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Repeated measures studies are frequently performed in patient-derived xenograft (PDX) models to evaluate drug activity or compare effectiveness of cancer treatment regimens. Linear mixed effects regression models were used to perform statistical modeling of tumor growth data. Biologically plausible structures for the covariation between repeated tumor burden measurements are explained. Graphical, tabular, and information criteria tools useful for choosing the mean model functional form and covariation structure are demonstrated in a Case Study of five PDX models comparing cancer treatments. Power calculations were performed via simulation. Linear mixed effects regression models applied to the natural log scale were shown to describe the observed data well. A straight growth function fit well for two PDX models. Three PDX models required quadratic or cubic polynomial (time squared or cubed) terms to describe delayed tumor regression or initial tumor growth followed by regression. Spatial(power), spatial(power) + RE, and RE covariance structures were found to be reasonable. Statistical power is shown as a function of sample size for different levels of variation. Linear mixed effects regression models provide a unified and flexible framework for analysis of PDX repeated measures data, use all available data, and allow estimation of tumor doubling time.

Objective: Chimeric antigen receptor (CAR)-T cell strategies ideally target a surface antigen that is exclusively and uniformly expressed by tumors; however, no such antigen is known for high-grade serous ovarian carcinoma (HGSC). A potential solution involves combinatorial antigen targeting with AND or OR logic-gating. Therefore, we investigated co-expression of CA125, Mesothelin (MSLN) and Folate Receptor alpha (FOLRA) on individual tumor cells in HGSC., Methods: RNA expression of CA125, MSLN, and FOLR1 was assessed using TCGA (HGSC) and GTEx (healthy tissues) databases. Antigen expression profiles and CD3+, CD8+ and CD20+ tumor-infiltrating lymphocyte (TIL) patterns were assessed in primary and recurrent HGSC by multiplex immunofluorescence and immunohistochemistry., Results: At the transcriptional level, each antigen was overexpressed in >90% of cases; however, MSLN and FOLR1 showed substantial expression in healthy tissues. At the protein level, CA125 was expressed by the highest proportion of cases and tumor cells per case, followed by MSLN and FOLRA. The most promising pairwise combination was CA125 and/or MSLN (OR gate), with 51.9% of cases containing ≥90% of tumor cells expressing one or both antigens. In contrast, only 5.8% of cases contained ≥90% of tumor cells co-expressing CA125 and MSLN (AND gate). Antigen expression patterns showed modest correlations with TIL. Recurrent tumors retained expression of all three antigens and showed increased TIL densities., Conclusions: An OR-gated CAR-T cell strategy against CA125 and MSLN would target the majority of tumor cells in most cases. Antigen expression and T-cell infiltration patterns are favorable for this strategy in primary and recurrent disease., Competing Interests: Declaration of Competing Interest MSB discloses involvement in clinical trials supported by the following companies: Merck, Marker Therapeutics, Transgene, Immune Design, Bristol-Myers Squib, Genentech, and Pharmacyclics. MM discloses involvement with the following companies: Celgene, Genentech, Kite Pharma, Morphosys, Nanostring, and Pfizer. BHN discloses involvement with Virogin Biotech, Akamara Therapeutics, IMV Technologies and Innovakine Therapuetics. None of these relationships influence or are influenced by the results presented in this study., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Purpose: Clear cell ovarian carcinoma (CCOC) is an aggressive disease that often demonstrates resistance to standard chemotherapies. Approximately 25% of patients with CCOC show a strong APOBEC mutation signature. Here, we determine which APOBEC3 enzymes are expressed in CCOC, establish clinical correlates, and identify a new biomarker for detection and intervention., Experimental Designs: APOBEC3 expression was analyzed by IHC and qRT-PCR in a pilot set of CCOC specimens ( n = 9 tumors). The IHC analysis of APOBEC3B was extended to a larger cohort to identify clinical correlates ( n = 48). Dose-response experiments with platinum-based drugs in CCOC cell lines and carboplatin treatment of patient-derived xenografts (PDXs) were done to address mechanistic linkages., Results: One DNA deaminase, APOBEC3B, is overexpressed in a formidable subset of CCOC tumors and is low or absent in normal ovarian and fallopian tube epithelial tissues. High APOBEC3B expression associates with improved progression-free survival ( P = 0.026) and moderately with overall survival ( P = 0.057). Cell-based studies link APOBEC3B activity and subsequent uracil processing to sensitivity to cisplatin and carboplatin. PDX studies extend this mechanistic relationship to CCOC tissues., Conclusions: These studies demonstrate that APOBEC3B is overexpressed in a subset of CCOC and, contrary to initial expectations, associated with improved (not worse) clinical outcomes. A likely molecular explanation is that APOBEC3B-induced DNA damage sensitizes cells to additional genotoxic stress by cisplatin. Thus, APOBEC3B is a molecular determinant and a candidate predictive biomarker of the therapeutic response to platinum-based chemotherapy. These findings may have broader translational relevance, as APOBEC3B is overexpressed in many different cancer types., (©2020 American Association for Cancer Research.)

