The association of histopathologic features after neoadjuvant chemo-immunotherapy with clinical outcome: Sub-analyses from the randomized double-blinded, placebo-controlled, Phase III IMagyn050/GOG3015/ENGOT-ov39 study.

OBJECTIVE: Histopathologic characteristics after neoadjuvant chemotherapy (NACT) may correlate with outcome. This study evaluates histopathologic features after immunotherapy and NACT/bevacizumab, and associated clinical outcomes. METHODS: Evaluable tissue from IMagyn050/GOG3015/ENGOT-ov39 patients from prespecified anatomic sites from interval cytoreductive surgery (ICS) after NACT/bevacizumab plus atezolizumab/placebo underwent central histopathologic scoring and analyzed with clinical outcomes. RESULTS: The predefined population had 243 evaluable NACT patients, with 48.1% tumors being PD-L1-positive. No statistically significant differences in PFS (16.9 months vs. 19.2 months, p = 0.21) or OS (41.5 months vs. 45.1 months, p = 0.67) between treatment arms were seen. Substantial residual tumor (RT) (3+) was identified in 26% atezolizumab vs. 24% placebo arms (p = 0.94). Most showed no (1+) necrosis (82% vs. 96%, respectively, p = 0.69), moderate (2+) to severe (3+) fibrosis (71% vs. 75%, respectively, p = 0.82), and extensive (2+) inflammation (53% vs. 47% respectively, p = 0.48). No significant histopathologic differences were identified by tissue site or by arm. Multivariate analyses showed increased risk for progression with moderate and substantial RT (13.6 mon vs. 21.1 mon, hazard ratio 2.0, p < 0.01; 13.6 mon vs. 21.1 mon, HR 1.9, p < 0.01, respectively); but decreased risk for death with extensive inflammation (46.9 mon vs. 36.3 mon, HR 0.65, p = 0.02). Inflammation also correlated with greater likelihood of response to NACT/bevacizumab plus immunotherapy (odds ratio 2.9, p < 0.01). Modeling showed inflammation as a consistent but modest predictor for OS. CONCLUSIONS: Detailed histologic assessment of ICS specimens appear to identify characteristics, such as inflammation and residual tumor, that may provide insight to certain clinical outcomes. Future work potentially leveraging emerging tools may provide further insight into outcomes.

Two staged phase II clinical trial of Eribulin monotherapy in advanced or recurrent cervical cancer.

BACKGROUND: Eribulin a microtubule targeting agent and analog of Halichondrin B, a natural product isolated from marine sponge H. okadai, has proven clinical efficacy in metastatic pretreated breast cancer and liposarcoma. We conducted a 2-stage Phase II study of eribulin in patients with advanced/recurrent cervical cancer to examine its clinical activity and evaluate biomarkers for predictors of response. METHODS: Women with advanced/recurrent cervical cancer after ≤1 prior chemotherapy regimen, measurable disease and ECOG performance status ≤2 were treated with eribulin (1.4 mg/m2 IV day 1 and 8, every 21 days) with tumor assessments every 2 cycles. Primary endpoint was 6-month progression-free survival (PFS6); secondary were best overall response (RECISTv1.1), toxicity (CTCAEv4.03) and overall survival (OS). Exploratory endpoints were associations of biomarkers with clinical activity. Immunohistochemistry was performed on archival tumor samples. Overexpression was defined when both intensity and distribution scores were ≥ 2. RESULTS: 32 patients enrolled from 11/2012-5/2017. 29/32 patients had prior chemotherapy with cisplatin/paclitaxel/bevacizumab (n = 12) or cisplatin/gemcitabine (n = 12) as the most common regimens. 14 patients received prior paclitaxel. 1 (3%) had a complete response, 5 (16%) had a partial response and 13 (41%) had stable disease for ORR of 19% (95% CI 8, 37). Those who are paclitaxel naïve experienced the greatest benefit with a 29% ORR (95% CI 12, 54). Patients who received prior paclitaxel responded less favorably than those who did not (p = .002) and had a shorter PFS and OS. Grade 3/4 adverse events occurring in >10% of patients were anemia (n = 12, 38%), neutropenia (n = 7, 22%) and leukopenia (n = 6, 19%). Analysis of correlative predictors of response revealed that patients who did not overexpress ßII and BAX were significantly more likely to respond to e`ribulin. PFS was significantly shorter in patients with ßII and BAX overexpression, OS was significantly shorter in those with ßIII and BAX overexpression. These associations remained after multivariate analysis. CONCLUSIONS: Eribulin shows modest activity in patients with recurrent/advanced cervical cancer with a favorable toxicity profile. Prior paclitaxel exposure is associated with decreased eribulin response. ßII, ßIII tubulin subtypes and BAX are predictors of response and survival. Eribulin may be an option for women with paclitaxel-naïve recurrent/advanced cervical cancer.

