Generation of an Inhibitory NK Cell Subset by TGF-ß1/IL-15 Polarization.

NK cells have been shown to exhibit inflammatory and immunoregulatory functions in a variety of healthy and diseased settings. In the context of chronic viral infection and cancer, distinct NK cell populations that inhibit adaptive immune responses have been observed. To understand how these cells arise and further characterize their immunosuppressive role, we examined in vitro conditions that could polarize human NK cells into an inhibitory subset. TGF-ß1 has been shown to induce regulatory T cells in vitro and in vivo; we therefore investigated if TGF-ß1 could also induce immunosuppressive NK-like cells. First, we found that TGF-ß1/IL-15, but not IL-15 alone, induced CD103+CD49a+ NK-like cells from peripheral blood NK cells, which expressed markers previously associated with inhibitory CD56+ innate lymphoid cells, including high expression of GITR and CD101. Moreover, supernatant from ascites collected from patients with ovarian carcinoma also induced CD103+CD49a+ NK-like cells in vitro in a TGF-ß-dependent manner. Interestingly, TGF-ß1/IL-15-induced CD103+CD56+ NK-like cells suppressed autologous CD4+ T cells in vitro by reducing absolute number, proliferation, and expression of activation marker CD25. Collectively, these findings provide new insight into how NK cells may acquire an inhibitory phenotype in TGF-ß1-rich environments.

Maximizing cancer prevention through genetic navigation for Lynch syndrome detection in women with newly diagnosed endometrial and nonserous/nonmucinous epithelial ovarian cancer.

BACKGROUND: Despite recommendations for reflex immunohistochemistry (IHC) for mismatch repair (MMR) proteins to identify Lynch syndrome (LS), the uptake of genetic assessment by those who meet referral criteria is low. The authors implemented a comprehensive genetic navigation program to increase the uptake of genetic testing for LS in patients with endometrial cancer (EC) or nonserous/nonmucinous ovarian cancer (OC). METHODS: Participants with newly diagnosed EC or OC were prospectively recruited from 3 cancer centers in Ontario, Canada. Family history questionnaires were used to assess LS-specific family history. Reflex IHC for MMR proteins was performed with the inclusion of clinical directives in pathology reports. A trained genetic navigator initiated a genetic referral on behalf of the treating physician and facilitated genetic referrals to the closest genetics center. RESULTS: A total of 841 participants (642 with EC, 172 with OC, and 27 with synchronous EC/OC) consented to the study; 194 (23%) were MMR-deficient by IHC. Overall, 170 women (20%) were eligible for a genetic assessment for LS: 35 on the basis of their family history alone, 24 on the basis of their family history and IHC, 82 on the basis of IHC alone, and 29 on the basis of clinical discretion. After adjustments for participants who died (n = 6), 149 of 164 patients (91%) completed a genetic assessment, and 111 were offered and completed genetic testing. Thirty-four women (4.0% of the total cohort and 30.6% of those with genetic testing) were diagnosed with LS: 5 with mutL homolog 1 (MLH1), 9 with mutS homolog 2 (MSH2), 15 with mutS homolog 6 (MSH6), and 5 with PMS2. CONCLUSIONS: The introduction of a navigated genetic program resulted in a high rate of genetic assessment (>90%) in patients with gynecologic cancer at risk for LS.

Evaluation of treatment effects in patients with endometrial cancer and POLE mutations: An individual patient data meta-analysis.