Background: Poly(adenosine diphosphate ribose) polymerase (PARP) inhibitors exhibit promising activity against ovarian cancers, but their efficacy can be limited by acquired drug resistance. This study explores the role of autophagy in regulating the sensitivity of ovarian cancer cells to PARP inhibitors., Methods: Induction of autophagy was detected by punctate LC3 fluorescence staining, LC3I to LC3II conversion on Western blot analysis, and electron microscopy. Enhanced growth inhibition and apoptosis were observed when PARP inhibitors were used with hydroxychloroquine, chloroquine (CQ), or LYS05 to block the hydrolysis of proteins and lipids in autophagosomes or with small interfering RNA against ATG5 or ATG7 to prevent the formation of autophagosomes. The preclinical efficacy of the combination of CQ and olaparib was evaluated with a patient-derived xenograft (PDX) and the OVCAR8 human ovarian cancer cell line., Results: Four PARP inhibitors (olaparib, niraparib, rucaparib, and talazoparib) induced autophagy in a panel of ovarian cancer cells. Inhibition of autophagy with CQ enhanced the sensitivity of ovarian cancer cells to PARP inhibitors. In vivo, olaparib and CQ produced additive growth inhibition in OVCAR8 xenografts and a PDX. Olaparib inhibited PARP activity, and this led to increased reactive oxygen species (ROS) and an accumulation of γ-H2AX. Inhibition of autophagy also increased ROS and γ-H2AX and enhanced the effect of olaparib on both entities. Treatment with olaparib increased phosphorylation of ATM and PTEN while decreasing the phosphorylation of AKT and mTOR and inducing autophagy., Conclusions: PARP inhibitor-induced autophagy provides an adaptive mechanism of resistance to PARP inhibitors in cancer cells with wild-type BRCA, and a combination of PARP inhibitors with CQ or other autophagy inhibitors could improve outcomes for patients with ovarian cancer., (© 2019 American Cancer Society.)

Objectives: Inhibition of pregnancy-associated plasma protein-A (PAPP-A), an upstream activator of the insulin-like growth factor (IGF) pathway, is known to augment sensitivity to platinum-based chemotherapy. This study further tests the efficacy of PAPP-A inhibition with a monoclonal antibody inhibitor (mAb-PA) in ovarian cancer (OC) platinum-resistant patient-derived xenograft (PDX) models., Methods: PAPP-A expression was quantitated in platinum-resistant PDX models by ELISA. A subset with High (n = 5) and Low (n = 2) expression were revived in female SCID/beige mice for studies with either saline, carboplatin/paclitaxel (CP) + mAb-PA, or CP + IgG2a. The primary endpoint was tumor area by ultrasound on day 28 relative to baseline. Conversion to platinum-sensitive was defined by average tumor regression below baseline. Statistical analyses included linear mixed effects modeling and Kaplan Meier curves. Response to therapy was correlated with changes in the ratio of phosphorylated/total AKT and ERK 1/2 using Wes analysis., Results: The addition of mAb-PA to CP induced tumor regression below baseline in one High PAPP-A PDX model; another three models exhibited notable growth inhibition relative to CP + IgG2a. None of the Low PAPP-A PDX models regressed below baseline. The PDX model with the greatest magnitude of tumor regression from baseline after combination therapy was maintained on single agent mAb-PA or IgG2a, but no benefit was observed. Decreased phosphorylation of ERK1/2 correlated with conversion to platinum-sensitive., Conclusions: The addition of mAb-PA to CP overcame platinum-resistance in one of five High PAPP-A PDX models; three other models demonstrated improved platinum-response. This supports further clinical development of this novel therapeutic., Competing Interests: Dr. Cheryl A. Conover holds a patent(US 8,653,020) on the monoclonal antibody which can selective exosite inhibition of PAPP-A activity against IGFBP-4). This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Objective: Poly(ADP-ribose) polymerase (PARP) inhibitors have shown substantial activity in homologous recombination- (HR-) deficient ovarian cancer and are undergoing testing in other HR-deficient tumors. For reasons that are incompletely understood, not all patients with HR-deficient cancers respond to these agents. Preclinical studies have demonstrated that changes in alternative DNA repair pathways affect PARP inhibitor (PARPi) sensitivity in ovarian cancer