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High‐grade neuroendocrine carcinoma of gynecologic origin (NEC‐GYN) is a highly aggressive cancer that often affects young women. The clinical management of NEC‐GYN is typically extrapolated from its counterpart, small cell carcinoma of the lung (SCLC), but, unfortunately, available therapies have limited benefit. In our NEC‐GYN cohort, median progression‐free survival (PFS) and overall survival (OS) were 1 and 12 months, respectively, indicating the highly lethal nature of this cancer. Our comprehensive genomic analyses unveiled that NEC‐GYN harbors a higher mutational burden with distinct mutational landscapes from SCLC. We identified 14 cancer driver genes, including the most frequently altered KMT2C (100%), KNL1 (100%), NCOR2 (100%), and CCDC6 (93%) genes. Transcriptomic analysis identified several novel gene fusions; astonishingly, the MALAT1 lincRNA gene was found in ˜ 20% of all fusion events in NEC‐GYN. Furthermore, NEC‐GYN exhibited a highly immunosuppressive state, intact RB1 expression, and was uniquely enriched with the YAP1 high molecular subtype. Our study identifies several potential therapeutic targets and suggests an urgent need to re‐evaluate the treatment options for NEC‐GYN., The neuroendocrine carcinoma of gynecologic origin (NEC‐GYN) is a deadly cancer that often affects young women. Treatment of NEC‐GYN is typically extrapolated from small cell lung cancer (SCLC), but, unfortunately, available treatments are inadequate. We report that NEC‐GYN is genomically distinct from SCLC, identify several potential therapeutic targets, and suggest an urgent need to re‐evaluate the treatment options for NEC‐GYN.

Poly-ADP Ribose Polymerase (PARP) targeted therapy is clinically approved for the treatment of homologous recombination (HR) repair deficient tumors. The remarkable success of this therapy in the treatment of HR repair deficient cancers has not translated to HR-proficient cancers. Our studies identify the novel role of non-receptor lymphocyte-specific protein tyrosine kinase (LCK) in the regulation of HR repair in endometrioid epithelial ovarian cancer (eEOC) model. We show that DNA damage leads to direct interaction of LCK with the HR repair proteins RAD51 and BRCA1 in a kinase dependent manner RAD51 and BRCA1 stabilization. LCK expression is induced and activated in the nucleus in response to DNA damage insult. Disruption of LCK expression attenuates RAD51, BRCA1, and BRCA2 protein expression by hampering there stability and results in inhibition of HR-mediated DNA repair including suppression of RAD51 foci formation, and augmentation of gH2AX foci formation. In contrast LCK overexpression leads to increased RAD51 and BRCA1 expression with a concomitant increase in HR DNA damage repair. Importantly, attenuation of LCK sensitizes HR-proficient eEOC cells to PARP inhibitor in cells and pre-clinical mouse studies. Collectively, our findings identify a novel therapeutic strategy to expand the utility of PARP targeted therapy in HR proficient ovarian cancer.

Das neuroendokrine Karzinom der Cervix uteri ist eine seltene und aggressive Form von Gebärmutterhalskrebs, die durch häufige Metastasierung schon bei Diagnosestellung und durch hohe Rezidivraten gekennzeichnet ist. Die Primärtherapie ist multimodal und beinhaltet oft eine Chemotherapie, zum Teil in Verbindung mit einer Strahlentherapie. Es liegen keine Daten vor, die Orientierung für das Vorgehen bei Rezidiven geben; Zweitlinientherapien werden aus Daten zum kleinzelligen Bronchialkarzinom abgeleitet. Die engmaschige Überwachung dieser Patientinnen im Hinblick auf Rezidive ist von größter Bedeutung. Dafür stellt die Bestimmung der zirkulierenden Tumor-DNA (ctDNA) im peripheren Blut eine attraktive, weil nicht-invasive Option dar. Mit dem ULP-WGS-Verfahren (ultra-low-pass whole-genome sequencing) lassen sich die Tumorlast und das Tumoransprechen beurteilen und Prognosen zur Rezidivierung abgeben; allerdings mangelt es bisher noch an Daten zur Rolle der ULP-WGS beim kleinzelligen Zervixkarzinom. Die vorliegende Arbeit stellt eine Patientin vor, deren Chemotherapie-Ansprechen und Tumor-Rezidivierung durch die ctDNA-Analyse korrekt überwacht wurden, wie radiologische Befunde bestätigten.

FIGO grade 3 endometrioid endometrial carcinoma (EEC) is a heterogenous group of tumors with variable molecular and clinicopathologic characteristics but is treated clinically as a single entity. There is a need for additional objective markers to help guide management. The aim of this study was to evaluate a cohort of FIGO grade 3 EEC to validate the prognostic impact of molecular classification using POLE mutation (POLE-mut) analysis and immunohistochemistry for p53 and mismatch repair proteins. A secondary aim was to assess for any morphologic or immunophenotypic correlates among the molecular groups. Ninety-five cases of FIGO grade 3 EEC who underwent a hysterectomy at our institution were identified. Ten tumors (11%) harbored POLE-mut, 35 tumors (37%) showed mismatch repair deficiency, 18 tumors (19%) showed aberrant p53 staining (p53-ab), and 26 cases (27%) lacked all of these findings and were classified as no specific molecular profile. Six separate cases harbored >1 abnormality (multiple classifier), 5 of which had POLE-mut. The POLE-mut group and multiple classifier group showed excellent clinical outcomes, the p53-ab group showed the worst clinical outcomes and the 2 remaining groups showed intermediate prognosis. While the POLE-mut tumors showed a statistically significant enrichment for morphologic features including serous-like atypia and lymphocytic infiltrates, these findings were seen across all 4 molecular groups. There was no correlation between molecular grouping and tumor immunophenotypic findings, but overall 18% and 24% of tumors were completely negative for PAX-8 and estrogen receptor, respectively. Five CTNNB1 mutations were identified, 3 of which occurred in the context of a POLE-mut (including 1 multiple classifier case with MLH1/PMS2 loss). Thus our study corroborates the prognostic impact of molecular classification of high-grade endometrioid carcinoma of the uterus, achieved by readily available immunohistochemical stains in addition to POLE-mut analysis.

