-catenin or by germline APC mutations. As the VEGFR has an important impact on the pathogenesis of JNA, we suggest that a targeted therapy with monoclonal antibodies against VEGFR might lead to a specific chemoprevention and treatment of these tumors and their recurrences.
2007
- A mononucleotide markers panel to identify hMLH1/hMSH2 germline mutations.
[Articolo su rivista]
Pedroni, Monica; Roncari, B; Maffei, S; Losi, Lorena; Scarselli, A; Di Gregorio, C; Marino, M; Roncucci, Luca; Benatti, Piero; Ponti, Giovanni; Rossi, G; Menigatti, M; Viel, A; Genuardi, M; PONZ DE LEON, Maurizio
abstract
Hereditary NonPolyposis Colorectal Cancer (Lynch syndrome) is an autosomal dominant disease caused by germline mutations in a class of genes deputed to maintain genomic integrity during cell replication, mutations result in a generalized genomic instability, particularly evident at microsatellite loci (Microsatellite Instability, MSI). MSI is present in 85-90% of colorectal cancers that occur in Lynch Syndrome. To standardize the molecular diagnosis of MSI, a panel of 5 microsatellite markers was proposed (known as the "Bethesda panel"). Aim of our study is to evaluate if MSI testing with two mononucleotide markers, such as BAT25 and BAT26, was sufficient to identify patients with hMLH1/hMSH2 germline mutations. We tested 105 tumours for MSI using both the Bethesda markers and the two mononucleotide markers BAT25 and BAT26. Moreover, immunohistochemical evaluation of MLH1 and MSH2 proteins was executed on the tumours with at least one unstable microsatellite, whereas germline hMLH1/hMSH2 mutations were searched for all cases showing two or more unstable microsatellites. The Bethesda panel detected more MSI(+) tumors than the mononucleotide panel (49.5% and 28.6%, respectively). However, the mononucleotide panel was more efficient to detect MSI(+) tumours with lack of expression of Mismatch Repair proteins (93% vs 54%). Germline mutations were detected in almost all patients whose tumours showed MSI and no expression of MLH1/MSH2 proteins. No germline mutations were found in patients with MSI(+) tumour defined only through dinucleotide markers. In conclusion, the proposed mononucleotide markers panel seems to have a higher predictive value to identify hMLH1 and hMSH2 mutation-positive patients with Lynch syndrome. Moreover, this panel showed increased specificity, thus improving the cost/effectiveness ratio of the biomolecular analyses.
2007
- BRAF mutations in multiple sebaceous hyperplasias of patients belonging to MYH-asociated polyposis pedigrees
[Articolo su rivista]
Ponti, Giovanni; T., Venesio; Losi, Lorena; Pellacani, Giovanni; L., Bertario; P., Sala; Pedroni, Monica; C., Petti; S., Maffei; L., Varesco; E., Lerch; A., Baggio; Bassoli, Sara; Longo, Caterina; Seidenari, Stefania
abstract
The characteristics of sebaceous gland hyperplasia (SGH) consist of yellowish or skin-colored papules and nodules. Chronic sun exposure and immunosuppressed conditions are the main environmental risk factors, whereas chronological aging regulated by hormones and molecular changes are the intrinsic risk factors. We have evaluated the contribution of BRAF, K-Ras, and N-Ras mutations to the pathogenesis of SGHs in four patients belonging to three MYH-associated polyposis (MAP) pedigrees. MAP is an autosomal-recessive disease characterized by multiple colorectal adenomas and cancer. Immunohistochemistry of mismatch repair and APC proteins was performed. DNA isolated from blood lymphocytes and formalin-fixed or paraffin-em bedded SGHs was PCR amplified and sequenced. In the SGH patients, we detected T1796A heterozygous substitution (V600E) in the BRAF gene. Compound biallelic germline MYH mutations (Y165C/G382D, R168H/379delC, and Y90X/ delGGA464) were detected in the MAP patients. In contrast to the majority of melanocytic lesions, activating hotspot mutations in BRAF have not been involved so far in the pathogenesis of SGH. BRAF mutation is not a specific marker of melanocytic cancerogenesis, and it can also be involved in SGHs. In both melanocytic and non-melanocytic skin tumors, BRAF mutation is linked to early tumorigenesis events.
2007
- Frequency of constitutional MSH6 mutations in a consecutive series of families with clinical suspicion of HNPCC.
[Articolo su rivista]
Roncari, Barbara; Pedroni, Monica; Maffei, S; Di Gregorio, C; Ponti, Giovanni; Scarselli, A; Losi, Lorena; Benatti, Piero; Roncucci, Luca; De Gaetani, C; Camellini, L; Lucci Cordisco, E; Tricarico, E; Genuardi, M; PONZ DE LEON, Maurizio
abstract
A large majority of constitutional mutations in hereditary non-polyposis colorectal cancer (HNPCC) are because of the MHL 1 or MSH 2 genes. In a lower fraction of cases, another gene of the mismatch repair (MMR) machinery, MSH6, may be responsible. Families with MSH6 mutations are difficult to recognize, as microsatellite instability (MSI) may not be detectable and immunohistochemistry (IHC) may give ambiguous results. In the present study, we proposed (i) to determine the frequency of MSH6 mutations in a selected population of colorectal cancer patients obtained from a tumor registry, (ii) to assess whether IHC is a suitable tool for selecting and identifying MSH6 mutation carriers. One hundred neoplasms of the large bowel from suspected HNPCC families were analyzed for MSI (BAT 25 and BAT 26 markers) and immunohistochemical expression of the MSH6 protein. We found on 12 tumors (from different families) showing instability or lack of MSH6 expression. Among these, four potentially pathogenic MSH6 mutations were detected (del A at 2984; del TT at 3119; del AGG cod 385; and del CGT cod 1242) by direct gene sequencing. These represented 12.9% of all families with constitutional mutations of the DNA MMR genes. Thus, some 5% of all HNPCC families are featured by constitutional mutation of the MSH6 gene. This appears, however, as a minimum estimate; routine use of IHC and the study of large numbers of individuals and families with little or no evidence of Lynch syndrome might reveal that mutation of this gene account for a large fraction of HNPCC.
2007
- Genotype-phenotype correlations in individuals with a founder mutation in the MLH1 gene and hereditary non-polyposis colorectal cancer
[Articolo su rivista]
PONZ DE LEON, Maurizio; Benatti, P; Di Gregorio, C; Losi, Lorena; Pedroni, Monica; Ponti, Giovanni; Genuardi, M; Lucci Cordisco, E; Roncucci, Luca
abstract
The results of the study underline the difficulty in discriminating between Lynch I and Lynch II syndromes on the basis of specific molecular changes.
2007
- Loss of Smad4 expression predicts liver metastasis in human colorectal cancer
[Articolo su rivista]
Losi, Lorena; Bouzourene, H; Benhattar, J.
abstract
Distant metastases represent the major cause of death after curative surgery of colorectal cancer. The aim of this study was to evaluate the role of Smad4 and KRAS genetic alterations in colorectal metastases taking into account both the site (hepatic versus extrahepatic) and the time (synchronous versus metachronous) of recurrence. We examined the immunohistochemical expression of Smad4 and frequency of KRAS mutation in primary colorectal tumors and in their corresponding metastatic tissues. Loss of Smad4 expression was noted in 37% (26/71) of the primary tumors and the corresponding metastases. Absence of Smad4 protein was more frequently observed in hepatic metastases, whether they were metachronous or synchronous, than in extrahepatic metastases (p<0.005). The frequency of KRAS mutations was high in the synchronous and extrahepatic metachronous metastases (68-80%), but was significantly lower in the hepatic metachronous metastases (11%). Our results indicate that absence of Smad4 expression correlated significantly with liver metastases regardless of the time of their occurrence and represents a promising new biomarker to predict liver metastasis in colorectal cancer patients. Therefore, this group of patients could benefit from a specific and appropriate pre- and/or post-operative therapy.
2007
- O6-methylguanine-DNA methyltransferase promoter hypermethylation in colorectal carcinogenesis
[Articolo su rivista]
Menigatti, Mirco; Pedroni, Monica; Verrone, Am; Borghi, F; Scarselli, A; Benatti, Piero; Losi, L; Di Gregorio, C; Schär, P; Marra, G; PONZ DE LEON, Maurizio; Roncucci, Luca
abstract
Epigenetic alterations have been reported in colorectal neoplasia which can either complement or in some cases be predisposed to genetic alterations such as K-ras mutations. We examined the promoter methylation status of the CDKN2A and O6-methylguanine-DNA methyltransferase (MGMT) genes, after sodium bisulfite conversion and DNA amplification with methylation specific PCR. Moreover, we searched for G to A transitions in codons 12 and 13 of the K-ras oncogene in normal colorectal mucosae, aberrant crypt foci (ACF, early premalignant lesions) and carcinomas. CDKN2A hypermethylation was an infrequent event in ACF (2 of 26, 7.7%). On the contrary, MGMT hypermethylation was found in the normal mucosae (3 of the 12 samples, 25%), in 14 of the 26 ACF (53.8%) and in 7 of the 9 (77.8%) carcinomas examined. K-ras mutations were evident in 6 ACF (23%) and in 3 carcinomas (33.3%), mostly associated with MGMT promoter hypermethylation. These findings strongly support the hypothesis that epigenetic mechanisms play an important role in the early steps of colorectal carcinogenesis.
2007
- Preoperative Therapy for Lower Rectal Cancer and Modifications in Distance From Anal Sphincter
[Articolo su rivista]
Gavioli, M; Losi, Lorena; Luppi, G; Iachetta, F; Zironi, S; Bertolini, F; Falchi, Am; Bertoni, F; NATALINI G. INT J., RADIAT ONCOL BIOL PHYS
abstract
Purpose:To assess the frequency and magnitude of changes in lower rectal cancer resulting from preoperative therapy and its impact on sphincter-saving surgery. Preoperative therapy can increase the rate of preserving surgery by shrinking the tumor and enhancing its distance from the anal sphincter. However, reliable data concerning these modifications are not yet available in published reports.
Methods and Materials; A total of 98 cases of locally advanced cancer of the lower rectum (90 Stage uT3-T4N0-N+ and 8 uT2N+M0) that had undergone preoperative therapy were studied by endorectal ultrasonography. The maximal size of the tumor and its distance from the anal sphincter were measured in millimeters before and after preoperative therapy. Surgery was performed 6–8 weeks after therapy, and the histopathologic margins were compared with the endorectal ultrasound data.
Results: Of the 90 cases, 82.5% showed tumor downsizing, varying from one-third to two-thirds or more of the original tumor mass. The distance between the tumor and the anal sphincter increased in 60.2% of cases. The median increase was 0.73 cm (range, 0.2–2.5). Downsizing was not always associated with an increase in distance. Preserving surgery was performed in 60.6% of cases. It was possible in nearly 30% of patients in whom the cancer had reached the anal sphincter before the preoperative therapy. The distal margin was tumor free in these cases.
Conclusion: The results of our study have shown that in very low rectal cancer, preoperative therapy causes tumor downsizing in >80% of cases and in more than one-half enhances the distance between the tumor and anal sphincter. These modifications affect the primary surgical options, facilitating or making sphincter-saving surgery possible.
2006
- Immunohistochemical expression of MYH protein can be used to identify patients with MYH-associated polyposis
[Articolo su rivista]
C., Di Gregorio; M., Frattini; S., Maffei; Ponti, Giovanni; Pedroni, Monica; Venesio, T; Bertario, L; Varesco, L; Risio, M; PONZ DE LEON, Maurizio; Losi, Lorena
abstract
BACKGROUND & AIMS: MYH-associated polyposis is a recently described, autosomal-recessive disease characterized by multiple colorectal adenomas and cancer. There are only few immunohistochemical studies of the MYH protein. We investigated the expression pattern of the MYH protein to evaluate whether a immunohistochemical approach could be used in clinical practice to screen patients for germline mutations in the MYH gene.METHODS: The expression of MYH, MSH2, MLH1, and MSH6 proteins was studied by immunohistochemistry in 20 samples (colorectal adenomas or cancer) from 18 patients with biallelic MYH mutation, in 11 samples from patients with germline adenomatous polyposis coli (APC) mutations, in 20 samples from patients with sporadic colorectal cancers, and in 10 samples from patients with normal colonic mucosa without malignancies.RESULTS: In all cases the mismatch repair proteins were expressed normally. Nuclear and cytoplasmic immunoreactivity for the MYH protein were observed in normal colorectal mucosa, in sporadic colorectal carcinomas, and in adenomas and carcinomas from patients carrying APC germline mutations. Adenomas and carcinomas from patients with MYH biallelic mutation showed a different pattern of expression: a strong granular cytoplasmic staining was observed without any nuclear expression. The same immunophenotype was observed in the surrounding normal mucosa.CONCLUSIONS: Patients with biallelic MYH mutations showed disappearance of staining from the nucleus, and segregation of immunoreactivity in the cytoplasm, both in neoplastic and surrounding healthy mucosa. Because this pattern of expression seems to be specific for biallelic mutations, it follows that immunohistochemistry might be used in clinical practice to screen patients at risk for MYH-associated polyposis.
2006
- Low rectal cancer and distance from anal sphincter before and after preoperative therapy
[Abstract in Rivista]
M., Gavioli; Losi, Lorena; Iachetta, Francesco; G., Luppi; A., Schiavone; Mauro, Addolorata; M. C., Gallo; A., Rinaldi; G., Natalini
abstract
Preoperative therapy increased the distance of the low rectal cancer from the anal sphincter until the near 50% of cases and can make possible or facilitate an anal preserving operation.