models. This has not previously been assessed in the clinical setting., Methods: Clonogenic and plasmid-based HR repair assays were performed to compare BRCA1-mutant COV362 ovarian cancer cells with or without 53BP1 gene deletion. Archival biopsies from ovarian cancer patients in the phase I, open-label clinical trial of PARPi ABT-767 were stained for PARP1, RAD51, 53BP1 and multiple components of the nonhomologous end-joining (NHEJ) DNA repair pathway. Modified histochemistry- (H-) scores were determined for each repair protein in each sample. HRD score was determined from tumor DNA., Results: 53BP1 deletion increased HR in BRCA1-mutant COV362 cells and decreased PARPi sensitivity in vitro. In 36 women with relapsed ovarian cancer, responses to the PARPi ABT-767 were observed exclusively in cancers with HR deficiency. In this subset, 7 of 18 patients (39%) had objective responses. The actual HRD score did not further correlate with change from baseline tumor volume (r = 0.050; p = 0.87). However, in the HR-deficient subset, decreased 53BP1 H-score was associated with decreased antitumor efficacy of ABT-767 (r = -0.69, p = 0.004)., Conclusion: Differences in complementary repair pathways, particularly 53BP1, correlate with PARPi response of HR-deficient ovarian cancers., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Purpose: Here we assess whether molecular subtyping identifies biological features of tumors that correlate with survival and surgical outcomes of high-grade serous ovarian cancer (HGSOC). Experimental Design: Consensus clustering of pooled mRNA expression data from over 2,000 HGSOC cases was used to define molecular subtypes of HGSOCs. This de novo classification scheme was then applied to 381 Mayo Clinic HGSOC patients with detailed survival and surgical outcome information. Results: Five molecular subtypes of HGSOC were identified. In the pooled dataset, three subtypes were largely concordant with prior studies describing proliferative, mesenchymal, and immunoreactive tumors (concordance > 70%), and the group of tumors previously described as differentiated type was segregated into two new types, one of which (anti-mesenchymal) had downregulation of genes that were typically upregulated in the mesenchymal subtype. Molecular subtypes were significantly associated with overall survival ( P < 0.001) and with rate of optimal surgical debulking (≤1 cm, P = 1.9E-4) in the pooled dataset. Among stage III-C or IV Mayo Clinic patients, molecular subtypes were also significantly associated with overall survival ( P = 0.001), as well as rate of complete surgical debulking (no residual disease; 16% in mesenchymal tumors compared with >28% in other subtypes; P = 0.02). Conclusions: HGSOC tumors may be categorized into five molecular subtypes that associate with overall survival and the extent of residual disease following debulking surgery. Because mesenchymal tumors may have features that were associated with less favorable surgical outcome, molecular subtyping may have future utility in guiding neoadjuvant treatment decisions for women with HGSOC. Clin Cancer Res; 23(15); 4077-85. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Purpose: Recent progress in understanding the molecular biology of epithelial ovarian cancer has not yet translated into individualized treatment for these women or improvements in their disease outcome. Gene expression has been utilized to identify distinct molecular subtypes, but there have been no reports investigating whether or not molecular subtyping is predictive of response to bevacizumab in ovarian cancer. Experimental Design: DASL gene expression arrays were performed on FFPE tissue from patients enrolled on the ICON7 trial. Patients were stratified into four TCGA molecular subtypes. Associations between molecular subtype and the efficacy of randomly assigned therapy with bevacizumab were assessed. Results: Molecular subtypes were assigned as follows: 122 immunoreactive (34%), 96 proliferative (27%), 73 differentiated (20%), and 68 mesenchymal (19%). In univariate analysis patients with tumors of proliferative subtype obtained the greatest benefit from bevacizumab with a median PFS improvement of 10.1 months [HR, 0.55 (95% CI, 0.34-0.90), P = 0.016]. For the mesenchymal subtype, bevacizumab conferred a nonsignificant improvement in PFS of 8.2 months [HR 0.78 (95% CI, 0.44-1.40), P = 0.41]. Bevacizumab conferred modest