Preclinical evaluation of olaparib and metformin combination in BRCA1 wildtype ovarian cancer.

OBJECTIVES: BRCA mutated ovarian cancers show increased responsiveness to PARP inhibitors. PARP inhibitors target DNA repair and provide a second hit to BRCA mutated tumors, resulting in "synthetic lethality". We investigated a combination of metformin and olaparib to provide "synthetic lethality" in BRCA intact ovarian cancer cells. METHODS: Ovarian cancer cell lines (UWB1.289, UWB1.289.BRCA, SKOV3, OVCAR5, A2780 and C200) were treated with a combination of metformin and olaparib. Cell viability was assessed by MTT and colony formation assays. Flow cytometry was used to detect cell cycle events. In vivo studies were performed in SKOV3 or A2780 xenografts in nude mice. Animals were treated with single agent, metformin or olaparib or combination. Molecular downstream effects were examined by immunohistochemistry. RESULTS: Compared to single drug treatment, combination of olaparib and metformin resulted in significant reduction of cell proliferation and colony formation (p<0.001) in ovarian cancer cells. This treatment was associated with a significant S-phase cell cycle arrest (p<0.05). Combination of olaparib and metformin significantly inhibited SKOV3 and A2780 ovarian tumor xenografts which were accompanied with decreased Ki-index (p<0.001). Metformin did not affect DNA damage signaling, while olaparib induced adenosine monophosphate activated kinase activation; that was further potentiated with metformin combination in vivo. CONCLUSION: Combining PARP inhibitors with metformin enhances its anti-proliferative activity in BRCA mutant ovarian cancer cells. Furthermore, the combination showed significant activity in BRCA intact cancer cells in vitro and in vivo. This is a promising treatment regimen for women with epithelial ovarian cancer irrespective of BRCA status.

Endometrial cancer in morbidly obese women: do racial disparities affect surgical or survival outcomes?

OBJECTIVE: Endometrial cancer mortality disproportionately affects black women and whether greater prevalence of obesity plays a role in this disparity is unknown. We examine the effect of race on post-surgical complications, length of stay, and mortality specifically in a morbidly obese population. METHODS: Black and white women with endometrial cancer diagnosed from 1996 to 2012 were identified from the University Pathology Group database in Detroit, Michigan, and records were retrospectively reviewed to obtain clinicopathological, demographic, and surgical information. Analysis was limited to those with a body mass index of 40kg/m(2) or greater. Differences in the distribution of variables by race were assessed by chi-squared tests and t-tests. Kaplan-Meier and Cox regression analyses were performed to examine factors associated with mortality. RESULTS: 97 white and 89 black morbidly obese women were included in this analysis. Black women were more likely to have type II tumors (33.7% versus 15.5% of white women, p-value=0.003). Hypertension was more prevalent in black women (76.4% versus 58.8%, p-value=0.009), and they had longer hospital stays after surgery despite similar rates of open vs minimally invasive procedures and lymph node dissection (mean days=5.4) compared to whites (mean days=3.5, p-value=0.036). Wound infection was the most common complication (16.5% in whites and 14.4% in blacks, p-value=0.888). Blacks were more likely to suffer other complications, but overall the proportions did not differ by race. In univariate analyses, black women had higher risk of endometrial cancer-related death (p-value=0.090). No racial differences were noted in adjusted survival analyses. CONCLUSION: A more complete investigation, incorporating socio-demographic factors, is warranted to understand the effects of morbid obesity and race on endometrial cancer.

Prognostic impact of lymphadenectomy in uterine serous cancer.