2006
- Prognostic significance of histological features and biological parameters in stage I (pT1 and pT2) colorectal adenocarcinoma
[Articolo su rivista]
Losi, Lorena; Ponti, Giovanni; Di Gregorio, Carmela; Marino, M; Rossi, Giuseppina; Pedroni, Monica; Benatti, Piero; Roncucci, Luca; Ponz De Leon, Maurizio
abstract
Patients with stage I colorectal cancer have a good prognosis, however, a small fraction of them die of local or distant recurrence after curative resection. The aggressive behavior reflects some biological properties of these tumors. In this study, we evaluated the prognostic role of some histopathological and biological parameters in stage I colorectal carcinomas. From the Colorectal Cancer Registry of Modena, we selected two series of patients; the first included all patients who had died of disease progression, the second included patients with a favorable outcome. The histopathological parameters assessed were grade of differentiation, growth pattern at the invasive tumor front, peritumoral lymphocytic infiltration, tumor budding and vascular invasion. The biological variables were proliferative activity (using Ki-67 nuclear antigen), overexpression of p53 protein and altered expression of the mismatch repair proteins (MLH1 and MSH2). The results showed that an infiltrating growth pattern, absent or sparse peritumoral lymphocytic infiltration, the presence of tumor budding and vascular invasion are significantly related to the risk of recurrence. Among the biological parameters, p53 overexpression was significantly correlated with a poor clinical outcome. Our study showed that the histopathologial features are relevant prognostic indicators and might be used as markers for an appropriate treatment strategy in patients with stage I carcinomas.
2006
- Prognostic value of Dworak grade of regression (GR) in patients with rectal carcinoma treated with preoperative radiochemotherapy
[Articolo su rivista]
Losi, Lorena; G., Luppi; M., Gavioli; Iachetta, Francesco; Bertolini, Federica; D'Amico, Roberto; Jovic, Gordana; F., Bertoni; Falchi, Anna Maria; Conte, Pierfranco
abstract
Preoperative radiochemotherapy improves local control in locally advanced rectal cancer; however, its role in prolonging survival is still controversial. In order to better define the subset in patients who might benefit from this multimodal treatment, we have evaluated the correlation between grade of regression (GR) to preoperative treatment and disease-free survival (DFS). METHODS: We reviewed retrospectively the surgical specimens of 106 patients with locally advanced T3/T4 N0/ M0 rectal cancer. All patients were treated preoperatively with radiotherapy and 5-fluorouracil-based regimen chemotherapy. We evaluated ypTNM stage, and tumor regression was graded using the Dworak system that varies from GR 0 (absence of regression) to GR 4 (complete regression). RESULTS: GR was as follows: GR 4, 16 patients (15%); GR 3, 25 patients (23.6%), GR 2, 30 patients (28.4%), GR 1, 32 patients (30.2%) and GR 0, 3 patients (2.8%). A significant correlation was found between GR and DFS. Three-year DFS was 100, 85, 82, 66 and 33% in GR 4, 3, 2, 1 and 0, respectively (p=0.01). DFS was significantly lower in patients with advanced stages at diagnosis and in patients without down-staging. Moreover, in postoperative stage II and III cases, GR 3 correlated with a better DFS than GR 2-0 (p=0.2 and p=0.4, respectively). CONCLUSIONS: The GR was a significant prognostic factor in locally advanced rectal carcinoma treated with preoperative chemoradiotherapy. The pathological stage and down-staging also have prognostic value. The use of a standardized system to evaluate GR in rectal cancer can allow for comparisons between different institutions and can identify patients at worse prognosis to be treated with adjuvant therapy
2006
- Subcellular localization of beta-catenin and APC proteins in colorectal preneoplastic and neoplastic lesions RID B-2583-2012
[Articolo su rivista]
Sena, Paola; Saviano, Massimo; Monni, Sebastiano Graziano; Losi, Lorena; Roncucci, Luca; Marzona, Laura; DE POL, Anto
abstract
Adenomatous polyposis coli (APC) is a tumor suppressor gene whose main function is the destabilization of beta-catenin, a key effector of the Wnt signaling pathway. This gene is defective in familial adenomatous polyposis (FAP), a dominantly inherited disease, but inactivation of APC has been reported also in most sporadic colorectal tumors and it is considered an early event in colorectal tumorigenesis. The aim of the present study was to evaluate the intracellular ultrastructural distribution of beta-catenin and APC proteins in epithelial cells of normal colorectal mucosa, aberrant crypt foci (ACF, an early premalignant lesion) and cancer. We used the immunogold electron microscopic method to identify both proteins. Normal colonic epithelial cells showed a strong membranous expression of beta-catenin and lacked cytoplasmic and nuclear expression. Normal cells showed APC localization pattern characterized by diffuse nuclear expression and along the plasma membrane. In ACF and in carcinoma an absent or reduced membranous expression of beta-catenin was associated with an increased nuclear and cytoplasmatic expression. In aberrant crypt foci and carcinoma, APC was evident inside the nucleus and at the level of cell-cell junctions, but it was decreased in the cytoplasm. This method allowed the accurate localization of proteins of the Writ signaling pathway in the early steps of colorectal carcinogenesis. The similar pattern of subcellular distribution of APC and beta-catenin in dysplastic ACF and colorectal cancer suggests that ACF are precursor lesions of sporadic and FAP-associated colorectal carcinoma. (c) 2005 Elsevier Ireland Ltd. All rights reserved.
2006
- Value of MLH1 and MSH2 mutations in the appearance of Muir-Torre syndrome phenotype in HNPCC patients presenting sebaceous gland tumors or Keratoacanthomas
[Articolo su rivista]
Ponti, Giovanni; Losi, Lorena; Pedroni, Monica; E., Lucci Cordisco; C., Di Gregorio; Pellacani, Giovanni; Seidenari, Stefania
abstract
Hereditary non-polyposis colorectal cancer (HNPCC) is an autosomal-dominant disorder characterized by predisposition to colorectal cancer and extracolonic malignancies, frequent multiple primary tumors in the same patient, and early age of cancer onset. A main clinical variant of Lynch syndrome, Muir-Torre syndrome (MTS) is characterized by the association between one or more visceral malignancies, with at least one sebaceous skin tumor or keratoacanthoma. In our study, we have screened a cohort of 538 HNPCC patients, related to 57 HNPCC families, to detect sebaceous skin tumors and keratoacanthomas and the role of mismatch repair (MMR) genes, MLH1, MSH2, and MSH6, in their pathogenesis. Among the 57 HNPCC families, we have identified four MTS families and one suspected MTS family, in which sebaceous carcinoma was found in one HNPCC mutation carrier subject who did not show visceral malignancy. In four of these families, linked to two MLH1 mutations and to two MSH2 mutations, biomolecular characterization showed concordance among immunohistochemistry analysis and gene mutations. The evidences of our investigations show that MLH1 and MSH2 gene mutations have an equivalent etiopathological role both for Lynch syndrome and for MTS; hence, we propose a broadened clinical criteria for definition of Lynch syndrome that will include sebaceous adenoma, carcinoma, and keratoacanthoma.
2005
- Attenuated familial adenomatous polyposis and Muir-Torre syndrome linked to compound biallelic constitutional MYH gene mutations.
[Articolo su rivista]
Ponti, Giovanni; PONZ DE LEON, Maurizio; Maffei, S; Pedroni, Monica; Losi, L; Di Gregorio, C; Gismondi, V; Scarselli, A; Benatti, Piero; Roncari, B; Seidenari, S; Pellacani, Giovanni; Varotti, C; Prete, E; Varesco, L; Roncucci, Luca
abstract
Attenuated familial adenomatous polyposis and Muir-Torre syndrome linked to compound biallelic constitutional MYH gene mutations.Peculiar dermatologic manifestations are present in several heritable gastrointestinal disorders. Muir-Torre syndrome (MTS) is a genodermatosis whose peculiar feature is the presence of sebaceous gland tumors associated with visceral malignancies. We describe one patient in whom multiple sebaceous gland tumors were associated with early onset colon and thyroid cancers and attenuated polyposis coli. Her family history was positive for colonic adenomas. She had a daughter presenting with yellow papules in the forehead region developed in the late infancy. Skin and visceral neoplasms were tested for microsatellite instability and immunohistochemical status of mismatch repair (MMR), APC and MYH proteins. The proband colon and skin tumors were microsatellite stable and showed normal expression of MMR proteins. Cytoplasmic expression of MYH protein was revealed in colonic cancer cells. Compound heterozygosity due to biallelic mutations in MYH, R168H and 379delC, was identified in the proband. The 11-year-old daughter was carrier of the monoallelic constitutional mutation 379delC in the MYH gene; in the sister, the R168H MYH gene mutation was detected. This report presents an interesting case of association between MYH-associated polyposis and sebaceous gland tumors. These findings suggest that patients with MTS phenotype that include colonic polyposis should be screened for MYH gene mutations.
2005
- Different phenotypes in Muir-Torre Syndrome: clinical and biomolecular characterization in two italian families
[Articolo su rivista]
Ponti, Giovanni; PONZ DE LEON, Maurizio; Losi, Lorena; C., Di Gregorio; Benatti, Piero; Pedroni, Monica; A., Scarselli; G., Riegler; L., Lembo; Pellacani, Giovanni; Seidenari, Stefania; Rossi, Giorgio; Roncucci, Luca
abstract
The Muir-Torre syndrome (MTS) is an autosomal dominant genodermatosis characterized by the presence of sebaceous gland tumours, with or without keratoacanthomas, associated with visceral malignancies. We describe and characterize two families in which the ample phenotypic variability of MTS was evident. After clinical evaluation, the skin and visceral tumours of one member of a family with 'classic' MTS and one member of a family with a 'peculiar' MTS phenotype without sebaceous lesions, but with only multiple keratoacanthomas, were analysed for microsatellite instability (MSI) and by immunohistochemistry. Tumours of both individuals showed MSI, with a concomitant lack of MSH2 immunostaining in all evaluated skin and visceral lesions; moreover, in the proband of family 2 a constitutional mutation (C -> T substitution leading to a stop codon) in the MSH2 gene was identified. We conclude that the diagnosis of MTS, which is mainly clinical, should take into account an ample phenotypic variability, which includes both cases with typical cancer aggregation in families and cases characterized by the association of visceral malignancies with multiple keratoacanthomas (without sebaceous lesions), without an apparent family history of cancer.
2005
- Evolution of intratumoral genetic heterogeneity during colorectal cancer progression
[Articolo su rivista]
Losi, Lorena; B., Baisse; H., Bouzourene; J., Benhattar
abstract
Evolution of intratumoral genetic heterogeneity during colorectal tumor progression has not been investigated so far. Multiple sample areas in colorectal adenocarcinoma at early and advanced stages and in metastases were studied for the well-known genetic alterations: K-ras and p53 point mutations and loss of heterozygosity (LOH) on chromosomes 5q and 18q. In primary colorectal cancers (CRCs), intratumoral genetic heterogeneity was more often observed in early than in advanced stages, at 90 and 67%, respectively. All but one of the advanced CRCs were composed of one predominant clone and other minor clones, whereas no predominant clone has been identified in half of the early cancers. At the early stage, the last events that were produced, the p53 mutation and LOH of 18q, were also the most heterogeneous. At the advanced stage, the LOH of 5q and 18q were the most frequent heterogeneous events (67 and 58%, respectively). The intratumoral heterogeneity for mutations was significantly reduced, from the early to the advanced stages (from 60 to 20% for K-ras and from 70 to 20% for p53). On the other hand, a quasi absence of intratumoral genetic heterogeneity was observed for K-ras and p53 in distant metastasis. In conclusion, colorectal adenocarcinomas are characterized by marked intratumoral genetic heterogeneity. A reduction of the intratumoral genetic heterogeneity for point mutations and a relative stability of the heterogeneity for allelic losses indicate that, during the progression of CRC, clonal selection and chromosome instability continue, while an increase cannot be proven.
2005
- Identification of Muir-Torre Syndrome among patients with sebaceous tumors and keratoacanthomas: role of clinical features, microsatellite instability and immunohistochemistry
[Articolo su rivista]
Ponti, Giovanni; Losi, Lorena; Di Gregorio, C; Roncucci, Luca; Pedroni, Monica; Scarselli, A; Benatti, Piero; Seidenari, S; Pellacani, Giovanni; Lembo, L; Rossi, G; Marino, M; Lucci Cordisco, E; PONZ DE LEON, Maurizio
abstract
BACKGROUND: The Muir-Torre syndrome (MTS) is an autosomal-dominant genodermatosis characterized by the presence of sebaceous gland tumors, with or without keratoacanthomas, associated with visceral malignancies. A subset of patients with MTS is considered a variant of the hereditary nonpolyposis colorectal carcinoma, which is caused by mutations in mismatch-repair genes. The objective of the current study was to evaluate whether a combined clinical, immunohistochemical, and biomolecular approach could be useful for the identification of Muir-Torre syndrome among patients with a diagnosis of sebaceous tumors and keratoacanthomas.
METHODS: The authors collected sebaceous skin lesions and keratoacanthomas recorded in the files of the Pathology Department of the University of Modena during the period 1986-2000. Through interviews and examination of clinical charts, family trees were drawn for 120 patients who were affected by these skin lesions.
RESULTS: Seven patients also were affected by gastrointestinal tumors, thus meeting the clinical criteria for the diagnosis of MTS. In the MTS families, a wide phenotypic variability was evident, both in the spectrum of visceral tumors and in the type of skin lesions. Microsatellite instability was found in five MTS patients: These patients showed concordance with immunohistochemical analysis; moreover, a constitutional mutation in the MSH2 gene was found in 1 patient. Lack of expression of MSH2/MSH6 or MLH1 proteins was evident in the skin lesions and in the associated internal malignancies of 3 patients and 2 patients with MTS, respectively.
CONCLUSIONS: The clinical, biomolecular, and immunohistochemical characterization of sebaceous skin lesions and keratoacanthomas may be used as screening for the identification of families at risk of MTS, a disease that is difficult to recognize and diagnose.