improvements in PFS for patients with immunoreactive subtype (3.8 months; P = 0.08) or differentiated subtype (3.7 months; P = 0.61). Multivariate analysis demonstrated significant PFS improvement in proliferative subtype patients only [HR, 0.45 (95% CI, 0.27-0.74), P = 0.0015]. Conclusions: Ovarian carcinoma molecular subtypes with the poorest survival (proliferative and mesenchymal) derive a comparably greater benefit from treatment that includes bevacizumab. Validation of our findings in an independent cohort could enable the use of bevacizumab for those patients most likely to benefit, thereby reducing side effects and healthcare cost. Clin Cancer Res; 23(14); 3794-801. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Purpose: APOBEC3 DNA cytosine deaminase family members normally defend against viruses and transposons. However, deregulated APOBEC3 activity causes mutations in cancer. Because of broad expression profiles and varying mixtures of normal and cancer cells in tumors, including immune cell infiltration, it is difficult to determine where different APOBEC3s are expressed. Here, we ask whether correlations exist between APOBEC3 expression and T-cell infiltration in high-grade serous ovarian cancer (HGSOC), and assess whether these correlations have prognostic value., Experimental Design: Transcripts for APOBEC3G, APOBEC3B, and the T-cell markers, CD3D, CD4, CD8A, GZMB, PRF1, and RNF128 were quantified by RT-qPCR for a cohort of 354 HGSOC patients. Expression values were correlated with each other and clinical parameters. Two additional cohorts were used to extend HGSOC clinical results. Immunoimaging was used to colocalize APOBEC3G and the T-cell marker CD3. TCGA data extended expression analyses to additional cancer types., Results: A surprising positive correlation was found for expression of APOBEC3G and several T cell genes in HGSOC. Immunohistochemistry and immunofluorescent imaging showed protein colocalization in tumor-infiltrating T lymphocytes. High APOBEC3G expression correlated with improved outcomes in multiple HGSOC cohorts. TCGA data analyses revealed that expression of APOBEC3D and APOBEC3H also correlates with CD3D across multiple cancer types., Conclusions: Our results identify APOBEC3G as a new candidate biomarker for tumor-infiltrating T lymphocytes and favorable prognoses for HGSOC. Our data also highlight the complexity of the tumor environment with respect to differential APOBEC family gene expression in both tumor and surrounding normal cell types. Clin Cancer Res; 22(18); 4746-55. ©2016 AACR., Competing Interests: R.S.H. is a co-founder of ApoGen Biotechnologies Inc. The other authors have no conflicts of interest to disclose., (©2016 American Association for Cancer Research.)

The immune system constitutes one of the host factors modifying outcomes in ovarian cancer. Regulatory T cells (Tregs) are believed to be a major factor in preventing the immune response from destroying ovarian cancers. Understanding mechanisms that regulate Tregs in the tumor microenvironment could lead to the identification of novel targets aimed at reducing their influence. In this study, we used immunofluorescence-based microscopy to enumerate Tregs, total CD4 T cells, and CD8(+) cytotoxic T cells in fresh frozen tumors from over 400 patients with ovarian cancer (>80 % high-grade serous). We sought to determine whether Tregs were associated with survival and genetic variation in 79 genes known to influence Treg induction, trafficking, or function. We used Cox regression, accounting for known prognostic factors, to estimate hazard ratios (HRs) associated with T cell counts and ratios. We found that the ratios of CD8 T cells and total CD4 T cells to Tregs were associated with improved overall survival (CD8/Treg HR 0.84, p = 0.0089; CD4/Treg HR 0.88, p = 0.046) and with genetic variation in IL-10 (p = 0.0073 and 0.01, respectively). In multivariate analyses, the associations between the ratios and overall survival remained similar (IL-10 and clinical covariate-adjusted CD8/Treg HR 0.85, p = 0.031; CD4/Treg HR 0.87, p = 0.093), suggesting that this association was not driven by variation in IL-10. Thus, integration of novel tumor phenotyping measures with extensive clinical and genetic information suggests that the ratio of T cells to Tregs may be prognostic of outcome in ovarian cancer, regardless of inherited genotype in genes related to Tregs.