BackgroundWe present a computational approach (ArcTIL) for quantitative characterization of the architecture of tumor-infiltrating lymphocytes (TILs) and their interplay with cancer cells from digitized H&E-stained histology whole slide images and evaluate its prognostic role in three different gynecological cancer (GC) types and across three different treatment types (platinum, radiation and immunotherapy).MethodsIn this retrospective study, we included 926 patients with GC diagnosed with ovarian cancer (OC), cervical cancer, and endometrial cancer with available digitized diagnostic histology slides and survival outcome information. ArcTIL features quantifying architecture and spatial interplay between immune cells and the rest of nucleated cells (mostly comprised cancer cells) were extracted from the cell cluster graphs of nuclei within the tumor epithelial nests, surrounding stroma and invasive tumor front compartments on H&E-stained slides. A Cox proportional hazards model, incorporating ArcTIL features was fit on the OC training cohort (N=51), yielding an ArcTIL signature. A unique threshold learned from the training set stratified the patients into a low and high-risk group.ResultsThe seven feature ArcTIL classifier was found to significantly correlate with overall survival in chemotherapy and radiotherapy-treated validation cohorts and progression-free survival in an immunotherapy-treated validation cohort. ArcTIL features relating to increased density of TILs in the epithelium and invasive tumor front were found to be associated with better survival outcomes when compared with those patients with an increased TIL density in the stroma. A statistically significant association was found between the ArcTIL signature and signaling pathways for blood vessel morphogenesis, vasculature development, regulation of cell differentiation, cell-substrate adhesion, biological adhesion, regulation of vasculature development, and angiogenesis.ConclusionsThis study reveals that computationally-derived features from the spatial architecture of TILs and tumor cells are prognostic in GCs treated with chemotherapy, radiotherapy, and checkpoint blockade and are closely associated with central biological processes that impact tumor progression. These findings could aid in identifying therapy-refractory patients and further enable personalized treatment decision-making.

Aims: We investigated immunogenomic signatures and correlated them with survival in ovarian cancer (OV) and endometrial cancer (EC). Materials & method: We used whole transcriptome sequencing data from uterine serous cancer and The Cancer Genome Atlas data of OV and EC (n = 719). Gene expression score was calculated. Population abundance of immune cells were estimated. Results: TGF-β, myeloid cells, IFN-γ, T cells, B cells and endothelial cells predicted overall survival. Whereas CD47, neutrophils and endothelial cells predicted progression-free survival. In multivariate analyses, TGF-β, CD47 and monocytic cells predicted survival in high levels of microsatellite instability (MSI-H) EC whereas high IFN-γ trended toward improved survival in the MSI-S EC. High IFN-γ/low TGF-β and high IFN-γ/low CD47 signatures predicted longer overall survival. Low TGF-β/low CD47 signature predicted longer overall survival only in the MSI-H EC. Conclusion: Our data support the role of immune markers in predicting survival in OV/EC.

Highlights • No partial or complete responses to pembrolizumab and lenvatinib therapy were observed in advanced or recurrent UCS. • Median PFS and OS following pembrolizumab and lenvatinib therapy is similar to traditional cytotoxic regimens. • Pembrolizumab and lenvatinib therapy was only used in patients who had failed two or more lines of therapy., Objective(s) To investigate the efficacy of pembrolizumab plus lenvatinib as a second-line or later-line therapy in women with advanced or recurrent uterine carcinosarcoma (UCS). Methods A single-institution pharmacy database was queried for women with advanced or recurrent UCS who were prescribed concurrent pembrolizumab and lenvatinib. Patient demographic, oncologic, and immunotherapy outcomes data were recorded. Univariate analysis summarized progression-free survival (PFS) and overall survival (OS). Results Seven patients with advanced or recurrent UCS were treated with combination pembrolizumab and lenvatinib, with a median age of 63.0 years. The majority had stage III or IV disease (n = 6, 85.7%) and had failed two or more lines of therapy (n = 7, 100.0%), and a minority were MMR deficient (n = 1, 14.3%) or PD-L1+ (n = 1, 14.3%). No partial or complete responses were observed. The median PFS was 2.6 months (95% CI, 0.9–11.2 months), and the median OS was 2.8 months (95% CI, 2.4-NE). Conclusions In this small, retrospective series, we demonstrate that pembrolizumab and lenvatinib combination therapy may not be highly active in UCS and may be associated with similar PFS and OS as traditional cytotoxic regimens. Further study is warranted to assess the efficacy of this regimen in more targeted cohorts of women with advanced or recurrent UCS.

Malignant Brenner tumor (MBT) is diagnosed in the setting of invasive high-grade carcinoma with urothelial-like morphology and the presence of an adjacent benign Brenner tumor (BBT) or borderline Brenner tumor (BLBT). MDM2 amplification was recently detected by next-generation sequencing on a small number of MBTs, potentially significant for future targeted therapy. Experience is limited, however, and evaluation of widely available MDM2 immunohistochemistry (IHC) has not been performed to determine clinical utility. After confirming all diagnoses morphologically and immunohistochemically, we performed MDM2 IHC on 4 MBTs, 3 BLBTs, 26 BBTs, 142 high-grade serous carcinomas (HGSC), 6 ovarian endometrioid carcinomas (OEC) with urothelial-like morphology, and 49 high-grade urothelial carcinomas (HGUC). MDM2 IHC was considered positive with diffuse (25%) nuclear reactivity; in cases of patchy staining (10-25% nuclear reactivity), MDM2 was considered equivocal. Positive staining in10% of cells was considered negative. In cases with positive or equivocal staining, MDM2 amplification was evaluated by fluorescence in-situ hybridization (FISH). Three MBTs (75%) showed diffuse nuclear reactivity for MDM2 by IHC, a finding corroborated by amplification of MDM2 in all three cases. One MBT and 2 BLBTs showed equivocal MDM2 IHC, but all three were negative for MDM2 amplification. The final BLBT, as well as all BBTs, HGSC, OEC, and HGUC, were negative for MDM2. In conclusion, our limited cohort confirms MDM2 amplification in MBT and suggests that MDM2 IHC may have an influence in rare diagnostically challenging cases.