2005
- Incidence and clinical impact of sterilized disease and minimal residual disease after preoperative radiochemotherapy for rectal cancer
[Articolo su rivista]
Gavioli, M; Luppi, G; Losi, Lorena; Bertolini, Federica; Santantonio, M; Falchi, Anna Maria; D'Amico, Roberto; Conte, Pierfranco; Natalini, G.
abstract
In advanced rectal cancer, chemoradiation can induce downstaging until complete disappearance of the tumor or its persistence in minimal form. The complete sterilized and the minimal residual disease often are considered similar. We evaluated the specific incidence of these two conditions and analyzed their impact in terms of local recurrence, distant metastasis, and survival. METHODS: We studied 139 uT3/T4 N0/N+ rectal cancers, treated with preoperative chemoradiation and curative surgery after six to eight weeks. We evaluated ypTNM stage and tumoral regression, according to the five degrees proposed by Dworak, with special attention to 4 and 3 (sterilized and minimal residual disease). RESULTS: Tumor downstaging occurred in 65 patients (46.7 percent), including 25 sterilized lesions (17.9 percent) and 24 minimal residual disease (17.2 percent). In median follow-up of 30 months, none of the patients with sterilized disease developed local or distant recurrence. Among patients with minimal residual disease, none developed local recurrence, whereas two (8.3 percent) developed distant metastasis, and one died from disease. In patients with gross residual disease (Grade 2, 1, 0) the percentage of local recurrence was 8.8 percent, distant recurrence 26.6 percent, and 13.3 percent died from disease. The difference between three groups is statistically significant as regards local and distant recurrence. CONCLUSIONS: After preoperative therapy, the sterilized disease shows an excellent prognosis. The minimal residual disease has an important numeric incidence. Its outcome is different, with a not-negligible risk of distant recurrence. The minimal residual disease has a much better prognosis in comparison with the gross residual disease
2005
- Incidence and survival of patients with Dukes' A (stages T1 and T2) colorectal carcinoma: a 15-year population-based study.
[Articolo su rivista]
Benatti, Piero; Roncucci, Luca; Rossi, G; Ponti, Giovanni; Marino, M; Pedroni, Monica; Scarselli, A; Roncari, Barbara; PONZ DE LEON, Maurizio; Di Gregorio, C; Losi, Lorena
abstract
BACKGROUND AND AIMS: Patients with stage I (Dukes' A) colorectal carcinoma tend to show a good prognosis; however, recurrences can be observed in some patients. Through a specialized colorectal cancer Registry, we attempted to investigate the epidemiological and clinical features of individuals with Dukes' A neoplasms.PATIENTS AND METHODS: From 1984 to 1998, 295 individuals were diagnosed with Stage I /Dukes' A tumors; 150 of these had lesions infiltrating the muscular wall (T2), while 145 had neoplasms limited to the submucosa (T1).RESULTS: Dukes' A tumors represented 13.8% of all registered neoplasms; the percentage doubled over the study period (8.1% in the first year vs. 16.8% in the final year). In each year of observation, the preferential locations were the rectum and sigmoid colon (75% of all lesions). Most patients required surgery, but only 21.3% could be managed by endoscopic polypectomy. Overall 5-year survival was 81.0% (82.1% in T1, 80.0% in T2). Recurrences were seen in 6.8% (2.8% in T1, 10.7% in T2), while 36 patients (12.2%) died of causes unrelated to colorectal cancer. In 17 out of 20 patients who died of cancer, the lesions were localized in the rectosigmoid region. Survival analysis showed a significantly better prognosis (P<0.007) for patients with T1 tumors.CONCLUSIONS: The proportion of stage I colorectal tumors tended to increase over time. Although the overall prognosis is good in four-fifths of the cases, approximately one-fifth of these patients die of recurrent disease or of other causes. As expected, the prognosis was significantly more favorable for patients with T1 lesions. For patients with T2 tumors, radical surgery is the most appropriate approach.
2005
- Microsatellite instability and colorectal cancer prognosis.
[Articolo su rivista]
Benatti, Piero; Gafà, R; Barana, D; Marino, M; Scarselli, A; Pedroni, Monica; Maestri, I; Guerzoni, L; Roncucci, Luca; Menigatti, M; Roncari, B; Maffei, S; Rossi, G; Ponti, Giovanni; Santini, A; Losi, Lorena; Di Gregorio, C; Oliani, C; PONZ DE LEON, Maurizio; Lanza, G.
abstract
PURPOSE: Many studies have evaluated the role of high levels of microsatellite instability (MSI) as a prognostic marker and predictor of the response to chemotherapy in colorectal cancer (CRC); however, the results are not conclusive. The aim of this study was to analyze the prognostic significance of high levels of MSI (MSI-H) in CRC patients in relation to fluorouracil-based chemotherapy.EXPERIMENTAL DESIGN: In three different institutions, 1,263 patients with CRC were tested for the presence of MSI, and CRC-specific survival was then analyzed in relation to MSI status, chemotherapy, and other clinical and pathologic variables.RESULTS: Two hundred and fifty-six tumors were MSI-H (20.3%): these were more frequently at a less advanced stage, right-sided, poorly differentiated, with mucinous phenotype, and expansive growth pattern than microsatellite stable carcinomas. Univariate and multivariate analyses of 5-year-specific survival revealed stage, tumor location, grade of differentiation, MSI, gender, and age as significant prognostic factors. The prognostic advantage of MSI tumors was particularly evident in stages II and III in which chemotherapy did not significantly affect the survival of MSI-H patients. Finally, we analyzed survival in MSI-H patients in relation to the presence of mismatch repair gene mutations. MSI-H patients with hereditary non-polyposis colorectal cancer showed a better prognosis as compared with sporadic MSI-H; however, in multivariate analysis, this difference disappeared.CONCLUSIONS: The type of genomic instability could influence the prognosis of CRC, in particular in stages II and III. Fluorouracil-based chemotherapy does not seem to improve survival among MSI-H patients. The survival benefit for patients with hereditary non-polyposis colorectal cancer is mainly determined by younger age and less advanced stage as compared with sporadic MSI-H counterpart.
2005
- Molecular genetic alterations and clinical features in early-onset colorectal carcinomas and their role for the recognition of hereditary cancer syndromes.
[Articolo su rivista]
Losi, Lorena; Di Gregorio, C; Pedroni, Monica; Ponti, Giovanni; Roncucci, Luca; Scarselli, A; Genuardi, M; Baglioni, S; Marino, M; Rossi, G; Benatti, Piero; Maffei, S; Menigatti, M; Roncari, Barbara; PONZ DE LEON, Maurizio
abstract
OBJECTIVES: Colorectal cancer (CRC) occurs rarely in young individuals (<45 yr) and represents one of the criteria for suspecting hereditary cancer families. In this study we evaluated clinical features and molecular pathways (chromosomal instability [CIN] and microsatellite instability [MSI]) in early-onset CRC of 71 patients.METHODS: Detailed family and personal history were obtained for each patient. Expression of APC, beta-catenin, p53, MLH1, MSH2, and MSH6 genes was evaluated by immunohistochemistry. MSI analysis was performed and constitutional main mutations of the mismatch repair (MMR) genes were searched by gene sequencing.RESULTS: Fourteen (19.7%) out of the 71 cases showed both MSI and altered expression of MMR proteins. In the 57 MSI-negative (MSI-) lesions altered expression of APC, beta-catenin, and p53 genes were found more frequently than in MSI-positive(MSI+) tumors. Seven (50%) out of the 14 patients with MSI+ tumors presented clinical features of Lynch syndrome (hereditary non-polyposis colorectal cancer [HNPCC]) and in all but one, constitutional mutations in MLH1 or MSH2 genes could be detected. The same mutations were also found in other family members.CONCLUSIONS: Our study demonstrates the involvement of CIN in a majority of early-onset colorectal tumors. Furthermore, we identified Lynch syndromes in seven cases (50%) of early-onset colorectal carcinomas with impairment of the MMR system. These results suggest that patients with early-onset CRC should be screened for hereditary cancer syndrome through clinical and molecular characterizations.
2005
- Prognostic value of "Dworak" tumor regression grade (TRG) after pre-operative chemo-radiotherapy in rectal cancer (RC)
[Abstract in Rivista]
Bertolini, Federica; Losi, Lorena; Iachetta, Francesco; Giacobazzi, Patrizia; E., Romagnani; Dealis, Cristina; F., Bertoni; G., Luppi; C., Bengala; Conte, Pierfranco
abstract
Our study confirms the prognostic value of pathological stage and Dworak TRG after pre-operative treatment. Moreover in post- operative stage I II-III our analysis has demonstrated a trend in disease free survival improvement for patients with a high Dworak TRG.
2004
- Aetiology of colorectal cancer and relevance of monogenic inheritance.
[Articolo su rivista]
PONZ DE LEON, Maurizio; Benatti, Piero; Borghi, Francesca; Pedroni, Monica; Scarselli, Alessandra; DI GREGORIO, C; Losi, Lorena; Viel, A; Genuardi, M; Abbati, G; Rossi, Giuseppina; Menigatti, Mirco; Lamberti, Igor; Ponti, Giovanni; Roncucci, Luca
abstract
BACKGROUND AND AIMS: Although diet and lifestyle are associated with the development of colorectal malignancies, the only clearly identified aetiological factors in colorectal cancer are inheritance (hereditary non-polyposis colorectal cancer (HNPCC) and familial polyposis), inflammatory bowel diseases, papillomavirus, and acquired immunodeficiency syndrome (AIDS). Our aim was to determine what proportion of colorectal neoplasms could be attributed to these specific factors.PATIENTS AND METHODS: Data from a colorectal cancer registry were analysed over a 15 year period, during which nearly 2500 cases were recorded. In patients with suspected HNPCC, microsatellite instability and immunohistochemical expression of proteins encoded by the main DNA mismatch repair genes were assessed. In families with unstable neoplasms, constitutional mutations of the mismatch repair genes hMSH2, hMLH1, and hMSH6 were evaluated by single strand conformation polymorphism analysis and sequencing.RESULTS: Inflammatory bowel diseases, familial polyposis, and AIDS were rare causes of colorectal cancer (three, three, and one case, respectively). Anal squamous carcinoma developed in 27 patients (1.0%) and could be attributed to papillomavirus infection. In 58 patients (from 34 families) a clinical diagnosis of HNPCC was established (2.4%). In total, cases with a known aetiology were 92 (3.7% of all patients). Microsatellite instability was detected in 15 cancers from HNPCC families, and germline mutations in six families (12 patients, 0.5% of the total). Families with unstable tumours, with or without mutations, were clinically similar, suggesting the involvement of the mismatch repair system even when mutations were not detected.CONCLUSIONS: The study suggests that the aetiology of colorectal malignancies remains elusive in the large majority of cases. Among specific causes, HNPCC represents the most frequent. However, with a population based approach, constitutional mutations of the main genes involved in HNPCC can be detected in only 20% of cases.
2004
- Assessment of K-ras, Smad4 and p53 gene alterations in colorectal metastases and their role in the metastatic process
[Articolo su rivista]
Losi, Lorena; G., Luppi; J., Benhattar
abstract
To date there is no genetic marker that gives accurate information on the prognostic impact for patients with colorectal cancer. A particular clone, not detected in the tumor, could be responsible for the metastatic process. To overcome this problem, genetic alterations were analyzed in metastatic tissues from 58 patients who developed metastases after curative surgery for colorectal cancer. K-ras, p53 and Smad4 alterations were observed in respectively 38, 60 and 27% of the metastases. These frequencies are similar to the ones reported in primary colorectal tumors. Thus, these genetic alterations cannot be used as prognostic biomarkers in patients with colorectal cancer. The metastases were stratified into 3 groups, according to the metastatic localization. K-ras mutations were detected in respectively 75, 26 and 11% of the distant, peritoneal and liver metastases. Loss of Smad4 expression was observed more frequently in the liver (62%) than in other metastases (13%). These results suggest that the genetic changes of the tumor cells indicate the location of the metastases and thus, the route of metastatic spread.
2004
- CDX-2 homeobox gene and MUC2 expression identifies true mucinous (so-called colloid) carcinoma of the lung and justifies its indolent outcome
[Abstract in Rivista]
L., Longo; Marchioni, Alessandro; B., Murer; A., Cavazza; Losi, Lorena; Natali, Pamela; M., Bavieri; Migaldi, Mario; G., Rossi
abstract
CDX-2/MUC2 coordinated expression may be helpful in distinguishing mucinous carcinoma from mucinous bronchioloalveolar carcinoma, a lung cancer frequently associated with recurrences and multicentricity.
2004
- Diagnosis of hereditary non-polyposis colorectal cancer (HNPCC) - Reply
[Articolo su rivista]
Roncucci, Luca; PONZ DE LEON, Maurizio; Benatti, Piero; Borghi, F; Pedroni, Monica; Scarselli, A; di Gregorio, C; Losi, Lorena; Viel, A; Genuardi, M; Abbati, G; Rossi, G; Menigatti, M; Ponti, Giovanni
abstract
NOTHING
2004
- Diagnosis of hereditary non-polyposis colorectal cancer (HNPCC) [2] (multiple letters)
[Articolo su rivista]
Jass, J. R; Roncucci, Luca; PONZ DE LEON, Maurizio; Benatti, Piero; Borghi, F.; Pedroni, Monica; Scarselli, A.; DI GREGORIO, Carmela; Losi, Lorena; Viel, A.; Genuardi, M.; Abbati, Gian Luca; Rossi, G.; Menigatti, Mirco; Ponti, Giovanni
abstract
Letter to the Editor
2004
- Genetic testing among high-risk individuals in families with hereditary non polyposis colorectal cancer
[Articolo su rivista]
PONZ DE LEON, Maurizio; Benatti, P.; Di Gregorio, C.; Pedroni, Monica; Losi, L.; Genuardi, M.; Viel, A.; Fornasarig, M.; Lucci Cordisco, E.; Anti, M.; Ponti, Giovanni; Borghi, F.; Lamberti, I.; Roncucci, Luca
abstract
Hereditary nonpolyposis colorectal cancer (HNPCC) is frequently associated with constitutional mutations in a class of genes involved in DNA mismatch repair. We identified 32 kindreds, with germline mutations in one of three genes hMSH2, hMLH1 or hMSH6. In this study, we purposed to evaluate how many high-risk individuals in each family underwent genetic testing: moreover, we assessed how many mutation-positive unaffected individuals accepted colonoscopic surveillance and the main findings of the recommended follow-up. Families were identified through a population-based registry, or referred from other centres. Members of the families were invited for an education session with two members of the staff. When a kindred was consistent with HNPCC, neoplastic tissues were examined for microsatellite instability (MSI) and immunohistochemical expression of MSH2, MLH1 and MSH6 proteins. Moreover, constitutional mutations were searched by SSCP or direct sequencing of the whole genomic region. Of the 164 subjects assessed by genetic testing, 89 were gene carriers (66 affected - that is, with HNPCC-related cancer diagnosis - and 23 unaffected) and 75 tested negative. Among the 23 unaffected gene carriers, 18 (78.3%) underwent colonoscopy and four declined. On a total of 292 first degree at risk of cancer, 194 (66.4%) did not undergo genetic testing. The main reasons for this were: (a) difficulty to reach family members at risk, (b) lack of collaboration, (c) lack of interest in preventive medicine or 'fatalistic' attitude towards cancer occurrence. The number of colorectal lesions detected at endoscopy in gene carriers was significantly (P<0.01) higher than in controls (noncarriers). We conclude that a large fraction of high-risk individuals in mutation-positive HNPCC families does not undergo genetic testing, despite the benefits of molecular screening and endoscopic surveillance. This clearly indicates that there are still barriers to genetic testing in HNPCC, and that we are unable to provide adequate protection against cancer development in these families.
