Citations

249 Background: Serum CGA has been identified as a candidate prognostic biomarker for mCRPC. In a two cohort study, we compared the prognostic value of serum CGA with a validated CTC assay. Methods: In the discovery cohort (DC), blood samples were collected from 256 men with mCRPC. In an independent validation cohort (VC), 92 men with mCRPC were enrolled in a biospecimen collection study. In both cohorts, men receiving proton pump inhibitors and those with non-castrate levels of testosterone ( > 50ng/dl) were excluded. Serum CGA was measured in a homogeneous automated immunofluorescent assay using time-resolved amplified cryptate emission. In the VC, CTC enumeration was performed using the FDA cleared CELLSEARCH CTC test prior to treatment with abiraterone acetate/ prednisone. Cox proportional hazard regression and Kaplan-Meier analysis were performed for associations with elevated CGA (above reference range), unfavorable (≥ 5) CTCs, and overall survival (OS). Results: In the DC, 200 men were eligible for analysis. The median age was 72 years (yrs), 81/200 pts had a Gleason score (GS) ≥ 8, 34/200 had an elevated CGA. At a median follow up of 2.2 yrs, 156/200 were deceased. In the subset of men with GS ≥ 8, elevated CGA was associated with shorter OS [hazard ratio (HR) 2.19, p = 0.017]. In the VC, 71 men were eligible for analysis. The median age was 71 yrs, 36/71 tumors were GS ≥ 8, 31/71 pts had an elevated CGA, 26/66 had unfavorable CTCs (≥ 5). At a median follow up of 1.8 yrs, 31/71 were deceased. Elevated CGA (HR 1.91, p = 0.043) and unfavorable CTC counts (HR 2.97, p = 0.0012) were adversely associated with OS. In the high GS group, both CTCs and CGA had the same area under the curve (AUC) of 0.72. Pts with elevated CTC and CGA had the poorest OS (HR 3.76, p = 0.008). Conclusions: Elevated serum CGA was negatively associated with OS in men with mCRPC. Serum CGA represents a prognostic biomarker that may complement CTC enumeration. Clinical trial information: NCT#01953640.

Background Docetaxel has recently been found to improve survival in patients with metastatic androgen-independent prostate cancer (AIPC). Chemotherapy as a first-line option leaves room for improvement, while second-line options are multiple and somewhat controversial. Methods Clinically relevant articles focusing on chemotherapy drugs for metastatic prostate cancer and their mechanism of action and efficacy were reviewed from January 2004 through April 2006. Results Docetaxel is the standard of care for AIPC. However, for doublets with docetaxel or second-line chemotherapy, multiple studies have shown interesting and promising results with calcitriol, thalidomide, bevacizumab, satraplatin, vaccines, ixabepilone, and atrasentan. Conclusions Docetaxel should be considered for first-line treatment of metastatic AIPC. Due to its progression-free survival of only 6 months, more effective drugs and drug combinations need to be developed to treat patients with AIPC. Combination treatments with docetaxel and other new agents are promising, but adequately powered phase III trials need to be conducted with survival as the principal endpoint for these promising drug combinations.

Abstract Purpose: Androgen receptor (AR) variant AR-V7 is a ligand-independent transcription factor that promotes prostate cancer resistance to AR-targeted therapies. Accordingly, efforts are under way to develop strategies for monitoring and inhibiting AR-V7 in castration-resistant prostate cancer (CRPC). The purpose of this study was to understand whether other AR variants may be coexpressed with AR-V7 and promote resistance to AR-targeted therapies. Experimental Design: We utilized complementary short- and long-read sequencing of intact AR mRNA isoforms to characterize AR expression in CRPC models. Coexpression of AR-V7 and AR-V9 mRNA in CRPC metastases and circulating tumor cells was assessed by RNA-seq and RT-PCR, respectively. Expression of AR-V9 protein in CRPC models was evaluated with polyclonal antisera. Multivariate analysis was performed to test whether AR variant mRNA expression in metastatic tissues was associated with a 12-week progression-free survival endpoint in a prospective clinical trial of 78 CRPC-stage patients initiating therapy with the androgen synthesis inhibitor, abiraterone acetate. Results: AR-V9 was frequently coexpressed with AR-V7. Both AR variant species were found to share a common 3′ terminal cryptic exon, which rendered AR-V9 susceptible to experimental manipulations that were previously thought to target AR-V7 uniquely. AR-V9 promoted ligand-independent growth of prostate cancer cells. High AR-V9 mRNA expression in CRPC metastases was predictive of primary resistance to abiraterone acetate (HR = 4.0; 95% confidence interval, 1.31–12.2; P = 0.02). Conclusions: AR-V9 may be an important component of therapeutic resistance in CRPC. Clin Cancer Res; 23(16); 4704–15. ©2017 AACR.