BACKGROUND: Endometrial cancers (ECs) with somatic mutations in DNA polymerase epsilon (POLE) are characterized by unfavorable pathological features, which prompt adjuvant treatment. Paradoxically, women with POLE-mutated EC have outstanding clinical outcomes, and this raises concerns of overtreatment. The authors investigated whether favorable outcomes were independent of treatment. METHODS: A PubMed search for POLE and endometrial was restricted to articles published between March 1, 2012, and March 1, 2018, that provided individual patient data (IPD), adjuvant treatment, and survival. Following the Preferred Reporting Items for Systematic Review and Meta-Analysis (PRISMA) reporting guidelines for IPD, the authors used univariate and multivariate one-stage meta-analyses with mixed effects Cox models (random effects for study cohorts) to infer the associations of treatment, traditional prognostic factors, and outcome, which was defined as the time from first diagnosis to any adverse event (progression/recurrence or death from EC). RESULTS: Three hundred fifty-nine women with POLE-mutated EC were identified; 294 (82%) had pathogenic mutations. Worse outcomes were demonstrated in patients with nonpathogenic POLE mutations (hazard ratio, 3.42; 95% confidence interval, 1.47-7.58; log-rank P < .01). Except for stage (P < .01), traditional prognosticators were not associated with progression/recurrence or death from disease. Adverse events were rare (11 progressions/recurrences and 3 disease-specific deaths). Salvage rates in patients who experienced recurrence were high and sustained, with 8 of 11 alive without evidence of disease (range, 5.5-14.2 years). Adjuvant treatment was not associated with outcome. CONCLUSIONS: Clinical outcomes for ECs with pathogenic POLE mutations are not associated with most traditional risk parameters, and patients do not appear to benefit from adjuvant therapy. The observed low rates of recurrence/progression and the high and sustained salvage rates raise the possibility of safely de-escalating treatment for these patients. LAY SUMMARY: Ten percent of all endometrial cancers have mutations in the DNA repair gene DNA polymerase epsilon (POLE). Women who have endometrial cancers with true POLE mutations experience almost no recurrences or deaths from their cancer even when their tumors appear to have very unfavorable characteristics. Additional therapy (radiation and chemotherapy) does not appear to improve outcomes for women with POLE-mutated endometrial cancer, and this supports the move to less therapy and less associated toxicity. Diligent classification of endometrial cancers by molecular features provides valuable information to inform prognosis and to direct treatment/no treatment.

Understanding the clinical implication of mismatch repair deficiency in endometrioid endometrial cancer through a prospective study.

OBJECTIVES: Findings on impact of mismatch repair deficiency (MMRd) on patient outcomes in endometrial cancer (EC) have been inconsistent to date. The objective of this study was to compare the oncologic outcomes and recurrence patterns between MMRd and MMR-intact (MMRi) endometrioid EC (EEC). METHODS: Between 2015 and 2018, we prospectively recruited 492 EEC cases from three cancer centers in Ontario, Canada. Tumors were reflexively assessed for MMR protein expression by immunohistochemistry (IHC). Clinicopathological, survival and recurrence patterns were compared between MMRd and MMRi cases. RESULTS: Of 492 EEC, 348 were MMRi (71%) and 144 were MMRd (29%) with median follow-up of 16.8 months (0-69.6). MMRd tumors tended to be grade 2 or 3 (56% vs. 29%, p < 0.001), with propensity for lymphovascular space invasion (28% vs. 18%, p = 0.024), lymph node involvement (7% vs. 5%, p < 0.001) and received more adjuvant treatment (46% vs. 33%, p = 0.027). This group also had significantly lower 3-year recurrence-free survival (78% vs. 90%, p = 0.014) although there was no difference in OS (p = 0.603). MMRd cases were more likely to recur in retroperitoneal lymph nodes (p = 0.045). Upon subgroup analysis, MLH1 methylated tumors had the worst prognostic features and survival outcomes. CONCLUSIONS: MLH1 methylated EECs exhibit more aggressive features compared to other MMRd and MMRi EECs. This may indicate an inherent difference in tumor biology, suggesting the importance of individualized management based on EC molecular phenotype.

Equivalent Survival of p53 Mutated Endometrial Endometrioid Carcinoma Grade 3 and Endometrial Serous Carcinoma.