2004
- Identification of Muir-Torre syndrome among patients with sebaceous tumors and keratoacantomas: role of clinical features, microsatellite instability and immunohistochemistry
[Abstract in Rivista]
Losi, Lorena; Ponti, Giovanni; Scarselli, A.; Roncucci, Luca; Pedroni, Monica; P., Benatti; PONZ DE LEON, Maurizio; C., Di Gregorio
abstract
The clinical, biomolecular and immunohistochemical characterization of sebaceous skin lesions and keratoakantomas migh be used in population-based screening for the identification of families at risk of Muir-Torre syndrome, a rare disease difficult to recognize and diagnose.
2004
- Primary mucinous (so-called colloid) carcinomas of the lung - A clinicopathologic and immunohistochemical study with special reference to CDX-2 homeobox gene and MUC2 expression
[Articolo su rivista]
Rossi, G; Murer, B; Cavazza, A; Losi, Lorena; Natali, P; Marchioni, A; Migaldi, Mario; Capitanio, G; Brambilla, E.
abstract
Herein we describe the clinicopathologic and immunohistochemical features of 13 primary mucinous (colloid) carcinomas (MCs) of the lung, an uncommon and controversial tumor. The patients, 7 males and 6 females, ranged in age from 50 to 79 years (mean, 64.5 years). All the tumors presented as a peripheral solitary nodule with gelatinous cut-surface and well circumscribed but lacking a complete fibrous wall. The size ranged from 1 to 5.5 cm. Microscopically, they consisted of neoplastic elements floating in large mucin pools and focally lining the alveolar spaces. Eleven cases were predominantly composed of tall, columnar goblet cells (goblet cell-type MC), while 2 consisted of signet-ring tumor cells (signet-ring cell-type MC). Five tumors were incidentally discovered by chest radiographs, while the others were symptomatic. All patients underwent complete surgical resection (six lobectomies and seven wedge resections). Postoperative chemotherapy was performed in 3 cases. Overall, the median follow-up was 26 months (mean 33 months; range 9-95 months). All patients with goblet cell-type MC were alive and well, while the 2 patients with signet-ring cell-type MC died of disease. Immunohistochemically, all the 11 goblet cell-type MCs were strongly stained with CDX-2 and MUC2, 8 reacted with TTF-1, 6 with cytokeratin 20 (CK20), 9 with cytokeratin 7 (CK7), and 2 with MUC-5AC. Conversely, the two signet-ring cell-type MCs were stained with TTF-1, CK7, and MUC5AC but were negative for CDX-2, MUC2, and CK20. Surfactant apoprotein-A (SP-A) was positive in four goblet cell-type and one signet-ring cell-type MC. When compared with 10 mucinous bronchioloalveolar carcinomas (m-BAC), the latter reacted with CK7, CK20, MUC5AC, TTF-1, SP-A, CDX-2, and MUC2 in 100%, 90%, 100%, 30%, 10%, 0%, and 0% of the cases, respectively. In summary, MC of the lung represents an entity with two distinct clinicopathologic and immunophenotypic variants: 1) the goblet cell-type, presenting a more indolent clinical behavior and frequently co-expressing markers of intestinal and pulmonary differentiation; and 2) the more aggressive signet-ring cell-type, which retains only markers of pulmonary origin. On morphologic and immunohistochemical grounds, MCs are easily distinguishable from m-BAC. Since goblet cell-type MC strongly stains with CDX2, MUC2, and CK20, differential diagnosis with metastatic colorectal carcinoma is very challenging and requires appropriate clinical correlation.
2004
- Prognostic value of total tumor regression after pre-operative chemo-radiotherapy in rectal cancer (RC)
[Abstract in Rivista]
Gavioli, M; Luppi, G; Losi, Lorena; Bertolini, Federica; Giacobazzi, Patrizia; Zironi, S; Natalini, G; Bertoni, F; Conte, Pierfranco
abstract
The main objective of this study is to determine the prognostic value of complete tumor regression after pre-operative chemoradiotherapy in RC. Our study confirms that a complete pathological response on T (Dworak 4) after pre-operative treatment is an important prognostic factor. The persistance of microscopic residual disease (Dworak 3) is associated with a favourable local control.
2004
- Relationship between MUC5AC and altered expression of MLH1 protein in mucinous and non-mucinous colorectal carcinomas
[Articolo su rivista]
Losi, Lorena; Scarselli, A; Benatti, Piero; PONZ DE LEON, Maurizio; Roncucci, Luca; Pedroni, Monica; Borghi, F; Lamberti, Igor; Rossi, Giorgio; Marino, M; Ponti, Giovanni; Zangardi, G; Menigatti, M; Di Gregorio, C.
abstract
The main purpose of this study was to examine the expression of mucins and mismatch repair proteins in colorectal carcinomas. The immunohistochemical distribution of apomucins MUC2, MUC5AC, and the expression of MLH1 and MSH2 proteins were examined in 76 mucinous and 60 non-mucinous colorectal carcinomas. MUC2 was noted in all mucinous carcinomas, whereas MUC5AC was present in 41 cases only (54%). In non-mucinous carcinomas, MUC2 was expressed in 61.7% of the tumors; by contrast, MUC5AC was present in 20% of the cases. The expression level of apomucins was significantly different in mucinous and non-mucinous lesions (p < 0.001). Twenty-seven (35.5%) of the mucinous carcinomas showed no MLH1 expression, whereas I I (18.3%) of the non-mucinous tumors did. This difference was statistically significant (p < 0.005). Altered expression of MSH2 protein was never observed. The lack of MLH1 expression was considerably more frequent in carcinomas with secretion of MUC5AC (p<0.005). Our study has demonstrated this close relationship by immunohistochemical methods. In summary, our data show: (1) differences in the expression of mucins between mucinous and non-mucinous tumors; (2) a high frequency of altered MLH1 protein expression (35.5%) in mucinous carcinomas; (3) a significant relationship between the presence of MUC5AC and the altered expression of MLH1 protein in colorectal carcinomas.
2004
- Trend of incidence, subsite distribution and staging of colorectal neoplasms in the 15-year experience of a specialised cancer registry.
[Articolo su rivista]
PONZ DE LEON, Maurizio; Marino, M; Benatti, P; Rossi, G; Menigatti, M; Pedroni, Monica; Di Gregorio, C; Losi, Lorena; Borghi, F; Scarselli, A; Ponti, Giovanni; Roncari, B; Zangardi, G; Abbati, G; Ascari, E; Roncucci, Luca
abstract
BACKGROUND: Two-thirds of colorectal malignancies are localised in the left colon and rectum. Recent studies suggest a trend towards an increase of right-sided tumours which might have important implications for screening and surveillance. A colorectal cancer registry was set up in Modena, northern Italy, with the purpose of examining incidence, subsite distribution and staging of colorectal malignancies over a 15-year period.PATIENTS AND METHODS: From 1984 to 1998, 2517 tumours in 2462 patients were detected and staged with the tumour node metastasis (TNM) system. The 'right colon' was considered from caecum to splenic flexure; the 'left colon' included descending and sigmoid colon; and the 'rectum' included rectosigmoid junction, ampulla and anus.RESULTS: Cancer incidence showed an overall increase. Considering the various subsites, an increase of 33.7% in all colonic segments was shown whereas rectal tumours tended to decline. TNM staging showed a gradual increase of localised lesions (41.2% in 1984 versus 53.3% in 1998), with a proportional reduction of advanced tumours.CONCLUSIONS: Our study indicates an increase of tumour incidence in all colonic segments more than a shift to the right colon. TNM staging tended to improve with an appreciable increase of localised lesions. These findings could be consequent to a more extensive use of colonoscopy.
2003
- Caratterizzazione immunoistochimica e biomolecolare del carcinoma colorettale giovanile
[Abstract in Rivista]
Losi, Lorena; Di Gregorio, C; Pedroni, Monica; Lamberti, I; Roncucci, Luca; Ponti, Giovanni; Rossi, G; PONZ DE LEON, Maurizio; Benatti, Piero
abstract
Rivista della Società Italiana di Anatomia Patologica e Citopatologia Diagnostica
2003
- Different involvement of target genes mutations in hereditary and sporadic colorectal cancer with Microsatellite Instability.
[Poster]
Borghi, F; Pedroni, Monica; Lamberti, I; Menigatti, M; Ponti, Giovanni; Rossi, G; Di Gregorio, C; Losi, Lorena; Scarselli, A; Benatti, P; Roncucci, Luca; PONZ DE LEON, Maurizio
abstract
Functional inactivation of Mismatch Repair (MMR) genes by mutations or epigenetic mechanisms favors the acquisition of mutations in 'target genes'. We analysed 94 colorectal cancer (CRC) with microsatellite instability-high (MSI-H) for the presence of mutations in microsatellites located in the coding regiobns (CDRs) of 6 geens: TGF; RII, BAX, hMLH3, hMSH6, MBD4 and BLM.
2003
- L'immunoistochimica delle proteine del mismatch repair può essere un utile test per identificare i pazienti HNPCC?
[Abstract in Atti di Convegno]
Di Gregorio, C; Scarselli, A; Pedroni, Monica; Borghi, F; Lamberti, I; Ponti, Giovanni; Roncucci, Luca; Losi, Lorena; PONZ DE LEON, Maurizio; Benatti, P.
abstract
abstract
2003
- Mutations of the hMSH6 gene as a possible cause of hereditary nonpolyposis colorectal cancer
[Abstract in Atti di Convegno]
PONZ DE LEON, Maurizio; Scarselli, A; Benatti, Piero; Roncucci, Luca; Ponti, Giovanni; Losi, Lorena; Pedroni, Monica; Borghi, F; Menigatti, M; Di Gregorio, C.
abstract
Not avaivable
2002
- Alternative marker panel for Microsatellite Instability analysis in deection of contitutional MLH1 and MSH2 mutations.
[Abstract in Atti di Convegno]
Pedroni, Monica; Borghi, F; Lamberti, I; Scarselli, A; Menigatti, M; Ponti, Giovanni; Benatti, P; Losi, Lorena; Di Gregorio, C; Abbati, G; Rossi, G; Viel, A; Genuardi, M; Roncucci, Luca; PONZ DE LEON, Maurizio
abstract
Atti del convegno
2002
- Alternative marker panel for microsatellite instability analysis in detection of constitutional MLH1 and MSH2 mutations.
[Monografia/Trattato scientifico]
Pedroni, Monica; Borghi, F.; Lamberti, I.; Scarselli, A.; Menigatti, M.; Ponti, Giovanni; Benatti, Piero; Losi, Lorena; Di Gregorio, C.; Abbati, G.; Rossi, Giorgio; Viel, A.; Genuardi, M.; Roncucci, Luca; PONZ DE LEON, Maurizio
abstract
Hereditary Nonpolyposis Colorectal Cancer (I-INPCC) is an autosomal dominant syndrome characterized by predisposition to develop a number of neoplasms including colorectal, endometrium, urinary, extracolonic gastrointestinal, brain and ovarian cancers. HNPCC is caused by inherited mutations in DNA Mismatch Repair (MMR) genes. Defective DNA Mismatch Repair results in genetic instability, which can easily be owerved inshort repetitive sequences as microsatellites (Microsatellite Instability, MSI). An international workshop on MSIheld in Bethesda in 1997 proposed a panel of live microsatellite markers to be used in MSI analysis. This panelincludes two mononucleotide repeats (BAT25, BAT26) and three dinucleotide repeats (D2SI23, D5S346 andDI7S250). In our laboratory we are currently using a di&`erent microsatellite panel composed by threemononucleotide repeats (BAT25, BAT26, BAT40) and two dimmleotide repeats (D2SI23 and Dl8S57). 'I`heaim of this study was to evaluate the specificity ofthe Bethesda markers, as compared with our panel, to idmtifyMLHI and MSIE mutation—positive IINPCC. We compared the results of MSI-analysis in cancer from 27HNPCC Iizmilies (according to the Amsterdam criteria II) and ii•om 75 families in which not all the Amsterdamcriteria were met (Suspected IINPCC), using both the Bethesda and the alternative panel. In addition,immunohistochemistry of MLIII and MSH2 proteins was pertbrmed in all tumors in order to study thecorrelation between the two MSI-panels and the expression of MLIII and MSIE proteins.Using the Bethesda markers, 49 (48%) of tumors showed MSI. On the other hand, using the alternative panel, 33(32,3%) of tumors and displayed MSI. Loss of MLHI or MSIE was evident in 25 of 49 (5I%) MSI tumorsaccording to the Bethesda panel, whereas with the alternative panel 25 of 33 (75,7%) MSI tumors showed noprotein expression. In this group, eleven patients were tested for germline mutations of MMR genes, and all ofthem showed constitutional alterations.Our data suggest that the Bethesda panel is more sensitive to define MSI tumors, but the proposed marker panelis more specific than the Bethesda one to identity MSI tumors with no expression of MLHI and MSIE proteins.'I`he proposed marker panel seems to have a higher predictive value in the identification of
2002
- Biological characterization of mucinous carcinoma of the colon and rectum
[Abstract in Rivista]
Losi, Lorena; Scarselli, A; Benatti, P; PONZ DE LEON, Maurizio; Roncucci, Luca; Pedroni, Monica; Borghi, F; Rossi, G; Ponti, Giovanni; Zangardi, G; Menigatti, M; Di Gregorio, C.
abstract
Atti del convegno
2002
- Contributo di un registro tumori sede specifico per il cancro colorettale nell'individuazione di Sindromi rare.
