Immunohistochemical and molecular pattern of p53 in epithelial ovarian cancers negative for germline BRCA1/2 variants.

Epithelial ovarian cancers (EOC) associated with germline or somatic BRCA pathogenetic variants have a significantly higher rate of TP53aberrations. The majority of TP53 mutations are detectable by immunohistochemistry and several studies demonstrated that an abnormal p53 pattern characterized high-grade EOCs. An abnormal p53 immunohistochemical staining in fallopian tube (serous tubal intraepithelial carcinoma (STIC) and "p53 signature" is considered as a precancerous lesion of high-grade EOCs and it is often found in fallopian tube tissues of BRCA germline mutated patients suggesting that STIC is an early lesion and the TP53 mutation is an early driver event of BRCA mutated high-grade EOCs. No relevant data are present in literature about the involvement of p53 abnormal pattern in EOC carcinogenesis of patients negative for germline BRCA variants. We describe TP53 mutation results in relationship to the immunohistochemical pattern of p53 expression in a series of EOCs negative for BRCA1 and BRCA2 germline mutations. In addition, we also investigated STIC presence and "p53 signature" in fallopian tube sampling of these EOCs. Our results demonstrate that TP53 alterations are frequent and early events in sporadic EOCs including also low-grade carcinomas. Also in this series, STIC is associated with an abnormal p53 pattern in fallopian tubes of high-grade EOCs. In summary, TP53 aberrations are the most frequent and early molecular events in EOC carcinogenesis independently from BRCA mutation status.

MLH1 Promoter Methylation Could Be the Second Hit in Lynch Syndrome Carcinogenesis.

(1) Background: MLH1 hypermethylation is an epigenetic alteration in the tumorigenesis of colorectal cancer (CRC) and endometrial cancer (EC), causing gene silencing, and, as a consequence, microsatellite instability. Commonly, MLH1 hypermethylation is considered a somatic and sporadic event in cancer, and its detection is recognized as a useful tool to distinguish sporadic from inherited conditions (such as, Lynch syndrome (LS)). However, MLH1 hypermethylation has been described in rare cases of CRC and EC in LS patients. (2) Methods: A total of 61 cancers (31 CRCs, 27 ECs, 2 ovarian cancers, and 1 stomach cancer) from 56 patients referred to cancer genetic counselling were selected for loss of MLH1 protein expression and microsatellite instability. All cases were investigated for MLH1 promoter methylation and MLH1/PMS2 germline variants. (3) Results: Somatic MLH1 promoter hypermethylation was identified in 16.7% of CRC and in 40% of EC carriers of MLH1 germline pathogenic variants. In two families, primary and secondary MLH1 epimutations were demonstrated. (4) Conclusions: MLH1 hypermethylation should not be exclusively considered as a sporadic cancer mechanism, as a non-negligible number of LS-related cancers are MLH1 hypermethylated. Current flow charts for universal LS screening, which include MLH1 methylation, should be applied, paying attention to a patient's family and personal history.

Similarities and differences in gene expression profiles of BRCA1 methylated and mutated epithelial ovarian cancers.

Introduction: BRCA1 methylated (BRCA1met) epithelial ovarian cancer (EOC) is a recently defined and not well-investigated subset of neoplasms. To date, no studies have focused on the transcriptional profiles of BRCA1met cases, and, as a matter of fact, we still do not know if this subset of EOCs is similar, and to what extent, to BRCA1 mutated (BRCA1mut) cases. Methods: We compared a group of 17 BRCA1met cases against 10 BRCA1mut cases using a subset of carefully selected 17 BRCAwt EOCs as a control group. Results: First, BRCA1met cases showed a downregulation of the relative transcript, while this association was not observed for BRCA1mut EOCs. The BRCA1met group exhibited a general upregulation of homologous recombination (HR)-related genes, as well as BRCA1mut. Overall, BRCA1met had a different gene expression profile, characterized by diffuse downregulation, whereas BRCA1mut showed a general upregulation (p < 0.0001). Both BRCA1-defective groups showed a slightly activated immune response mediated by interferon (IFN) gamma pathways. Discussion: In conclusion, even if the expression profile of many genes related to DNA damage and repair system is shared between BRCA1mut and BRCA1met EOCs supporting that BRCA1met EOCs may benefit from PARPi therapies, our data demonstrate that BRCA1mut and BRCA1met EOCs show different expression profiles, suggesting a different mechanism of carcinogenesis that can be reflected in different responses to therapies and disease recovery.