TP53 status is the most important prognostic biomarker in endometrial carcinoma. We asked the question whether p53 mutated endometrial endometrioid carcinomas grade 3 (EEC3) or endometrial serous carcinomas (ESC), the latter ubiquitously harboring TP53 mutation, have different outcomes. TP53 mutation status was assessed by surrogate p53 immunohistochemistry on 326 EEC3 and ESC from 2 major cancer centers in Canada. Mutant-type p53 expression, including overexpression, complete absence, or cytoplasmic expression, was distinguished from the wild-type pattern. Statistical associations with clinico-pathological parameter, other key biomarkers, and survival analyses were performed. P53 mutant-type immunohistochemistry was observed in all 126 ESC and in 47/200 (23.5%) EEC3. ESC and p53 mutated EEC3 had an unfavorable outcome compared with p53 wild-type EEC3 (hazard ratio=2.37, 95% confidence interval=1.48-3.80, P=0.003, hazard ratio=2.19, 95% confidence interval=1.16-4.12, P=0.016, respectively) in multivariable analyses adjusted for age, stage, center, and presence of lymph-vascular invasion. There was no significant difference in survival between ESC and p53 mutated EEC3 in multivariable analysis. Furthermore, p53 mutated EEC3 and ESC almost completely overlapped in univariate survival analysis when mismatch repair (MMR)-deficient cases were excluded, which suggests that EEC3 harboring combined MMR deficiency and TP53 mutations behave more according to the MMR status. Significant differences between p53 mutated MMR-proficient EEC3 and ESC in PTEN and p16 expression status remained. p53 mutated, MMR-proficient EEC3 and ESC have overlapping survival significantly different from p53 wild-type EEC3, which justifies a similar treatment with current non-targeted standard therapy. Although this is so, separate classification should continue due to biological differences that will become important for future targeted therapy.

The Value of DICER1 Mutation Analysis in "Subtle" Diagnostically Challenging Embryonal Rhabdomyosarcomas of the Uterine Cervix.

Embryonal rhabdomyosarcoma of the uterine cervix is a rare neoplasm which is almost invariably associated with pathogenic somatic or germline DICER1 mutations; patients with germline mutations have DICER1 syndrome. We report 2 subtle cervical embryonal rhabdomyosarcoma, one occurring in a 21-yr-old woman with a known history of DICER1 syndrome and the other in a 19-yr-old woman with no history of DICER1 syndrome or DICER1-associated neoplasms. Both neoplasms focally involved otherwise benign endocervical polyps and were characterized histologically by subtle areas of increased stromal cellularity, nuclear atypia and mitotic activity; there was focal nuclear staining of these areas with the skeletal muscle markers myogenin and myoD1. In both cases, demonstration of a somatic DICER1 RNase IIIb mutation in the tumor was instrumental in establishing the diagnosis. We believe these neoplasms represent the earliest discernible phase of cervical embryonal rhabdomyosarcoma. Pathologists should have a high index of suspicion when atypical stromal elements are present in endocervical polyps and immunohistochemistry together with DICER1 sequencing will assist in diagnosis.

Performance characteristics of screening strategies to identify Lynch syndrome in women with ovarian cancer.

BACKGROUND: For women with ovarian cancer (OC), the optimal screening strategy to identify Lynch syndrome (LS) has not been determined. In the current study, the authors compared the performance characteristics of various strategies combining mismatch repair (MMR) immunohistochemistry (IHC), microsatellite instability testing (MSI), and family history for the detection of LS. METHODS: Women with nonserous and/or nonmucinous ovarian cancer were recruited prospectively from 3 cancer centers in Ontario, Canada. All underwent germline testing for LS and completed a family history assessment. Tumors were assessed using MMR IHC and MSI. The sensitivity, specificity, and positive and negative predictive values of screening strategies were compared with the gold standard of a germline result. RESULTS: Of 215 women, germline data were available for 189 (88%); 13 women (7%) had pathogenic germline variants with 7 women with mutS homolog 6 (MSH6); 3 women with mutL homolog 1 (MLH1); 2 women with PMS1 homolog 2, mismatch repair system component (PMS2); and 1 woman with mutS homolog 2 (MSH2). A total of 28 women had MMR-deficient tumors (13%); of these, 11 had pathogenic variants (39%). Sequential IHC (with MLH1 promoter methylation analysis on MLH1-deficient tumors) followed by MSI for nonmethylated and/or MMR-intact patients was the most sensitive (92.3%; 95% confidence interval, 64%-99.8%) and specific (97.7%; 95% confidence interval, 94.2%-99.4%) approach, missing 1 case of LS. IHC with MLH1 promoter methylation analysis missed 2 patients of LS. Family history was found to have the lowest sensitivity at 55%. CONCLUSIONS: Sequential IHC (with MLH1 promoter methylation analysis) followed by MSI was found to be most sensitive. However, IHC with MLH1 promoter methylation analysis also performed well and is likely more cost-effective and efficient in the clinical setting. The pretest probability of LS is high in patients with MMR deficiency and warrants universal screening for LS.