Ovarian cancer represents the most lethal tumor type among malignancies of the female reproductive system. Overall survival rates remain low. In this study, we identify the serine protease inhibitor Kazal type 1 (SPINK1) as a potential therapeutic target for a subset of ovarian cancers. We show that SPINK1 drives ovarian cancer cell proliferation through activation of epidermal growth factor receptor (EGFR) signaling, and that SPINK1 promotes resistance to anoikis through a distinct mechanism involving protease inhibition. In analyses of ovarian tumor specimens from a Mayo Clinic cohort of 490 patients, we further find that SPINK1 immunostaining represents an independent prognostic factor for poor survival, with the strongest association in patients with nonserous histological tumor subtypes (endometrioid, clear cell, and mucinous). This study provides novel insight into the fundamental processes underlying ovarian cancer progression, and also suggests new avenues for development of molecularly targeted therapies.

Objectives: Inflammation is a common feature of epithelial ovarian cancer (EOC), and measurement of plasma markers of inflammation might identify candidate markers for use in screening or presurgical evaluation of patients with adnexal masses., Methods: Plasma specimens from cohorts of 100 patients with advanced EOC (AJCC Stage III and IV), 50 patients with early stage EOC (Stage I and II), and 50 patients with benign surgical conditions were assayed for concentrations of multiple cytokines, toll-like receptor agonists, and vascular growth factors via ELISA and electrochemiluminescence. Immune proteins were then analyzed for association with EOC. Differences in plasma protein levels between benign, early, and advanced EOC patient groups were assessed with and without adjustment for plasma cancer antigen 125 (CA-125) levels., Results: Out of 23 proteins tested, six-including interferon gamma (IFNγ), interleukin 6 (IL-6), IL-8, IL-10, tumor necrosis factor alpha (TNFα), and placental growth factor (PlGF)-were univariately associated with EOC (all p<0.005), and one-IL-6-was associated with early stage EOC (p<0.0001). Heat shock protein 90kDa beta member 1 (HSP90B1, gp96) was associated with EOC and early stage EOC with borderline statistical significance (p=0.039 and p=0.026, respectively). However, when adjusted for (CA-125), only HSP90B1 independently predicted EOC (p=0.008), as well as early stage EOC (p=0.014)., Conclusions: Multiple plasma cytokines, including IFNγ, IL-6, IL-8, IL-10, TNFα, PlGF, and HSP90B1 are associated with EOC. Of these, HSP90B1 is associated with EOC independent from the biomarker CA-125., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Objectives: Epithelial ovarian cancer (EOC) is an aggressive disease in which first line therapy consists of a surgical staging/debulking procedure and platinum based chemotherapy. There is significant interest in clinically applicable, easy to use prognostic tools to estimate risk of recurrence and overall survival. In this study we used a large prospectively collected cohort of women with EOC to validate currently published models and assess prognostic variables., Methods: Women with invasive ovarian, peritoneal, or fallopian tube cancer diagnosed between 2000 and 2011 and prospectively enrolled into the Mayo Clinic Ovarian Cancer registry were identified. Demographics and known prognostic markers as well as epidemiologic exposure variables were abstracted from the medical record and collected via questionnaire. Six previously published models of overall and recurrence-free survival were assessed for external validity. In addition, predictors of outcome were assessed in our dataset., Results: Previously published models validated with a range of c-statistics (0.587-0.827), though application of models containing variables which are not part of routine practice were somewhat limited by missing