Pyrosequencing Assay for BRCA1 Methylation Analysis: Results from a Cross-Validation Study.

Epithelial ovarian cancers (EOCs) harboring germline or somatic pathogenic variants in BRCA1 and BRCA2 genes show sensitivity to poly(ADP-ribose) polymerase inhibition. It has been suggested that BRCA1 promoter methylation is perhaps a better determinant of therapy response, because of its intrinsic dynamic feature, with respect to genomic scars or gene mutation. Conflicting evidence was reported so far, and the lack of a validated assay to measure promoter methylation was considered a main confounding factor in data interpretation. To contribute to the validation process of a pyrosequencing assay for BRCA1 promoter methylation, 109 EOCs from two Italian centers were reciprocally blindly investigated. By comparing two different pyrosequencing assays, addressing a partially overlapping region of BRCA1 promoter, an almost complete concordance of results was obtained. Moreover, the clinical relevance of this approach was also supported by the finding of BRCA1 transcript down-regulation in BRCA1-methylated EOCs. These findings could lead to the development of a simple and cheap pyrosequencing assay for diagnostics, easily applicable to formalin-fixed, paraffin-embedded tissues. This technique may be implemented in routine clinical practice in the near future to identify EOCs sensitive to poly(ADP-ribose) polymerase inhibitor therapy, thus increasing the subset of women affected by EOCs who could benefit from such treatment.

Targeted molecular profiling of epithelial ovarian cancer from Italian BRCA wild-type patients with a BRCA and PARP pathways gene panel.

Ovarian cancer (OC) is the fifth most common type of cancer in women and the fourth most common cause of cancer death in women. Identification of pathogenic variants in OC tissues has an important clinical significance for therapeutic and prevention purposes. This study aims to evaluate the mutational profile of a patient cohort, negative for BRCA1/2 germinal variants and Mismatch Repair defects, using next-generation sequencing (NGS) approach on DNA from formalin-fixed paraffin-embedded samples. We used a custom NGS panel, targeting 34 cancer-related genes, mainly of the BRCA and PARP pathways, and analyzed NGS data to identify somatic and germline variants in Italian patients affected by primary epithelial ovarian cancer. We analyzed 75 epithelial ovarian cancer tissues and identified 54 pathogenic variants and 56 variants of unknown significance. TP53 was characterized by the highest mutational rate, occurring in 55% of tested epithelial ovarian cancers (EOCs). Interestingly, a subset of 8 EOCs showed pathogenic variants of homologous recombination pathway, which could be sensitive to PARP-inhibitor therapies. Germline analysis of actionable genes revealed 4 patients carrier of pathogenic germline variants respectively of RAD51C (2 patients), RAD51D, and PALB2. Molecular profiling of EOCs using our custom NGS panel has enabled the detection of both somatic and germline variants, allowing the selection of patients suitable for targeted therapies, and the identification of high-risk OC families that can benefit from genetic counseling and testing.

Case Report: Male Lobular Breast Cancer in Hereditary Cancer Syndromes.

Background: Lobular breast carcinoma (LBC) is considered an exceptionally rare disease in men, including only 1% of all male breast malignancies. The majority of LBCs have negative immunohistochemical staining for E-cadherin (CDH1) expression, and the loss of CDH1 function was traditionally implicated in the tumorigenesis of diffuse gastric cancer as well as LBC. It is well recognized that LBC in women could be involved in both hereditary breast and ovarian cancer (HBOC) and hereditary diffuse gastric cancer (HDGC) syndromes; however, there are no data present in literature about the involvement of male LBC in these inherited conditions. Methods: BRCA1, BRCA2, and CDH1 genes were performed on DNA from peripheral blood using next-generation sequencing (NGS), Sanger sequencing, and multiplex ligation-dependent probe amplification analyses. BRCA2 and CDH1 somatic gene analyses were performed on breast tumoral DNA using the NGS sequencing approach. Results and conclusions: Here, we describe two men affected by LBC, the carriers of a pathogenic variant of BRCA2 and CDH1 genes, respectively. Our data, including somatic and germline results, demonstrate a strong relationship between male LBC and HBOC/HDGC syndromes, excluding a sporadic origin of LBC in these two patients. Male LBC could represent a sentinel cancer for inherited syndrome identification, and early identification of cancer susceptibility could improve cancer prevention both for men and women in these families. The history of the LBC patient carrier of the CDH1 variant suggests to include male LBC genetic testing criteria and male breast surveillance in HDGC guidelines.