[Relazione in Atti di Convegno]
Ponti, Giovanni; Di Gregorio, C; Scarselli, A; Rossi, G; Zangardi, G; Losi, Lorena; Roncucci, Luca; Pedroni, Monica; Borghi, F; Lamberti, I; Benatti, P; PONZ DE LEON, Maurizio
abstract
nd
2002
- HMSH6 immunohistochemistry in patients with clinical suspicion of Hereditary Non-Polyposis Colorectal Cancer.
[Monografia/Trattato scientifico]
Scarselli, A.; Benatti, Piero; Chichierchia, G.; Ponti, Giovanni; Lucci Cordisco, E.; Losi, Lorena; Menigatti, M.; Pedroni, Monica; Borghi, F.; Roncucci, Luca; Viel, A.; Genuardi, M.; PONZ DE LEON, Maurizio; Di Gregorio, C.
abstract
Colorectal Cancer (HNPCC) occurs in hMLHl or hMSH2. Recent observations have shown that mutations at hMSl·I6 also involved. Aim of our study is to investigate the role of hMSH6 gene in HNPCC by imnohistochernisny.Materials and methods. 28 colorectal cancer patients with clinical diagnosis of HNPCC or suspected HNPCC were inelected. Immunoliistochemical studies of hMSH6, hMLl—Il and hMSH2 were carried out on paraflin-embedded tumoursamples. lmrm1noperoxidase—staining using diarninobenzidinc as chromogen was carried out with the NEX-ES,Automatic Staining System (Ventana). Mouse monoclonal antibodies to 11MLI-I1 and hMSI-12 proteins (6163-15 andGI29-1129, Pharmingen) were used at 1:40 dilution, mouse monoclonal antibody to hMSH6 protein (clone 44,Transduction Laboratories) was used at 1:2000 dilution. All tumour samples were tested for MSI. Results. Lack of bMSH6 expression was detected in 7 out of 28 tumors. 4 of them also showed absence of hMSH2expression All 7 patients (mean age 56.6 yrs) were affected by right-sided colon cancer, most trequently mucinous and MSI+. 3 patients were from HNPCC families fulfilling Amsterdam Criteria I or I1, 2 were diagnosed as having Muir- Torre syndrome (MTS), and 2 had a diagnosis of suspected HNPCC. An excess of extracolonic tumours was observed in ali the pedigrees but one. interestingly, in both MTS patients, colorectal cancers and sebaceous dermatologic lesionsif thowed the same immunohistochemical pattern. At the moment, the complete MMR gene sequencing has been performed for 3 out of the 7 patients. 2 IJMSH6 and l hMSH2 ramcshifl mutations were detected- Conclusions. l1MSH6 mutations could be characteristic of a subset of HNPCC families. Altered hMSH6 immunohistochemical expression (although often associated with lack of hMSH2 protein), MSI positivity, proximal ` lllcalization, later age at diagnosis and association with extracolonic tumours, all seem to be prognostic features for the presence of this genetic alteration.
2002
- hMSH6 immunohistochemistry in patients with clinical suspicion of Hereditary Non-Polyposis Colorectal Cancer.
[Abstract in Atti di Convegno]
Scarselli, A; Benatti, Piero; Chichierchia, G; Ponti, Giovanni; Lucci Cordisco, E; Losi, Lorena; Menigatti, M; Pedroni, Monica; Borghi, F; Roncucci, Luca; Viel, A; Genuardi, M; PONZ DE LEON, Maurizio; Di Gregorio, C.
abstract
Congresso Italiano Patologia e Diagnostica Molecolare
2002
- Il registro dei tumori colorettali
[Monografia/Trattato scientifico]
PONZ DE LEON, Maurizio; Benatti, Piero; Di Gregorio, C.; Rossi, Giorgio; Losi, Lorena; Foroni, M.; Pedroni, Monica; Menigatti, M.; Zangardi, G.; Roncucci, Luca; Borghi, F.; Scarselli, A.; Percesepe, Antonio; Pasquale, C.
abstract
Not available
2002
- Management clinico-endoscopico di soggetti appartenti a famiglie con caratteristiche di sospetta HNPCC:la Sindrome di Muir-Torre.
[Abstract in Atti di Convegno]
Ponti, Giovanni; Benatti, Piero; Scarselli, A; Rossi, G; Zangardi, G; Abbati, G; Roncucci, Luca; Pedroni, Monica; Borghi, F; Lamberti, I; Riegler, G; Losi, Lorena; PONZ DE LEON, Maurizio
abstract
Atti del convegno
2002
- Two different marker panels for microsatellie instability analysis in detection of constitutional MLH1 and MSH2 mutations.
[Monografia/Trattato scientifico]
Pedroni, Monica; Borghi, F.; Lamberti, I.; Scarselli, A.; Menigatti, M.; Ponti, Giovanni; Benatti, Piero; Losi, Lorena; Di Gregorio, C.; Abbati, G.; Rossi, Giorgio; Roncucci, Luca; PONZ DE LEON, Maurizio
abstract
Hereditary Nonpolyposis Colorectal Cancer (HNPCC), an autosomal dominant susceptibility syndroriie, for approximately 5% of all colorectal tumours. Members of HNPCC families have a predisposition to the of colorectal cancer at an early age and an increased incidence of extracolonic cancer, e.g-, endometrium, ovaries, small bowel, biliary tract, renal pelvis and ureter. Gerrnline mutations in DNA mismatch repair genes, in particular MLI-Il and MSH2, are present in individuals with I-INPCC. Defective DNA mismatch results in genetic instability, which can easily be observed in short repetitive sequences such as microsatellites instability, MSI). MSI has been detected in most MMR-deficient tumours and it is considered the of I-INPCC. An international workshop on MSI held in Bethesda in 1997 proposed a panel of live markers to be used in MSI analysis. This panel, lmown as "Bethesda markers", includes two reats (BAT25, BAT26) and three dinucleotide repeats (D2Sl23, D5S346 and Dl7S250). ln our we are currently using a different microsatellite panel composed by three rnononucleotide repeats (BAT25, BAT40) and two dinucleoide repeats (D2Sl23 and Dl8S57). Aim of our study is to evaluate the speciticity ofr Q Btllhcsda markers, as compared with our panel, to identify MLHI and MSI-I2 mutation-positive HNPCC.We compared the results of MSI~analysis in tumours from 17 HNPCC families (according to and from 75 families in which not all the Amsterdam criteria were met (Suspected HNPCC),both the Bethesda panel and our alternative panel. MSI-H tumours were defined as having instability at two or markers (out of 5), while MSI-L tumours had instability at only one marker and MSS tumours were stable at all In addition, imrrninohistochemistry of MLH1 and MSH2 proteins was performed in all MSI-H and MSI-L to study the correlation between thc two MSI—panels and the expression of MLHI and MSH2 proteins. Results. Using the Bethesda markers, 27 (36%) out of the 75 Suspected HNPCC tumours and I2 (70,6%) out of the HNPCC tumours showed MSI. On the other hand, using our alternative panel, 16 (21.3%) Suspected HNPCC and 7 (41,2%) HNPCC tumours displayed MSI. In Suspected HNPCC tumours we found I5 (20%) md 5 (6.7%) MSI.,using the Bethesda panel and our alternative panel respectively, while in I-INPCC tumours no MSI-L was detected us either panel. Loss of MLHI or MSH2 was evident in 15 out of 39 (38,5%) MSI tumours according to the Beth panel, whereas with our alternative panel 15 out of 23 (65.2%) MSI tumours showed no protein expression.Conclusions. Our data suggest that the Bethesda panel is more sensitive to defne MSI tumours, but the propose marker panel is more sensitive to identify MSI tumours lacking expression of MLHI and MSI-I2 proteins. The marker panel seems to have higher predictive value in the identification of patients with ML!-Il and MSH2 mutations.
2001
- Epidemiologia dei tumori del colon-retto. Incidenza, mortalità, familiarità e sopravvivenza nella ex USL di Modena, 1984-1998.
[Monografia/Trattato scientifico]
PONZ DE LEON, Maurizio; Benatti, Piero; Rossi, Giorgio; Di Gregorio, C.; Roncucci, Luca; Losi, Lorena; Foroni, M.; Pedroni, Monica; Menigatti, M.; Zangardi, G.; Scarselli, A.; Percesepe, Antonio; Borghi, F.; Pasquale, C.
abstract
not available
2001
- Genotype-phenotype association in Italian HNPCC families:preliminary results and proposal of a National collaborative study.
[Monografia/Trattato scientifico]
Benatti, Piero; Lucci Cordisco, E.; Caluseriu, O.; Pedroni, Monica; Scarselli, A.; Losi, Lorena; Menigatti, M.; Borghi, F.; Roncucci, Luca; Viel, A.; Genuardi, M.; PONZ DE LEON, Maurizio
abstract
not available
2001
- hMSH6 immunohistochemistry in families with clinical suspicion of Hereditary Non Polyposis Colorectal Cancer
[Monografia/Trattato scientifico]
Scarselli, A.; Lucci Cordisco, E.; Benatti, Piero; Losi, Lorena; Menigatti, M.; Pedroni, Monica; Borghi, F.; Roncucci, Luca; Viel, A.; Genuardi, M.; PONZ DE LEON, Maurizio; Di Gregorio, C.
abstract
Not available
2001
- Kisten ras mutations in patients with colorectal cancer: the “RASCAL II” study.
[Articolo su rivista]
Andreyev, Hjn; Norman, Ar; Cunningham, D; Oates, J; Dix, Br; Iacopetta, Bj; Young, J; Walsh, T; Ward, R; Hawkins, N; Beranek, M; Jandik, P; Benamouzig, R; Jullian, E; LAURENT PUIG, P; Olschwang, S; Muller, O; Hoffmann, I; Rabes, Hm; Zietz, C; Troungos, C; Valavanis, C; Yuen, St; Ho, Jwc; Croke, Ct; Odonogue, Dp; Giaretti, W; Rapallo, A; Russo, A; Bazan, V; Tanaka, M; Omura, K; Onkusa, T; Fujimori, T; Ono, Y; Pauly, M; Faber, C; Glaesener, R; DE GOEIJ, Afpm; Arends, Jw; Andersen, Sn; Lovig, T; Breivik, J; Gaudernack, G; Clausen, Opf; DE ANGELIS, P; Meling, Gi; Rognum, To; Smith, R; Goh, H. S.; Font, A; Rosell, R; Sun, Xf; Zhang, H; Benhattar, J; Losi, Lorena; Lee, Jq; Wang, St; Clarke, Pa; Bell, S; Quirke, P; Bubb, Vj; Piris, J; Cruickshank, Nr; Morton, D; Fox, Jc; AL MULLA, F; Lees, N; Hall, Cn; Snary, D; Wilkinson, K; Dillon, D; Costa, J; Pricolo, Ve; Filkenstein, Sd; Thebo, Js; Senagore, Aj; Halter, Sa; Wadler, S; Malik, S; Krtolica, K; Urosevic, N.
abstract
This collaborative study suggests that not only is the presence of a codon 12 glycine to valine mutation important for cancer progression but also that it predispose to more aggressive biological behaviour in patients with advanced colorectal cancer.
2001
- Microsatellite instability and mismatch-repair protein expression in hereditary and sporadic colorectal carcinogenesis
[Articolo su rivista]
Pedroni, Monica; Sala, E; Scarselli, A; Borghi, F; Menigatti, M; Benatti, Piero; Percesepe, Antonio; Rossi, Giorgio; Foroni, M; Losi, Lorena; Di Gregorio, C; DE POL, Anto; Nascimbeni, R; Di Betta, E; Salerni, B; PONZ DE LEON, Maurizio; Roncucci, Luca
abstract
Aberrant crypt foci (ACF) are microscopic clusters of altered colonic crypts considered premalignant lesions in the large bowel. Genomic instability at short tandem repeats in the DNA, referred to as microsatellite instability (MSI) is the hallmark of hereditary nonpolyposis colorectal carcinoma (HNPCC) caused by mutations in DNA mismatch-repair genes, mostly hMLH1 and LMSH2. In this study, we evaluated for MSI ACF (n = 16), adenomas (n = 18), carcinomas (n = 22), and lymph node metastases (n = 3) from 17 patients with colorectal cancer positive for MSI, Ten patients were members of HNPCC families; 7 patients had no family history of cancer. MSI was found in 7 of 7 (100%) ACF and 11 of 12 (91%) adenomas from patients with HNPCC, MSI was not related to histology and size of ACF. A progressive increase in instability as estimated by the number of shifted bands was observed along the ACF-adenoma-carcinoma sequence, In contrast, two of nine (228) ACF and none of six adenomas from patients with MSI sporadic carcinoma were unstable at microsatellite loci. hMLH1 or hMSH2 protein expression was altered only in MSI-positive premalignant lesions (ACF and/or adenomas), but not in all MSI-positive lesions in patients with HNPCC. These observations provide evidence of the premalignant nature of ACF in HNPCC and suggest that MSI is a very early event both in HNPCC and in sporadic colorectal carcinogenesis, although in the latter it seems infrequent.
