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.

Two Cases of Carcinosarcomas of the Ovary Involved in Hereditary Cancer Syndromes.

Ovarian carcinosarcomas (OCS), also known as malignant mixed mesodermal/Müllerian tumors, are rare neoplasms (1%-4% of all malignant ovarian tumors) composed of high-grade malignant epithelial and mesenchymal elements. OCS occurs in older women. It is associated with a poor outcome and is usually not involved in inherited cancer syndromes. We present 2 cases of OCS; one arising in a patient with a pathogenetic BRCA1 mutation and the other in a woman affected by Lynch Syndrome (LS) carrying a MSH6 germline mutation. To the best of our knowledge, this is the first time that this second type of case has been reported. In this study, we investigated somatic impairment of the wild-type BRCA1 and MSH6 alleles in the OCS of these 2 patients. We also explored in both OCS, the occurrence of TP53 loss of function, which is a genetic alteration known to occur in BRCA-linked ovarian tumorigenesis but not in LS tumors. Moreover, we also provide further data about the histogenesis of OCS.

Somatic Testing on Gynecological Cancers Improve the Identification of Lynch Syndrome.

OBJECTIVE: Recent data from the literature indicate gynecological cancers (GCs) as sentinel cancers for a diagnosis of Lynch syndrome (LS). Clinical approaches to identifying LS have low sensitivity, whereas somatic tests on GCs may be a more sensitive and cost-effective strategy. METHODS: A series of 78 GCs belonging to 74 patients sent to the Genetic Counselling Service were investigated using microsatellite instability, immunohistochemical expression of mismatch repair (MMR) genes, and MLH1 promoter methylation. RESULTS: The presence of microsatellite instability was observed in 67.5% of GCs, and the absence of immunohistochemical expression of at least 1 of the 4 MMR proteins was observed in 71.4% of GCs, showing 96.1% concordance between the methods. Methylation analysis using methylation specific multiplex ligation-dependent probe amplification performed on 35 samples revealed MLH1 promoter hypermethylation in 18 cases (54%). Molecular analysis identified 36 LS carriers of MMR variants (27 pathogenetic and 9 variants of uncertain significance), and, interestingly, 3 LS patients had MLH1 methylated GC.With regard to histological features, LS-related GCs included endocervical cancers and also histological types different from the endometrioid cancers. The presence of peritumoral lymphocytes in GCs was statistically associated with LS tumors. CONCLUSIONS: Somatic analysis is a useful strategy to distinguish sporadic from LS GC. Our data allow the identification of a subset of LS patients otherwise unrecognized on the basis of clinical or family history alone. In addition, our results indicate that some clinicopathological features including age of GC diagnosis; presence of peritumoral lymphocytes; isthmic, endocervical sites, and body mass index value could be useful criteria to select patients for genetic counseling.

Systematic implementation of laparoscopic hysterectomy independent of uterus size: clinical effect.

STUDY OBJECTIVE: To investigate the effect of uterine weight on the mode of hysterectomy and on perioperative outcomes and to explore how the increasing experience in endoscopic techniques influenced our choice of surgical approach to hysterectomy to treat benign conditions. DESIGN: Retrospective analysis (Canadian Task Force classification II-2). SETTING: University-based department of obstetrics and gynecology. PATIENTS: A series of 1518 consecutive women with benign uterine conditions other than pelvic organ prolapse who underwent hysterectomy at our department between January 2000 and December 2011. INTERVENTIONS: Gradual implementation of the laparoscopic approach over years, with the goal of attempting endoscopic hysterectomy whenever possible and irrespective of uterine weight. Comparisons were made on the basis of various approaches to hysterectomy including vaginal hysterectomy (VH), abdominal hysterectomy (AH), and total laparoscopic hysterectomy (TLH) and on uterine weight. MEASUREMENTS AND MAIN RESULTS: Hysterectomies performed included 568 VH (37.4%), 234 AH (15.4%), and 716 TLH (47.2%). Postoperative complications were lower in the TLH group vs the AH group; no significant difference was observed between the VH vs TLH groups or the AH vs VH groups. A marked reduction in the need for open surgery was noted between 2000 and 2011 (p for trend <.001). Restricting the analysis to TLH, an increase in operative time and blood loss was observed, parallel to increasing uterine weight. Hospital stay and rate of intraoperative and postoperative complications were independent of uterine weight. In 45 women with uterus weight ≥1000 g, the initial approach was via laparoscopy, with a success rate of 95.6% (n = 43). A marked tendency toward reduction in the use of open surgery was observed through the years when uterine weight was ≥1 kg (p for trend <.001). CONCLUSION: Systematic implementation of laparoscopic hysterectomy enables a marked reduction in the need for AH. In experienced hands, even very large uteri (≥1 kg) can be safely removed via laparoscopy.