OncoPan®: An NGS-Based Screening Methodology to Identify Molecular Markers for Therapy and Risk Assessment in Pancreatic Ductal Adenocarcinoma.

Pancreatic cancer has a high morbidity and mortality with the majority being PC ductal adenocarcinomas (PDAC). Whole genome sequencing provides a wide description of genomic events involved in pancreatic carcinogenesis and identifies putative biomarkers for new therapeutic approaches. However, currently, there are no approved treatments targeting driver mutations in PDAC that could produce clinical benefit for PDAC patients. A proportion of 5-10% of PDAC have a hereditary origin involving germline variants of homologous recombination genes, such as Mismatch Repair (MMR), STK11 and CDKN2A genes. Very recently, BRCA genes have been demonstrated as a useful biomarker for PARP-inhibitor (PARPi) treatments. In this study, a series of 21 FFPE PDACs were analyzed using OncoPan®, a strategic next-generation sequencing (NGS) panel of 37 genes, useful for identification of therapeutic targets and inherited cancer syndromes. Interestingly, this approach, successful also on minute pancreatic specimens, identified biomarkers for personalized therapy in five PDAC patients, including two cases with HER2 amplification and three cases with mutations in HR genes (BRCA1, BRCA2 and FANCM) and potentially eligible to PARPi therapy. Molecular analysis on normal tissue identified one PDAC patient as a carrier of a germline BRCA1 pathogenetic variant and, noteworthy, this patient was a member of a family affected by inherited breast and ovarian cancer conditions. This study demonstrates that the OncoPan® NGS-based panel constitutes an efficient methodology for the molecular profiling of PDAC, suitable for identifying molecular markers both for therapy and risk assessment. Our data demonstrate the feasibility and utility of these NGS analysis in the routine setting of PDAC molecular characterization.

Strategies for Lynch syndrome identification in selected and unselected gynecological cancers.

BACKGROUND: Endometrial carcinoma represents a sentinel cancer for Lynch syndrome (LS) identification. It is crucial to highlight how other types of tumors can arise in the gynecological tract acting as sentinel tumors in LS patients.Up to now, no established LS patient management strategy has incorporated the presence of these additional candidate sentinel tumors to improve the prevention and management of LS tumors. METHODS: In order to investigate the involvement of the most frequent gynecological cancers in gynecological cancers, we studied different subsets of gynecological cancers using both somatic approaches, including mismatch repair (MMR) gene immunohistochemical expression, microsatellite instability, and germline analyses ofMSH2, MSH6, MLH1, PMS2 and EPCAM genes.A total of 261 patients referring to the Cancer Genetic Counselling Service of our institution were included in the study. In detail, our series was composed of 131 patients affected by uterus cancers including endometrial, isthmus and non-HPV endocervical carcinomas, 113 patients affected by ovarian cancers and 17 patients affected by synchronous endometrial/ovarian carcinomas (SEOC).In addition, we studied 115 cases of endometrial cancers identified by 2 years of universal testing (endometrial cancers/UTs) using IHC analysis of four MMR proteins. RESULTS AND CONCLUSIONS: The incidence of MMR defective gynecological cancers ranged from 7.1 to 47.1% depending on cancer site and selection. LS patients carriers of pathogenetic MMR variants were identified in 19.8% of uterus cancers, 35.3% of SEOC, 4.4% of ovarian cancers. In addition, pathogenetic MMR variants were identified in 4.3% of endometrial cancers/universal testing investigated with universal screening.In conclusion, gynecological cancers are heavily involved in LS and our study shows that MMR screening using immunohistochemical pattern and MSI analysis of endometrial and ovarian cancers as well as of rare entities such as non-HPV related endocervical cancers and synchronous endometrial and ovarian cancers are sentinels for LS.Tumor testing approach improves early identification of MMR defective gynecological cancers and this is an effective strategy to detect high-risk patients and to offer them and their relatives personalized cancer prevention.

Mixed Neuroendocrine/Non-neuroendocrine Neoplasm (MiNEN) of the Ovary Arising from Endometriosis: Molecular Pathology Analysis in Support of a Pathogenetic Paradigm.