Proteomic alterations associated with residual disease in neoadjuvant chemotherapy treated ovarian cancer tissues.

BACKGROUND: Optimal cytoreduction to no residual disease (R0) correlates with improved disease outcome for high-grade serous ovarian cancer (HGSOC) patients. Treatment of HGSOC patients with neoadjuvant chemotherapy, however, may select for tumor cells harboring alterations in hallmark cancer pathways including metastatic potential. This study assessed this hypothesis by performing proteomic analysis of matched, chemotherapy naïve and neoadjuvant chemotherapy (NACT)-treated HGSOC tumors obtained from patients who had suboptimal (R1, n = 6) versus optimal (R0, n = 14) debulking at interval debulking surgery (IDS). METHODS: Tumor epithelium was harvested by laser microdissection from formalin-fixed, paraffin-embedded tissues from matched, pre- and post-NACT treated tumors for twenty HGSOC patients and analyzed by quantitative mass spectrometry-based proteomics. RESULTS: Differential analysis of patient matched pre- and post-NACT treated tumors revealed proteins associated with cell survival and metabolic signaling to be significantly altered in post-NACT treated tumor cells. Comparison of pre-NACT treated tumors from suboptimal (R1) versus optimally (R0) debulked patients identified proteins associated with tumor cell viability and invasion signaling enriched in R1 patients. We identified five proteins altered between R1 and R0 patients in pre- NACT treated tumors that significantly correlated with PFS in an independent cohort of HGSOC patients, including Fermitin family homolog 2 (FERMT2), a protein elevated in R1 that correlated with disease progression in HGSOC patients (multivariate Cox HR = 1.65, Wald p = 0.022) and increased metastatic potential in solid-tumor malignancies. CONCLUSIONS: This study identified distinct proteome profiles in patient matched pre- and post-NACT HGSOC tumors that correlate with NACT resistance and that may predict residual disease status at IDS that collectively warrant further pre-clinical investigation.

Genomic heterogeneity in peritoneal implants: A differential analysis of gene expression using nanostring Human Cancer Reference panel identifies a malignant signature.

OBJECTIVES: Peritoneal implants of ovarian borderline serous tumors are diagnostically challenging. Distinguishing invasive from non-invasive cases is crucial for patient management. This study aims to develop a molecular signature to distinguish invasive implants with malignant potential from those with benign. METHODS: Archival formalin-fixed paraffin embedded tissues were retrieved from 3 institutions, with consensus histologic review. Lesions were classified as a non-invasive implant (n = 10), invasive implant (n = 9) or high grade (HG) peritoneal metastasis from HG serous ovarian carcinoma (n = 4). The nCounter® GX Human Cancer Gene Reference Assay was used to profile expression of 230 cancer genes and 6 control genes. The DEGs in HG peritoneal metastases compared to non-invasive implants were identified using T-tests performed in the NanoString Diff package, then used to cluster cases using the Eisen cluster 3.0 package. Lasso in glmnet package was used to select the subset of genes that most strongly correlate with a malignant potential. RESULTS: 37 genes were downregulated and 16 genes were upregulated in HG peritoneal metastases. Using this 53-gene signature, one of nine of the invasive implants clustered with the HG peritoneal metastasis. Expression of ABCB1, CDC2, CDKN1A, FAT1, MMP9, MSH2, NQO1 and TOP2A were sufficient to indicate malignant potential of implants. The HG peritoneal metastasis and one invasive implant exhibited a high malignant likelihood (>92%) whereas the non-invasive implants and eight invasive implants displayed a low malignant likelihood (≤0.1%). CONCLUSIONS: Invasive implants are heterogenous and often morphologically indistinguishable lesions with transcriptomes that may be classified as malignant or not. Additional research is needed to determine the importance of these genes as drivers and/or surrogates of malignant potential, and their utility for triaging invasive implants.

