Clinical and Emergent Biomarkers and Their Relationship to the Prognosis of Ovarian Cancer.

OBJECTIVE: Ovarian cancer is the most lethal gynecological malignancy, but information relevant to prognosis and outcomes remain unknown. Here, we used statistical methods to focus specifically on interactions between candidate prognostic variables. METHODS AND RESULTS: Univariate, multivariate, and elastic net modeling of 42 variables were applied to a cohort of 542 ovarian cancer patients with 393 episodes of cancer recurrence/death. In univariate analyses, overexpression of TFF3, MDM2, and p53 were associated with improved recurrence-free survival. In multivariate analyses adjusted for age, histology, stage, grade, ascites, and residual disease, overexpression of PR appeared to provide a protective effect [hazard ratio for >50% of cells positive, 0.64 (95% confidence interval 0.44-0.94) compared to <1%], and TFF3 showed a nonlinear association. Importantly, we observed no interactions among variables. However, patients with tumors with moderate TFF3 expression were at a marginally increased risk of recurrence, and patients with tumors with high expression were at a similar to slightly lower risk, compared to those with tumors with no TFF3 expression. CONCLUSIONS: Although no interactions among variables were observed, this study provides important precedent for seeking interactions between clinical and tumor variables in future studies.

In vivo anti-tumor activity of the PARP inhibitor niraparib in homologous recombination deficient and proficient ovarian carcinoma.

OBJECTIVE: Poly(ADP-ribose) polymerase (PARP) inhibitors have yielded encouraging responses in high-grade serous ovarian carcinomas (HGSOCs), but the optimal treatment setting remains unknown. We assessed the effect of niraparib on HGSOC patient-derived xenograft (PDX) models as well as the relationship between certain markers of homologous recombination (HR) status, including BRCA1/2 mutations and formation of RAD51 foci after DNA damage, and response of these PDXs to niraparib in vivo. METHODS: Massively parallel sequencing was performed on HGSOCs to identify mutations contributing to HR deficiency. HR pathway integrity was assessed using fluorescence microscopy-based RAD51 focus formation assays. Effects of niraparib (MK-4827) on treatment-naïve PDX tumor growth as monotherapy, in combination with carboplatin/paclitaxel, and as maintenance therapy were assessed by transabdominal ultrasound. Niraparib responses were correlated with changes in levels of poly(ADP-ribose), PARP1, and repair proteins by western blotting. RESULTS: Five PDX models were evaluated in vivo. Tumor regressions were induced by single-agent niraparib in one of two PDX models with deleterious BRCA2 mutations and in a PDX with RAD51C promoter methylation. Diminished formation of RAD51 foci failed to predict response, but Artemis loss was associated with resistance. Niraparib generally failed to enhance responses to carboplatin/paclitaxel chemotherapy, but maintenance niraparib therapy delayed progression in a BRCA2-deficient PDX. CONCLUSIONS: Mutations in HR genes are neither necessary nor sufficient to predict response to niraparib. Assessment of repair status through multiple complementary assays is needed to guide PARP inhibitor therapy, design future clinical trials and identify ovarian cancer patients most likely to benefit from PARP inhibition.

Regulatory T cells, inherited variation, and clinical outcome in epithelial ovarian cancer.

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.

Exposure to environmental chemicals and heavy metals, and risk of pancreatic cancer.

PURPOSE: Exposure to various chemicals and heavy metals has been associated with risk of different cancers; however, data on whether such exposures may increase the risk of pancreatic cancer (PC) are very limited and inconclusive. We examined PC risk with self-reported exposures to chemicals and heavy metals. METHODS: The design was a clinic-based, case-control study of data collected from 2000 to 2014 at Mayo Clinic in Rochester, Minnesota, USA. Cases were rapidly ascertained patients diagnosed with pancreatic ductal adenocarcinoma (n = 2,092). Controls were cancer-free patients in primary care clinics (n = 2,353), frequency-matched to cases on age, race, sex, and state/region of residence. Cases and controls completed identical risk factor questionnaires, which included yes/no questions about regular exposure to pesticides, asbestos, benzene, chlorinated hydrocarbons, chromium, and nickel. Unconditional logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CI) comparing those who affirmed exposure to each of the chemicals/heavy metals to those who reported no regular exposure, adjusting for potential confounders. RESULTS: Self-reported regular exposure to pesticides was associated with increased odds of PC (OR 1.21, 95% CI 1.02-1.44). Regular exposure to asbestos (OR 1.54, 95% CI 1.23-1.92), benzene (OR 1.70, 95% CI 1.23-2.35), and chlorinated hydrocarbons (OR 1.63, 95% CI 1.32-2.02) also was associated with higher odds of PC. Chromium and nickel exposures were not significantly associated with PC. CONCLUSIONS: These findings add to the limited data suggesting that exposure to pesticides, asbestos, benzene, and chlorinated hydrocarbons may increase PC risk. They further support the importance of implementing strategies that reduce exposure to these substances.