Significantly greater prevalence of DICER1 alterations in uterine embryonal rhabdomyosarcoma compared to adenosarcoma.

Embryonal rhabdomyosarcomas (ERMS) account for 2-3% of cancers in pediatric and adolescent populations. They are rarer in adults. We and others have reported that ERMS arising in the uterine cervix may harbor mutations in the gene encoding the microRNA biogenesis enzyme, DICER1, but a large series of cases has not been published. In the uterus, distinguishing ERMS from adenosarcoma can be very challenging, even for expert pathologists, and DICER1 alterations have been identified in a variable subset of uterine adenosarcomas. We hypothesized that DICER1 genetic testing may be useful in distinguishing between ERMS and adenosarcoma. We conducted a central pathology review-based study of 64 tumors initially thought to be uterine ERMS or adenosarcoma; 19 neoplasms had a consensus diagnosis of ERMS, 27 of adenosarcoma and for 18, no consensus diagnosis was reached. The median age at diagnosis was 30 years (range 2.5-69) for ERMS, 57.5 years (range 27-82) for adenosarcoma, and 65.5 years (range 32-86) for no consensus cases. In our series, the DICER1 mutation prevalence differed between the three groups: DICER1 alterations were present in 18/19 (95%) ERMS, 7/27 (26%) adenosarcomas (p < 0.001), and 4/18 (22%) no consensus cases. A germline alteration was present in 6/12 ERMS patients tested versus 0/6 adenosarcoma patients. Thus, although DICER1 mutations are near ubiquitous in uterine ERMS and are significantly less common in uterine adenosarcoma, DICER1 testing is only of value in distinguishing between the two neoplasms when a DICER1 mutation is absent, as this is helpful in excluding ERMS. On review of the clinical and radiological features of the single DICER1 wild-type cervical ERMS, this was thought most likely to be of vaginal origin. Given the significant prevalence of DICER1 germline pathogenic variants in uterine ERMS, all patients with this diagnosis should be referred to a genetics service.

Interlaboratory Concordance of ProMisE Molecular Classification of Endometrial Carcinoma Based on Endometrial Biopsy Specimens.

Molecular classifiers improve the consistency of categorization of endometrial carcinoma and provide valuable prognostic information. We aimed to evaluate the interlaboratory agreement in ProMisE assignment across 3 dedicated Canadian gynecologic oncology centers. Fifty cases of endometrial carcinoma diagnosed on biopsy were collected from 3 centers and 3 unstained sections were provided to each participating site so that immunohistochemistry for MSH6, PMS2, and p53 could be performed and interpreted at each center, blinded to the original diagnoses and the results from other centers. A core was taken for DNA extraction and POLE mutation testing. Overall accuracy and κ statistic were assessed. MSH6, PMS2, and p53 could be assessed for all 50 cases, with agreement for 140/150 results. There was a high level of agreement in molecular classification (κ=0.82), overall. Cases with a discordant result for one of the features used in classification (n=10) were reviewed independently and the most common reason for disagreement was attributable to the weak p53 staining in 1 laboratory (n=4). Interpretive error in PMS2 (n=1) and MSH6 (n=2) assessment accounted for 3 of the remaining disagreements. Interpretive error in the assessment of p53 was identified in 2 cases, with very faint p53 nuclear reactivity being misinterpreted as wild-type staining. These results show strong interlaboratory agreement and the potential for greater agreement if technical and interpretive factors are addressed. Several solutions could improve concordance: central quality control to ensure technical consistency in immunohistochemical staining, education to decrease interpretation errors, and the use of secondary molecular testing.

HPV-independent Vulvar Squamous Cell Carcinoma is Associated With Significantly Worse Prognosis Compared With HPV-associated Tumors.