Expression of the luteinizing hormone receptor (LHR) in ovarian cancer.

We investigated the association of LHR expression in epithelial ovarian cancer (OC) with clinical and pathologic characteristics of patients. LHR expression was examined immunohistochemically using tissue microarrays (TMAs) of specimens from 232 OC patients. Each sample was scored quantitatively evaluating LHR staining intensity (LHR-I) and percentage of LHR (LHR-P) staining cells in tumor cells examined. LHR-I was assessed as no staining (negative), weak (+ 1), moderate (+ 2), and strong positive (+ 3). LHR-P was measured as 1 to 5, 6 to 50% and >  50% of the tumor cells examined. Positive LHR staining was found in 202 (87%) patients' tumor specimens and 66% patients had strong intensity LHR expression. In 197 (85%) of patients, LHR-P was measured in > 50% of tumor cells. LHR-I was significantly associated with pathologic stage (p = 0.007). We found that 72% of stage III or IV patients expressed strong LHR-I in tumor cells. There were 87% of Silberberg's grade 2 or 3 patients compared to 70% of grade 1 patients with LHR expression observed in > 50% of tumor cells, p = 0.037. Tumor stage was significantly associated with overall survival and recurrence free survival, p < 0.001 for both analyses, even after adjustment for age, tumor grade and whether patient had persistent disease after therapy or not. Our study demonstrates that LHR is highly expressed in the majority of OC patients. Both LHR-I and LHR-P are significantly associated with either the pathologic stage or tumor grade.

Dysregulation of Placental Functions and Immune Pathways in Complete Hydatidiform Moles.

Gene expression studies of molar pregnancy have been limited to a small number of candidate loci. We analyzed high-dimensional RNA and protein data to characterize molecular features of complete hydatidiform moles (CHMs) and corresponding pathologic pathways. CHMs and first trimester placentas were collected, histopathologically examined, then flash-frozen or paraffin-embedded. Frozen CHMs and control placentas were subjected to RNA-Seq, with resulting data and published placental RNA-Seq data subjected to bioinformatics analyses. Paraffin-embedded tissues from CHMs and control placentas were used for tissue microarray (TMA) construction, immunohistochemistry, and immunoscoring for galectin-14. Of the 14,022 protein-coding genes expressed in all samples, 3,729 were differentially expressed (DE) in CHMs, of which 72% were up-regulated. DE genes were enriched in placenta-specific genes (OR = 1.88, p = 0.0001), of which 79% were down-regulated, imprinted genes (OR = 2.38, p = 1.54 × 10-6), and immune genes (OR = 1.82, p = 7.34 × 10-18), of which 73% were up-regulated. DNA methylation-related enzymes and histone demethylases were dysregulated. "Cytokine-cytokine receptor interaction" was the most impacted of 38 dysregulated pathways, among which 17 were immune-related pathways. TMA-based immunoscoring validated the lower expression of galectin-14 in CHM. In conclusion, placental functions were down-regulated, imprinted gene expression was altered, and immune pathways were activated, indicating complex dysregulation of placental developmental and immune processes in CHMs.

Significance of Lymphovascular Space Invasion by the Sarcomatous Component in Uterine Carcinosarcoma.

OBJECTIVE: The aim of this study was to examine the significance of lymphovascular space invasion (LVSI) with a sarcomatous component on the tumor characteristics and clinical outcomes of women with uterine carcinosarcoma (UCS). METHODS: This was a secondary analysis of a prior multicenter retrospective study that examined women with stage I-IV UCS who underwent primary hysterectomy. Archived histopathology slides were reviewed and LVSI was scored as follows: LVSI with a carcinomatous component alone (LVSI-carcinoma; n = 375, 76.8%) or LVSI containing a sarcomatous component with or without a carcinomatous component (LVSI-sarcoma; n = 113, 23.2%). Qualitative metrics of LVSI were correlated to clinicopathological factors and survival outcome. RESULTS: Tumors in the LVSI-sarcoma group were more likely to have sarcoma dominance (82.1 vs. 26.4%) heterologous sarcomatous component (51.3 vs. 37.9%), low-grade carcinoma (42.5 vs. 22.4%), and large tumor size (81.0 vs. 70.2%) in the primary tumor site compared with tumors in the LVSI-carcinoma group (all p < 0.05). On multivariate analysis, LVSI-sarcoma was independently associated with decreased progression-free survival (5-year rates: 34.9 vs. 40.8%, adjusted hazard ratio [HR] 1.84, 95% confidence interval [CI] 1.36-2.50, p < 0.001), and cause-specific survival (5-year rates: 41.8 vs. 55.9%, adjusted HR 1.95, 95% CI 1.39-2.75, p < 0.001) compared with LVSI-carcinoma. Postoperative radiotherapy for women with LVSI-sarcoma had a higher reduction rate of recurrence/progression of disease (54% reduction, p = 0.04) compared with postoperative radiotherapy for women with LVSI-carcinoma (26% reduction, p = 0.08). CONCLUSION: In UCS, the presence of a sarcomatous component in LVSI is particularly prevalent when a tumor has sarcoma dominance. Our study suggests that LVSI containing a sarcomatous component may be a predictor of decreased survival for women with UCS.

