Rochelle Payne Ondracek

The Thoc1 Ribonucleoprotein and Prostate Cancer Progression

BACKGROUND The majority of newly diagnosed prostate cancers will remain indolent, but distinguishing between aggressive and indolent disease is imprecise. This has led to the important clinical problem of overtreatment. THOC1 encodes a nuclear ribonucleoprotein whose expression is higher in some cancers than in normal tissue. The hypothesis that THOC1 may be a functionally relevant biomarker that can improve the identification of aggressive prostate cancer has not been tested. METHODS THOC1 protein immunostaining was evaluated in a retrospective collection of more than 700 human prostate cancer specimens and the results associated with clinical variables and outcome. Thoc1 was conditionally deleted in an autochthonous mouse model (n = 22 or 23 per genotype) to test whether it is required for prostate cancer progression. All statistical tests were two-sided. RESULTS THOC1 protein immunostaining increases with higher Gleason score and more advanced Tumor/Node/Metastasis stage. Time to biochemical recurrence is statistically significantly shorter for cancers with high THOC1 protein (log-rank P = .002, and it remains statistically significantly associated with biochemical recurrence after adjusting for Gleason score, clinical stage, and prostate-specific antigen levels (hazard ratio = 1.61, 95% confidence interval = 1.03 to 2.51, P = .04). Thoc1 deletion prevents prostate cancer progression in mice, but has little effect on normal tissue. Prostate cancer cells deprived of Thoc1 show gene expression defects that compromise cell growth. CONCLUSIONS Thoc1 is required to support the unique gene expression requirements of aggressive prostate cancer in mice. In humans, high THOC1 protein immunostaining associates with prostate cancer aggressiveness and recurrence. Thus, THOC1 protein is a functionally relevant molecular marker that may improve the identification of aggressive prostate cancers, potentially reducing overtreatment.

Selenomethionine and methyl selenocysteine: Multiple-dose pharmacokinetics in selenium-replete men

According to the Nutritional Prevention of Cancer (NPC) trial, a selenized yeast supplement containing selenium, 200 mcg/day, decreased the incidence of total cancer, cancers of the prostate, colon and lung, and cancer mortality. The active agent in the selenized yeast supplement was assumed to be selenomethionine (SEMET), although the supplement had not been well speciated. The SELECT study, largely motivated by the NPC trial, enrolling nearly 40 times as many subjects, showed unequivocally that selenium 200 mcg/day, with selenium in the form of SEMET, does not protect selenium-replete men against prostate or other major cancer. The agent tested by SELECT, pure SEMET, could have been different from the selenized yeast tested in NPC. One of the selenium forms suspected of having chemopreventive effects, and which may have been present in the NPC agent, is methyl selenocysteine (MSC). This study, with 29 selenium-replete patients enrolled in a randomized, double-blind trial, compared the multiple-dose toxicity, pharmacokinetics and pharmacodynamics of MSC and SEMET. Patients were on trial for 84 days. No toxicity was observed. Although SEMET supplementation increased blood selenium concentration more than MSC did, neither form had a more than minimal impact on the two major selenoproteins: selenoprotein P(SEPP1) and glutathione peroxidase(GPX).

Abstract 4998: Breast tumor FOXA1 protein expression and reproductive characteristics among African-American and European-American women