Vulvar squamous cell carcinomas (VSCC) represent the most common carcinoma of the female external genitalia, with increasing incidence. Although high-risk human papillomavirus (HPV) infection has long been implicated in the majority of cervical and anal squamous cell carcinomas, there is uncertainty about its prevalence and prognostic impact in VSCC. In this study, we conducted a retrospective integrated morphologic and multimodal HPV analysis of a cohort of 114 VSCC cases treated at the Princess Margaret Cancer Centre/University Health Network, Toronto, Canada between 2000 and 2010. VSCC histology was reviewed. We analyzed the cohort for HPV using polymerase chain reaction based method, and tissue microarray DNA and RNA in situ hybridization (ISH), and p16 immunohistochemistry. Among the 114 cases (age 70±16 yr), 36.7% of cases were classified as having histomorphology of HPV infection. HPV was detected in 31.9% (polymerase chain reaction), 14.0% (DNA ISH), and 27.3% (RNA ISH) of cases. p16 immunohistochemistry was positive in 37.8% of cases. On univariate analysis, HPV morphology (P=0.009), p16+ (P=0.00013), DNA ISH+ (P=0.021), and RNA ISH+ (P=0.00061) were associated with better 5-yr progression-free survival. DNA ISH+ (P=0.049) was associated with better 5-yr overall survival. On multivariate analysis, HPV morphology (P=0.033), p16+ (P=0.01), and RNA ISH+ (P=0.035) were associated with better 5-yr progression-free survival. In conclusion, a subset of VSCC is associated with HPV, which correlates with better outcome. Relatively inexpensive tests such as histomorphologic evaluation, p16 immunohistochemistry, and HPV RNA ISH can be used to predict outcome in VSCC. Therefore, routine reporting of HPV status in VSCC is recommended.

Expanding the morphological spectrum of ovarian microcystic stromal tumour.

AIMS: To expand the morphological spectrum of ovarian microcystic stromal tumour, a rare neoplasm considered to have a relatively constant morphology with microcysts, solid cellular regions and hyalinised fibrous stroma. METHODS AND RESULTS: We report four ovarian neoplasms in patients aged 45, 56, 61 and 71 years with the characteristic immunophenotype of microcystic stromal tumour (diffuse nuclear positivity with beta-catenin, cyclin D1 and WT1; diffuse cytoplasmic positivity with CD10; negative inhibin, calretinin, oestrogen receptor and progesterone receptor). The tumours had variant morphology (diffuse, nested and corded arrangements in three cases, including one with spindle cell elements; nested, corded and tubular in the other). A CTNNB1 point mutation in exon 3 (c.98C>G,p.S33C; c.100G>A,p.G34R; c.97T>G,p.S33A) was present in the three cases with material available for testing. CONCLUSIONS: We feel that the cases we report are related to microcystic stromal tumour but with variant morphology; as such, the morphological spectrum of ovarian microcystic stromal tumour is broader than hitherto reported.

Clinical, morphological and immunohistochemical evidence that small-cell carcinoma of the ovary of hypercalcaemic type (SCCOHT) may be a primitive germ-cell neoplasm.

AIMS: The histogenesis and cell lineage of small-cell carcinoma of the ovary of hypercalcaemic type (SCCOHT) is unknown. We aim to provide evidence that this may be a primitive germ-cell neoplasm arising from a teratoma. METHODS AND RESULTS: Following the identification of two cases of SCCOHT associated with germ-cell tumours (one dermoid cyst, one immature teratoma with a focus of yolk sac tumour), we undertook a literature review to look for any prior reports of SCCOHT in association with other neoplasms or elements. This revealed two cases associated with immature teratomas, one arising in an ovary where a cystectomy had been undertaken previously for a teratoma and another in association with a mucinous borderline tumour. Mucinous elements have also been reported in SCCOHT, this type of epithelium potentially being of teratomatous derivation. We stained whole tissue sections of nine cases of SCCOHT and a tissue microarray (TMA) containing 34 different SCCOHT with germ-cell markers SALL4, OCT3/4, alpha fetoprotein (AFP) and glypican 3. All except one of the whole tissue sections and approximately half the TMA cases were positive with SALL4, while all cases were OCT3/4-, AFP- and glypican 3-negative, except for focal glypican 3 staining in an occasional case. CONCLUSIONS: Our findings provide additional evidence to that proposed by others that SCCOHT is a primitive germ-cell neoplasm arising from a teratoma.