Proposal for a Risk-Based Categorization of Uterine Carcinosarcoma.

PURPOSE: To propose a categorization model of uterine carcinosarcoma (UCS) based on tumor cell types (carcinoma and sarcoma) and sarcoma dominance. METHODS: This secondary analysis of a prior multicenter retrospective study examined 889 cases of UCS with available histologic evaluation. Based on survival outcome, cases were clustered into three groups: low-grade carcinoma with nondominant homologous sarcoma [type A, n = 96 (10.8%)], (1) low-grade carcinoma with heterologous sarcoma or any sarcoma dominance and (2) high-grade carcinoma with nondominant homologous sarcoma [type B, n = 412 (46.3%)], and high-grade carcinoma with heterologous sarcoma or any sarcoma dominance [type C, n = 381 (42.9%)]. Tumor characteristics and outcome were examined based on the categorization. RESULTS: Women in type C category were more likely to be older, obese, and Caucasian, whereas those in type A category were younger, less obese, Asian, and nulligravid (all P < 0.01). Type C tumors were more likely to have metastatic implants, large tumor size, lymphovascular space invasion with sarcoma cells, and higher lymph node ratio, whereas type A tumors were more likely to be early-stage disease and small (all P < 0.05). On multivariate analysis, tumor categorization was independently associated with progression-free survival (5-year rates: 70.1% for type A, 48.3% for type B, and 35.9% for type C, adjusted P < 0.01) and cause-specific survival (5-year rates: 82.8% for type A, 63.0% for type B, and 47.1% for type C, adjusted P < 0.01). CONCLUSION: Characteristic differences in clinicopathological factors and outcomes in UCS imply that different underlying etiologies and biological behaviors may be present, supporting a new classification system.

Validity and prognostic significance of sperm protein 17 as a tumor biomarker for epithelial ovarian cancer: a retrospective study.

BACKGROUND: Prior small studies have shown increased expression of sperm protein 17 (Sp17) in epithelial ovarian cancer (EOC) tissue and suggest Sp17 as a potential biomarker for EOC. However, how Sp17 expression varies with histology, grade, and stage of EOC and its expression in other ovarian neoplasms has not been defined. It is unknown whether patients with EOC have elevated serum Sp17 levels or if Sp17 expression is associated with survival outcomes. METHODS: The study included 982 patients with benign, borderline, and malignant ovarian neoplasms and normal ovary. There were 878 patients with tissue only, 39 with serum only, and 65 with matching serum and tissue. Immunohistochemical (IHC) staining with anti-Sp17 antibody was performed on tissue specimens and the intensity scored as weak, moderate, or strong. A sandwich enzyme-linked immunosorbent assay (ELISA) was performed to measure Sp17 sera concentrations. RESULTS: Sp17 expression was most commonly seen in serous cystadenomas (83%) and serous borderline tumors (100%). Of the 773 EOC specimens, 223 (30%) expressed Sp17. Grade and histology were significantly associated with Sp17 expression among EOC specimens (p < 0.001) on both univariate and multivariable analysis, with grade 1 serous adenocarcinomas showing the highest expression (51%). Sp17 expression was limited in other benign and non-epithelial malignant neoplasms. Neither Sp17 tissue expression nor serum concentration correlated with survival outcomes. Serum concentrations were higher in patients with Sp17 tissue expression, and the highest concentrations were noted among patients with serous and clear cell adenocarcinomas. CONCLUSIONS: Sp17 is highly expressed in benign, borderline, and low grade malignant serous ovarian neoplasms and can be quantified in serum. Sp17 expression may have diagnostic significance in this subset of patients.