To assess oncologic outcomes in endometrial cancer patients with low-volume metastasis (LVM) in the sentinel lymph nodes (SLNs).Patients with endometrial cancer and SLN-LVM (≤2 mm) from December 3, 2009, to December 31, 2018, were retrospectively identified from 22 centers worldwide. Patients with International Federation of Gynecology and Obstetrics (FIGO) stage IV, adnexal involvement, or unknown adjuvant therapy (ATx) were excluded.Of 247 patients included, 132 had isolated tumor cell (ITC) and 115 had micrometastasis (MM). Overall 4-year recurrence-free survival (RFS) was 77.6% (95% CI, 70.2%-85.9%); median follow-up for patients without recurrence was 29.6 (interquartile range, 19.2-41.5) months. At multivariate analysis, Non-endometrioid (NE) (HR, 5.00; 95% CI, 2.50-9.99; P .001), lymphovascular space invasion (LVSI) (HR, 3.26; 95% CI, 1.45-7.31; P = .004), and uterine serosal invasion (USI) (HR, 3.70; 95% CI, 1.44-9.54; P = .007) were independent predictors of recurrence. Among 47 endometrioid ITC patients without ATx, 4-year RFS was 82.6% (95% CI, 70.1%-97.2). Considering 18 ITC patients with endometrioid grade 1 disease, without LVSI, USI, or ATx, only 1 had recurrence (median follow-up, 24.8 months).In patients with SLN-LVM, NE, LVSI, and USI were independent risk factors for recurrence. Patients with any risk factor had poor prognosis, even when receiving ATx. Patients with ITC and grade 1 endometrioid disease (no LVSI/USI) had favorable prognosis, even without ATx. Further analysis (with more patients and longer follow-up) is needed to assess whether ATx can be withheld in this low-risk subgroup.

Poly-ADP Ribose Polymerase (PARP) inhibitors are clinically approved for the treatment of homologous recombination (HR) repair deficient tumors. PARP targeted therapy has limited efficacy in HR-proficient cancer. In this study, we identified the non-receptor lymphocyte-specific protein tyrosine kinase (LCK) as a novel regulator of HR repair pathways in endometrioid epithelial ovarian cancer (eEOC). Inhibition of LCK attenuates the expression of RAD51, BRCA1, and BRCA2 proteins necessary for HR-mediated DNA repair. HR repair in eEOC cells is LCK dependent. Upon DNA damage LCK expression is increased, and autophosphorylated, activated LCK is localized in the nucleus. LCK inhibition impairs RAD51 foci formation but augments {gamma}H2AX formation during DDR indicating reduced ability to repair DNA damage. DNA damage leads to direct interaction of LCK with RAD51 and BRCA1. Finally, attenuation of LCK sensitized HR-proficient eEOC cells to PARP inhibitor. Collectively, the findings identify a mechanism for expanding utility of PARP inhibitors. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=161 SRC="FIGDIR/small/433791v1_ufig1.gif" ALT="Figure 1"> View larger version (94K): org.highwire.dtl.DTLVardef@1b9069corg.highwire.dtl.DTLVardef@612304org.highwire.dtl.DTLVardef@141fca1org.highwire.dtl.DTLVardef@130c330_HPS_FORMAT_FIGEXP M_FIG C_FIG In BriefDey and colleagues identify the nonreceptor tyrosine kinase LCK as a mediator of homologous recombination directed DNA repair in ovarian cancer. The studies show that LCK inhibition (LCKi) is sufficient to augment Poly (ADP-Ribose) Polymerase inhibitor efficacy in Homologous Recombination (HR) proficient endometrioid ovarian cancer. HighlightsO_LINonreceptor tyrosine kinase LCK regulates expression of HR repair proteins RAD51, BRCA1 and BRCA2. C_LIO_LILCKi induces HR deficiency in endometrioid epithelial ovarian cancer. C_LIO_LIDNA damage leads to autophosphorylation of LCK and co-immunoprecipitation with RAD51 and BRCA1. C_LIO_LILCKi potentiates PARP targeted therapy in HR proficient ovarian cancer and expands the utility of the highly successful PARP inhibitors in the clinic. C_LI Statement of significanceThis study identifies a novel regulator and signaling pathway for maintaining HR repair during DNA damage. It further demonstrates a new opportunity to increase the utility of PARP inhibitors in HR-proficient eEOC cells.

Endometriosis is a leading cause of pelvic pain and infertility. It is defined by the presence of endometrial tissue in extrauterine locations. The development of novel therapies and diagnostic tools for endometriosis has been limited due in part to challenges in studying the disease. Outside of primates, few mammals menstruate, and none develop spontaneous endometriosis. Rodent models are popular but require artificial induction of endometriosis, with many utilizing either immunocompromised mice or surgically induced disease. Recently, more attention has been given to models involving intraperitoneal injection. We present a murine model of endometriosis that integrates several features of existing endometriosis models into a novel, simplified system that relies on microscopic quantification in lieu of subjective grading. In this model, we perform hormonal stimulation of donor mice, intraperitoneal injection, systematic abdominal survey and tissue harvest, and histologic quantification that can be performed and verified at any time after necropsy. This model requires minimal resources and training; does not require expertise by lab technicians in murine survival surgery or in the identification of gross endometriotic lesions; can be used in immunocompromised, immunocompetent, and/or mutant mice; and reliably creates endometriotic lesions that are histologically consistent with human endometriotic disease.