Background: Forkhead box protein A1 (FOXA1) plays a key role in determining estrogen receptor (ER) function and mammary ductal development and may repress the basal cell phenotype during differentiation of breast epithelium. While reproductive factors are known to influence breast cancer risk depending on the subtypes, data on the association of tumor FOXA1 protein expression and reproductive characteristics are very limited. Methods: Tissue microarrays comprising surgical tumors from 638 women (466 African-American [AA] and 172 European-American [EA], aged 20-75 years) with primary breast cancer in the Women’s Circle of Health Study (WCHS) were analyzed for FOXA1 expression by immunohistochemistry and automated image analysis. In-person interviews were conducted to obtain data on demographics, medical and family histories, and reproductive and menstrual histories. Logistic regression was performed for FOXA1 positivity (>10% cells with strong staining) with menopausal status, age at menarche, age at first live birth, parity, and breastfeeding, adjusting for age at diagnosis, family history of breast cancer, and history of benign breast disease. Results: FOXA1 expression was higher in tumors from EA compared to those from AA women (80% vs. 70% positivity, P=0.011). FOXA1 expression was highly correlated with ER positivity (88% positive in ER+ and 26% in ER- breast cancer among AA women; 90% and 29% respectively among EA women; both P Conclusion: In AA women, a higher number of live births, which has been related to greater risk of ER- breast cancer, is associated with lower FOXA1 expression. Because FOXA1 promotes luminal and represses the basal differentiation, respectively, the effects of parity on expression of FOXA1 may be the link whereby it increases risk of ER- breast cancer. (Funding: NIH P01CA151135, R01CA100598, R01CA185623, P30CA072720, K07CA201334; US Army Medical Research and Material Command DAMD-17-01-1-0334; the Breast Cancer Research Foundation; and the Philip L. Hubbell family) Citation Format: Ting-Yuan David Cheng, Rochelle Payne Ondracek, Song Yao, Wiam Bshara, Thaer Khoury, Gary R. Zirpoli, Warren Davis, Elisa V. Bandera, Michael Higgins, Christine B. Ambrosone. Breast tumor FOXA1 protein expression and reproductive characteristics among African-American and European-American women [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4998. doi:10.1158/1538-7445.AM2017-4998

Abstract 4664: The effect of neoadjuvant androgen deprivation therapy on long-term outcome after radical prostatectomy

Neoadjuvant androgen deprivation therapy (NADT) was championed for men treated with radical prostatectomy (RP) for high risk prostate cancer (CaP) in the early 1990s. Although NADT improved neither surgical margin positivity nor progression-free survival, it continues to be part of some clinical trials. The effect of NADT on long term outcome was evaluated using modern aggressiveness and failure criteria among 720 patients treated with RP at Roswell Park Cancer Institute between 1993 and 2005. Among RP patients, 125 received NADT. NCCN pre-surgery risk categories were low in 36 (29%), intermediate in 54 (43%), and high or very high in 35 (28%). Clinical and pathological aggressiveness indicators included clinical Gleason sum, clinical tumor stage (2002 TNM), highest preoperative PSA, pathological Gleason sum, pathological tumor stage (2002 TNM) and surgical margin status. Ten post-RP recurrence definitions were considered: 1) at least 1 detectable PSA, 2) at least 2 detectable PSAs, 3) PSA>0.2 ng/ml, 4) PSA>0.4 ng/ml, 5) at least 1 post RP treatment (chemotherapy, hormone, radiation), 6) PSA doubling time Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4664. doi:10.1158/1538-7445.AM2011-4664

Tobacco use and outcome in radical prostatectomy patients

Cigarette smoking has been consistently associated with increased risk of overall mortality, but the importance of smoking for patients with prostate cancer (CaP) who are candidates for curative radical prostatectomy (RP) has received less attention. This retrospectively designed cohort study investigated the association of smoking history at RP with subsequent CaP treatment outcomes and overall mortality. A total of 1981 patients who underwent RP at Roswell Park Cancer Institute (RPCI) between 1993 and 2014 were studied. Smoking history was considered as a risk factor for overall mortality as well as for currently accepted CaP treatment outcomes (biochemical failure, treatment failure, distant metastasis, and disease‐specific mortality). The associations of smoking status with these outcomes were tested by Cox proportional hazard analyses. A total of 153 (8%) patients died during follow‐up. Current smoking at diagnosis was a statistically significant predictor of overall mortality after RP (current smokers vs. former and never smokers, hazards ratio 2.07, 95% confidence interval [CI]: 1.36–3.14). This association persisted for overall mortality at 3, 5, and 10 years (odds ratios 2.07 [95% CI: 1.36–3.15], 2.05 [95% CI: 1.35–3.12], and 1.8 [95% CI: 1.18–2.74], respectively). Smoking was not associated with biochemical failure, treatment failure, distant metastasis, or CaP‐specific mortality, and the association of smoking with overall mortality did not appear to be functionally related to treatment or biochemical failure, or to distant metastasis. Smoking is a non‐negligible risk factor for death among CaP patients who undergo RP; patients who smoke are far more likely to die of causes other than CaP.