Impact of lympho-vascular space invasion on tumor characteristics and survival outcome of women with low-grade serous ovarian carcinoma.

BACKGROUND AND OBJECTIVES: To examine association of lympho-vascular space invasion (LVSI) with clinico-pathological factors and to evaluate survival of women with low-grade serous ovarian carcinoma containing areas of LVSI. METHODS: This is a multicenter retrospective study examining consecutive cases of surgically treated stage I-IV low-grade serous ovarian carcinoma (n = 178). Archived histopathology slides for the ovarian tumors were reviewed, and LVSI was scored as present or absent. LVSI status was correlated to clinico-pathological findings and survival outcome. RESULTS: LVSI was seen in 79 cases (44.4%, 95% confidence interval [CI] 37.1-51.7). LVSI was associated with increased risk of omental metastasis (87.0% vs 64.9%, odds ratio [OR] 3.62, P = 0.001), high pelvic lymph node ratio (median 12.9% vs 0%, P = 0.012), and malignant ascites (49.3% vs 32.6%, OR 2.01, P = 0.035). On multivariable analysis, controlling for age, stage, and cytoreductive status, presence of LVSI in the ovarian tumor remained an independent predictor for decreased progression-free survival (5-year rates 21.0% vs 35.7%, adjusted-hazard ratio 1.57, 95%CI 1.06-2.34, P = 0.026). LVSI was significantly associated with increased risk of recurrence in lymph nodes (OR 2.62, 95%CI 1.08-6.35, P = 0.047). CONCLUSION: LVSI in the ovarian tumor is associated with adverse clinico-pathological characteristics and decreased progression-free survival in women with low-grade serous ovarian carcinoma.

High glucocorticoid receptor expression predicts short progression-free survival in ovarian cancer.

OBJECTIVE: To investigate the association of tumor glucocorticoid receptor (GR) expression and patient outcome in ovarian cancer. METHODS: GR expression was evaluated by immunohistochemistry using tissue microarrays of specimens from 481 patients with ovarian cancer and 4 patients with benign conditions. Low GR expression was defined as an intensity of 0 or 1+ and high GR as 2+ or 3+ in >1% of tumor cells. Analyses were performed to evaluate the relationship of GR expression with clinical characteristics, progression-free survival (PFS) and overall survival (OS). RESULTS: GR protein was highly expressed in 133 of 341 (39.0%) tumors from patients who underwent upfront cytoreduction surgery followed by adjuvant chemotherapy. High GR expression was more common in serous tumors (p<0.001), high grade tumors (p<0.001), and advanced stage tumors (p=0.037). Median PFS was significantly decreased in cases with high GR (20.4months) compared to those with low GR (36.0months, HR=1.66, 95% CI 1.29-2.14, p<0.001). GR remained an independent prognostic factor for PFS in multivariate analysis. OS was not associated with GR status. CONCLUSIONS: These data suggest that high GR expression correlates with poor prognosis and support the hypothesis that modulating GR activity in combination with chemotherapy may improve outcomes.

Nestin: A biomarker of aggressive uterine cancers.

OBJECTIVE: Evidence of potential prognostic and predictive value for nestin was investigated in well-annotated uterine cancers (UCs). METHODS: Nestin expression and previously-published biomarkers were evaluated by immunohistochemistry (IHC) in UC tissue microarrays. Biomarkers were categorized as low vs. high, and nestin was cut at 10% positive staining. Relationship between nestin and clinicopathologic factors, biomarkers and outcome were evaluated using exact/log-rank testing or logistic/Cox modeling. RESULTS: There were 323 eligible cases, 34% had advanced stage disease, 37% had type II disease, and 5% were carcinosarcomas. High nestin, observed in 19% of cases, was more common in advanced vs. early stage disease, type II cancers or uterine carcinosarcoma vs. type I cancers, grade 3 disease, positive lymphovascular space invasion (LVSI) and tumors >6cm (p<0.05). Nestin was inversely correlated with ER, PR and TFF3, and correlated with p53 and IMP3. Women with high vs. low nestin had worse progression-free survival (PFS) and cancer-specific survival overall, and worse PFS in the subset who received no adjuvant therapy or radiation, or had early stage, type I disease or tumors with both low and high ER, PR, TFF3, PTEN, p53 or IMP3. The relationship between nestin and PFS was independent of stage, LVSI and risk categorization but not type of UC. CONCLUSIONS: High nestin was more common in UCs with aggressive features and poor outcome. Nestin may represent a predictive biomarker for treatment selection for patients previously considered to be lower risk and a candidate for no or radiation-based adjuvant therapy, and compliment ER/PR testing.