Neuroendocrine carcinoma of the cervix is a rare and aggressive form of cervical cancer that presents with frequent metastasis at diagnosis and high recurrence rates. Primary treatment is multimodal, which often includes chemotherapy with or without radiation therapy. There is no data available to guide treatment for recurrence, and second-line therapies are extrapolated from small cell lung carcinoma data. Close monitoring of these patients for recurrence is paramount. Evaluation of circulating tumor DNA (ctDNA) in the peripheral blood is an attractive approach due to its noninvasive nature. Ultra-low pass whole genome sequencing (ULP-WGS) can assess tumor burden, response to therapy, and predict recurrence; however, data are lacking regarding the role of ULP-WGS in small cell carcinoma of the cervix. This study demonstrates a patient whose response to chemotherapy and cancer recurrence was accurately monitored by ctDNA using ULP-WGS testing and confirmed with radiologic imaging findings.

Background: Epithelial Ovarian Cancer (EOC) is the second most common gynecologic malignancy in the United States, but the leading cause of gynecologic cancer death. Despite many achieving remission with first-line therapy, up to 80% of patients will recur and require additional treatment. Antibiotic therapy is frequently used during cancer treatments for both prophylaxis and treatment of infections, although this profoundly impacts the gut microbiome. Multiple studies suggest that an unperturbed gut microbiome may provide a protective microenvironment, and disruption may be permissive to tumor growth and chemotherapy resistance, including platinum agents. Experimental Design: We assessed whether antibiotic therapy would impact growth of EOC and sensitivity to cisplatin in murine models. Immune competent or compromised mice were given either metronidazole, ampicillin, vancomycin, and neomycin (ABX) containing or control water for two weeks before being intraperitoneally injected with murine ID8 or ID8-VEGF EOC cells. Tumors were monitored and cisplatin therapy was administered weekly until endpoint. Stool was collected throughout the study to asses for microbial population effects over time.Results: Both immune competent and immune compromised ID8 and ID8 VEGF tumor-bearing mice demonstrated a decreased response to cisplatin therapy in ABX treated groups with an increase in overall tumor burden. RNAseq analysis showed enrichment of multiple cell proliferation and stem cell pathways, and stem cell genes SOX2, WNT and PAX2. The self-renewal of ABX treated tumor cells was also increased.Conclusion: Collectively, these studies indicate an intact microbiome provides a tumor suppressive microenvironment and enhances sensitivity to cisplatin.

Objectives There is little information regarding sentinel lymph node (SLN) frozen-section examination in patients with a history of ductal carcinoma in situ (DCIS). We evaluated the usage, clinical impact, and pathology resources used for SLN cryosectioning in mastectomy cases with a DCIS history. Methods Mastectomies with SLNs submitted from 2012 to 2013 at a tertiary care center were analyzed. Medicare reimbursement was used to estimate pathology health care expenditures of intraoperative frozen sections. Results There was no difference in the rate of SLN frozen-section examination or parts submitted, total blocks frozen, total blocks submitted, or total SLNs identified per case between the DCIS (n = 139) and invasive (n = 369) groups. Nine patients with DCIS had SLN metastases (three macrometastases, two micrometastases, and four isolated tumor cells), all of which were examined by frozen section. Only the macrometastases were identified by cryosectioning, which led to two synchronous axillary lymph node dissections that did not yield any additional positive nodes. A total of $19,313 was spent for pathology per DCIS patient with surgical management affected, whereas only $1,019 was spent per invasive carcinoma patient affected. Conclusions Decreasing SLN frozen-section use in patients with a history of DCIS represents an opportunity for pathology cost containment.

Objective. Thyroid disease has been reported among the endocrinopathies that can occur after treatment with ipilimumab. Graves’ disease, however, has been rarely reported with this medication. Here we report a case of Graves’ disease diagnosed after initiation of ipilimumab in a patient with melanoma.Methods. We present the clinical presentation and management course of this patient followed by a related literature review.Results. A 67-year-old male with metastatic melanoma was started on ipilimumab. He developed hyperthyroidism after two doses of ipilimumab. The cause of hyperthyroidism was determined to be Graves’ disease. Ipilimumab was held and the patient was started on methimazole with return to euthyroid status. Ipilimumab was resumed and the patient continued methimazole during the course of ipilimumab therapy, with controlled hyperthyroidism. Restaging studies following four cycles of ipilimumab showed complete response in the lungs, with residual melanoma in the neck. The patient then underwent total thyroidectomy and left neck dissection as a definitive treatment for both hyperthyroidism and residual melanoma.Conclusion. Graves’ disease can develop after starting ipilimumab and methimazole can be an effective treatment. For patients whose hyperthyroidism is well-controlled on methimazole, ipilimumab may be resumed with close monitoring.