175 Background: Genome and transcriptome aberrations associated with primary resistance to abiraterone acetate/prednisone (AA/P) in mCRPC are not known. In a prospective trial (NCT#01953640) we assessed whole-exome and RNA-seq based aberrations in metastases of CRPC stage patients (pts) for identifying markers associated with primary resistance to AA/P. Methods: Whole-exome and RNA-seq of biopsies from metastases was performed followed by analyses for association between genomic aberrations & primary resistance. Primary resistance was defined by progression on AA/P after 12-weeks of therapy(non-responders) using PSA, RECIST, bone scan and symptom criteria (per PCWG2). Gene network analysis was performed in genes mutated more frequently in non-responders, and also in genes that were differentially expressed between non-responders and responders and a “risk ratio” (RR) was calculated thereof. Results: Between 6/2013 & 8/2015, 92 pts were enrolled of which 82 had complete whole-exome, RNA-seq and 12-week outcome data available for analysis. Median age was 72.5 yrs (IQR: 68.5-78); median PSA-18 ng/ml (IQR: 8.1- 46.6). At 12-weeks 33/82 had progressed. Using the RR of 2 as threshold, we identified 113 and 292 genes that were more frequently mutated in non-responders & responders respectively. Among the 113 genes, OBSCN, ADAM21, LPHN3, DOCK10 ( P = 0.08, RR= inf) & USP42 ( P = 0.16, RR = 5.71) were the top candidates.Gene network analysis revealed that in non-responders, genes involved in the Wnt/β-catenin pathway were frequently mutated and negative regulators of Wnt pathway ( DKK4, SFRP2 & LRP6) were also frequently deleted. Gene expression analyses revealed the expression levels of Wnt/β-catenin pathway inhibitors were significantly reduced while expression levels of cell cycle regulatory genes were significantly increased in non-responders. Conclusions: In this study we observed Wnt/β-catenin pathway activation to be associated with primary AA/P resistance. This finding offers a potential for the development of predictive biomarkers and modulation of targeted pathways to overcome AA/P resistance. Clinical trial information: NCT# 01953640.

152 Background: Plasma cfDNA ARcopy number variations (CNV) have been suggested to have prognostic significance in mCRPC stage. We compared plasma AR amplification ( ARamp) with CTC counts (the only FDA approved prognostic marker in mCRPC stage) in a prospective trial of patients (pts) progressing on androgen deprivation therapy. Methods: Plasma and CTC counts were obtained concurrently as part of a prospective clinical trial in mCRPC stage pts initiating abiraterone acetate (NCT# 01953640). Plasma cfDNA was purified using DNA Blood Mini Kit (Qiagen, Valencia, CA) and AR CNVs were detected using QuantStudio3D digital PCR system (Life Technologies, Carlsbad, CA, USA). CTC enumeration was performed using the FDA cleared CellSearch platform. The distributions of overall survival (OS) time based on ARamp status and CTC counts ( < 5 vs ≥ 5 cells) were estimated using the Kaplan-Meier method. Log-rank tests were used to test association of ARamp status and CTC counts with overall survival (OS). Univariate and multivariate logistic regression models and ROC evaluated association of OS at 24 months by ARamp status and CTCs ≥ 5. Results: Between 05/2013 and 09/2015, 92 pts were enrolled of which complete CTC, plasma and outcome data was available for 70 pts. Median age of the cohort (range) was 72 yrs (39-91); Median PSA–16.2 ng/ml (range: 0.9-2026.0); Median CTC count was 2.0 (range: 0-372); 58/70 pts had bone metastases; 38/70 had high volume and 32/70 had low volume metastatic disease at the time of sample collection. The median (range) follow up time at the time of this analysis was: 26.5 (7.9-37.0) months at which time 28/70 patients had died. ARamp was present in 19/70 pts while 51/70 did not have amplification. 24/70 had CTCs ≥ 5. A significant association of increased ARamp status with OS (p < 0.0001, log-rank test; AUC: 0.66) as well as with CTC counts ≥ 5 (p = 0.001, log-rank test; AUC: 0.67) was observed. Combining CTC status and plasma ARamp status the AUC was 0.75. Conclusions: Plasma cfDNA ARamp status and CTC counts have comparable and clinically meaningful prognostic marker value in mCRPC stage patients.