2001
- Molecular screening for Hereditary Non Polyposis Colorectal Cancer (HNPCC): a prospective, population-based study
[Articolo su rivista]
Percesepe, Antonio; F., Borghi; M., Menigatti; Losi, Lorena; M., Foroni; C., Di Gregorio; Rossi, Giuseppina; Pedroni, Monica; E., Sala; F., Vaccina; Roncucci, Luca; Benatti, Piero; A., Viel; M., Genuardi; G., Marra; P., Kristo; P., Peltomäki; PONZ DE LEON, Maurizio
abstract
PURPOSE: Germline mutations in mismatch repair genes predispose to hereditary nonpolyposis colorectal cancer (HNPCC). To address effective screening programs, the true incidence of the disease must be known. Previous clinical investigations reported estimates ranging between 0.5% and 13% of all the colorectal cancer (CRC) cases, whereas biomolecular studies in Finland found an incidence of 2% to 2.7% of mutation carriers for the disease. The aim of the present report is to establish the frequency of the disease in a high-incidence area for colon cancer.PATIENTS AND METHODS: Through the data of the local CRC registry, we prospectively collected all cases of CRC from January 1, 1996, through December 31, 1997 (N = 391). Three hundred thirty-six CRC cases (85.9% of the incident cases) were screened for microsatellite instability (MSI) with six to 12 mono- and dinucleotide markers. MSI cases were subjected to MSH2 and MLH1 germline mutation analysis and immunohistochemistry; the methylation of the promoter region was studied for MLH1.RESULTS: Twenty-eight cases (8.3% of the total) showed MSI. MSI cases differed significantly from microsatellite-stable (MSS) cases for their proximal location (P < .01), high mucinous component (P < .01), and poor differentiation (P = .002). Of MSI cases studied (n = 12), only one with a family history compatible with HNPCC had a germline mutation (in MSH2). Five other patients with a family history of HNPCC (two with MSI and three with MSS tumors) did not show germline mutations.CONCLUSION: We conclude that the incidence of molecularly confirmed HNPCC (one [0.3%] of 336) in a high-incidence area for CRC is lower than in previous biomolecular and clinical estimates.
2000
- Genomic instability and target gene mutations in colon cancers with different degrees of allelic shifts
[Articolo su rivista]
Percesepe, Antonio; Pedroni, Monica; Sala, E; Menigatti, M; Borghi, F; Losi, Lorena; Viel, A; Genuardi, M; Benatti, Piero; Roncucci, Luca; Peltomaki, P; PONZ DE LEON, Maurizio
abstract
Two grades (high and low) of microsatellite instability (MSI) are known, depending on the number of mutated markers and the amount of allelic shifts. Forty-two colorectal tumors, previously found to have high-degree MSI at dinucleotidic repeat loci, were revisited with BAT26, a mononucleotide marker, and the number of shifted bases were counted. Seven tumors, all with local stages at diagnosis, had less than or equal to 6-bp deletions and consistently displayed shorter shifts also with other intronic mononucleotide markers. Analysis of mononucleotide tracts in the coding regions of MSH3, MSH6, BAX, and TGF beta R11 in the groups with large (>6 bp) and short (less than or equal to 6 bp) allelic shifts showed specific patterns of involvement for the individual genes: TGF beta R11 displayed a uniformly high rate of mutations, while MSH3, MSH6, and BAX were less frequently altered in tumors with short shifts. Our findings suggest that microsatellite instability arises gradually, evenly involving loci with similar features of length and repetition. However, target genes have a specific timing of mutation in this process: TGF beta R11 is involved in the early phases, while BAX and MSH6 are frequently associated with big size shifts and tumors with more advanced stages.
2000
- Staging and survival of colorectal cancer: are we making progress? The 14-year experience of a Specialized cancer Registry
[Articolo su rivista]
PONZ DE LEON, Maurizio; Benatti, Piero; Di Gregorio, C; Fante, R; Rossi, Giorgio; Losi, Lorena; Pedroni, Monica; Percesepe, Antonio; Roncucci, Luca
abstract
Background and Aims. It is still unclear whether recent advancements in colorectal cancer research have led to an improvement in management and prognosis of the disease. Through the data of a specialized colorectal cancer Registry we aimed at analysing pathological staging and 5-year survival of ail patients with malignancies of large bowel diagnosed between 1984 and 1997. Main objective was to ascertain whether or not we are making progress in the control of this common neoplasm. Patients and Methods. During the 14-year period 1984-97, a total of 2,240 colorectal cancer patients were registered, for a crude incidence rate of 64.5 and 55.2/100,000/year in males and females, respectively Tumours were staged with Tumour Node, Metastasis system, corresponding to Dukes' classification, into four main groups. Survival was assessed with Life Table analysis, and statistical significance - between various subgroups - evaluated with Log-Rank Test. Results. Crude incidence rates of colorectal neoplasms showed minor fluctuations during initial period of registration, increasing sharply after 1990 mainly due to localized (stage I and II) lesions and, to a lesser degree, to stage ill tumours. Number of advanced (stage IV and unstaged) malignancies remained virtually stable. When results were expressed as percent of total cases, the fraction of localized lesions increased from 39% in the biennium 1984-5 to 51.6% in 1986-97, and the proportion of advanced tumours fail from 39% to 21.6% (p for trend <0.001). As expected, 5-year survival was significantly (p<0.002) more favourable for individuals diagnosed in 1990-91 than for patients registered in 1984-89. Conclusions. In Northern Italy, incidence rates of colorectal carcinoma are rising. This trend is associated with a sharp increase of newly detected localized lesions and with a significant improvement of overall 5-year survival The result may be attributed to several concomitant factors, such as: A) wider use of colonoscopy, B) increased education of patients, C) more attention given to symptoms.
1999
- Epidemiology of cancer of the large bowel - The 12-year experience of a specialized registry in Northern Italy
[Articolo su rivista]
PONZ DE LEON, Maurizio; Benatti, Piero; Percesepe, Antonio; C., Di Gregorio; R., Fante; Losi, Lorena; Rossi, Giorgio; Pedroni, Monica; Roncucci, Luca
abstract
Background. In 1984 a specialized colorectal cancer registry was instituted in Modena; aims of the Registry were: the evaluation of incidence and mortality, the study of morphological aspects, staging, staging and familiarity of the registered patients. Aims. Purpose of the research was to provide an updated description of the main findings (in particular incidence, staging, morphology and survival) observed in the 12-year registration period. Patients and methods. Between January 1984 and December 1995, 1,899 malignancies of the large bowel in 1,831 patients were registered. Tumours were classified according to the International Classification of the Diseases for Oncology (ICD-O) and staged with the TNM system Cancer specific srm il nl was assessed with life table analysis and Log-Rank tests. Results. Crude incidence rare showed minor fluctuations between 1984 and 1989, but tended to rise in the following pears. Tumours were mostly located distal to the splenic flexure (73.3% of the total), with a slight tendency over time to a gradual shift to the right colon. Staging became progressively more favourable throughout the registration; in 1984 both stages I, II and stage IV + unstaged lesions represented 40% of the total, but in 1995 the former rose to 50% whereas the latter fell to 21.6% (p<0.001). This more to earlier stages resulted in an improved survival of patients registered in 1990-91 versus 1983-85 (Log-Rank 14.3 p<0.002). Factors associated with a poor survival were the advanced age of patients at diagnosis (>74) and clinical stage. Conclusions. Main aspects of the investigation were the increasing crude incidence rates of colorectal turnovers and the gradual increase of neoplasms diagnosed irt a more favourable staging. It is like likely that the improvement of staging and survival observed in the 12 fears of registration can be attributed to the improved attitude to health care of the population, and possibly to the improvement of surgical techniques.
1999
- Microsatellite instability in multiple colorectal tumors
[Articolo su rivista]
Pedroni, Monica; Mg, Tamassia; Percesepe, Antonio; Roncucci, Luca; Benatti, Piero; G., Lanza; R., Gafa; C., Di Gregorio; R., Fante; Losi, Lorena; L., Gallinari; PONZ DE LEON, Maurizio; F., Scorcioni; F., Vaccina; Rossi, Giuseppina; A. M., Cesinaro
abstract
Tumor multiplicity is a hallmark of hereditary cancers: in the colon-rectum multiple tumors represent 5-10% of all colorectal cancer cases. A portion of these cases belongs to hereditary non-polyposis colorectal cancer (HNPCC), a genetic cancer syndrome due to mismatch repair (MMR) gene mutations, phenotypically expressed as microsatellite instability (MSI); the majority of multiple tumors, however, is apparently without any family history. We analyzed 78 (38 synchronous and 40 metachronous) neoplasms from 37 patients with multiple tumors of the large bowel, both HNPCC and sporadic, with the aim of identifying a common genetic basis in multiple tumors. DNA was extracted from normal and cancerous formalin-fixed tissue and was analyzed for MSI using 6 markers. Tumors showing MSI in at least 2 of 6 microsatellite loci were defined as MSI(+). The overall number of MSI(+) tumors was 22 (28.2% of the total). A significant difference in the rate of MSI(+) between HNPCC and sporadic tumors was observed (85% vs. 17%). In the same patients, the MSI phenotype of synchronous tumors (both HNPCC and sporadic) tended to be more concordant than that of the metachronous ones. The higher frequency of MSI in HNPCC than in sporadic tumors, even when multiple, suggests that the involvement of MMR genes in the pathogenesis of the sporadic cases may be uncommon, thus confirming that screening for MSI in multiple colorectal tumors could be a useful tool in the identification of HNPCC in the general population.
1998
- Aberrant crypt foci in patients with colorectal cancer
[Articolo su rivista]
Roncucci, Luca; S., Modica; Pedroni, Monica; Mg, Tamassia; M., Ghidoni; Losi, Lorena; R., Fante; C., Di Gregorio; Manenti, Antonio; L., Gafa; PONZ DE LEON, Maurizio
abstract
Aberrant crypt foci (ACF) are clusters of abnormally large colonic crypts identified on the mucosal surface of the human colon. They are thought to be preneoplastic lesions. The aim of the present study was to compare density (number of ACF per square cm of mucosal surface), crypt multiplicity (number of crypts per ACF) and histology of ACF in colonic resections of colorectal cancer patients resident in two Italian provinces with a twofold difference in colorectal cancer incidence rates. Thirty-two and 26 colonic resections were collected after operation in Ragusa (Southern Italy) and Modena (Northern Italy), respectively, and fixed in 10% formalin. Mucosal layers were observed under a light microscope at 25x after staining with methylene blue. Density of ACF was significantly higher in Modena (median 0.101 ACF cm(-2)) than in Ragusa (0.049, P= 0.001), whereas there was no difference in crypt multiplicity. ACF were classified into three groups according to histological features. ACF with mild alterations (hypertrophic ACF, 73%), ACF with hyperplasia (hyperplastic ACF, 17%) and ACF with dysplasia (microadenomas, 10%). The proportions of ACF in the three groups were similar in the two provinces, Density of ACF was higher and crypt multiplicity lower proceeding from proximal to distal large bower. Microadenomas were observed only in the colon, whereas hyperplastic ACF were more frequent in the rectum, In conclusion, density of ACF correlates with colorectal cancer rates in two Italian provinces, and shows a positive gradient from proximal to distal large bowel. Histology of ACF suggests that they may be precursors of both hyperplastic and adenomatous polyps. These data provide further evidence of the role of ACF in human colorectal carcinogenesis.
1997
- Colorectal carcinoma in different age groups: A population based investigation
[Articolo su rivista]
R., Fante; Benatti, Piero; S., Depietri; Pedroni, Monica; Mg, Tamassia; Percesepe, Antonio; Losi, Lorena; Roncucci, Luca; PONZ DE LEON, Maurizio; DI GREGORIO, Carmela; Rossi, Giuseppina
abstract
Although colorectal cancer is a disease of the older population, these tumors are not infrequent before the age of 55. Through the data of a population-based registry, we proposed giving a description of the clinical features of three groups of patients in whom the disease occurred at a relatively early age of onset (group I: < 40 yr; group II: 41-50 yr; group III: 51-55 yr). There were only 14 patients under the age of 40 yr (1.1% of total registered patients, n = 1298 in the period 1984-1992). Group II and III represented 5.9% and 6.0%, respectively (n = 76 and 78), with minor fluctuations throughout the 9-yr period of registration. Inherited colorectal tumors [hereditary nonpolyposis colorectal cancer (HNPCC), adenomatosis coli, and suspected HNPCC] accounted for 38.4% of group I patients (5 of 14), 17.1% of group II, 10.2% of group III, and only 3.5% of individuals older than 55 (p, for trend, < 0.001). Thus, hereditary colorectal tumors were detected significantly more often in younger individuals. The majority of colorectal malignancies were localized in the left colon or rectum in all three groups,,vith a tendency (not significant) to a preferential localization in the right colon for tumors developed in group I (37% vs 18% and 14% in groups II and III, respectively). Pathological stage and main histological types did not differ among the three groups. Finally, life-table analysis did not show significant differences in 5-yr survival among the three groups; however, when considered together, early onset cases showed a more favorable prognosis than older individuals (log-rank 11.6; p < 0.001). In conclusion, colorectal cancer is diagnosed very rarely before the age of 40 yr, whereas about 12% of all cases belong to the age group 41 to 55 yr of age. Hereditary tumors were found more frequently in younger patients, with a well-defined inverse relationship between age of onset and frequency of genetically determined tumors. Finally, the clinical outcome was more favorable in the whole series of early onset cases than in older registered patients.
1997
- Histology of aberrant crypt foci in the human colon
[Articolo su rivista]
Di Gregorio, C; Losi, Lorena; Fante, R; Modica, S; Ghidoni, M; Pedroni, Monica; Tamassia, Mg; Gafà, R; PONZ DE LEON, Maurizio; Roncucci, Luca
abstract
Aberrant crypt foci (ACF) have been identified in the methylene-blue stained mucosa of the human colon, Some lines of evidence suggest that ACF may be precursors of colon cancer, The objective of the present study was to establish morphological criteria able to define and classify ACF in histological sections, Twenty four colectomy specimens were collected after operation for colorectal cancer and fixed in 10% formalin, Strips of grossly normal mucosa were stained in a 0.2% solution of methylene blue in saline for 5-10min. The strips were measured, put on a glass slide and observed under a light microscope at x25, One hundred and fourteen ACF identified by topology were sectioned parallel to the muscularis mucosae. Eighty-four ACF were evident at histological examination and could be classed into three main groups: group A (61 ACF 72.6%) including foci whose epithelial cells had regular nuclei, with only mild or focal crowding but no stratification, no mucin depletion and no dysplasia; group B (16 ACF 19.1%), in which features of hyperplasia were evident; and group C (seven ACF 8.3%) including foci with enlarged, crowded and stratified nuclei, mucin depletion, frequent mitoses, and evident dysplasia, diffuse or focal (mild in five cases, moderate in two) representing microadenomas, Finally, hyperplastic foci were significantly larger than foci of group A and C, Group B ACF were also more frequent in the rectum than in the colon. In conclusion, selected histological features allow the definition of groups of ACF, which may represent sequential steps in the development of human colorectal tumours.