OBJECTIVE: To estimate the survival impact of lymphadenectomy in women diagnosed with uterine serous cancer. DESIGN: Women with a diagnosis of uterine serous cancer were identified from the Surveillance, Epidemiology and End Results Program (SEER) from 1988 to 2007. Only surgically treated women were included. SETTING: The Surveillance, Epidemiology and End Results Program database provided data from 17 registries. POPULATION: The study population comprised 4178 women. METHODS: Statistical analyses using Student's t-test, Kaplan-Meier survival methods and Cox proportional hazards regression were performed. MAIN OUTCOME MEASURE: Overall survival. RESULTS: Three thousand, one hundred and ninety-four (67.7%) women underwent lymphadenectomy. Older women (≥65 years) and Caucasian women (relative to Asian) were less likely to have lymphadenectomy (P < 0.001). The prevalence of nodal metastasis in women having lymphadenectomy was 32%. Of the 1997 women who had disease grossly confined to the uterus and underwent lymphadenectomy, 387 (19%) were found to have nodal metastasis. Lymphadenectomy was associated with improved survival; women who underwent lymphadenectomy were 41% (hazard ratio [HR], 0.59; 95% confidence interval [CI], 0.54-0.64; P < 0.001) less likely to die than women who did not have the procedure. Moreover, more extensive lymphadenectomy correlated positively with survival. Compared with women with 1-10 nodes removed, those with more extensive lymphadenectomy (>10 nodes removed) were 26% (HR, 0.74; 95% CI, 0.67-0.83; P < 0.001) less likely to die. The impact of the extent of lymphadenectomy on survival was significant in both node-negative and node-positive women. CONCLUSION: Age and race influenced the prevalence of lymphadenectomy in our cohort. This study suggests that the extent of lymphadenectomy is associated with significant improvement in survival of women with uterine serous cancer.

Correlation of tumor size with other prognostic factors in uterine serous carcinoma: a large multi-institutional study.

OBJECTIVE: Uterine serous carcinoma (USC) constitutes 10% of uterine cancers but ~40% of deaths. Tumor size is a known prognostic factor in other solid tumors. In endometriod cancers it is one element used to identify the need for complete staging, while its significance in USC is debated. Therefore tumor size was examined as an independent prognostic factor. METHODS: Clinical and pathologic variables were recorded for 236 institutional patients, and those patients in the SEER database with USC. Chi-square and Fisher exact t-tests were utilized and survival data generated via Kaplan-Meier method; multivariate analysis was performed via cox-regression. RESULTS: The patients' mean age was 67.2 years (range 40-91). Survival ranged from 0 to 184 months (mean 42.8). We used a tumor size cut-off of 1cm and noted significant associations with myometrial invasion (p<0.0001), angiolymphatic invasion (p<0.0001), peritoneal washings (p=0.03), stage (p=0.015) and positive lymph nodes (p=0.05). Furthermore, recurrence was associated with larger tumors (p=0.03). In multivariate analysis, extra-uterine disease was the only factor associated with both recurrence and survival. Review of the SEER database noted association of larger tumors with lymph node involvement and a significant survival advantage with tumors <1cm in both univariate and multivariate analysis. CONCLUSIONS: Treatment options for USC are often predicated on the surgical stage and therefore components of the staging are vitally important. The 1cm tumor-size cut-off should be studied prospectively as a prognostic indicator of survival and recurrence in USC and considered for inclusion in USC staging.

The molecular genomics of metastatic brain tumours.

INTRODUCTION: Metastatic brain tumours remain an intractable clinical problem despite notable advances in the treatment of the primary cancers. It is estimated that 30-40% of breast and lung cancer patients will develop brain metastases. Typically, brain lesions are not diagnosed until patients exhibit neurological symptoms because there are currently no tests that can predict which patients will be afflicted. Brain metastases are resistant to current chemotherapies, and despite surgical resection and radiotherapy, the prognosis for these patients remains very poor with an average survival of only 6-9 months. Cancer is ultimately a genetic disease, involving patient genetics and aberrant tumour genomics; therefore the pursuit of an explanation for why or how brain metastases occur requires investigation of the associated somatic mutations. In this article, we review the current literature surrounding the molecular and genome-based mechanistic evidence to indicate driver oncogenes that hold potential biomarkers for risk, or therapeutic targets for treatment of brain metastases. CONCLUSION: Patients afflicted with metastatic brain tumours are in dire need of more effective therapies, and clinicians need predictive laboratory tests to identify patients at risk of developing metastatic brain tumours. The as yet unrealized comprehensive analysis of metastatic brain tumour genomics is necessary to meet these needs. Moreover, without improved understanding of the genomic aberrations that drive metastatic brain tumours, development of biomarkers and molecularly targeted therapies will remain stalled and patient outcomes will continue to be dismal.