Highlights • Previously treated gynecologic clear cell tumors had a CPI response rate of 57% despite MSI stable status. • Immune related adverse events occurred in 40.3% of patients. • Patients who developed immune toxicity had higher ORR, PFS and OS. • In chemo-naïve MSI-H recurrent endometrial carcinoma ORR to CPI was 83%., Prognostic factors for immune checkpoint inhibitor (CPI) response in gynecologic cancer are limited. This retrospective study aimed to identify prognostic factors associated with improved overall response rate (ORR) and progression free survival (PFS) in gynecologic cancer patients receiving at least two cycles of CPI. PFS was compared by univariate cox regressions. Univariate and multivariable analyses were used for prognostic factors of PFS and ORR. 72 patients were identified (20 ovarian, 36 endometrial, 13 cervix, 1 vaginal, 2 others). Immune related adverse events (IRAE) occurred in 40.3% of patients (29/72). IRAE was associated with higher ORR (44.8% IRAE vs 20.9% no IRAE, OR 3.1, p = 0.024), improved PFS (12.9 m IRAE vs 4.7 m no IRAE, HR 0.43, p = 0.004) and improved OS (22.9 m IRAE vs 12.2 m no IRAE, HR 0.47, p = 0.021). Additionally, Clear cell histology had superior ORR compared to MSI stable endometrial and ovarian cancers (ORR 57.1% vs 11.8%, OR 10.0, p = 0.032). Responders more often had ARIDIA mutation, PI3K/PTEN alteration and less often had a P53 mutation. In a subset of six MSI-H, recurrent, chemo-naive endometrial cancer ORR was 83.3%. Overall, we found favorable outcomes after CPI for clear cell tumors and patients who developed IRAE. Additionally, first-line systemic therapy with CPI in recurrent MSI-H endometrial cancer had encouraging ORR with durable responses.

e18092 Background: There is limited data on outcomes for gynecologic cancer patients treated with immune checkpoint inhibitors (ICI) outside the scope of clinical trials. Here we present our Institutional experience with a cohort of endometrial (EC) and ovarian cancer (OC) treated with ICI. Methods: 59 patients who received ICI were included (23 OC and 36 EC). Progression-free (PFS) and Overall survivals (OS) were determined by Kaplan-Meier (KM) curve and log rank test. Comparison of duration of response (DOR) and stable disease (DOSD) was done with unpaired t-test or one-way ANOVA. Rates of objective response (ORR) including partial response (PR) and complete response (CR), and stable disease (SD) were compared by Fischer’s exact test. Results: Median age was 66 years. 23 patients were microsatellite stable (MSS), 23 microsatellite instability high (MSI-H). Median number of prior lines was 2 (0-11). PFS and OS for EC and OC were overlapping; therefore outcomes for both were combined [(PFS 6.4m OC vs 7.3 m EC, p = 0.61), (OS 15.9 m OC vs 14.2 m EC, p = 0.78)]. Response rates consisted of 20.3% PR, 8.5% CR, 37.3% SD. Differences in responses were noted for clear cell carcinoma (CC) (33.3% PR, 11.1% CR, 33.3% SD) and MSI-H (36.4% PR, 18.2% CR, 22.7% SD) compared to MSS (11.8% PR, 0% CR, 47% SD). MSI-H had higher ORR vs. MSS (54.1% vs 11.8%, p = 0.0078). CC trended toward improved ORR vs. MSS (44.4% vs 11.8%, p = 0.14). PFS was improved for MSI-H vs. MSS (10m v 5.0m, p = 0.03). OS for CC compared to any other histology was improved (NR vs 12.8m respectively, p = 0.009). 5 recurrent MSI-H EC patients received ICI as first line monotherapy. Responses included 4 PR and 1 SD (80% ORR, 100% clinical benefit). PFS was 9.2m (3.3-13.3). 80% remained progression-free at last follow up. Overall, 38.9% experienced toxicity: hypothyroidism (15%), dermatitis (5%), pneumonitis (10%), LFT elevation (2%), amylase/lipase elevation (3%), colitis or diarrhea (5%), uveitis (2%) or nephritis (5%). 10% of patients required discontinuation of ICI secondary to toxicity. Trends for PFS and OS favored improved outcomes in patients with toxicity vs. no toxicity [(PFS 12.9m vs 5.6m, p = 0.07), (OS 22.9m vs 13.1m, p = NS)] respectively. Conclusions: In this study, immunotherapy with ICI outcomes favor MSI-H and CC compared to MSS disease. CC had promising OS compared to other histology types. ICI showed promising efficacy in MSI-H EC with 100% clinical benefit rate in chemonaive patients. First line ICI should be investigated in these patients. Positive correlation between toxicity and outcome is noted and will be further investigated.

6093 Background: High-grade neuroendocrine carcinoma (NEC) of the cervix and other gynecologic origins are rare and aggressive cancers that unfortunately affect young women with high mortality. Treatment recommendations are often extrapolated from their counterpart, small cell carcinoma of the lung (SCLC). In the present study, we have performed comprehensive genomic and transcriptomic analyses in a cohort of patients with NEC of the cervix and other gynecologic origins and compared them to SCLC. Methods: We have identified 27 patients diagnosed with NEC of gynecologic origin from 1998-2019. Of them, we were able to obtain archival tissue from 14 patients (17 samples), including seven cervical NEC, three ovarian NEC, and two endometrial NEC. Two ovarian and one cervical patient have recurrent tumors. Whole exome sequencing (WES) and RNA-seq were successfully performed in 14 and 13 samples, respectively. BWA-MEM was used for mapping (hg38), and GATK Haplotype Caller was used for WES analysis. Ensemble-VEP was used for variant annotation, and SIFT and PolyPhen-2 were used for pathogenic prediction of missense mutations. Stranded pair-end RNA-seq data were analyzed using STAR and DESeq2. SCLC data from cBioPortal and EBI (accession number EGAS00001000334) were used to compare mutation and transcriptomic profiles. Results: We found that TP53 is not mutated in our cohort, and RB1 is mutated only in 1 (7%) tumor. TP53 and RB1 are the most frequently mutated genes in small cell lung carcinoma. However, LRP1B, the third most mutated gene in SCLC was also mutated in 4 (29%) tumors. We observed that WNT, RTK-RAS, NOTCH, and MYC are among the top mutated pathways in our cohort. Among the top 20 mutated genes, only four genes, MUC4, KMT2C, MAP3K1, and HLA-A, were common in cervical, ovarian, and endometrial tumors suggesting a high diversity within the NEC of gynecologic origin. Our RNA-seq analysis revealed a distinct transcriptional signature when compared to SCLC, especially the expression pattern of SCLC molecular subtypes defining ASCL1, NEUROD1, POU3F2, and YAP1 genes. Surprisingly, we observed a high expression of the YAP1 gene in all of the 13 tumors. However, the YAP1+ subtype represents a minority in SCLC and known to predict chemotherapy resistance and lower survival. Conclusions: Our results suggest a unique mutational profile and transcriptional signature that are distinct compared to SCLC tumors. Therefore, there is an urgent need to re-evaluate the therapeutic options and targets for NEC of gynecologic origin.

