Castration-resistant Prostate Cancer Patients Research Articles

Efficacy of Systemic Treatment in Prostate Cancer Patients With Visceral Metastasis: A Systematic Review, Meta-analysis, and Network Meta-analysis.

There are limited pooled data showing the impact of visceral metastasis on oncologic outcomes in metastatic prostate cancer patients treated with combination systemic therapies. We aimed to analyze and compare the efficacy of combination systemic therapies in metastatic hormone-sensitive prostate cancer and metastatic castration-resistant prostate cancer with or without visceral metastasis. Three databases were queried in July 2022 for randomized, controlled trials analyzing metastatic prostate cancer patients treated with combination systemic therapy (androgen receptor signaling inhibitor and/or docetaxel plus androgen deprivation therapy) to standard of care. We analyzed the association between presence of visceral metastases and efficacy of systemic therapies in metastatic hormone-sensitive prostate cancer and metastatic castration-resistant prostate cancer patients. The main and secondary outcomes of interest were overall survival and progression-free survival, respectively. Formal meta-analysis using fixed-effect model and network meta-analysis using random-effect model with a frequentist approach were conducted. We followed PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-analyses) and AMSTAR (A MeaSurement Tool to Assess systematic Reviews) guidelines. Overall, 12 and 8 randomized, controlled trials were included for systematic review and meta-analyses/network meta-analyses, respectively. In metastatic hormone-sensitive prostate cancer patients, adding androgen receptor signaling inhibitor to standard of care improved overall survival in patients with visceral metastasis (pooled HR: 0.77, 95% CI: 0.64-0.94) as well as in those without (pooled HR: 0.66, 95% CI: 0.60-0.72; no differences in both across- and within-trial approach; P = .13 and P = .06, respectively). On the other hand, the progression-free survival benefit from androgen receptor signaling inhibitor + androgen deprivation therapy was significantly lower in patients with visceral metastasis using across-trial approach (P = .03), while it did not reach statistical significance using within-trial approach (P = .14). Analysis of treatment ranking in metastatic hormone-sensitive prostate cancer showed that darolutamide + docetaxel + androgen deprivation therapy had the highest likelihood of improved overall survival irrespective of visceral metastasis. In post-docetaxel metastatic castration-resistant prostate cancer patients, adding androgen receptor signaling inhibitor to androgen deprivation therapy significantly improved overall survival in both patients with visceral metastasis (pooled HR: 0.79, 95% CI: 0.63-0.98) and those without (pooled HR: 0.63, 95% CI: 0.55-0.72). No randomized, controlled trials reported the differential oncologic outcomes stratified by lung vs liver metastases. Despite aggressive clinical behavior and worse trajectory of metastatic hormone-sensitive prostate cancer and metastatic castration-resistant prostate cancer with visceral metastasis, the effectiveness of novel systemic therapies is similar in both metastatic hormone-sensitive prostate cancer and metastatic castration-resistant prostate cancer patients with and without visceral metastasis. Further well-designed studies with detailed visceral metastatic sites and number will enrich the clinical decision-making.

Personalized [177Lu]Lutetium-PSMA Therapy for Patients with Pre-Treated Castration-Resistant Prostate Cancer: A Single Institution Experience from a Comprehensive Cancer Centre.