Ovarian hilar proliferations resembling Sertoli cell tumours: microscopic neoplasms or non-neoplastic remnants?

AIMS: To describe four cases of incidentally detected microscopic ovarian hilar Sertoli cell proliferation, a phenomenon that, as far as we are aware, has not been previously reported, and to discuss the differential diagnosis of these lesions. METHODS AND RESULTS: The proliferations occurred in women aged 35-62 years, and were composed of solid and hollow tubules that were morphologically and immunohistochemically in keeping with Sertoli cells. Two cases were bilateral. CONCLUSIONS: We feel that these uncommon proliferations are likely to represent non-neoplastic embryological remnants rather than hilar Sertoli cell tumours.

Loss of SMARCA4 (BRG1) protein expression as determined by immunohistochemistry in small-cell carcinoma of the ovary, hypercalcaemic type distinguishes these tumours from their mimics.

AIMS: Molecular investigation of small-cell carcinoma of the ovary, hypercalcaemic type (SCCOHT) has revealed that it is a monogenetic tumour characterized by alteration of SMARCA4 (BRG1), encoding a member of the switch/sucrose non-fermentable (SWI/SNF) chromatin remodelling complex. A large majority of cases show loss of expression of the corresponding SMARCA4/BRG1 protein. Furthermore, three cases of SCCOHT with retained SMARCA4 protein expression showed loss of SMARCB1/INI1 expression. The aim of this study was to assess the sensitivity and specificity of loss of SMARCA4 expression as a diagnostic test for SCCOHT. METHODS AND RESULTS: We performed SMARCA4 and SMARCB1 staining in 245 tumours, many of which were potentially in the differential diagnosis of SCCOHT. We also stained 56 cases of SCCOHT for SMARCA4 and 37 of these for SMARCB1. Fifty-four of the SCCOHT cases showed complete absence of SMARCA4 expression. The two cases with retained expression showed molecular alteration of SMARCA4. Of the 217 other neoplasms with interpretable staining, all retained SMARCA4 expression. Although the majority showed diffuse, strong nuclear expression, a heterogeneous, typically weak staining pattern was present in 13% of cases. All 37 cases of SCCOHT tested and all other neoplasms, apart from three malignant rhabdoid tumours, showed retained nuclear SMARCB1 expression. Loss of SMARCA4 expression had a sensitivity of 96.55% and specificity of 100%. CONCLUSIONS: Loss of SMARCA4 expression is sensitive and specific for SCCOHT. Although some mimics show heterogeneous expression, there is retention of nuclear staining in at least a part of the tumour; therefore, only complete loss of staining should be regarded as being supportive of SCCOHT.

Progesterone receptor expression is associated with longer overall survival within high-grade histotypes of endometrial carcinoma: A Canadian high risk endometrial cancer consortium (CHREC) study.

OBJECTIVE: To assess the association of hormone receptor expression with outcome in high-grade endometrial carcinomas. METHODS: This study included three sites participating in the Canadian High Risk Endometrial Cancer (CHREC) consortium. Sections from tissue microarrays containing cases with a diagnosis of endometrioid grade 3 (EC3) and endometrial serous carcinoma (ESC) were assessed for estrogen (ER) and progesterone receptor (PR) expression by immunohistochemistry. Expression was considered present if >1% of tumor cell nuclei were labeled. Associations with overall survival were assessed. RESULTS: ER expression was present in 168/216 (78%) of EC3 and 124/192 (65%) of ESC. PR expression was present in 148/212 (70%) of EC3 and 83/196 (42%) of ESC. PR expression was significantly associated with favorable overall survival in EC3 and ESC (log rank, p=0.018 and p=0.0024) but ER expression was not. PR expression was significantly associated with favorable overall survival in EC3 independent of age, stage, center and lymph-vascular invasion (hazard ratio=0.457, 95% CI 0.257-0.811, p=0.0075) as well as in stage I and II ESC (hazard ratio=0.266, 95% CI 0.094-0.750, p=0.0123). CONCLUSION: Our data provide support for the assessment of the PR expression status in EC3 and ESC. Future work will be required to determine how PR expression may be incorporated into management of patients with EC3 and ESC.