1997
- K-ras and p53 mutations in hereditary non-polyposis colorectal cancers
[Articolo su rivista]
Losi, Lorena; PONZ DE LEON, Maurizio; Jiricny, J; Di Gregorio, C; Benatti, Piero; Percesepe, Antonio; Fante, R; Roncucci, Luca; Pedroni, Monica; Benhattar, J.
abstract
Genetic instability related to defective DNA mismatch repair genes may be involved in the pathogenesis of carcinoma in Hereditary Non-Polyposis Colorectal Cancer (HNPCC). To test that the targets of genetic instability could include critical transforming genes involved in colon tumor progression, we examined 23 colorectal carcinomas in patients with HNPCC in order to detect somatic mutations in K-ros and p53 genes. Using single strand conformation polymorphism followed by direct DNA sequencing, we detected 4 mutations in K-ros gene (17%) and 3 in p53 gene (13%) which change the aminoacid sequence of the protein p53. This is significantly lower than in sporadic cancer. Our data suggest that colon cancer in HNPCC might partly involve a distinct pathogenetic mechanism that involves other genes than those altered in sporadic tumors.
1997
- Myxoid metastases of melanoma: report of three cases and review of the literature
[Articolo su rivista]
G., Collina; Losi, Lorena; G. L., Taccagni; Maiorana, Antonino
abstract
We report three cases of melanoma whose metastases to skin and regional lymph nodes showed myxoid foci, findings absent in their cutaneous primary tumors.
1996
- Clinical features, frequency and prognosis of Dukes' a colorectal carcinoma: A population-based investigation
[Articolo su rivista]
C., Digregorio; R., Fante; Roncucci, Luca; Mg, Tamassia; Losi, Lorena; Benatti, Piero; Pedroni, Monica; Percesepe, Antonio; S., Depietri; PONZ DE LEON, Maurizio
abstract
The main aim of this study was, through the data of a population-based Registry, to establish the incidence of Dukes´ A lesions by year of registration and the main clinical features, and to assess cancer-specific survival. One hundred and eighteen Dukes´ A colorectal tumours were diagnosed (in 117 patients) out of 1337 registered between 1984 and 1992 in the Health Care District of Modena, Northern Italy; 94 patients were treated with surgery and 23 with endoscopic polypectomy. The frequency of Dukes´ A tumours ranged between 4.8% and 18% by year of registration. Dukes´ A carcinomas were significantly more frequent in the distal colon. Only 5 patients (4%) died of their cancer, and in all patients the tumour was localised in the rectum. Carcinomas associated with a poor prognosis did not show any of the biological variables usually associated with an unfavourable outcome, but, our data suggest the possibility of incomplete removal of tumours at surgery. Copyright
1996
- Clinico-pathological correlation and prognostic significance of nuclear p53 protein in colorectal cancer
[Articolo su rivista]
R., Fante; C., Digregorio; Losi, Lorena; Roncucci, Luca; PONZ DE LEON, Maurizio
abstract
Overexpression of p53 protein was studied in neoplastic specimens of 150 patients registered for colorectal adenocarcinoma in the Health Care District 16 of Modena, Italy, from 1984 to 1986, We selected Dukes' stage B (92) and C (58) patients whose survival and recurrence rates are not easily predictable, with the purpose of defining subgroups of patients at high risk of recurrence, Monoclonal antibody PAb 1801 was used on formalin-fixed, paraffin embedded specimens, Nuclear staining was assessed to divide tumours into three groups: a) negative, b) low expressors, c) high expressors, Histomorphological variables of tumours, major clinical features of the patients and 5-year specific survival, were evaluated and related to p53 status. p53 was found in 71 out of 150 cases (47.3%); 50 tumours were high and 21 low expressors, No correlation was found between p53 overexpression and clinico-pathological variables. No difference in survival was found between patients with p53 positive and negative tunours in the entire series or within Dukes' stage B and C patients, However, the subgroup of patients with stage C rectal cancer seemed to have a better prognosis if the tumour was p53 negative (of borderline significance, p=0.15), The same results were obtained by grouping low expressor tunours alternatively with negative or high expressors. We conclude that p53 nuclear overexpression does not seem to influence the prognosis of Dukes' stage B or C colorectal cancer patients.
1996
- Frequency and clinical features of multiple tumors of the large bowel in the general population and in patients with hereditary colorectal carcinoma
[Articolo su rivista]
Fante, R; Roncucci, Luca; Di Gregorio, C; Tamassia, Mg; Losi, Lorena; Benatti, Piero; Pedroni, Monica; Percesepe, Antonio; De Pietri, A; PONZ DE LEON, Maurizio
abstract
BACKGROUND. Reports on the frequency of multiple carcinomas of the colon and rectum have varied from 3-4% to more than 10% of all tumors of the large bowel. METHODS. We reviewed the files of a specialized colorectal cancer registry with chronous or metachronous) in the general population; b) to compare these values with those observed in patients with hereditary nonpolyposis colorectal carcinoma the following objectives: a) to determine the frequency of multiple tumors (synchronous or metachronous) in the general population; b) to compare these values with those observed in patients with hereditary nonpolyposis colorectal carcinoma (HNPCC); and c) to evaluate the clinical outcome of patients with multiple tumors and the role of other clinical parameters in the development of these neoplasms. RESULTS. From 1984 to 1992, 53 patients with multiple tumors (of 1298 registered patients, 4%) had large bowel carcinoma; 33 (2.5%) were synchronous and 20 (1.5%) metachronous. The total number of multiple colorectal carcinomas was 95, which was 7% of all registered colorectal carcinomas (1337 carcinomas in 1298 patients). Multiple tumors occurred significantly more often in patients with HNPCC than in those with sporadic carcinomas (P < 0.001); this increased prevalence was more marked for metachronous lesions, which occurred almost 4 times more often in patients with HNPCC (5.8% vs. 1.3%; P < 0.001). The average interval of time between the first and the second malignancy was 8.7 years; there was no significant difference between hereditary and sporadic tumors. Patients with synchronous tumors did not show appreciable differences in survival when compared with individuals who had single neoplasms. In contrast, a poor clinical outcome was observed in patients with metachronous tumors after the development of the second carcinoma. Finally, polypoid adenomas of the large bowel were found significantly more often in patients with multiple primary tumors than in those with a single tumor. CONCLUSIONS. These results emphasize the importance of preoperative pancolonoscopy for the identification of possible synchronous tumors (both benign and malignant) and long-lasting endoscopic follow-up for the detection of recurrent or metachronous lesions. The conclusions are even more pertinent for patients with HNPCC, whose risk of metachronous tumors is significantly higher than that of patients with sporadic carcinoma.
1996
- Identification of subjects at risk for colorectal carcinoma through a test based on K-ras determination in the stool
[Articolo su rivista]
Villa, Erica; A., Dugani; Am, Rebecchi; A., Vignoli; Grottola, Antonella; P., Buttafoco; Losi, Lorena; M., Perini; P., Trande; A., Merighi; R., Lerose; Manenti, Federico
abstract
Background & Aims: The gold standard for screening for colorectal carcinoma is colonoscopy. The aim of this study was to compare endoscopic results with those obtained using the noninvasive screening test of K-ras determination in the stool in a large population of patients undergoing colonoscopy. Methods: Two hundred thirty consecutive patients were studied by K-ras amplification on stool-derived DNA using polymerase chain reaction and oligomer-specific hybridization. Results: Wild-type K-ras was amplified in 103 of 230 patients (44.8%), the rate of amplification being directly proportional to the presence of an organic disease of the intestine characterized by hyperproliferating mucosa. In 30 of these 103 patients (29.1%), a K-ras mutation was found. Four of 5 with early colorectal carcinoma, all who had K-ras mutations in the tumor, were identified. In first-degree relatives of patients with colorectal carcinoma, all subjects either carrying adenomas >1 cm in diameter or multiple smaller adenomas were identified. In patients with inflammatory bower disease, the test identified the only patient with neoplastic transformation. Conclusions: The sensitivity and specificity of K-ras determination on stool-derived DNA in patients with colorectal carcinoma, in first-degree relatives of patients with colorectal carcinoma, and in patients with inflammatory bowel disease support the opportunity of a large-scale trial to validate its use as a screening test.
1996
- K-ras and p53 mutations in human colorectal aberrant crypt foci
[Articolo su rivista]
Losi, Lorena; Roncucci, Luca; Di Gregorio, C; PONZ DE LEON, Maurizio; Benhattar, J.
abstract
Aberrant crypt foci (ACF) are putative precursor lesions of colon cancer, recently identified on the methylene blue-stained mucosal surface of human colon. No mutations in K-ras or p53 genes were found by non-radioactive single-strand conformation polymorphism analysis in 14 ACF collected from five patients. Using the more sensitive method of allele-specific polymerase chain reaction (PCR) for K-ras, 8 of 14 ACF were found to contain K-ras mutations, suggesting that mutated cells are present in minute clones in ACF, No dysplasia was observed in any of the ACF containing a mutated clone. The presence of K-ras mutations in ACF suggests that these lesions occur at a very early stage in human colorectal carcinogenesis.
1996
- Survival for colon and rectal cancer in a population-based cancer registry
[Articolo su rivista]
Roncucci, Luca; R., Fante; Losi, Lorena; C., Digregorio; A., Micheli; Benatti, Piero; N., Madenis; D., Ganazzi; Mt, Cassinadri; P., Lauriola; PONZ DE LEON, Maurizio
abstract
Dukes' stage is the most powerful indicator of patient outcome for colorectal cancer. Several cancer survival studies have considered other prognostic variables, but results are often conflicting. We sought to assess the independent value of several clinical and morphological variables in defining colorectal cancer specific survival. 397 colorectal cancer patients diagnosed from 1984 to 1986, and registered in a large bowel cancer registry instituted in a local health district of Northern Italy, were actively followed-up until 31 December 1991. Univariate and multivariate survival analyses were carried out in colon and rectal cancer cases, separately, using the actuarial life-table method and Cox proportional hazard regressions. Crude and specific 5-year survival rates were 37.5 and 41.4%. In univariate analysis, TNM (tumour, nodes and metastases) stage was the strongest predictor of prognosis in both sites. Other variables significantly related to survival were age of patient at diagnosis and pattern of tumour growth in colon cancer, type of differentiation and pattern of tumour growth in rectal cancer. In multivariate analyses, after adjusting for stage, age had a weak but significant negative effect on colon cancer survival, whereas rectal tumours with the infiltrating type of growth had a significantly worse prognosis than those with the expanding type. Colorectal cancer survival should be analysed in the main large bowel subsites in order to define high-risk groups within each TNM stage category.
1995
- Argyrophilic nucleolar organizer regions and bromodeoxyuridine and h3-thymidine labelling indices in colorectal cancer
[Articolo su rivista]
Losi, Lorena; C., Di Gregorio; R., Fante; Migaldi, Mario; Roncucci, Luca; Pedroni, Monica; PONZ DE LEON, Maurizio; G. P., Trentini
abstract
The count of argyrophilic nucleolar organizer regions (AgNORs) has been proposed as a useful method for evaluating cell replication in human tumours. The current study was undertaken to compare AgNOR values in colorectal cancers with two better established methods for investigating cell proliferation such as bromodeoxyuridine (BrdUrd) and (3)[H]-thymidine ((3)[H]dT) labelling indices (LIs). Because some concern still exists regarding accuracy and reproducibility of AgNOR quantifying methods, we carried out a control study by independently repeating the same measurements (number, area and area per silver-stained NOR particle) in two centres with different operators and computer-assisted image analysers on 40 colorectal carcinomas. AgNOR values recorded in the two centres were strictly correlated (r = 0.75; P < 0.001 for number; r = 0.62, P < 0.01 for area; r = 0.63, P < 0.001 for area per silver-stained NOR particle) and the range of values were almost identical, Then, AgNOR values were compared with BrdUrd and (3)[H]dT LIs, respectively obtained by in vivo incorporation and in vitro incubation in the same series of colorectal carcinomas. No correlation was found between AgNOR values and BrdUrd or (3)[H]dT LIs. BrdUrd and (3)[H]dT LIs were instead reciprocally significantly correlated, No evident correlation was seen between LIs or AgNOR values and clinico-pathological parameters of the tumour. In conclusion, in colorectal neoplasms, AgNOR values did not appear to relate with more direct parameters of cell proliferation. It follows that AgNOR reliability as a biomarker of cell proliferation remains questionable.
1995
- Biologic characterization of Hereditary Non-Polyposis Colorectal Cancer. Nuclear ploidy, AgNOR count, microvessel distribution, oncogene expression, and grade-related parameters.
[Articolo su rivista]
Losi, Lorena; R., Fante; C., DI GREGORIO; M. L., Aisoni; G., Lanza; I., Maestri; Roncucci, Luca; Pedroni, Monica; PONZ DE LEON, Maurizio
abstract
The identification of hereditary non-polyposis colorectal cancer (HNPCC) is important not only for the patient, but also for family members who are at increased risk of developing cancer. To determine if measuring various pathobiologic features of the colon carcinomas is useful in separating sporadic from HNPCC tumors, the authors studied tumor tissues from 46 patients with HNPCC and compared them to 70 with sporadic colorectal carcinoma. Parameters investigated included DNA ploidy (flow cytometry), AgNOR count (by silver staining), microvessel density (immunohistochemistry), p53 and K-vas expression, and grade-related parameters. Diploid tumors were more frequent in patients with HNPCC (65% vs 40%, P <.02), thus confirming previous observations concerning such an association. Higher AgNOR counts and greater AgNOR areas were observed in sporadic tumors than in HNPCC (5.2 +/- 1.5 vs 4.5 +/- 1.8, P <.01). Hereditary tumors tended to be less vascularized, whereas oncogene expression and grade-related parameters did not show appreciable differences between the two types of tumors. In conclusion, some of the investigated parameters may contribute to defining the biologic profile of HNPCC. In addition, these findings support the clinical impression of a more favorable outcome that is frequently seen in HNPCC patients.