data; utilization of all applicable models and comparison of results are suggested. Examination of prognostic variables identified only the presence of ascites and ASA score to be independent predictors of prognosis in our dataset, albeit with marginal gain in prognostic information, after accounting for stage and debulking., Conclusions: Existing prognostic models for newly diagnosed EOC showed acceptable calibration in our cohort for clinical application. However, modeling of prospective variables in our dataset reiterates that stage and debulking remains the most important predictors of prognosis in this setting., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Edmonston vaccine strains of measles virus (MV) have significant antitumor activity in mouse xenograft models of ovarian cancer. MV engineered to express the sodium iodide symporter gene (MV-NIS) facilitates localization of viral gene expression and offers a tool for tumor radiovirotherapy. Here, we report results from a clinical evaluation of MV-NIS in patients with taxol- and platinum-resistant ovarian cancer. MV-NIS was given intraperitoneally every 4 weeks for up to 6 cycles. Treatment was well tolerated and associated with promising median overall survival in these patients with heavily pretreated ovarian cancer; no dose-limiting toxicity was observed in 16 patients treated at high-dose levels (10(8)-10(9) TCID50), and their median overall survival of 26.5 months compared favorably with other contemporary series. MV receptor CD46 and nectin-4 expression was confirmed by immunohistochemistry in patient tumors. Sodium iodide symporter expression in patient tumors after treatment was confirmed in three patients by (123)I uptake on SPECT/CTs and was associated with long progression-free survival. Immune monitoring posttreatment showed an increase in effector T cells recognizing the tumor antigens IGFBP2 and FRα, indicating that MV-NIS treatment triggered cellular immunity against the patients' tumor and suggesting that an immune mechanism mediating the observed antitumor effect. Our findings support further clinical evaluation of MV-NIS as an effective immunovirotherapy., (©2014 American Association for Cancer Research.)

The presence of regulatory T cells (Treg) in solid tumors is known to play a role in patient survival in ovarian cancer and other malignancies. We assessed inherited genetic variations via 749 tag single-nucleotide polymorphisms (SNP) in 25 Treg-associated genes (CD28, CTLA4, FOXP3, IDO1, IL10, IL10RA, IL15, 1L17RA, IL23A, IL23R, IL2RA, IL6, IL6R, IL8, LGALS1, LGALS9, MAP3K8, STAT5A, STAT5B, TGFB1, TGFB2, TGFB3, TGFBR1, TGRBR2, and TGFBR3) in relation to ovarian cancer survival. We analyzed genotype and overall survival in 10,084 women with invasive epithelial ovarian cancer, including 5,248 high-grade serous, 1,452 endometrioid, 795 clear cell, and 661 mucinous carcinoma cases of European descent across 28 studies from the Ovarian Cancer Association Consortium (OCAC). The strongest associations were found for endometrioid carcinoma and IL2RA SNPs rs11256497 [HR, 1.42; 95% confidence interval (CI), 1.22-1.64; P = 5.7 × 10(-6)], rs791587 (HR, 1.36; 95% CI, 1.17-1.57; P = 6.2 × 10(-5)), rs2476491 (HR, = 1.40; 95% CI, 1.19-1.64; P = 5.6 × 10(-5)), and rs10795763 (HR, 1.35; 95% CI, 1.17-1.57; P = 7.9 × 10(-5)), and for clear cell carcinoma and CTLA4 SNP rs231775 (HR, 0.67; 95% CI, 0.54-0.82; P = 9.3 × 10(-5)) after adjustment for age, study site, population stratification, stage, grade, and oral contraceptive use. The rs231775 allele associated with improved survival in our study also results in an amino acid change in CTLA4 and previously has been reported to be associated with autoimmune conditions. Thus, we found evidence that SNPs in genes related to Tregs seem to play a role in ovarian cancer survival, particularly in patients with clear cell and endometrioid epithelial ovarian cancer.