6074 Background: Immune checkpoint inhibitors (ICI) have demonstrated success in solid tumors. In gynecologic cancers (GC), the response rate is still low (~10-15%) except in MSI-H endometrial cancer (~ 50%). Current biomarkers (e.g. PDL1 expression) have limited utility in identifying benefit from ICI in GC. In this work we evaluated the ability of computational measurements of spatial arrangement of tumor infiltrating lymphocytes (TIL) from H&E slide images in predicting overall survival (OS) and response to ICI in ovarian, cervical and endometrial cancers. Methods: The study included 151 patients, including 102 ovarian carcinomas treated with surgery and chemotherapy (D1) and another set (D2) of n=49 patients (n=14 ovarian, n=27 endometrial and n=8 cervical), treated with different ICI agents (Pembrolizumab, Nivolumab, Ipilimumab, Avelumab) in the second line setting. Progressors and non-progressors in D2 were classified according to clinical improvement and radiologic assessment by RECIST. A machine learning approach was employed to identify tumor regions on the diagnostic slides from D1 and D2 and then used to automatically identify TILs within the tumor regions. Subsequently machine learning was used to define TIL clusters based on TIL proximity, and graph network theory was used to capture measurements relating to spatial arrangement of TIL clusters. The multivariable Cox regression model (MCRM) was trained on n=51 patients from D1 to predict OS and then independently evaluated in predicting (1) OS on the hold-out n=51 patients in D1 and (2) response and progression-free survival (PFS) in D2. Results: Statistical analysis identified 7 prognostic features relating to interaction of TIL clusters with cancer nuclei. MCRM was prognostic of OS on the n=51 hold out patients in D1 (hazard ratio (HR)=2.06, 95% confidence interval [1.04- 4.07], p=0.008) and predictive of PFS in D2 (HR=2.24, CI=[1.13-4.44], p=0.03). The AUC for MCRM in predicting progression in D2 was 82%. Conclusions: Computerized features of spatial arrangement of TILs on H&E images were prognostic of OS and PFS and predicted response to ICI in three gynecological cancers. These findings need to be validated in larger, multi-site validation sets. [Table: see text]

ObjectiveThis study aims to determine the prevalence of programmed death ligand 1 (PD-L1) expression in endometrial carcinoma (EC) and determine clinical and pathological associations.MethodsImmunohistochemistry for PD-L1 was performed on sections of a triple-core tissue microarray of 700 ECs. Positive PD-L1 expression, defined as 1% of cells staining positive, was evaluated in tumor and stromal compartments. Using age-adjusted logistic regression, we estimated odds ratios and 95% confidence intervals for associations between PD-L1 expression (overall and by staining compartment) with clinical and tumor characteristics. Kaplan-Meier plots and log-rank tests were used to evaluate associations between PD-L1 expression and EC-specific survival.ResultsPD-L1 expression was observed in 100 cases (14.3%), including 27 (3.9%) with expression in tumor cells only, 35 (5.0%) with expression in both tumor cells and stroma, and 38 (5.4%) with expression in stroma only. Expression was observed in ECs of different histologic types. Tumors characterized by loss of mismatch repair proteins were significantly associated with tumoral PD-L1 expression (P< 0.0001), but not with stromal PD-L1 expression. Both tumoral and stromal PD-L1 expressions were associated with high-grade endometrioid histology, nonendometrioid histology, and lymphovascular space invasion. We observed no significant associations between PD-L1 expression and EC-specific survival.ConclusionsPD-L1 is expressed in a significant proportion of EC and is associated with mismatch repair deficiency, potentially representing a mechanism of tumor immune evasion and a therapeutic target in EC.

The topic of hereditary gynecologic malignancies readily evokes associations between Lynch syndrome and endometrial adenocarcinoma, or between BRCA mutations and tubo-ovarian serous carcinoma, but other familial associations are less well-known. Two hereditary syndromes are known to be related to uterine mesenchymal tumors: hereditary leiomyomatosis and renal cell carcinoma syndrome and the tuberous sclerosis complex. In the following review, we describe the current literature on these syndromes, summarizing their clinical, morphologic, immunophenotypic, and genetic data. It is possible that the surgical pathologic diagnosis is the first indication of a familial syndrome, thus emphasizing the importance of a pathologist's familiarity with these potentially suggestive lesions.

Background Pituitary carcinoma is a rare entity requiring the presence of metastasis to confirm its malignant potential. We report a case of pituitary carcinoma and discuss the diagnosis and management of this lesion in relation to the existing literature. Case presentation The patient is a 51-year-old woman with Cushing’s disease and intact adrenal glands who was diagnosed with metastatic pituitary carcinoma to the liver, 29 months after initial resection of an ACTH-secreting primary atypical pituitary adenoma (APA). Prior to detection of this metastasis the patient underwent repeat resection and radiotherapy for residual cavernous sinus disease. The metastatic lesion was detected by interval surveillance of serum ACTH and 24-hour urine cortisol, which despite stable pituitary MRI, were significantly elevated. These abnormalities prompted a PET scan that demonstrated hypermetabolic liver parenchyma, which was suspicious for metastasis on abdominal MRI. An ultrasound-guided liver biopsy demonstrated nests of moderately-differentiated cells with intermediate-sized, monotonous nuclei, distinct nucleoli, and abundant basophilic cytoplasm, confirmed by immunohistochemistry to represent metastatic pituitary carcinoma. The liver lesion was subsequently successfully removed by wedge resection. One year later, the patient’s residual cavernous sinus disease grew markedly, and she was placed on dual-agent chemotherapy consisting of oral temozolomide and capecitabine, with stabilization of her intracranial disease to present, although liver metastases recurred. Conclusions Pituitary carcinoma is a rare entity impossible to recognize as a primary tumor because its diagnosis by definition requires the presence of metastasis. Maintaining awareness of the entity and its precursor lesion APA is essential for its accurate pathologic diagnosis and appropriate management.