Clinical outcomes of adjuvant chemotherapy and vaginal brachytherapy with or without pelvic radiation for surgical stage I-II uterine serous carcinoma.

OBJECTIVE(S): To evaluate the benefit of adding pelvic radiation treatment (EBRT) to vaginal cuff brachytherapy (VB) for women with early stage uterine serous carcinoma (USC) treated with adjuvant chemotherapy. MATERIALS AND METHODS: After institutional review board (IRB) approval, the authors retrospectively identified 56 patients with 2009 International Federation of Gynecology and Obstetrics (FIGO) Stage I-II USC treated with hysterectomy, bilateral oophorectomy +/- lymphadenectomy, adjuvant chemotherapy, and radiation therapy with either VB alone (n = 33) or VB + EBRT (n = 23) between July 1998 and August 2009. RESULTS: Median age and follow-up were 68.5 years and 54 months respectively. Median VB alone surface dose was 37.5 Gy and median pelvic EBRT dose was 45 Gy. The prevalence of lower uterine segment involvement, > 50% myometrial invasion, and Stage II disease were higher for patients receiving VB + EBRT. Overall, only one vaginal recurrence was observed. Pelvic recurrence rate was 26% for VB + EBRT compared to 12% for VB alone (p = 0.179). The five-year recurrence-free survival (RFS) was 80.5% for VB vs 67.3% for VB + EBRT (p = 0.3847), and the five-year overall survival (OS) was 65.9% for VB vs 66.7% for VB + EBRT (p = 0.7159). On univariate and multivariate analysis, radiation treatment modality was not a predictor for local control or survival. CONCLUSIONS: In this cohort, there was no significant clinical benefit of adding pelvic EBRT to the adjuvant management of early stage uterine serous carcinoma. The higher prevalence of high-risk features in the VB + EBRT group may underestimate the value of this treatment. Further investigation is warranted to identify the optimal radiation treatment regiment for early stage USC treated with surgery and adjuvant chemotherapy.

Prognostic impact of lymphadenectomy in clinically early stage malignant germ cell tumour of the ovary.

BACKGROUND: The aim of this study was to determine the impact of lymphadenectomy and nodal metastasis on survival in clinical stage I malignant ovarian germ cell tumour (OGCT). METHODS: Data were obtained from the National Cancer Institute registry from 1988 to 2006. Analyses were performed using Student's t-test, Kaplan-Meier and Cox proportional hazard methods. RESULTS: In all, 1083 patients with OGCT who have undergone surgical treatment and deemed at time of the surgery to have disease clinically confined to the ovary were included 590 (54.48%) had no lymphadenectomy (LND-1) and 493 (45.52%) had lymphadenectomy. Of the 493 patients who had lymphadenectomy, 441 (89.5%) were FIGO surgical stage I (LND+1) and 52 (10.5%) were upstaged to FIGO stage IIIC due to nodal metastasis (LND+3C). The 5-year survival was 96.9% for LND-1, 97.7% for LND+1 and 93.4% for LND+3C (P=0.5). On multivariate analysis, lymphadenectomy was not an independent predictor of survival when controlling for age, histology and race (HR: 1.26, 95% CI: 0.62-2.58, P=0.5). Moreover, the presence of lymph node metastasis had no significant effect on survival (HR: 2.7, 95% CI: 0.67-10.96, P=0.16). CONCLUSION: Neither lymphadenectomy nor lymph node metastasis was an independent predictor of survival in patients with OGCT confined to the ovary. This probably reflects the highly chemosensitive nature of these tumours.

Genomic evolution in Barrett's adenocarcinoma cells: critical roles of elevated hsRAD51, homologous recombination and Alu sequences in the genome.