Primary ovarian neuroendocrine neoplasms (Ov-NENs) are infrequent and mainly represented by well-differentiated forms (neuroendocrine tumors - NETs - or carcinoids). Poorly differentiated neuroendocrine carcinomas (Ov-NECs) are exceedingly rare and only few cases have been reported in the literature. A subset of Ov-NECs are admixed with non-neuroendocrine carcinomas, as it occurs in other female genital organs, as well (mostly endometrium and uterine cervix), and may be assimilated to mixed neuroendocrine/non-neuroendocrine neoplasms (MiNENs) described in digestive and extra-digestive sites. Here, we present a case of large cell Ov-NEC admixed with an endometrioid carcinoma of the ovary, arising in the context of ovarian endometriosis, associated with a uterine endometrial atypical hyperplasia (EAH). We performed targeted next-generation sequencing analysis, along with a comprehensive immunohistochemical study and FISH analysis for TP53 locus, separately on the four morphologically distinct lesions (Ov-NEC, endometrioid carcinoma, endometriosis, and EAH). The results of our study identified molecular alterations of cancer-related genes (PIK3CA, CTNNB1, TP53, RB1, ARID1A, and p16), which were present with an increasing gradient from preneoplastic lesions to malignant proliferations, both neuroendocrine and non-neuroendocrine components. In conclusion, our findings underscored that the two neoplastic components of this Ov-MiNEN share a substantially identical molecular profile and they progress from a preexisting ovarian endometriotic lesion, in a patient with a coexisting preneoplastic proliferation of the endometrium, genotypically and phenotypically related to the ovarian neoplasm. Moreover, this study supports the inclusion of MiNEN in the spectrum ovarian and, possibly, of all gynecological NENs, among which they are currently not classified.

Detection of Germline Variants in 450 Breast/Ovarian Cancer Families with a Multi-Gene Panel Including Coding and Regulatory Regions.

With the progress of sequencing technologies, an ever-increasing number of variants of unknown functional and clinical significance (VUS) have been identified in both coding and non-coding regions of the main Breast Cancer (BC) predisposition genes. The aim of this study is to identify a mutational profile of coding and intron-exon junction regions of 12 moderate penetrance genes (ATM, BRIP1, CDH1, CHEK2, NBN, PALB2, PTEN, RAD50, RAD51C, RAD51D, STK11, TP53) in a cohort of 450 Italian patients with Hereditary Breast/Ovarian Cancer Syndrome, wild type for germline mutation in BRCA1/2 genes. The analysis was extended to 5'UTR and 3'UTR of all the genes listed above and to the BRCA1 and BRCA2 known regulatory regions in a subset of 120 patients. The screening was performed through NGS target resequencing on the Illumina platform MiSeq. 8.7% of the patients analyzed is carriers of class 5/4 coding variants in the ATM (3.6%), BRIP1 (1.6%), CHEK2 (1.8%), PALB2 (0.7%), RAD51C (0.4%), RAD51D (0.4%), and TP53 (0.2%) genes, while variants of uncertain pathological significance (VUSs)/class 3 were identified in 9.1% of the samples. In intron-exon junctions and in regulatory regions, variants were detected respectively in 5.1% and in 32.5% of the cases analyzed. The average age of disease onset of 44.4 in non-coding variant carriers is absolutely similar to the average age of disease onset in coding variant carriers for each proband's group with the same cancer type. Furthermore, there is not a statistically significant difference in the proportion of cases with a tumor onset under age of 40 between the two groups, but the presence of multiple non-coding variants in the same patient may affect the aggressiveness of the tumor and it is worth underlining that 25% of patients with an aggressive tumor are carriers of a PTEN 3'UTR-variant. This data provides initial information on how important it might be to extend mutational screening to the regulatory regions in clinical practice.

Lynch syndrome and Muir-Torre phenotype associated with a recurrent variant in the 3'UTR of the MSH6 gene.