Association of fatty-acid synthase polymorphisms and expression with outcomes after radical prostatectomy

Background:Fatty-acid synthase (FASN), selectively overexpressed in prostate cancer (PCa) cells, has been described as linked to the aggressiveness of PCa. Constitutional genetic variation of the FASN gene and the expression levels of FASN protein in cancer cells could thus be expected to predict outcome after radical prostatectomy (RP). This study evaluates the associations of malignant tissue status, neoadjuvant androgen deprivation therapy (NADT) and single-nucleotide polymorphisms (SNPs) of FASN with FASN protein expression in prostate tissue. The study then examines the associations of FASN SNPs and gene expression with three measures of post-prostatectomy outcome.Methods:Seven tagging FASN SNPs were genotyped in 659 European American men who underwent RP at Roswell Park Cancer Institute between 1993 and 2005. FASN protein expression was assessed using immunohistochemistry. The patients were followed for an average of 6.9 years (range: 0.1–20.6 years). Outcome was assessed using three end points: biochemical failure, treatment failure and development of distant metastatic PCa. Cox proportional hazards analyses were used to evaluate the associations of the tagging SNPs and FASN expression with these end points. Bivariate associations with outcomes were considered; the associations also were controlled for known aggressiveness indicators.Results:Overall, no SNPs were associated with any known aggressiveness indicators. FASN staining intensity was stronger in malignant than in benign tissue, and NADT was associated with decreased FASN staining in both benign and malignant tissue. The relationships of FASN SNPs and staining intensity with outcome were less clear. One SNP, rs4246444, showed a weak association with outcome. FASN staining intensity also showed a weak and seemingly contradictory relationship with outcome.Conclusions:Additional study with longer follow-up and populations that include more metastatic patients is warranted.

Abstract 4852: Comparison of manual and automated scoring of androgen receptor staining in prostate tissue.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Although manual scoring is generally used to evaluate the presence of androgen receptor (AR) protein as detected by immunohistochemistry in tissue, manual evaluation is tedious, inefficient and subjective. Therefore, manual scoring is not feasible for the evaluation of large numbers of patients. In this study, a technician (CP) rated AR staining in tissue sections of 3 tissue microarrays (TMAs) constructed from 3 cores each of benign and malignant tissue from 134 patients who underwent radical prostatectomy between 1993 and 2006 at Roswell Park Cancer Institute. The manual scorer randomly sampled 50 nuclei from each core and scored each nuclei as not stained (score =0), or lightly (score = 1), moderately (score=2) or darkly (score = 3) stained. The average staining intensity was recorded per patient across the benign cores, and across the tumor cores. Automated staining used the Image Pro image processing program, in which the entire core was scanned, all nuclei segmented and average intensity values per between 0 (completely black) and 255 (completely white) were determined for each nucleus. The mean staining score for each core is calculated by taking the average of all the identified nuclei in the core, and an average intensity is calculated per patient across the benign cores, and across the malignant cores. The Pearson correlation between manual and automated scoring was 0.76. When stratified by benign or malignant tissue, the Pearson correlation between manual and automated scoring was 0.75 (benign) and 0.78 (malignant). Automated scoring of AR staining is comparable to manual scoring of AR in prostate tissue. In light of the greater efficiency of automated scoring, future evaluations of immunohistochemical staining may be usefully conducted using automated scoring of nuclei. Citation Format: Rochelle P. Ondracek, James L. Mohler, Catherine Pilarz, Sarah McEvoy, Karin Kasza, Carl Morrison, James Marshall. Comparison of manual and automated scoring of androgen receptor staining in prostate tissue. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4852. doi:10.1158/1538-7445.AM2013-4852