Endoplasmic reticulum stress in complex atypical hyperplasia as a possible predictor of occult carcinoma and progestin response.

Glucose-regulated protein (GRP)-78, the key regulator of endoplasmic reticulum (ER) stress, is associated with endometrial cancer (EC) development and progression. However, its role in the continuum from complex atypical hyperplasia (CAH) to EC is unknown and the focus of this study. METHODS: 252 formalin-fixed, paraffin-embedded endometrial biopsies from patients with CAH diagnosed between 2003 and 2011 were evaluated for GRP78 expression by immunohistochemistry. Expression was also evaluated in subsequent biopsies from those patients treated with progestins. Differences in GRP78 expression were assessed using standard statistical methods. RESULTS: GRP78 expression was undetectable in 45(18%) patients with CAH, while 120(48%) CAH cases showed moderate/strong expression. Among women who ultimately underwent hysterectomy for CAH (n=134), 54(40%) had occult EC while 57(43%) had persistent CAH. Those with occult EC upon hysterectomy had significantly stronger GRP78 expression than those who did not have occult EC (p=0.007). Greater GRP78 expression within CAH remained independently associated with the presence of an occult EC (p=0.017). Thirty-four of 54 (63%) patients with occult EC had moderate/strong GRP78 expression compared to 36 of 80 (45%) patients with persistent CAH, benign or non-atypical hyperplastic endometrium. In those treated with progestins, samples with persistent CAH and EC were more likely to have high levels of GRP78 expression in the initial biopsies than those who responded (p=0.014). CONCLUSIONS: Increased GRP78 expression in untreated CAH correlates with the presence of an occult EC. In addition, CAH specimens with greater GRP78 expression may identify patients who are less likely to respond to progestin therapy.

Microfocus of Anaplastic Carcinoma Arising in Mural Nodule of Ovarian Mucinous Borderline Tumor With Very Rapid and Fatal Outcome.

A 36-yr-old woman presented with abdominal discomfort. A computed tomography scan revealed a large left cystic and solid pelvic mass without evidence of metastatic disease. Total hysterectomy with bilateral salpingo-oophorectomy and tumor staging was performed. Grossly, the ovarian mass measured 20×18 cm and the cut surface was multiloculated with 1 single mural nodule measuring 2×1.5 cm. The histologic diagnosis of ovarian mucinous borderline tumor with a microfocus of anaplastic carcinoma arising in sarcoma-like mural nodule, FIGO Stage IA was rendered. After 3 mo, the patient returned with symptomatic anemia. A computed tomography scan showed enlarged retroperitoneal and pelvic lymph nodes. Image-guided biopsy of the pelvic lymph node showed a metastatic anaplastic carcinoma from her primary ovarian carcinoma. Chemotherapy was initiated, but the patient developed fulminant disseminated intravascular coagulation within <1 wk of her presentation which was fatal.

NPPB is a novel candidate biomarker expressed by cancer-associated fibroblasts in epithelial ovarian cancer.

Most solid tumors contain cancer-associated fibroblasts (CAFs) that support tumorigenesis and malignant progression. However, the cellular origins of CAFs in epithelial ovarian cancers (EOCs) remain poorly understood, and their utility as a source of clinical biomarkers for cancer diagnosis has not been explored in great depth. Here, we report establishing in vitro and in vivo models of CAFs in ovarian cancer development. Normal ovarian fibroblasts and mesenchymal stem cells cultured in the presence of EOC cells acquired a CAF-like phenotype, and promoted EOC cell migration in vitro. CAFs also promoted ovarian cancer growth in vivo in both subcutaneous and intraperitoneal murine xenograft assays. Molecular profiling of CAFs identified gene expression signatures that were highly enriched for extracellular and secreted proteins. We identified novel candidate CAF-specific biomarkers for ovarian cancer including NPPB, which was expressed in the stroma of 60% primary ovarian cancer tissues (n = 145) but not in the stroma of normal ovaries (n = 4). NPPB is a secreted protein that was also elevated in the blood of 50% of women with ovarian cancer (n = 8). Taken together, these data suggest that the tumor stroma is a novel source of biomarkers, including NPPB, that may be of clinical utility for detection of EOC.