Castration resistant prostate cancer (CRPC) is characterized by an aggressive biological behavior with a relatively short survival time, especially in progressive tumors pretreated with new hormonal agents and taxane chemotherapy. [177Lu]-Lutetium-PSMA (Lu-PSMA) treatment has proven efficacy in these patients. However, around 30% of the CRPC patients do not benefit from Lu-PSMA treatment, and little is known about predictive factors for treatment success if Lu-PSMA is offered in an individualized approach based on clinical and laboratory features. In this monocentric retrospective study, 86 CRPC patients receiving Lu-PSMA treatment were evaluated. The focus of the study was to describe clinical factors at baseline and during early treatment that are related to overall survival (OS). In addition, PSMA PET/CT-, PSA-response, and safety and tolerability (CTCAE adverse event reporting) were assessed. Efficacy endpoints were calculated using stratified Kaplan-Meier methods and Cox regression models. Mean applied dose was 17.7 GBq (mean 5.3 ± 1.1 GBq per cycle) with an average of 3.6 (range 1-8) therapy cycles. Patients were followed up for a mean of 12.4 months (range 1-39). The median OS was 15 months (95% CI 12.8-17.2). The best overall response rate in patients assessed with PSMA PET/CT and PSA response was 27.9%, and 50.0% had at least stable disease. Nine patients had a ≥grade 3 adverse event with anemia being the most frequent adverse event. Positive predictors for prolonged OS from baseline parameters were pre-treatment hemoglobin level of ≥10 g/dL and a lower PSA values at treatment start, while the presence of visceral or liver metastases were not significantly associated with worse prognoses in this cohort. With careful patient selection, an individualized Lu-PSMA treatment approach is feasible and patients with dose-limiting factors or visceral metastases should be included in prospective trials.

Mental health care utilization among men with castration-resistant prostate cancer receiving abiraterone or enzalutamide.

Abiraterone and enzalutamide are castration-resistant prostate cancer (CRPC) therapies with potentially distinct associations with mental health symptoms given their differing antiandrogen targets. We used national Veterans Health Administration data to identify patients with CRPC who received first-line abiraterone or enzalutamide from 2010 to 2017. Using Poisson regression, we compared outpatient mental health encounters per 100 patient-months on drug between the abiraterone and enzalutamide cohorts adjusting for patient factors (e.g., age). We compared mental health encounters in the year before versus after starting therapy using the McNemar test. We identified 2902 CRPC patients who received abiraterone (n = 1992) or enzalutamide (n = 910). We found no difference in outpatient mental health encounters between the two groups (adjusted incident rate ratio [aIRR] 1.04, 95% confidence interval [CI] 0.95-1.15). However, men with preexisting mental health diagnoses received 81.3% of the outpatient mental health encounters and had higher rates of these encounters with enzalutamide (aIRR 1.21, 95% CI 1.09-1.34). Among patients with ≥1 year of enrollment before and after starting abiraterone (n = 1139) or enzalutamide (n = 446), there was no difference in mental health care utilization before versus after starting treatment (17.0% of patients vs. 17.6%, p = 0.60, abiraterone; 16.4% vs. 18.4%, p = 0.26, enzalutamide). We found no overall differences in mental health care utilization between CRPC patients who received first-line abiraterone versus enzalutamide. However, men with preexisting mental health diagnoses received the majority of mental health care and had more mental health visits with enzalutamide.

Conditional survival does not improve over time in metastatic castration-resistant prostate cancer patients undergoing docetaxel.

To investigate the conditional overall survival (OS) ofmetastatic castration-resistant prostate cancer (mCRPC) patients receiving docetaxel chemotherapy. We used deidentified patient-level data from the Prostate Cancer DREAM Challenge database and the control arm of the ENTHUSE 14 trial. We identified 2158 chemonaïve mCRPC patients undergoing docetaxel chemotherapy in the five randomized clinical trials. The 6-month conditional OS was calculated at times 0, 6, 12, 18, and 24 months from randomization. Survival curves of each group were compared using the log-rank test. Patients were then stratified into low- and high-risk groups based on the median predicted value of our recently published nomogram predicting OS in mCRPC patients. Nearly half (45%) of the study population was aged between 65 and 74 years. Median interquartile range prostate-specific antigen for the overall cohort was 83.2 (29.6-243) ng/mL, and 59% of patients had bone metastasis with or without lymph node involvement. The 6-month conditional survival rates at 0, 6, 12, 18, and 24 months for the entire cohort were 93% (95% confidence interval [CI]: 92-94), 82% (95% CI: 81-84), 76% (95% CI: 73-78), 75% (95% CI: 71-78), and 71% (95% CI: 65-76). These rates were, respectively, 96% (95% CI: 95-97), 92% (95% CI: 90-93), 84% (95% CI: 81-87), 81% (95% CI: 77-85), and 79% (95% CI: 72-84) in the low-risk group and 89% (95% CI: 87-91), 73% (95% CI: 70-76), 65% (95% CI: 60-69), 64% (95% CI: 58-70), and 58% (95% CI: 47-67) in the high-risk group. The conditional OS for patients undergoing docetaxel chemotherapy tends to plateau over time, with the main drop in conditional OS happening during the first year from initiating docetaxel treatment. That is the longer a patient survives, the more likely they are to survive further. This prognostic information could be a useful tool for a more accurate tailoring of both follow-up and therapies. In this report, we looked at the future survival in months of patients with metastatic castration resistant prostate cancer on chemotherapy who have already survived a certain period. We found that the longer time that a patient survives, the more likely they will continue to survive. We conclude that this information will help physicians tailor follow-ups and treatments for patients for a more accurate personalized medicine.