Purpose: Ovarian cancer has a high recurrence and mortality rate. A barrier to improved outcomes includes a lack of accurate models for preclinical testing of novel therapeutics., Experimental Design: Clinically relevant, patient-derived tumorgraft models were generated from sequential patients and the first 168 engrafted models are described. Fresh ovarian, primary peritoneal, and fallopian tube carcinomas were collected at the time of debulking surgery and injected intraperitoneally into severe combined immunodeficient mice., Results: Tumorgrafts demonstrated a 74% engraftment rate with microscopic fidelity of primary tumor characteristics. Low-passage tumorgrafts also showed comparable genomic aberrations with the corresponding primary tumor and exhibit gene set enrichment of multiple ovarian cancer molecular subtypes, similar to patient tumors. Importantly, each of these tumorgraft models is annotated with clinical data and for those that have been tested, response to platinum chemotherapy correlates with the source patient., Conclusions: Presented herein is the largest known living tumor bank of patient-derived, ovarian tumorgraft models that can be applied to the development of personalized cancer treatment., (©2014 AACR)

Ovarian cancer is a clinically and molecularly heterogeneous disease. The driving forces behind this variability are unknown. Here, we report wide variation in the expression of the DNA cytosine deaminase APOBEC3B, with elevated expression in the majority of ovarian cancer cell lines (three SDs above the mean of normal ovarian surface epithelial cells) and high-grade primary ovarian cancers. APOBEC3B is active in the nucleus of several ovarian cancer cell lines and elicits a biochemical preference for deamination of cytosines in 5'-TC dinucleotides. Importantly, examination of whole-genome sequence from 16 ovarian cancers reveals that APOBEC3B expression correlates with total mutation load as well as elevated levels of transversion mutations. In particular, high APOBEC3B expression correlates with C-to-A and C-to-G transversion mutations within 5'-TC dinucleotide motifs in early-stage high-grade serous ovarian cancer genomes, suggesting that APOBEC3B-catalyzed genomic uracil lesions are further processed by downstream DNA "repair" enzymes including error-prone translesion polymerases. These data identify a potential role for APOBEC3B in serous ovarian cancer genomic instability., (©2013 AACR.)

Ovarian cancer is an immune reactive malignancy with a complex immune suppressive network that blunts successful immune eradication. This suppressive microenvironment may be mediated by recruitment or induction of CD4(+) regulatory T cells (Tregs). Our study sought to investigate the association of tumor-infiltrating CD4(+)CD25(+)FOXP3(+) Tregs, and other immune factors, with clinical outcome in serous ovarian cancer patients. We performed immunofluorescence and quantification of intraepithelial tumor-infiltrating triple positive Tregs (CD4(+)CD25(+)FOXP3(+)), as well as CD4(+)CD25(+)FOXP3(-), CD3(+) and CD8(+) T cells in tumor specimens from 52 patients with high stage serous ovarian carcinoma. Thirty-one of the patients had good survival (i.e. > 60 months) and 21 had poor survival of < 18 months. Total cell counts as well as cell ratios were compared among these two outcome groups. The total numbers of CD4(+)CD25(+)FOXP3(+) Tregs, CD4(+)CD25(+)FOXP3(-), CD3(+) and CD8(+) cells were not significantly different between the groups. However, higher ratios of CD8(+)/CD4(+)CD25(+)FOXP3(+) Treg, CD8(+)/CD4(+) and CD8/CD4(+)CD25(+)FOXP3(-) cells were seen in the good outcome group when compared to the patients with poor outcome. These data show for the first time that the ratios of CD8(+) to both CD4(+)CD25(+)FOXP3(+) Tregs and CD4(+)CD25(+)FOXP3(-) T cells are associated with disease outcome in ovarian cancer. The association being apparent in ratios rather than absolute count of T cells suggests that the effector/suppressor ratio may be a more important indicator of outcome than individual cell count. Thus, immunotherapy strategies that modify the ratio of CD4(+)CD25(+)FOXP3(+) Tregs or CD4(+)CD25(+)FOXP3(-) T cells to CD8(+) effector cells may be useful in improving outcomes in ovarian cancer.