Recurrent somatic DICER1 mutations in nonepithelial ovarian cancers.

BACKGROUND: Germline truncating mutations in DICER1, an endoribonuclease in the RNase III family that is essential for processing microRNAs, have been observed in families with the pleuropulmonary blastoma-family tumor and dysplasia syndrome. Mutation carriers are at risk for nonepithelial ovarian tumors, notably sex cord-stromal tumors. METHODS: We sequenced the whole transcriptomes or exomes of 14 nonepithelial ovarian tumors and noted closely clustered mutations in the region of DICER1 encoding the RNase IIIb domain of DICER1 in four samples. We then sequenced this region of DICER1 in additional ovarian tumors and in certain other tumors and queried the effect of the mutations on the enzymatic activity of DICER1 using in vitro RNA cleavage assays. RESULTS: DICER1 mutations in the RNase IIIb domain were found in 30 of 102 nonepithelial ovarian tumors (29%), predominantly in Sertoli-Leydig cell tumors (26 of 43, or 60%), including 4 tumors with additional germline DICER1 mutations. These mutations were restricted to codons encoding metal-binding sites within the RNase IIIb catalytic centers, which are critical for microRNA interaction and cleavage, and were somatic in all 16 samples in which germline DNA was available for testing. We also detected mutations in 1 of 14 nonseminomatous testicular germ-cell tumors, in 2 of 5 embryonal rhabdomyosarcomas, and in 1 of 266 epithelial ovarian and endometrial carcinomas. The mutant DICER1 proteins had reduced RNase IIIb activity but retained RNase IIIa activity. CONCLUSIONS: Somatic missense mutations affecting the RNase IIIb domain of DICER1 are common in nonepithelial ovarian tumors. These mutations do not obliterate DICER1 function but alter it in specific cell types, a novel mechanism through which perturbation of microRNA processing may be oncogenic. (Funded by the Terry Fox Research Institute and others.).

Data set for reporting of ovary, fallopian tube and primary peritoneal carcinoma: recommendations from the International Collaboration on Cancer Reporting (ICCR).

A comprehensive pathological report is essential for optimal patient management, cancer staging and prognostication. In many countries, proforma reports are used but these vary in their content. The International Collaboration on Cancer Reporting (ICCR) is an alliance formed by the Royal College of Pathologists of Australasia, the Royal College of Pathologists of the United Kingdom, the College of American Pathologists, the Canadian Partnership Against Cancer and the European Society of Pathology, with the aim of developing an evidence-based reporting data set for each cancer site. This will reduce the global burden of cancer data set development and reduplication of effort by different international institutions that commission, publish and maintain standardised cancer reporting data sets. The resultant standardisation of cancer reporting will benefit not only those countries directly involved in the collaboration but also others not in a position to develop their own data sets. We describe the development of a cancer data set by the ICCR expert panel for the reporting of primary ovarian, fallopian tube and peritoneal carcinoma and present the 'required' and 'recommended' elements to be included in the report with an explanatory commentary. This data set encompasses the recent International Federation of Obstetricians and Gynaecologists staging system for these neoplasms and the updated World Health Organisation Classification of Tumours of the Female Reproductive Organs. The data set also addresses issues about site assignment of the primary tumour in high-grade serous carcinomas and proposes a scoring system for the assessment of tumour response to neoadjuvant chemotherapy. The widespread implementation of this data set will facilitate consistent and accurate data collection, comparison of epidemiological and pathological parameters between different populations, facilitate research and hopefully will result in improved patient management.

Microscopic extraovarian sex cord proliferations: an undescribed phenomenon.