Elevated Body Mass Index Is Associated with Improved Overall Survival in Castration-Resistant Prostate Cancer Patients Undergoing Prostate-Specific Membrane Antigen-Directed Radioligand Therapy.

In patients with prostate cancer scheduled for systemic treatment, being overweight is linked to prolonged overall survival (OS), whereas sarcopenia is associated with shorter OS. We investigated fat-related and body composition parameters in patients undergoing prostate-specific membrane antigen (PSMA)-directed radioligand therapy (RLT) to assess their predictive value for OS. Methods: Body mass index (BMI, in kg/m2) and CT-derived body composition parameters (total, subcutaneous, visceral fat area, and psoas muscle area at the L3-L4 level) were determined for 171 patients scheduled for PSMA-directed RLT. After normalization for stature, the psoas muscle index was used to define sarcopenia. Outcome analysis was performed using Kaplan-Meier curves and Cox regression including fat-related and other clinical parameters (Gleason score, C-reactive protein [CRP], lactate dehydrogenase [LDH], hemoglobin, and prostate-specific antigen levels). The Harrell C-index was used for goodness-of-fit analysis. Results: Sixty-five patients (38%) had sarcopenia, and 98 patients (57.3%) had increased BMI. Relative to the 8-mo OS in normal-weight men (BMI < 25), overweight men (25 ≥ BMI > 30) and obese men (BMI ≥ 30) achieved a longer OS of 14 mo (hazard ratio [HR], 0.63; 95% CI, 0.40-0.99; P = 0.03) and 13 mo (HR, 0.47; 95% CI, 0.29-0.77; P = 0.004), respectively. Sarcopenia showed no impact on OS (11 vs. 12 mo; HR, 1.4; 95% CI, 0.91-2.1; P = 0.09). Most of the body composition parameters were tightly linked to OS on univariable analyses, with the highest C-index for BMI. In multivariable analysis, a higher BMI (HR, 0.91; 95% CI, 0.86-0.97; P = 0.006), lower CRP (HR, 1.09; 95% CI, 1.03-1.14; P < 0.001), lower LDH (HR, 1.08; 95% CI, 1.03-1.14; P < 0.001), and longer interval between initial diagnosis and RLT (HR, 0.95; 95% CI, 0.91-0.99; P = 0.02) were significant predictors of OS. Conclusion: Increased fat reserves assessed by BMI, CRP, LDH, and interval between initial diagnosis and RLT, but not CT-derived body composition parameters, were relevant predictors for OS. As BMI can be altered, future research should investigate whether a high-calorie diet before or during PSMA RLT may improve OS.

The Impact of PSMA PET-Based Eligibility Criteria Used in the Prospective Phase II TheraP Trial in Metastatic Castration-Resistant Prostate Cancer Patients Undergoing Prostate-Specific Membrane Antigen-Targeted Radioligand Therapy.