A MSH6 3'UTR variant (c.*23_26dup) was found in 13 unrelated families consulted for Lynch/Muir-Torre Syndrome. This variant, which is very rare in the genomic databases, was absent in healthy controls and strongly segregated with the disease in the studied pedigrees. All tumors were defective for MSH2/MSH6/MSH3 proteins expression, but only MSH2 somatic pathogenic mutations were found in 5 of the 12 sequenced tumors. Moreover, we had no evidence of MSH6 transcript decrease in carriers, whereas MSH2 transcript was downregulated. Additional evaluations performed in representative carriers, including karyotype, arrayCGH and Linked-Reads whole genome sequencing, failed to evidence any MSH2 germline pathogenic variant. Posterior probability of pathogenicity for MSH6 c.*23_26dup was obtained from a multifactorial analysis incorporating segregation and phenotypic data and resulted >0.999, allowing to classify the variant as pathogenic (InSiGHT Class 5). Carriers shared a common haplotype involving MSH2/MSH6 loci, then a cryptic disease-associated variant, linked with MSH6 c.*23_26dup, cannot be completely excluded. Even if it is not clear whether the MSH6 variant is pathogenic per se or simply a marker of a disease-associated MSH2/MSH6 haplotype, all data collected on patients and pedigrees prompted us to manage the variant as pathogenic and to offer predictive testing within these families.

BRCA Methylation Testing Identifies a Subset of Ovarian Carcinomas without Germline Variants That Can Benefit from PARP Inhibitor.

Homologous Recombination Deficiency (HRD) is a frequent feature of high-grade epithelial ovarian carcinoma (EOC), associated with sensitivity to PARP-inhibitors (PARPi). The best characterized causes of HRD in EOCs are germline or somatic mutations in BRCA1 and BRCA2 genes. Although promoter methylation is a well-known mechanism of gene transcriptional repression, few data have been published about BRCA gene methylation in EOCs. In this retrospective study, we quantitatively analyzed by pyrosequencing a selected series of 90 formalin-fixed (FFPE) primary EOCs without BRCA germline mutations. We identified 20/88 (22.7%) EOCs showing BRCA promoter methylation, including 17/88 (19.3%) in BRCA1 and 4/86 (4.6%) in BRCA2 promoters, one of which showing concomitant BRCA1 methylation. Mean methylation levels were 49.6% and 45.8% for BRCA1 and BRCA2, respectively, with methylation levels ≥50% in 10/20 methylated EOCs. Constitutive BRCA methylation was excluded by testing blood-derived DNA. In conclusion, pyrosequencing methylation analysis of BRCA genes is a robust, quantitative and sensitive assay applicable to FFPE samples. Remarkably, a considerable subset of germline BRCA-negative EOCs showed somatic methylation and, likely, HRD. A subpopulation of women with BRCA methylation, even without BRCA mutations, could potentially benefit from PARP-inhibitors; further clinical studies are needed to clarify the predictive role of somatic BRCA methylation of PARP-therapy response.

HPV nonrelated endocervical adenocarcinoma in hereditary cancer syndromes.

INTRODUCTION: The relationship between endocervical cancer and cancer susceptibility syndromes is not yet fully understood. We present 2 cases of endocervical cancer: 1 arising in a patient carrier with a pathogenic BRCA1 variant and the second detected in a Lynch syndrome family carrying the MSH2 germline pathogenic variant. CASE DESCRIPTION: Somatic analyses including loss of heterozygosity and fluorescent in situ hybridization demonstrated that the second hit in patient 1 is BRCA1-related. Mismatch repair somatic analyses in the second family demonstrated that the endocervical cancers of patient 2 and of her sister are MSH2-related. These data confirm the relationship between the pathogenesis of endocervical cancer and the presence of germline BRCA1 and MSH2 mutations. CONCLUSIONS: Our study confirms that gynecologic cancers including rare entities such as non-human papillomavirus-related endocervical cancer (NHPVA) are sentinels for inherited cancer syndromes. Endocervical cancer NHPVAs might be considered for cancer genetic counseling in order to improve cancer prevention. For this reason, the role of pathologists is particularly important for the correct identification of the cervical tumor site.

STELO: a new tool for family physicians for the correct identification of inherited cancer syndromes.

BACKGROUND: The hereditary cancer syndromes represent overall <10% of all cancers. These syndromes are not irrelevant for public health because all the cancers typical of these syndromes affected young people and many members of the same family and the cancers are more aggressive than the sporadic ones and need specific surgery and medical therapy. We developed a new family assessment tool: STELO designed for family physicians to identify patients could benefit from Cancer Genetic Counselling. OBJECTIVE: Test the sensitivity and specificity of a new assessment tool for the correct identification of inherited cancer syndromes. METHODS: Retrospectively we tested the new tool on a subset of patients who had already undergone genetic counselling at the Cancer Genetic Counselling Service of ASST (Azienda Socio Sanitaria Territoriale) Settelaghi Varese, to investigate sensitivity, specificity and applicability of this new tool in routine genetic screening. STELO responses were matched against the opinion of two cancer geneticists (i.e. gold standard) who blinded each other decided if the history of these patients was properly suspected as a hereditary cancer syndrome. RESULTS: The Genetic Counselling Service followed 546 subjects from 2014 to 2015. STELO tool was tested retrospectively on these clinical records and resulted positive in 418 cases, out of 546 (76.5%). STELO reported, towards the gold standard, 88.5% and 52.3% of sensitivity and specificity, respectively. CONCLUSIONS: STELO has demonstrated to have a good sensitivity. The specificity was expectedly low given that STELO has been developed for general medicine, so it needs to be simple, practical, of rapid consultation and effectively used in clinical practice.