Perioperative allogenic blood transfusions and the risk of endometrial cancer recurrence.

PURPOSE: To evaluate the effect of perioperative blood transfusions on the risk of recurrence of endometrial cancer. METHODS: This study is a retrospective analysis of 358 consecutive patients, without a history of other tumors, who underwent surgery for endometrial cancer between January 2000 and April 2010. RESULTS: Women who did not need any transfusion (N = 331) and patients who received allogenic blood donations (N = 27) were compared in terms of risk of cancer recurrence. The surgical standard procedure included peritoneal washing for cytologic examination, total hysterectomy + bilateral adnexectomy (N = 358), and pelvic lymphadenectomy (N = 227). The two groups were homogeneous in term of age, BMI, previous abdominal surgery, type of intervention, operative time, nodal count, and hospital stay. The median (range) estimated blood loss was higher in the transfusion group, 400 mL (100-2,000 mL), than in the non-transfusion group, 150 mL (10-1,000 mL). Median (range) follow-up was 67.5 months (6-132.4 months). Blood transfusions were associated with a higher relapse rate (P = 0.0021). At multivariate analysis, administration of packed red blood cells remained independently associated with recurrence (OR 4.64; CI 95 % 1.45-14.9), as well as myometrial invasion ≥50 % (OR 2.88; CI 95 % 1.18-7.07) and stage >1 (OR 4.24; CI 95 % 1.75-10.3). CONCLUSIONS: The use of allogenic blood transfusions is associated with a higher risk of recurrence. We hypothesize that this could be due to a transitory perioperative immunodepression that promotes the spread of neoplastic cells.

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.

Radiation-induced bowel complications: laparoscopic versus open staging of gynecologic malignancy.

PURPOSE: To evaluate whether the type of surgical approach used to stage gynecologic malignancies influences the risk of developing nonrectal radiation-induced intestinal injury (NRRIII) in patients who subsequently receive adjuvant radiotherapy. METHODS: A prospectively entered database was queried for all women with either primary or recurrent gynecologic malignancy who underwent external-beam radiation therapy ± brachytherapy and who had prior abdominopelvic surgery at our institution. Univariate and multivariate analysis of variables potentially affecting the risk of developing significant bowel toxicity (defined as grade 2 or more according to Radiation Therapy Oncology Group scoring) were performed. RESULTS: One hundred fifty-nine patients were identified. The site of primary tumor was the cervix in 61 (38%) patients and the corpus uteri in the remaining patients (98, 62%). Treatment was delivered with a combination of external-beam and intracavitary irradiation to 50 (31.4%) patients, and 109 (68.6%) patients received only external-beam irradiation. Staging procedures were performed by open surgery in 93 (58.5%) patients, whereas laparoscopy was the surgical approach of choice in 66 (41.5%) women. Fifteen patients (9.4%) developed grade 2 or greater NRRIII, at median latency of 10 months (range 3-64 months); six were diagnosed as grade 3 complications requiring surgery, and three developed grade 4 complication. Multiple regression revealed an independent protective effect of pretreatment laparoscopic staging against the risk of developing both grade ≥2 and grade ≥3 NRRIII. CONCLUSIONS: Notwithstanding potential limitations of nonrandomized study design, our findings suggest that the benefits of minimal-access surgery used to perform staging procedures may translate into long-term reduction in radiation-induced bowel injury.

Prognostic significance of preoperative plasma fibrinogen in endometrial cancer.