Assessment of published models and prognostic variables in epithelial ovarian cancer at Mayo Clinic.

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.

Detection of endometrial cancer via molecular analysis of DNA collected with vaginal tampons.

OBJECTIVE: We demonstrate the feasibility of detecting EC by combining minimally-invasive specimen collection techniques with sensitive molecular testing. METHODS: Prior to hysterectomy for EC or benign indications, women collected vaginal pool samples with intravaginal tampons and underwent endometrial brushing. Specimens underwent pyrosequencing for DNA methylation of genes reported to be hypermethylated in gynecologic cancers and recently identified markers discovered by profiling over 200 ECs. Methylation was evaluated individually across CpGs and averaged across genes. Differences between EC and benign endometrium (BE) were assessed using two-sample t-tests and area under the curve (AUC). RESULTS: Thirty-eight ECs and 28 BEs were included. We evaluated 97 CpGs within 12 genes, including previously reported markers (RASSF1, HSP2A, HOXA9, CDH13, HAAO, and GTF2A1) and those identified in discovery work (ASCL2, HTR1B, NPY, HS3ST2, MME, ADCYAP1, and additional CDH13 CpG sites). Mean methylation was higher in tampon specimens from EC v. BE for 9 of 12 genes (ADCYAP1, ASCL2, CDH13, HS3ST2, HTR1B, MME, HAAO, HOXA9, and RASSF1) (all p<0.05). Among these genes, relative hypermethylation was observed in EC v. BE across CpGs. Endometrial brush and tampon results were similar. Within tampon specimens, AUC was highest for HTR1B (0.82), RASSF1 (0.75), and HOXA9 (0.74). This is the first report of HOXA9 hypermethylation in EC. CONCLUSION: DNA hypermethylation in EC tissues can also be identified in vaginal pool DNA collected via intravaginal tampon. Identification of additional EC biomarkers and refined collection methods are needed to develop an early detection tool for EC.

Self-Perceived Improvement in Bladder Health After Viewing a Novel Tutorial on Knack Use: A Randomized Controlled Trial Pilot Study.

Purpose: To test a novel bladder health tutorial on use of the Knack for overcoming bladder control challenges. The Knack-tutorial is a self-administered vignette-based instructional program on preempting bladder challenges in daily life (urgency, stress-leakage, or urge-leakage) through anticipatory, well-timed pelvic floor muscle contraction at the moment of challenge. Materials and Methods: This is a randomized controlled trial pilot test of 108 women with stress or mixed urinary incontinence. The Knack-tutorial group saw a 15-minute slide show with 10 vignettes portraying use of the Knack in daily life. The slide show format used inserted narrated videos, dubbed and animation enhanced pictures and cartoons, and automatic slide advancement. A control group saw a similarly constructed slide show on incorporating good diet/exercise habits. Outcomes were self-perceived improvement (yes/no, and as 0%-100%) 1 month after viewing the tutorial. Results: We enrolled 123 women, randomizing 64 to Knack-tutorial group and 59 to diet/exercise tutorial group. Eleven and one participant, respectively, did not return. Three did not fill out the self-perceived improvement report. Significant improvement was reported by 71% in the Knack-tutorial group compared to 25% in the diet/exercise group (p < 0.001). Self-perceived improvement was 21%-22% higher (Model I Est: 21.01, SE: 4.25, p < 0.001) in the Knack-tutorial group. Conclusions: An electronic tutorial viewed independent of a health care provider with vignettes showing Knack application to manage the everyday bladder challenges women face shows benefit of a magnitude that warrants more widespread use and rigorous testing. A professional remake of the intervention is now available (www.myconfidentbladder.com).