Identification of novel candidate biomarkers of epithelial ovarian cancer by profiling the secretomes of three-dimensional genetic models of ovarian carcinogenesis.

Epithelial ovarian cancer (EOC) is still considered the most lethal gynecological malignancy and improved early detection of ovarian cancer is crucial to improving patient prognoses. To address this need, we tested whether candidate EOC biomarkers can be identified using three-dimensional (3D) in vitro models. We quantified changes in the abundance of secreted proteins in a 3D genetic model of early-stage EOC, generated by expressing CMYC and KRAS(G) (12) (V) in TERT-immortalized normal ovarian epithelial cells. Cellular proteins were labeled in live cells using stable isotopic amino acid analogues, and secreted proteins identified and quantified using liquid chromatography-tandem mass spectrometry. Thirty-seven and 55 proteins were differentially expressed by CMYC and CMYC+KRAS(G) (12) (V) expressing cells respectively (p < 0.05; >2-fold). We evaluated expression of the top candidate biomarkers in ∼210 primary EOCs: CHI3L1 and FKBP4 are both expressed by >96% of primary EOCs, and FASN and API5 are expressed by 86 and 75% of cases. High expression of CHI3L1 and FKBP4 was associated with worse patient survival (p = 0.042 and p = 0.002, respectively). Expression of LGALS3BP was positively associated with recurrence (p = 0.0001) and suboptimal debulking (p = 0.018) suggesting that these proteins may be novel prognostic biomarkers. Furthermore, within early stage tumours (I/II), high expression of API5, CHI3L1 and FASN was associated with high tumour grade (p = 3 × 10(-4) , p = 0.016, p = 0.010, respectively). We show in vitro cell biology models of early-stage cancer development can be used to identify novel candidate biomarkers for disease, and report the identification of proteins that represent novel potential candidate diagnostic and prognostic biomarkers for this highly lethal disease.

Prediction of concurrent endometrial carcinoma in women with endometrial hyperplasia.

OBJECTIVE: Although a fraction of endometrial hyperplasia cases have concurrent endometrial carcinoma, patient characteristics associated with concurrent malignancy are not well described. The aim of our study was to identify predictive clinico-pathologic factors for concurrent endometrial carcinoma among patients with endometrial hyperplasia. METHODS: A case-control study was conducted to compare endometrial hyperplasia in both preoperative endometrial biopsy and hysterectomy specimens (n=168) and endometrial carcinoma in hysterectomy specimen but endometrial hyperplasia in preoperative endometrial biopsy (n=43). Clinico-pathologic factors were examined to identify independent risk factors of concurrent endometrial carcinoma in a multivariate logistic regression model. RESULTS: The most common histologic subtype in preoperative endometrial biopsy was complex hyperplasia with atypia [CAH] (n=129) followed by complex hyperplasia without atypia (n=58) and simple hyperplasia with or without atypia (n=24). The majority of endometrial carcinomas were grade 1 (86.0%) and stage I (83.7%). In multivariate analysis, age 40-59 (odds ratio [OR] 3.07, p=0.021), age≥60 (OR 6.65, p=0.005), BMI≥35kg/m(2) (OR 2.32, p=0.029), diabetes mellitus (OR 2.51, p=0.019), and CAH (OR 9.01, p=0.042) were independent predictors of concurrent endometrial carcinoma. The risk of concurrent endometrial carcinoma rose dramatically with increasing number of risk factors identified in multivariate model (none 0%, 1 risk factor 7.0%, 2 risk factors 17.6%, 3 risk factors 35.8%, and 4 risk factors 45.5%, p<0.001). Hormonal treatment was associated with decreased risk of concurrent endometrial cancer in those with ≥3 risk factors. CONCLUSIONS: Older age, obesity, diabetes mellitus, and CAH are predictive of concurrent endometrial carcinoma in endometrial hyperplasia patients.

Efficacy of vaccination with recombinant vaccinia and fowlpox vectors expressing NY-ESO-1 antigen in ovarian cancer and melanoma patients.