1995
- Migration cholécysto-cholédocienne de végétations adénomateuses papillaires
[Articolo su rivista]
Manenti, Antonio; Gandolfo, C; Miselli, A; Losi, Lorena; Miselli, R.
abstract
L'adénome de la vésicule biliaire peut se cancériser; il entraine exceptionnellement une obstruction du canal cystique ou du choledoque par migration de ses végétations papillaires. Notre observation en est un exemple typique.
1995
- Variant estrogen receptor messenger RNA species detected in human primary hepatocellular carcinoma
[Articolo su rivista]
Villa, Erica; L., Camellini; A., Dugani; F., Zucchi; Grottola, Antonella; A., Merighi; P., Buttafoco; Losi, Lorena; Manenti, Federico
abstract
The development of hepatocellular carcinoma (HCC) in addition to cirrhosis affects males in a significantly higher proportion than females. Liver estrogen receptors increase when HCC develops in males; however, these tumors usually respond poorly to antiestrogens. We have, therefore, hypothesized that, similar to breast cancer, estrogen receptors in males with HCC may be mutated. Variant estrogen receptor transcripts (lacking exon 5 of the hormone binding domain) were investigated by reverse transcription-PCR in 14 patients (7 males and 7 females) with HCC. While females mostly displayed the wild-type transcript (both in peritumoral and in tumor Liver tissue), males showed both transcripts in the cirrhotic tissue and almost only the variant in the tumor. ris the variant ER transcripts when translated could give rise to truncated receptors still able to constitutively activate transcription, they may be key factors in favoring deregulated proliferation in the male liver.
1994
- Expression of p53 protein in prostate cancers of different histologic types.
[Articolo su rivista]
Losi, Lorena; DI GREGORIO, C; Brausi, M; Fante, R; Hurlimann, J.
abstract
Mutations and overexpression of p53 gene in prostate carcinoma have been found but their significance in the development and progression of cancer is so far unknown. We investigated the prevalence of abnormalities of p53 protein in a heterogeneous group of prostate carcinoma to verify whether acinar and non acinar carcinomas have a different expression of p53 protein. Paraffin sections of 45 prostate carcinomas (39 acinar, 3 ductal papillary, 1 transitional cell, 1 mucinous and 1 pure small cell) were examined for the expression of p53 protein using a panel of antibodies (monoclonal antibodies Pab 1801, D07 and polyclonal antibody CM1). No p53 expression was observed in any acinar carcinomas independent of grade and stage. For non acinar carcinomas only small cell and transitional cell carcinomas exhibited detectable amounts of p53 protein in tumour cell nuclei. The prevalence of p53 overexpression in prostate carcinoma is relatively low compared with that found in many other tumours. In the present study, the overexpression of p53 in a small cell carcinoma and in a transitional cell carcinoma suggests that the loss of suppressing role of p53 gene may be an important mechanism in the genesis and in the development of these uncommon tumours.
1994
- Rare prostatic carcinomas: histogenesis and morphologic pattern.
[Articolo su rivista]
Losi, Lorena; Brausi, M; DI GREGORIO, C.
abstract
Most prostate cancers (90%) are acinar adenocarcinomas originating in the peripheral or other prostatic regions. The pathological finding, clinical course and immunohistochemical studies, indicate that the small cell carcinoma of the prostate is most likely to be a neuroendocrine neoplasm.
1993
- Genetic events in sporadic colorectal adenomas: K-ras and p53 heterozygous mutations are not sufficient for malignant progression.
[Articolo su rivista]
DE BENEDETTI, L; Varesco, L; Pellegata, Ns; Losi, Lorena; Gismondi, V; Casarino, L; Sciallero, S; Bonelli, L; Biticchi, R; Bafico, A; Masetti, E; James, R; Heouaine, A; Ranzani, Gn; Aste, H; Ferrara, G.
abstract
Twenty-four sporadic colorectal adenomas were analysed for the presence of allelic loss on the short arm of chromosome 17 as well as mutations in the K-ras and p53 genes. Chromosome 17p13 allelic loss was not present in 14 out of 14 informative cases. K-ras mutations were observed in 15 out of 24 cases. A p53 gene mutation (GGC --> GAC at codon 245) was detected in two biopsies taken at a four year interval from a recurrent rectal villous adenoma. Both biopsies also contained the same K-ras gene mutation (GGT --> GTT at codon 12). The data from the recurrent rectal adenoma provide in vivo evidence that K-ras and p53 heterozygous mutations confer a proliferative advantage but together are not sufficient for malignant transformation.
1993
- Prognostic significance of K-ras mutations in colorectal carcinoma.
[Articolo su rivista]
Benhattar, J; Losi, Lorena; Chaubert, P; Givel, Jc; Costa, J.
abstract
In Dukes' B and C primary tumours, mutations other than GGT to GAT identify patients at very high risk of recurrence. Our reslults indicate that determining the K-ras mutations provides a good prognostic factor in patients with advanced colorectal carcinoma.
1993
- Rare frequence of point mutations for codon 12, 13 and 61 of ras gene in Italian neuroblastoma.
[Articolo su rivista]
Iolascon, A; Badiali, M; Pession, A; Basso, G; Losi, Lorena; MIRAGLIA DEL GIUDICE, E; Perrotta, S; Cutillo, S; Tonini, Gp
abstract
Single point mutations of ras oncogenes are found in many tumors and contribute to the pathogenesis of the cancer. The product of the ras gene, p21 protein, was found expressed in several neuroblastoma tissues. However, the role of ras gene in this tumor has yet to be clarified. To contribute to the understanding of the ras activation, 79 fresh biopsies of neuroblastoma were studied to investigate the possibility that ras would be activated by point mutation. Analysis of H-ras and N-ras was performed by means of PCR and SSO, while K-ras mutations were detected by multiplex-ASPCR. None of the neuroblastomas examined showed H- or K-ras activation, while N-ras mutations were demonstrated in only three patients (3,7%). N-myc oncogene is amplified in a substantial number of patients with neuroblastoma. N-myc amplification was studied by Southern blot technique. N-myc amplification was demonstrated in 13.2% of patients less than 1 year of age at diagnosis and 23% of older children. Two of the patients (one stage I and one stage IVs) with N-ras mutation and without N-myc amplification had a good outcome, while the third (stage IVs) with N-myc amplification had a poor prognosis. These results suggest that ras activation is a rare event in both amplified and non-amplified neuroblastoma tumors and that N-ras activation was not involved in the clinical outcome of these patients. Moreover, our data suggest that p21 expression is induced by a post-transcriptional activation.
1993
- Tumour spectrum in hereditary non-polyposis colorectal cancer (HNPCC) and in families with "suspected HNPCC". A population-based study in Northern Italy. Colorectal Cancer Study Group.
[Articolo su rivista]
Benatti, Piero; R., Sassatelli; Roncucci, Luca; Pedroni, Monica; R., Fante; C., DI GREGORIO; Losi, Lorena; Gelmini, Roberta; PONZ DE LEON, Maurizio
abstract
Hereditary non-polyposis colorectal cancer (HNPCC or Lynch syndrome) is characterized by the early onset of colorectal neoplasms, frequently localized in the right colon, increased occurrence of multiple primaries, vertical transmission and aggregation of tumours in families in accordance to a Mendelian dominant type of inheritance. The syndrome accounts for approximately 5% of all colorectal cancers. The purpose of the present study was to describe the tumour spectrum and the most relevant clinical features of 28 kindreds with HNPCC, classified according to the guidelines of the international Collaborative Study Group, and of 61 ''suspected'' HNPCC. These families were observed during a 6-year registration of colorectal neoplasms in a health-care district of Northern Italy. Colorectal cancer was by far the most frequent malignancy; gastric cancer was the second. Uterine carcinoma was only slightly more frequent than expected. Lung- and breast-tumour rates were lower than expected. Cancer distribution in the large bowel showed that about two fifths of the tumours developed in the right colon. The occurrence of cancer before the age of SO to 60 was much more frequent in HNPCC. Multiple tumours developed in 25 patients with HNPCC and in 32 with ''suspected'' HNPCC. Pancolonoscopy remains the procedure of choice for surveillance; other examinations, such as gastroscopy, gynaecological investigations, urography and cholangiography, are suggested only to selected families. One of the main features of the study was the inclusion of 61 ''suspected'' HNPCC, a heterogeneous group of families which nonetheless deserves careful follow-up.
1992
- Cell kinetics evaluation of colorectal tumours after in vivo administration of bromodeoxyuridine
[Articolo su rivista]
Roncucci, Luca; Pedroni, Monica; A., Scalmati; Ml, Bormioli; R., Sassatelli; R., Fante; Losi, Lorena; C., DI GREGORIO; B., Petocchi; PONZ DE LEON, Maurizio
abstract
Although several biomarkers have been tested, Dukes' (or TNM) stage at diagnosis is still considered the only prognostic factor of clinical relevance in colorectal cancer. Among the various biomarkers, the fraction of cells engaged in DNA synthesis has been extensively investigated as an indicator of tumor aggressiveness. Bromodeoxyuridine (BUdR) is a non-radioactive thymidine analogue which is incorporated into DNA during the S-phase of cycling cells. In order to evaluate the relationships between cell kinetics and morphologic variables, 500 mg of BUdR were given i.v. to patients with colorectal cancer prior to surgery. After operation, a large tumor sample was taken and processed for immunohistochemical detection of BUdR-labeled cells in various regions of the neoplasm and in normal colorectal mucosa. Smaller superficial tumor specimens were also incubated with H-3-thymidine (H-3-TdR) for the autoradiographic identification of labeled cells. In the 43 evaluable tumors, the overall BUdR labeling index (BLI, percent of labeled cells) was significantly higher in carcinoma (20.30 +/- 0.86%, SEM) than in normal colonic mucosa (6.51 +/- 0.49%). BLIs in central and peripheral regions of carcinoma were closely correlated (r = 0.48, p = 0.003). In 21 neoplasms a high correlation between overall BUdR and H-3-TdR labeling index in the same tumor was observed (r = 0.57, p = 0.007). No evident association between overall BU and clinical or morphologic parameters of the tumor was seen, including number of capillaries and ras-p21 protein expression. We conclude that BUdR immunostaining after in vivo administration of BUdR is a simple method for studying cell kinetics in various regions of colorectal cancer. BUdR labeling data are comparable to those obtained with in vitro incorporation of H-3-TdR.
1992
- Paneth and argyrophil cells in prostatic carcinoma.
[Articolo su rivista]
Botticelli, Ar; Digregorio, C; Fano, Rita Adriana; Losi, Lorena; Manenti, Antonio
abstract
The presence of Paneth and argyrophil cells in prostatic carcinomas supports the existence of mixed and/or metaplastic carcinoma. We believe that these cells derive from local precursor or common stem cells.
1992
- Stability of K-ras mutations throughout the natural history of human colorectal cancer.
[Articolo su rivista]
Losi, Lorena; Benhattar, J; Costa, J.
abstract
We have studied 35 patients who developed recurrent tumour. In 71% of these patients a ras mutation in codons 12 or 13 was observed in the primary tumour. For each of these cases an identical ras mutation was found in the DNA from the local or distant recurrence. Our results indicate that K-ras mutation provides a stable tumour marker throughout the natural history of colorectal cancer.
1991
- Nucleolar organizer regions in malignant melanoma and melanocytic nevi. Comparison of two counting methods.
[Articolo su rivista]
DI GREGORIO, C; Losi, Lorena; Annessi, G; Botticelli, A.
abstract
Using a silver staining technique, we studied nucleolar organizer regions (AgNOR) in paraffin sections of junctional nevi, compound nevi, intradermal nevi, blue nevi, dysplastic nevi, Spitz nevi, lentigo maligna, malignant melanomas in nevus, superficial spreading melanomas, and nodular melanomas. Two methods of counting black dots within nuclei were employed. One method was to count the discrete black dots within the nuclei, including the tiny black dots seen within the nucleolus; the second method did not take into account the subsidary cluster of tiny black dots seen within the nucleolus, instead treating these dots as a single structure. Whichever method we used, a significant difference was found between the pooled mean AgNOR numbers for benign and malignant lesions. We found an overlap, however, between benign, in particular Spitz and dysplastic nevi, and malignant lesions when considering individual counts of AgNOR using both methods. We conclude that studying AgNOR does not seem to be a useful technique to differentiate Spitz and dysplastic nevi from malignant melanomas.
1991
- Pathological findings in perinatal autopsies
[Capitolo/Saggio]
Criscuolo, M; Migaldi, Mario; Botticelli, Annibale Renzo; Losi, Lorena
abstract
The study of perinatal mortality rates has recently be entered by pathologists, because of their capacity to discover the true cause of death at necroscopy.
1989
- MELANOSIS (PIGMENTED MELANOCYTOSIS) OF THE PROSTATE GLAND.
[Articolo su rivista]
Botticelli, Ar; DI GREGORIO, C; Losi, Lorena; Fano, Ra; Manenti, A.
abstract
A case of stromal melanosis of the prostate gland is reported. Ultrastructurally, dendritic processes and melanosomes, but no basal lamina were observed in melanin-containing cells. In similar cases, characterized by the diffuse distribution of stromal pigmented melanocytes, the term "pigmented melanocytosis" rather than "blue naevus" appears more appropriate.
1989
- ONCOGENES AND GROWTH FACTORS IN OVARIAN EPITHELIAL TUMORS.
[Articolo su rivista]
Trentini, Gp; DE GAETANI, C; Losi, Lorena
abstract
The present data underline the existence of over-expression of several oncogenes in common epithelial tumours. The detection of oncogene expression in ovarian tumours may be useful in predicting tumor prognosis, and in determining genetic relationships between histologically similar but clinically dissimilar tumor types.