A prominent feature of most cancers including Barrett's adenocarcinoma (BAC) is genetic instability, which is associated with development and progression of disease. In this study, we investigated the role of recombinase (hsRAD51), a key component of homologous recombination (HR)/repair, in evolving genomic changes and growth of BAC cells. We show that the expression of RAD51 is elevated in BAC cell lines and tissue specimens, relative to normal cells. HR activity is also elevated and significantly correlates with RAD51 expression in BAC cells. The suppression of RAD51 expression, by short hairpin RNA (shRNA) specifically targeting this gene, significantly prevented BAC cells from acquiring genomic changes to either copy number or heterozygosity (P<0.02) in several independent experiments employing single-nucleotide polymorphism arrays. The reduction in copy-number changes, following shRNA treatment, was confirmed by Comparative Genome Hybridization analyses of the same DNA samples. Moreover, the chromosomal distributions of mutations correlated strongly with frequencies and locations of Alu interspersed repetitive elements on individual chromosomes. We conclude that the hsRAD51 protein level is systematically elevated in BAC, contributes significantly to genomic evolution during serial propagation of these cells and correlates with disease progression. Alu sequences may serve as substrates for elevated HR during cell proliferation in vitro, as they have been reported to do during the evolution of species, and thus may provide additional targets for prevention or treatment of this disease.

The racial disparity in outcomes in endometrial cancer: could this be explained on a molecular level?

OBJECTIVE: The racial disparities among patients with endometrial carcinoma have been previously reported. The objective of this study is to analyze and compare the molecular profiles in endometrial cancer in Caucasian and African American patients using a number of known molecular markers. MATERIALS AND METHODS: 147 patients diagnosed with endometrial cancer between 1995 and 2001 were included in the study. Patients' demographics, clinical and pathological data were reviewed. Immunohistochemical staining for p53, VEGF, Ki-67 and HIF-1alpha was performed on tissue micro array sections. Tumors' expression of p53, VEGF, Ki-67, and HIF-1alpha was compared based on ethnicity and tumor type (Type I = endometrioid carcinomas and Type II = non-endometrioid carcinomas). Spearman's correlation and Fisher's Exact Tests were used for statistical analysis and Kaplan-Meier, log-rank and Cox regression were used for survival analysis. RESULTS: 97 patients were Caucasian and 50 patients were African American. The mean age was 62 (33-91) years for Caucasian patients and 63.5 (24-89) years for the African American patients. African American patients had more Type II carcinoma than Caucasian patients (P = 0.055). High p53 expression was statistically significant among the African American patients (49% vs. 30%, P = 0.035) versus Caucasian patients. There was no significant difference demonstrated when comparing the VEGF, Ki-67, and HIF-1alpha expression between the racial groups. Survival analysis showed a trend toward a shorter survival in the African American patients compared to the Caucasian patients; median survival 62 versus 77 months (P = 0.061). On the other hand, we did not find a significant difference in survival by ethnicity when we adjusted for tumor histology. CONCLUSION: While African American patients with endometrial cancer seem to show a trend toward a shorter survival, this seems to be mainly due to the fact that they have a higher proportion of Type II tumors. The molecular profiles for p53, Ki-67, VEGF and HIF-1alpha expression of histologically matched tumors were similar between the two ethnic groups.

Maspin expression and localization impact on angiogenesis and prognosis in ovarian cancer.

INTRODUCTION: The goal of this study is to evaluate the relation of maspin expression and its cellular localization to markers of angiogenesis in epithelial ovarian serous carcinoma (OSC). MATERIALS AND METHODS: We identified 118 patients with high-grade advanced stage OSC who were treated at our institution. Clinical data were collected, and immunohistochemistry (IHC) with antibodies to VEGF, CD34, COX-2, and maspin was performed on paraffin-embedded tumor blocks. CD34 immunostaining was used to determine microvessel density. The correlation between the various molecular markers was assessed using the Chi-square test. Survival analysis was computed using the Kaplan-Meier model, and various prognostic variables were compared using Cox regression analysis. RESULTS: Maspin expression was noted in 81.4% (96/118) of tumors. Expression was localized to the nuclear compartment in 21.2% of cases, whereas 60.2% of cases showed evidence of cytoplasmic +/- nuclear expression. Tumors that exhibited nuclear maspin expression had lower VEGF and COX-2 expression than tumors with negative or cytoplasmic expression. Tumors with high nuclear maspin expression had lower mean MVD than those with low or negative expression. The median survival based on localization of maspin was 1146 days for those with negative tumors, 1803 days for those with nuclear maspin, and 637 days for those with cytoplasmic maspin (P < 0.001). In a Cox regression analysis, maspin localization was an independent prognostic factor. CONCLUSION: Maspin expression and localization seem to play a role in ovarian cancer angiogenesis and progression. High nuclear expression was associated with reduced markers of angiogenesis and prolonged survival.