Background: Epithelial ovarian cancer (EOC) is the fifth leading cause of cancer-related death in females and leading gynecologic cause of cancer-related death. Despite the identification of a number of serum biomarkers, methods to identify early-stage disease and predict prognosis remain scarce. We have evaluated two biologically connected serum biomarkers, serum leukocyte protease inhibitor (SLPI) and progranulin (PGRN)., Methods: Two-hundred frozen plasma samples were acquired from the Mayo Clinic Biospecimen Repository for Ovarian Cancer Research. Samples were obtained from 50 patients with benign conditions, 50 with American Joint Committee on Cancer (AJCC) stage I and II EOC, and 100 with AJCC stage III and IV EOC. Samples were obtained before surgical resection of a mass and were analyzed for absolute levels of SLPI and PGRN using ELISA assays. Receiver-operator characteristic curves were generated for SLPI and PGRN. Median follow-up was 48 months., Results: Absolute levels of SLPI were significantly elevated in patients with EOC compared with benign disease and predicted the presence of EOC (AUC of 0.812; P = 0.04); SLPI remained elevated in the subset of patients with normal CA-125. PGRN levels were not significantly increased in patients with early-stage or late-stage EOC as a whole, but an increase in PGRN levels was associated with decreased overall survival in advanced EOC., Conclusions: SLPI levels are elevated in EOC, and SLPI shows promise as a diagnostic biomarker for patients with both elevated and normal CA-125 levels. An increase in PGRN is associated with decreased overall survival., Impact: SLPI is elevated in EOC and warrants investigation in a screening study in women at risk for EOC.

There is a significant variation of outcome among ovarian cases. Clinical features such as age, stage, comorbidities, or degree of debulking are known prognostic factors for the disease. However, additional variation remains unexplained, some of which may be due to inherited factors. Here, we describe identification of survival-associated inherited variants in ovarian cancer that can enhance our current prognostic capabilities.

Although ovarian cancer is the most lethal of gynecologic malignancies, wide variation in outcome following conventional therapy continues to exist. The presence of tumor-infiltrating regulatory T cells (Tregs) has a role in outcome of this disease, and a growing body of data supports the existence of inherited prognostic factors. However, the role of inherited variants in genes encoding Treg-related immune molecules has not been fully explored. We analyzed expression quantitative trait loci (eQTL) and sequence-based tagging single nucleotide polymorphisms (tagSNPs) for 54 genes associated with Tregs in 3,662 invasive ovarian cancer cases. With adjustment for known prognostic factors, suggestive results were observed among rarer histological subtypes; poorer survival was associated with minor alleles at SNPs in RGS1 (clear cell, rs10921202, p=2.7×10(-5)), LRRC32 and TNFRSF18/TNFRSF4 (mucinous, rs3781699, p=4.5×10(-4), and rs3753348, p=9.0×10(-4), respectively), and CD80 (endometrioid, rs13071247, p=8.0×10(-4)). Fo0r the latter, correlative data support a CD80 rs13071247 genotype association with CD80 tumor RNA expression (p=0.006). An additional eQTL SNP in CD80 was associated with shorter survival (rs7804190, p=8.1×10(-4)) among all cases combined. As the products of these genes are known to affect induction, trafficking, or immunosuppressive function of Tregs, these results suggest the need for follow-up phenotypic studies.

Purpose: Due to variation of outcome among cases, we sought to examine whether overall survival in ovarian cancer was associated with common inherited variants in 227 candidate genes from ovarian cancer-related pathways including angiogenesis, inflammation, detoxification, glycosylation, one-carbon transfer, apoptosis, cell cycle regulation, and cellular senescence., Experimental Design: Blood samples were obtained from 325 women with invasive epithelial ovarian cancer diagnosed at the Mayo Clinic from 1999 to 2006. During a median follow-up of 3.8 years (range, 0.1-8.6 years), 157 deaths were observed. Germline DNA was analyzed at 1,416 single nucleotide polymorphisms (SNP). For all patients, and for 203 with serous subtype, we assessed the overall significance of each gene and pathway, and estimated risk of death via hazard ratios (HR) and 95% confidence intervals (CI), adjusting for known prognostic factors., Results: Variation within angiogenesis was most strongly associated with survival time overall (P = 0.03) and among patients with serous cancer (P = 0.05), particularly for EIF2B5 rs4912474 (all patients HR, 0.69; 95% CI, 0.54-0.89; P = 0.004), VEGFC rs17697305 (serous subtype HR, 2.29; 95% CI, 1.34-3.92; P = 0.003), and four SNPs in VHL. Variation within the inflammation pathway was borderline significant (all patients, P = 0.09), and SNPs in CCR3, IL1B, IL18, CCL2, and ALOX5 which correlated with survival time are worthy of follow-up., Conclusion: An extensive multiple-pathway assessment found evidence that inherited differences may play a role in outcome of ovarian cancer patients, particularly in genes within the angiogenesis and inflammation pathways. Our work supports efforts to target such mediators for therapeutic gain.