Lynch syndrome-related non-endometrioid endometrial cancer: analysis of outcomes.

OBJECTIVE: Women with Lynch syndrome have a risk up to 40-60% of developing endometrial cancer, which is higher than their risk of developing colorectal or ovarian cancer. To date, no data on the outcomes of patients with Lynch syndrome diagnosed with non-endometrioid endometrial cancer are available. The goal of this study was to evaluate the outcome of patients with Lynch syndrome diagnosed with non-endometrioid endometrial cancer. METHODS: Data from consecutive patients diagnosed with Lynch syndrome and with a histological diagnosis of non-endometrioid endometrial cancer were retrospectively collected in two referral institutes in Italy. A case-control comparison (applying a propensity matching algorithm) was performed in order to compare patients with proven Lynch syndrome and controls. Inclusion criteria were: (a) histologically-proven endometrial cancer; (b) detection of a germline pathogenic variant in one of the MMR genes; (c) adequate follow-up. Only carriers of pathogenic or likely pathogenic variants (ie, class 5 and 4 according to the InSiGHT classification) were included in the study. Survival outcomes were assessed using KaplanMeier and Cox models. RESULTS: Overall, 137 patients with Lynch syndrome were collected. Mean patient age was 49.2 (10.9) years. Genes involved in the Lynch syndrome included MLH1, MSH2, and MSH6 in 43%, 39%, and 18% of cases, respectively. The study population included 27 patients with non-endometrioid endometrial cancer, who were matched 1:2 with patients with sporadic cancers using a propensity matching algorithm. After a median follow-up of 134 months (range 1-295), 2 (7.4%) of the 27 patients developed recurrent disease (3 and 36 months) and subsequently died of disease (7 and 91 months). Patients diagnosed with Lynch syndrome experienced better disease-free survival (HR 7.86 (95% CI 1.79 to 34.5); p=0.006) and overall survival (HR 5.33 (95% CI 1.18 to 23.9); p=0.029) than controls. CONCLUSIONS: Non-endometrioid endometrial cancer occurring in patients with Lynch syndrome might be associated with improved oncologic outcomes compared with controls. Genetic/molecular profiling should be investigated in order to better understand the mechanism underlying the prognosis.

Association of Genomic Domains in BRCA1 and BRCA2 with Prostate Cancer Risk and Aggressiveness.

Pathogenic sequence variants (PSV) in BRCA1 or BRCA2 (BRCA1/2) are associated with increased risk and severity of prostate cancer. We evaluated whether PSVs in BRCA1/2 were associated with risk of overall prostate cancer or high grade (Gleason 8+) prostate cancer using an international sample of 65 BRCA1 and 171 BRCA2 male PSV carriers with prostate cancer, and 3,388 BRCA1 and 2,880 BRCA2 male PSV carriers without prostate cancer. PSVs in the 3' region of BRCA2 (c.7914+) were significantly associated with elevated risk of prostate cancer compared with reference bin c.1001-c.7913 [HR = 1.78; 95% confidence interval (CI), 1.25-2.52; P = 0.001], as well as elevated risk of Gleason 8+ prostate cancer (HR = 3.11; 95% CI, 1.63-5.95; P = 0.001). c.756-c.1000 was also associated with elevated prostate cancer risk (HR = 2.83; 95% CI, 1.71-4.68; P = 0.00004) and elevated risk of Gleason 8+ prostate cancer (HR = 4.95; 95% CI, 2.12-11.54; P = 0.0002). No genotype-phenotype associations were detected for PSVs in BRCA1. These results demonstrate that specific BRCA2 PSVs may be associated with elevated risk of developing aggressive prostate cancer. SIGNIFICANCE: Aggressive prostate cancer risk in BRCA2 mutation carriers may vary according to the specific BRCA2 mutation inherited by the at-risk individual.