OBJECTIVE: To investigate the prognostic significance of preoperative plasma fibrinogen concentration, with particular focus on tumor dissemination and nodal involvement, in a substantial cohort of patients with endometrial cancer. METHODS: The study population comprised 336 women with endometrial cancer who underwent surgical staging at two tertiary institutions, from 2000 to 2009. Pretreatment plasma samples from the study cohort were assayed for fibrinogen by the Clauss assay. Information on demographics, laboratory testing, histopathology and follow-up was gathered from databases of prospectively collected data. Factors associated with survival were identified in a Cox proportional hazards model. Univariate and multivariate analyses were used to evaluate predictors of extrauterine disease and nodal metastasis. RESULTS: One-hundred-thirty-seven (40.8%) patients exhibited preoperative hyperfibrinogenemia. Univariate analysis demonstrated that histological type, tumor grade, depth of myometrial invasion, surgical stage, patient age, and hyperfibrinogenemia affect disease-free (DFS) and overall survival rates significantly. When these variables were entered simultaneously into a Cox regression model, raised preoperative levels of plasma fibrinogen retained significance as poor prognosticator of DFS (HR 2.0, 95%CI 1.1-3.6) and overall survival (HR 2.7, 95%CI 1.3-5.5). Preoperative hyperfibrinogenemia was an independent determinant of extrauterine disease (OR 2.7, 95%CI 1.3-5.6). In the subcohort of women with endometrioid histology, increased fibrinogen concentration at presentation was predictive of pelvic nodal involvement (OR 3.6, 95%CI 1.1-11.7). CONCLUSION: Plasma fibrinogen level may be of value in the prediction of outcome, improve the stratification of endometrial cancer patient, at diagnosis, based on their risk of recurrence, and possibly alter their treatment accordingly.

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.

Risk factors for cervical intraepithelial neoplasia recurrence after conization: a 10-year study.

OBJECTIVE: To evaluate the risk factors potentially involved in the development of cervical intraepithelial neoplasia (CIN) recurrence after cervical conization in a long-term follow-up period. STUDY DESIGN: Consecutive patients with histologically proven CIN who had undergone either cold knife conization or a loop electrosurgical excision procedure were enrolled and scheduled for serial follow-up examinations over a 10-year period. Data were stored in a digital database. Multivariate analysis was performed to identify factors for recurrence. RESULTS: Between January 1999 and December 2009, 282 patients fulfilled the inclusion criteria and were included in the final statistical analysis. After a median follow-up of 26.7 months (range 6-100), 64 (22.7%) women developed histologically confirmed recurrence. The 2-year recurrence-free survival was 83.7% and 66.7% for women with negative and positive margins, respectively (p=0.008). The 5-year recurrence-free survival was 75.4% and 50.3% for patients with negative and positive margins, respectively (p=0.0004). Positive surgical margin was the most important independent predictor of recurrence [HR 2.5 (95%CI 1.5-4.5), p=0.0007; Wald 11.338]. After multinomial logistic regression the indication for conization based on persistent CIN1 was the only independent predictor for negative margin [OR 0.3 (95%CI 0.1-0.7), p=0.008]. CONCLUSIONS: Our study demonstrated that the surgical margin status represents the most important predictor for CIN recurrence after conization. After excisional therapy, close follow-up is mandatory for the early detection of recurrent disease. The identification of risk factors for recurrence may guide clinical decision-making on expectant management versus re-intervention.

Carcinosarcoma of the uterine cervix: case report and discussion.

BACKGROUND: Cervical carcinosarcomas are rare neoplasms; optimal treatment is unclear. CASE 1: A 42-year-old woman underwent abdominal hysterectomy because of bleeding, anaemia and uterine fibromatosis. Histology showed a homologous carcinosarcoma of the cervix. Laparoscopic re-staging (pelvic lymphadenectomy, bilateral salpingo-oophorectomy) was negative for neoplasia. Adjuvant chemotherapy with ifosfamide and cisplatin was performed. At 48 months of follow-up, the patient is NED. CASE 2: A 74-year-old woman reporting vaginal bleeding, with carcinosarcoma on the cervical biopsy, underwent radical hysterectomy, bilateral salpingo-oophorectomy, pelvic and paraortic lymphadenectomy. Histology confirmed a homologous carcinosarcoma of the cervix, stage IIb. Whole-pelvis irradiation and brachytherapy were carried out. Nine months later, the patient developed systemic recurrence and died of disease. Aggressive primary therapy can result in cure of early-stage cervical carcinosarcomas. Extracervical disease is associated with a poor prognosis.