MELF invasion has been associated with non-vaginal recurrences and lymph node (LN) metastases in multi-institutional case control studies, but has not been well examined in large single institution cohorts. Hysterectomy specimens with FIGO 1 endometrioid endometrial carcinoma (EEC) and lymphadenectomies from 2007 to 2012 were identified. Electronic medical records and histologic slides were reviewed. Of 464 identified cases, 163 (35.1%) were noninvasive, 60 (12.9%) had MELF, 222 (47.8%) had a component of the infiltrative invasion pattern without MELF, 13 (2.8%) had pure pushing borders of invasion, 5 (1.1%) had pure adenomyosis-like invasion, and 1 (0.2%) had pure adenoma malignum-like invasion. Sixteen cases had LN metastases. Significantly more MELF cases had positive LNs than non-MELF cases overall (18.3% vs 1.2%, p

Background Pleomorphic xanthoastrocytoma (PXA) is an uncommon, primary neoplasm of the central nervous system with a relatively favorable prognosis. Most patients are managed with surgery alone and experience significant long-term survival. PXAs occur most commonly along the superficial surfaces of the temporal lobes. Although these tumors may occur in other regions of the brain, their origin within the spinal cord is rare, and it is unclear whether spinal cord PXAs should be managed differently from their intracranial counterparts. Case Description We describe a 31-year-old patient with a PXA of spinal cord origin who despite surgery, radiation, and multiple chemotherapy regimens experienced anaplastic transformation of his tumor and died of extensive leptomeningeal progression. Conclusions To our knowledge, our patient represents the seventh reported case of PXA of the spinal cord but is the first described to have a BRAF mutation. Specifically, both the initial and recurrent tumors of the patient showed the same BRAF V600E mutation, which refutes previous suggestions that BRAF mutations may be limited to intracranial PXAs and also shows that BRAF mutations may occur earlier in PXA tumorigenesis.

Background Follicular lymphoma (FL) is one of the most common lymphoid malignancies in the western world. FL cases are stratified into three histological grades based on the average centroblast count per high power field (HPF). The centroblast count is performed manually by the pathologist using an optical microscope and hematoxylin and eosin (H&E) stained tissue section. Although this is the current clinical practice, it suffers from high inter- and intra-observer variability and is vulnerable to sampling bias. Methods In this paper, we present a system, called Follicular Lymphoma Grading System (FLAGS), to assist the pathologist in grading FL cases. We also assess the effect of FLAGS on accuracy of expert and inexperienced readers. FLAGS automatically identifies possible HPFs for examination by analyzing H&E and CD20 stains, before classifying them into low or high risk categories. The pathologist is first asked to review the slides according to the current routine clinical practice, before being presented with FLAGS classification via color-coded map. The accuracy of the readers with and without FLAGS assistance is measured. Results FLAGS was used by four experts (board-certified hematopathologists) and seven pathology residents on 20 FL slides. Access to FLAGS improved overall reader accuracy with the biggest improvement seen among residents. An average AUC value of 0.75 was observed which generally indicates “acceptable” diagnostic performance. Conclusions The results of this study show that FLAGS can be useful in increasing the pathologists’ accuracy in grading the tissue. To the best of our knowledge, this study measure, for the first time, the effect of computerized image analysis on pathologists’ grading of follicular lymphoma. When fully developed, such systems have the potential to reduce sampling bias by examining an increased proportion of HPFs within follicle regions, as well as to reduce inter- and intra-reader variability. Electronic supplementary material The online version of this article (doi:10.1186/s12911-015-0235-6) contains supplementary material, which is available to authorized users.

Pituitary carcinoma is a rare entity requiring the presence of metastasis to confirm its malignant potential. Most examples secrete prolactin or adrenocorticotropic hormone (ACTH). Reported sites of metastasis include to the cerebral cortex, cerebellum, spinal cord, leptomeninges, eyes, heart, lung, lymph nodes, pancreas, liver, kidney, ovary, myometrium and bone. Liver metastasis is most common with prolactin-secreting tumors. In ACTH-secreting tumors it is frequently associated with Nelson's syndrome (rapid enlargement of a pituitary adenoma after bilateral adrenalectomy to palliate Cushing's syndrome). Here we report a case of an ACTH-secreting atypical pituitary adenoma metastasizing to the liver in a 51-year-old woman with Cushing's syndrome and intact adrenal glands 2.5 years after resection of the primary pituitary lesion with subsequent radiotherapy for residual cavernous sinus disease. Surveillance of ACTH and 24 hour urine cortisol levels revealed that they were significantly elevated to 142 pg/mL (normal range 10-60 pg/mL) and 541 µg/day (normal range 3.5-45 µg/day), respectively. These abnormalities prompted a PET scan which demonstrated a hypermetabolic liver lesion with a maximal SUV of 3.3, which was suspicious for metastasis on MRI. An ultrasound-guided liver biopsy demonstrated involvement by a moderately differentiated carcinoma consistent with a primary pituitary lesion. Nests of tumor cells showed a monotonous epithelioid appearance, abundant cytoplasm and small nucleoli. Immunostains for AE1/3, synaptophysin, and ACTH diffusely highlighted the lesional cells. This case thus represents an additional rare example of an ACTH-producing pituitary carcinoma metastatic to the liver in a patient with Cushing's syndrome and intact adrenal glands.