Gut Microbiota Analysis in Postoperative Lynch Syndrome Patients.

Lynch syndrome (LS) is a dominantly inherited condition with incomplete penetrance, characterized by high predisposition to colorectal cancer (CRC), endometrial and ovarian cancers, as well as to other tumors. LS is associated with constitutive DNA mismatch repair (MMR) gene defects, and carriers of the same pathogenic variants can show great phenotypic heterogeneity in terms of cancer spectrum. In the last years, human gut microbiota got a foothold among risk factors responsible for the onset and evolution of sporadic CRC, but its possible involvement in the modulation of LS patients' phenotype still needs to be investigated. In this pilot study, we performed 16S rRNA gene sequencing of bacterial DNA extracted from fecal samples of 10 postoperative LS female patients who had developed colonic lesions (L-CRC) or gynecological cancers (L-GC). Our preliminary data show no differences between microbial communities of L-CRC and L-GC patients, but they plant the seed of the possible existence of a fecal microbiota pattern associated with LS genetic background, with Faecalibacterium prausnitzii, Parabacteroides distasonis, Ruminococcus bromii, Bacteroides plebeius, Bacteroides fragilis and Bacteroides uniformis species being the most significantly over-represented in LS patients (comprising both L-CRC and L-GC groups) compared to healthy subjects.

Large scale multifactorial likelihood quantitative analysis of BRCA1 and BRCA2 variants: An ENIGMA resource to support clinical variant classification.

The multifactorial likelihood analysis method has demonstrated utility for quantitative assessment of variant pathogenicity for multiple cancer syndrome genes. Independent data types currently incorporated in the model for assessing BRCA1 and BRCA2 variants include clinically calibrated prior probability of pathogenicity based on variant location and bioinformatic prediction of variant effect, co-segregation, family cancer history profile, co-occurrence with a pathogenic variant in the same gene, breast tumor pathology, and case-control information. Research and clinical data for multifactorial likelihood analysis were collated for 1,395 BRCA1/2 predominantly intronic and missense variants, enabling classification based on posterior probability of pathogenicity for 734 variants: 447 variants were classified as (likely) benign, and 94 as (likely) pathogenic; and 248 classifications were new or considerably altered relative to ClinVar submissions. Classifications were compared with information not yet included in the likelihood model, and evidence strengths aligned to those recommended for ACMG/AMP classification codes. Altered mRNA splicing or function relative to known nonpathogenic variant controls were moderately to strongly predictive of variant pathogenicity. Variant absence in population datasets provided supporting evidence for variant pathogenicity. These findings have direct relevance for BRCA1 and BRCA2 variant evaluation, and justify the need for gene-specific calibration of evidence types used for variant classification.

Performance of tumor testing for Lynch syndrome identification in patients with colorectal cancer: A retrospective single-center study.

OBJECTIVE:: To investigate the performance of tumor testing approaches in the identification of Lynch syndrome (LS) in a single-center cohort of people with colorectal cancer (CRC). METHODS:: A retrospective analysis of data stored in a dedicated database was carried out to identify patients with CRC suspected for LS who were referred to Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy, between 1999 and 2014. The sensitivity and specificity of immunohistochemistry (IHC) for mismatch repair (MMR) proteins and microsatellite instability (MSI) analysis (alone or combined) were calculated with respect to the presence of causative MMR germline variants. RESULTS:: A total of 683 patients with CRC suspected for LS were identified. IHC results of MMR protein analysis and MSI were assessed in 593 and 525 CRCs, respectively, while germline analysis was performed in 418 patients based on the IHC or MSI test result and/or clinical features. Univariate and multivariate analysis revealed a significant correlation of pathogenic MMR germline variants with all clinicopathologic features including Amsterdam criteria, presence of endometrial cancer, CRC site, age at onset, stage, and grade. The highest odds ratio values were observed for IHC and MSI (17.1 and 8.8, respectively). The receiver operating characteristic curve and area under the curve values demonstrated that IHC alone or combined with other clinicopathologic parameters was an excellent test for LS identification. CONCLUSIONS:: This study confirms the effectiveness of tumor testing to identify LS among patients with CRC. Although IHC and MSI analysis were similarly effective, IHC could be a better strategy for LS identification as it is less expensive and more feasible.