260 Background: The purpose of this study was to evaluate cell free nucleic acid based biomarkers for prognosis in mCRPC stage. Methods: Uniformly collected blood from mCRPC patients was processed for plasma which was used for exosomal RNA extraction. RNA sequencing was applied to examine microRNA (miRNA) profiles initially in a discovery cohort of 23 mCRPC patients. The sequences from fastq files were mapped to human miRBase (release 19) for known miRNA read counts. A software package (miRDeep2) was used for novel miRNA prediction. Cox regression was used for association of candidate miRNAs with overall survival (OS), defined as time from development of mCRPC to death or last follow up after adjusting for Gleason Score (GS) and time on androgen deprivation (AD) for hormone sensitive disease. miRNAs significantly associated with OS (FDR<0.05) were then validated in an independent cohort of 100 mCRPC subjects using univariate Cox regression. Results: RNA sequencing generated >10 million mappable reads per specimen in the discovery cohort. Among these reads ~50% were known miRNAs which included 1237 unique RNA sequences. Due to low abundance of most RNA transcripts, only miRNAs (n=250) with >/= 32 reads (on average) were analyzed. Four mRNAs (miR-1290, -1246, -375 and a predicted miRNA on chr. 12) were associated with OS(FDR<0.05). These were evaluated as prognotic markers in the independent cohort (N=100) using real time RT-PCR. Median time on AD for this cohort prior to progression to mCRPC was 35.7 (range: 3.5-136) months; median GS was 7; median follow up after mCRPC development was 12.5 (1-41) months at which time 28 patients had died. Elevated expression for miR-1290 and -375 were associated with shorter OS after adjusting for GS and AD time with a Hazard Ratio (HR) for miR-375 of 1.19 (CI: 1.006-1.18); (p=0.04) and a HR for miR1290 of 1.58 (CI:1.01-2.4); (p=0.04). A stronger association with shorter OS was observed for elevated expression of both miRs-HR of 2.02 (CI: 1.1-3.6) (p=0.01). Conclusions: Blood-based nucleic acid assays are easily accessible for biomarker development in mCRPC. Results from this study suggest specific exosomal miRs are of prognostic value in mCRPC and should be validated in larger cohorts.

249 Background: Evaluating tumor specific pathologic and genetic profiles in mCRPC stage is difficult due to the limited availability of mCRPC tissue. We describe findings from a prospective cohort study performed to obtain concomitant histopathology and genetic information in mCRPC. Methods: Patients with mCRPC initiating abiraterone acetate therapy underwent 2 serial (3 months apart) metastatic (met) site needle core biopsies (NCB1/2). Bone lesions were biopsied using 11-13G core needles and 18G core biopsy devices were used for non-osseous masses. Up to 4 cores were obtained at each biopsy with the 1st core (S1) sent for DNA sequencing, the 2nd (S2) for RNA sequencing, and the 3rd/4th (X3, X4) submitted for xenograft implantation. From each, 1-2 mm segments were separated and formalin-fixed for histopathologic examination (HPE). Results: A total of 54 patients, enrolled between June 2013 and July 2014, underwent 94 NCB (54 NCB1, 40 NCB2), rendering a total of 259 samples (94 S1; 75 S2; 59 X3 and 31 X4) of which 85 (32%) were positive (pos) for tumor by HPE. Positivity for tumor in S1, S2, X3, and X4 cores was 42%, 33%, 22%, and 19%, respectively. At least one core pos for met tumor was observed in 62% NCB1 and 52% NCB2 (overall 52/94; 56%). Met sites biopsied include bone (71), lymph nodes (18), liver (3), penile (1) and pelvic (1) soft tissues. HPE revealed met adenocarcinoma in 44/52, and poorly differentiated carcinoma in 8/52. Gleason grade applied to the met ranged from 3+4 to 5+5. 45/85 pos samples (53%) had > 50% tumor cellularity. Pos NCB in bone lesions was observed in 23/71 (32%), compared to 17/23 (73%) of non-bone sites (p=.0004). 85 S1 samples yielded DNA material, of which 66 (77%) had ≥10% tumoral DNA. 24 cases with negative HPE had ≥10% tumoral DNA; 7 cases with pos HPE had <10 tumoral DNA. In all, 76/94 (81%) NCB yielded tumor material either by HPE or DNA analysis. Conclusions: NCB of mCRPC is feasible and provides adequate tissue for pursuing HPE and sequencing studies. HPE of extracted material correlates with DNA sequencing data and provides complementary information on tumor features. Bone lesions yield significantly less tumoral material than non-osseous sites.