AIMS: To report a previously undescribed phenomenon of incidentally detected microscopic proliferations of sex cord cells, often mimicking adult granulosa cell tumour or sex cord tumour with annular tubules, in extraovarian locations. METHODS AND RESULTS: The six cases were in patients aged 23-58 years. The proliferations were located in the fallopian tube in three cases, and in paraovarian connective tissues, the pelvic side wall, and appendiceal serosa (one case each). Microscopically, they were typically composed of well-demarcated nests of regular cells with round/ovoid vesicular nuclei, some containing grooves. Microfollicular and/or cribriform arrangements were present in three cases. In five cases, the sex cord lineage was confirmed by positive staining with inhibin and/or calretinin and other sex cord markers. FOXL2 mutation analysis was performed in one case, but was inconclusive. Bilateral oophorectomies and bilateral cystectomies were performed in three cases and one case respectively; there was no sex cord-stromal neoplasm in the removed ovaries. In the two cases in which the ovaries were not removed, imaging showed no suspicious features. Follow-up in four cases (11 months-6 years) has been uneventful. CONCLUSIONS: The pathogenesis of these microscopic extraovarian sex cord proliferations is unknown, but they may represent non-neoplastic proliferations of embryonic remnants.

Chromosomal instability as a prognostic marker in cervical cancer.

BACKGROUND: Cervical cancer is the third most common cancer in women globally, and despite treatment, distant metastasis and nodal recurrence will still develop in approximately 30% of patients. The ability to predict which patients are likely to experience distant relapse would allow clinicians to better tailor treatment. Previous studies have investigated the role of chromosomal instability (CIN) in cancer, which can promote tumour initiation and growth; a hallmark of human malignancies. In this study, we sought to examine the published CIN70 gene signature in a cohort of cervical cancer patients treated at the Princess Margaret (PM) Cancer Centre and an independent cohort of The Cancer Genome Atlas (TCGA) cervical cancer patients, to determine if this CIN signature associated with patient outcome. METHODS: Cervical cancer samples were collected from 79 patients, treated between 2000-2007 at the PM, prior to undergoing curative chemo-radiation. Total RNA was extracted from each patient sample and analyzed using the GeneChip Human Genome U133 Plus 2.0 array (Affymetrix). RESULTS: High CIN70 scores were significantly related to increased chromosomal alterations in TCGA cervical cancer patients, including a higher percentage of genome altered and a higher number of copy number alterations. In addition, this same CIN70 signature was shown to be predictive of para-aortic nodal relapse in the PM Cancer Centre cohort. CONCLUSIONS: These findings demonstrate that chromosomal instability plays an important role in cervical cancer, and is significantly associated with patient outcome. For the first time, this CIN70 gene signature provided prognostic value for patients with cervical cancer.

A phase II study of single-agent RO4929097, a gamma-secretase inhibitor of Notch signaling, in patients with recurrent platinum-resistant epithelial ovarian cancer: A study of the Princess Margaret, Chicago and California phase II consortia.

PURPOSE: A phase II study was performed to evaluate the efficacy and safety of single-agent RO4929097 (a gamma-secretase inhibitor) in patients with recurrent platinum-resistant ovarian cancer. EXPERIMENTAL DESIGN: Women with progressive platinum-resistant ovarian cancer treated with ≤2 chemotherapy regimens for recurrent disease were enrolled in this trial. Patients received oral RO4929097 at 20 mg once daily, 3 days on/4 days off each week in a three week cycle. The primary endpoint was progression-free survival (PFS) rate at the end of 4 cycles. Secondary objectives included assessment of the safety of RO4929097 and exploration of molecular correlates of outcome in archival tumor tissue and serum. RESULTS: Of 45 patients enrolled, 40 were evaluable for response. Thirty-seven (82%) patients had high-grade ovarian cancer. No objective responses were observed. Fifteen patients (33%) had stable disease as their best response, with a median duration of 3.1 months. The median PFS for the whole group was 1.3 months (1.2-2.5). Treatment was generally well tolerated with 10% of patients discontinuing treatment due to an adverse event. In high grade serous ovarian cancer patients, the median PFS trended higher when the expression of intracellular Notch (NICD) protein by immunohistochemistry was high versus low (3.3 versus 1.3 months, p=0.09). No clear relationship between circulating angiogenic factors and PFS was found despite a suggestion of an improved outcome with higher baseline VEGFA levels. CONCLUSIONS: RO4929097 has insufficient activity as a single-agent in platinum-resistant ovarian cancer to warrant further study as monotherapy. Future studies are needed to explore the potential for cohort enrichment using NICD expression.