Smoking, Sex, and Non–Small Cell Lung Cancer: Steroid Hormone Receptors in Tumor Tissue (S0424)

Background To what extent steroid hormones contribute to lung cancer in male and female never smokers and smokers is unclear. We examined expression of hormone receptors in lung tumors by sex and smoking. Methods Patients with primary non-small cell lung cancer were recruited into an Intergroup study in the United States and Canada, led by SWOG (S0424). Tumors from 813 cases (450 women and 363 men) were assayed using immunohistochemistry for estrogen receptor (ER)-α, ER-β, progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). Linear regression was used to examine differences in expression by sex and smoking status. Cox proportional hazard models were used to estimate survival associated with the receptors. All statistical tests were two-sided. Results In ever smokers, postmenopause and oral contraceptive use were associated with lower nuclear ER-β (P = .02) and total (nuclear + cytoplasmic) PR expression (P = .02), respectively. Women had lower cytoplasmic ER-α (regression coefficient [β], or differences in H-scores = -15.8, P = .003) and nuclear ER-β (β = -12.8, P = .04) expression than men, adjusting for age, race, and smoking. Ever smokers had both higher cytoplasmic ER-α (β = 45.0, P < .001) and ER-β (β = 25.9, P < .001) but lower total PR (β = -42.1, P < .001) than never smokers. Higher cytoplasmic ER-α and ER-β were associated with worse survival (hazard ratio = 1.73, 95% confidence interval [CI] = 1.15 to 2.58, and HR = 1.59, 95% CI = 1.08 to 2.33, respectively; quartiles 4 vs 1). Conclusions Lower expression of nuclear ER-β in women supports the estrogen hypothesis in lung cancer etiology. Increasing cytoplasmic ER-α and ER-β and decreasing PR protein expression may be mechanisms whereby smoking disrupts hormone pathways.

Frequency of breast cancer subtypes among African American women in the AMBER consortium

BackgroundBreast cancer subtype can be classified using standard clinical markers (estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2)), supplemented with additional markers. However, automated biomarker scoring and classification schemes have not been standardized. The aim of this study was to optimize tumor classification using automated methods in order to describe subtype frequency in the African American Breast Cancer Epidemiology and Risk (AMBER) consortium.MethodsUsing immunohistochemistry (IHC), we quantified the expression of ER, PR, HER2, the proliferation marker Ki67, and two basal-like biomarkers, epidermal growth factor receptor (EGFR) and cytokeratin (CK)5/6, in 1381 invasive breast tumors from African American women. RNA-based (prediction analysis of microarray 50 (PAM50)) subtype, available for 574 (42%) cases, was used to optimize classification. Subtype frequency was calculated, and associations between subtype and tumor characteristics were estimated using logistic regression.ResultsRelative to ER, PR and HER2 from medical records, central IHC staining and the addition of Ki67 or combined tumor grade improved accuracy for classifying PAM50-based luminal subtypes. Few triple negative cases (< 2%) lacked EGFR and CK5/6 expression, thereby providing little improvement in accuracy for identifying basal-like tumors. Relative to luminal A subtype, all other subtypes had higher combined grade and were larger, and ER-/HER2+ tumors were more often lymph node positive and late stage tumors. The frequency of basal-like tumors was 31%, exceeded only slightly by luminal A tumors (37%).ConclusionsOur findings indicate that automated IHC-based classification produces tumor subtype frequencies approximating those from PAM50-based classification and highlight high frequency of basal-like and low frequency of luminal A breast cancer in a large study of African American women.

Impact of devascularization and tissue procurement on cell number and RNA integrity in prostatectomy tissue.

Minimizing the time between tissue devascularization in robot‐assisted laparoscopic radical prostatectomy (RALP) and tissue procurement should produce the highest quality tissue for research study. This study examines the relationship between intra‐operative time and two indicators of tissue integrity: number of epithelial cells per gram of tissue and RNA integrity numbers (RINs). The study also compares the RIN values of tissue obtained intra‐operatively by biopsy, before and after devascularization, to those from RALP specimen tissue, obtained through the routine research tissue procurement process.