Clinicopathologic analysis of breast lesions associated with multiple papillomas.

We performed a retrospective clinicopathologic study of 28 patients with breast lesions characterized by the presence of multiple (at least 5) papillomas (MPs) in at least 2 nonconsecutive blocks. All histologic sections were assessed for the presence of coexisting fibrocystic lesions, including atypical hyperplasia (atypical ductal hyperplasia [ADH] or atypical lobular hyperplasia [ALH]), lobular carcinoma in situ (LCIS), and papillary atypia (defined as nuclear hyperchromatism, stratification, and architectural complexity of a lesser degree than in papillary carcinoma). All of the lesions were compared with a set of cases in which ductal carcinoma in situ (DCIS) (n = 20) or invasive carcinoma (INV)(n = 13) was accompanied by MPs. The MP cases had a characteristic morphologic appearance, typically presenting as a mass comprising multiple adjacent ducts filled by papillomas, accompanied by dense fibrosis and intermingled with various proliferative fibrocystic lesions, particularly florid adenosis. Atypical hyperplasia was a frequent finding (in 12 of 28 cases; 43%), particularly in cases with atypical papillomas (7 of 11; 63.6%). Although contralateral lesions occurred in 4 of 28 patients (14.2%; 3 MPs and 1 INV), only 1 patient (4%) has developed ipsilateral breast carcinoma (mean follow-up, 47 months). DCIS associated with MP was typically low grade (17 of 20; 85%) and arose from areas within or immediately adjacent to preexisting benign lesions. None has recurred (mean follow-up, 41 months), although 1 patient has contralateral MP and 3 patients (23%) have developed carcinomas in the opposite breast. INVs developing in a background of (ipsilateral) MPs were mostly small (8 of 11 <2.0 cm), node negative (7 of 10), and estrogen receptor (ER) positive (8 of 8). Only 1 of 13 patients (8%) has died from disease (mean follow-up, 59 months), but 5 (38%) have developed contralateral breast lesions (including 1 MP, 1 MP-DCIS, 1 DCIS, 1 LCIS, and 1 INV). We conclude that the frequent associations with ADH, ALH/LCIS, malignant lesions, and bilaterality imply that MP may represent a marker of constitutionally increased breast cancer risk. Because carcinomas arose within or close to areas involved by preexisting benign MP lesions, it may also be appropriate to excise segments of tissue involved by MP, particularly cases with atypia, and closely monitor for contralateral disease.

Use of ThinPrep monolayer technique and cytospin preparation in urine cytology: a comparative analysis.

We compared the ThinPrep (TP) technique to the cytospin (CS) preparation in the cytological diagnosis of urine by processing 79 specimens by these two techniques. Ten cases were positive for malignancy (six high grade (HG)/carcinoma in situ; four low grade (LG) transitional cell carcinomas (TCC)). Forty-eight cases were within normal limits (59%) and 21 cases had atypical cytological features (19%). The TP technique was better in terms of a cleaner background with fewer obscuring inflammatory cells and blood and with a more even distribution of cells. In general, the cytomorphology was comparable in both techniques. However, in cases with malignancy, CS was relatively superior in the cytomorphologic details; in TP, the diagnostic cells were mostly dispersed as single cells with loss of architectural features and were difficult to find. Artifactual empty spaces and air-drying were more frequently present in TP. In cases contaminated with squamous cells, the urothelial cells were difficult to find in TP. Screening time was comparable for both techniques. In conclusion, to avoid false-negative diagnosis, CS would be complementary to the TP technique in malignant cases and, in particular, those with low cellularity.