Recombinant poxviruses (vaccinia and fowlpox) expressing tumor-associated antigens are currently being evaluated in clinical trials as cancer vaccines to induce tumor-specific immune responses that will improve clinical outcome. To test whether a diversified prime and boost regimen targeting NY-ESO-1 will result in clinical benefit, we conducted two parallel phase II clinical trials of recombinant vaccinia-NY-ESO-1 (rV-NY-ESO-1), followed by booster vaccinations with recombinant fowlpox-NY-ESO-1 (rF-NY-ESO-1) in 25 melanoma and 22 epithelial ovarian cancer (EOC) patients with advanced disease who were at high risk for recurrence/progression. Integrated NY-ESO-1-specific antibody and CD4(+) and CD8(+) T cells were induced in a high proportion of melanoma and EOC patients. In melanoma patients, objective response rate [complete and partial response (CR+PR)] was 14%, mixed response was 5%, and disease stabilization was 52%, amounting to a clinical benefit rate (CBR) of 72% in melanoma patients. The median PFS in the melanoma patients was 9 mo (range, 0-84 mo) and the median OS was 48 mo (range, 3-106 mo). In EOC patients, the median PFS was 21 mo (95% CI, 16-29 mo), and median OS was 48 mo (CI, not estimable). CD8(+) T cells derived from vaccinated patients were shown to lyse NY-ESO-1-expressing tumor targets. These data provide preliminary evidence of clinically meaningful benefit for diversified prime and boost recombinant pox-viral-based vaccines in melanoma and ovarian cancer and support further evaluation of this approach in these patient populations.

Expression of DNA repair proteins in endometrial cancer predicts disease outcome.

OBJECTIVE: The consequences of defective homologous recombination and other DNA repair pathways are important in disease outcomes of numerous tumor types. The objective of this study was to explore BRCA1, PARP, FANCD2, PTEN, H2AX, and ATM protein expression in endometrial cancer (EC). METHODS: PARP1, γH2AX, ATM, FANCD2, PTEN, BRCA1, and p53 proteins were evaluated in EC tissue microarray (TMA) and their expressions were correlated with clinical and pathological parameters in 357 patients. RESULTS: In type I EC, PARP1(+), ATM(+), and FANCD2(+) were associated with high tumor grade (p 0.031, p 0.0045, p 0.0062 respectively); γH2AX(+) and FANCD2(+) with advanced tumor stage (p 0.0004, p 0.0085 respectively); γH2AX(+), FANCD2(+) and p53(+) with the presence of lympho-vascular invasion (p 0.0004, p 0.0042, p 0.0098 respectively); and γH2AX(+) and ATM(+) with tumor recurrence (p 0.0203, p 0.0465) respectively. In type II EC, only PARP1(+) was associated with tumor stage (p 0.0499). EC patients with p53(+) or FANCD2(+) were more likely to recur with 5year recurrence free survival (RFS) probability of 71.4% in comparison to 85.5% for the other patients and they were more likely to have shorter 5year overall survival (OS) of 66.46% in comparison to 78.5% of those other patients Finally, patients with ATM(+) and p53(+) or FANCD2(+) were more likely to recur with 5year RFS probability of 68% versus 80.3% for the other patients. CONCLUSION: DNA repair proteins seemed to play an important role in EC, and their expressions can forecast for poor outcomes.

Proteogenomic analysis of enriched HGSOC tumor epithelium identifies prognostic signatures and therapeutic vulnerabilities.

We performed a deep proteogenomic analysis of bulk tumor and laser microdissection enriched tumor cell populations from high-grade serous ovarian cancer (HGSOC) tissue specimens spanning a broad spectrum of purity. We identified patients with longer progression-free survival had increased immune-related signatures and validated proteins correlating with tumor-infiltrating lymphocytes in 65 tumors from an independent cohort of HGSOC patients, as well as with overall survival in an additional 126 HGSOC patient cohort. We identified that homologous recombination deficient (HRD) tumors are enriched in pathways associated with metabolism and oxidative phosphorylation that we validated in independent patient cohorts. We further identified that polycomb complex protein BMI-1 is elevated in HR proficient (HRP) tumors, that elevated BMI-1 correlates with poor overall survival in HRP but not HRD HGSOC patients, and that HRP HGSOC cells are uniquely sensitive to BMI-1 inhibition.