Objective To evaluate the viability of glomeruli in the peritumor parenchyma of partial nephrectomy specimens removed for renal cell carcinoma (RCC) and relate it to kidney function, to better understand the contribution of peritumor parenchyma to renal function. Materials and Methods A retrospective analysis of 53 partial nephrectomies containing RCC was performed. Glomeruli within 0.25-cm increments from the tumor were quantified and histologically assessed for viability. Tumor size, minimum and maximum margin size, and pre- and postoperative estimated glomerular filtration rate (eGFR) were obtained. Results Glomerular viability positively correlated with distance from tumor with mean viable glomeruli in successive 0.25-cm increments of 0-0.25 cm, 58%; 0.25-0.5 cm, 80%; 0.5-0.75 cm, 90%; and 0.75-1.0 cm, 92%. Glomerular viability near the tumor did not correlate with preoperative eGFR, whereas decreased viability further from the tumor did correlate with worse preoperative eGFR. Tumor size showed a nonstatistically significant positive trend with minimum (median 0.15 cm) and maximum margin (median 0.7 cm) sizes. Percent change of glomerular filtration rate did not correlate with margin size ( P = .190). Conclusion Renal parenchyma immediately adjacent to RCC contains fewer viable glomeruli compared with the parenchyma further from the tumor. Based on this information, attempts to preserve all non-neoplastic renal parenchyma via a surgical margin approaching zero may not necessarily result in clinically relevant differences in the amount of viable glomeruli remaining or the renal function preserved.

Objective The aim of this study is to determine the expression of caspase-14, a key protein in maturation of squamous epithelia, in archival malignant and premalignant vulvar squamous lesions and examine in-vitro effects of a black raspberry extract (BRB-E) on a vulvar squamous cell carcinoma (VSCC) cell line. Methods VSCC cell cultures were exposed to different BRB-E concentrations and used to create cell blocks. Immunohistochemistry for caspase-14 was performed on cell block sections, whole tissue sections, and a tissue microarray consisting of normal vulvar skin, lichen sclerosus (LS), classic and differentiated vulvar intraepithelial neoplasia (cVIN and dVIN respectively), and VSCC. Results LS demonstrated abnormal full thickness (5/11) or absent (1/11) caspase-14 staining. dVIN often showed markedly reduced expression (4/7), and cVIN occasionally demonstrated either absent or reduced caspase-14 (6/22). VSCC predominantly had absent or markedly reduced caspase-14 (26/28). VSCC cell cultures demonstrated a significant increase in caspase-14 ( p =0.013) after BRB-E treatment: 7.3% (±2.0%) of untreated cells showed caspase-14 positivity, while 21.3% (±8.9%), 21.7% (±4.8%), and 22.6% (±5.3%) of cells were positive for caspase-14 after treatment with 200, 400, and 800μg/mL BRB-E, respectively. Pair-wise comparisons between the treatment groups and the control demonstrated significant differences between no treatment with BRB-E and each of these treatment concentrations (Dunnett's adjusted p -values: 0.024, 0.021, and 0.014, respectively). Conclusions Caspase-14 is frequently decreased in premalignant and malignant vulvar squamous lesions, and is upregulated in VSCC cell culture by BRB-E. BRB-E should be further explored and may ultimately be incorporated in topical preparations.

Links between obesity, with its attendant estrogen abnormalities, and the endometrial carcinoma (EC) DNA Mismatch Repair Protein (MMR) system have recently been proposed. We investigated relationships between body mass index (BMI) and clinicopathological correlates including MMR expression in a large single institution EC cohort.Clinical and pathological databases from 2007 to 2012 were used to identify consecutive hysterectomy specimens with EC. Univariate and multivariate analyses were used to explore relationships between BMI, age, stage, tumor type and immunohistochemical results for MLH1, PMS2, MSH2 and MSH6.1049 EC were identified. Overall, BMI was higher amongst women with normal MMR (p=0.002). However, when stratified by age and specific MMR, statistically significant differences localized exclusively to women50years old with loss of MSH2 and/or MSH6 (p=0.003 and p=0.005 respectively). Higher BMI correlated with endometrioid FIGO 1 and 2 tumors (p0.001) and with stage 1a (p0.001). Conversely, MMR abnormalities did not show significant associations with stage (p=0.302) or histologic grade (p=0.097).BMI showed statistically significant associations with MMR expression, tumor grade and stage amongst 1049 consecutive EC. Obesity correlates with lower grade and stage EC. A link between BMI and maintenance of the MMR system is not supported by our data because the only statistically significant association occurred in women50years old with MSH2 and/or MSH6 abnormalities where Lynch syndrome related cases are expected to cluster.

Cutaneous ciliated cysts (CCC) are exquisitely rare, benign cystic lesions demonstrating simple, ciliated epithelial linings reminiscent of fallopian tube epithelium. Most commonly, CCC show a predilection for the lower extremities of young reproductive age women and demonstrate immunohistochemical positivity for estrogen and progesterone receptors, supporting the theory that they are derived from ectopic Mullerian rests. PAX-8 is a paired box gene, important in the development of Mullerian and thyroid organs and has utility in the identification of tumors of Mullerian, renal, and thyroid origin. Prompted by the precedent studies on PAX-8 immunohistochemical expression in tumors of Mullerian origin, this article aimed to explore the utility of this antibody in defining the histogenesis of 2 bona fide cases of CCC, both occurring in young reproductive age women. Herein, 2 prototypic index cases of CCC with strong nuclear positivity for estrogen and progesterone receptors are shown to also have positive nuclear staining for PAX-8, further supporting their likely Mullerian origin. These data support the designation of these lesions as cutaneous Mullerian cysts, distinct from potential ciliated cysts of eccrine origin.