190 Background: It is not known if biopsy of CRPC stage metastases alters CTC results. We evaluated pre & post biopsy CTC counts in a prospective cohort undergoing pre-chemotherapy abiraterone acetate/prednisone (AA/P). Methods: CTC enumeration was performed on 7.5mL blood using the FDA cleared immunomagnetic/immunofluorescence assay (CellSearch). Time of blood draw for CTC counts pre/post biopsy was recorded for each biopsy. Biopsies were performed using 11/13 gauge core biopsy device for bone lesions and 18 gauge core needle biopsy device for soft tissue/nodal metastases. All patients (pts) underwent two serial biopsies 3 months apart. Change in CTC counts pre/post biopsy for visits 1 and 2 were evaluated using Wilcoxon signed rank test (significance at p<0.05). Differences in CTC count change (visit1) pre/post biopsy based on metastatic site (skeletal vs. non-skeletal) was compared using Wilcoxon rank-sum test. Results: Median age of the cohort was 74 years (IQR: 68-78); median PSA at V1 was 16.4 ng/ml (IQR: 6.4-87.6); at V2 was 11.5 ng/ml (IQR: 1.8-23.7). Of 59 pts undergoing biopsies 42 were in bone and 17 in nodal/soft tissue masses. At V1, pre/post biopsy CTC counts were measured on 50 and 49 pts, respectively; at V2, pre/post CTC counts were measured on 43 and 36 pts. High volume metastatic disease at V1 was observed in 67.8 % of all pts. Table lists CTC counts and time of collection pre/post biopsy for both visits. Mean change in pre/post biopsy CTC counts for skeletal sites was 0.7 (range: -12 to 64) compared to 12.1 (range: -21 to 13) for non-skeletal sites (p=0.23). Conclusions: An increase in mean CTC counts after biopsy of CRPC metastases was observed. No significant differences in change of CTC counts based on site biopsied (skeletal versus non-skeletal) was noted. Correlation with clinical outcomes is on-going. Clinical trial information: NCT# 01953640. [Table: see text]

174 Background: AA/P is an FDA approved treatment for mCRPC. Since markers of early resistance to AA/P are unknown, we report initial findings of androgen receptor (AR) based associations with short term (12 week) progression on AA/P. Methods: mCRPC stage patients (pts) initiating pre-chemotherapy AA/P underwent metastatic site biopsies at baseline (pre AA/P) and after 12 weeks. Baseline somatic whole exome DNA, tumor gene expression for AR full length (ARFL), AR splice variant 7 (ARV7) and ARV7/ARFL ratios were compared in progessors versus non-progressors. Progression at or within 12 weeks of AA/P therapy was defined as death or disease progression by PCWG2 “composite progression (CP)” and/or “radiographic progression” endpoints. Wilcoxon rank-sum tests were used to test for differences in the two groups for comparing ARFL, ARV7 expressions and ARV7/ARFL ratios and chi square tests were used for differences in copy number variation. Results: Between 1/2013 and 6/2014, 59 pts were enrolled of which 44 have disease assessment data at the12-week time point. CP was observed in 17/44 patients. DNA seq and clinical data was available for 42/44 pts. Using radiographic progression at 12 weeks, AR Amplification/Gain was observed in 20/26 non-progressors (13 with Amplification) and in 9/16 progressors (P-value = 0.19; OR 2.5). ARFL and ARV7 gene expressions in both groups is provided in table. Conclusions: A trend towards higher ARV7/ARFL gene expression ratio in metastases was observed with early progression on AA/P. AR gain/amplification is observed less often in pts with early progression. Validation of these findings is on-going in this prospective trial. Clinical trial information: NCT# 01953640. [Table: see text]