Prostate-specific membrane antigen (PSMA) radioligand therapy (RLT) has shown encouraging results for treatment of metastatic castration-resistant prostate cancer (mCRPC) in the prospective, multicenter, randomized phase II TheraP study. The inclusion criteria for that study comprised a pretherapeutic 68Ga-PSMA-11 PET scan showing sufficient tumor uptake using a predefined threshold and the absence of 18F-FDG-positive, PSMA ligand-negative tumor lesions. However, the prognostic value of these PET-based inclusion criteria remains unclear. Therefore, we evaluated the outcome of mCRPC patients treated with PSMA RLT using TheraP as well as other TheraP-based PET inclusion criteria. Methods: First, patients were dichotomized into 2 groups whose PSMA PET scans did (TheraP contrast-enhanced PSMA [cePSMA] PET-positive) or did not (TheraP cePSMA PET-negative) fulfill the inclusion criteria of TheraP. Notably, unlike in TheraP, 18F-FDG PET was not performed on our patients. Prostate-specific antigen (PSA) response (PSA decline ≥ 50% from baseline), PSA progression-free survival, and overall survival (OS) were compared. Additionally, patients were further dichotomized according to predefined SUVmax thresholds different from those used in TheraP to analyze their potential impact on outcome as well. Results: In total, 107 mCRPC patients were included in this analysis (TheraP cePSMA PET-positive, n = 77; TheraP cePSMA PET-negative, n = 30). PSA response rates were higher in TheraP cePSMA PET-positive patients than in TheraP cePSMA PET-negative patients (54.5% vs. 20%, respectively; P = 0.0012). The median PSA progression-free survival (P = 0.007) and OS (P = 0.0007) of patients were significantly longer in the TheraP cePSMA PET-positive group than in the TheraP cePSMA PET-negative group. Moreover, being in the TheraP cePSMA PET-positive group was identified as a significant prognosticator of longer OS (P = 0.003). The application of different SUVmax thresholds for a single hottest lesion demonstrated no influence on outcome in patients eligible for PSMA RLT. Conclusion: Patient selection for PSMA RLT according to the inclusion criteria of TheraP led to a better treatment response and outcome in our preselected patient cohort. However, a relevant number of patients not fulfilling these criteria also showed substantial rates of response.

Abstract B025: MicroRNA regulators of neuroendocrine differentiation of prostate cancer

Abstract Background: Therapy-induced neuroendocrine prostate cancer (NEPC) is an extremely aggressive variant of castration-resistant prostate cancer (CRPC) that is increasing in incidence with the widespread use of second generation of androgen receptor (AR)-pathway inhibitors (APIs) such as Enzalutamide (ENZ) and Abiraterone. This aggressive variant arises from CRPC-Adenocarcinomas (CRPC-Adeno) via a reversible trans-differentiation process, referred to as neuroendocrine differentiation (NED) wherein cells undergo a lineage switch and exhibit neuroendocrine (NE) features, characterized by expression of neuronal markers such as enolase 2 (ENO2), chromogranin A (CHGA) and synaptophysin (SYP). There is an urgent need of identifying novel molecular markers to assess emergence of NED in CRPC patients. We demonstrated that progression of advanced CRPC with adenocarcinoma characteristics (CRPC-Adeno) to therapy-induced, androgen-independent NE (CRPC-NE) states is associated with a characteristic set of miRNA alterations that promote plasticity of advanced prostate adenocarcinomas to NEPC (Bhagirath et al., Oncogene, 2020). Importantly, we could develop a ‘novel miRNA classifier’ to robustly stratify CRPC-NE tumors from CRPC-Adenocarcinomas. Here we further validate the classifier in independent clinical cohorts and deduce the optimal miRNA genes required for NEPC diagnosis. We further study the functional significance of miR-28-3p, a miRNA identified by sequencing, in NEPC. Methods: Human clinical samples with corresponding clinical information were procured from two independent sites of Prostate Cancer Biorepository Network (PCBN). Samples included human CRPC-adeno vs CRPC-NE. FFPE sections from these clinical samples were microdissected, RNA were extracted and small RNA sequencing was performed using an Illumina NextSeq 500 platform. Sequencing data were analyzed and machine learning algorithms were applied (random forest machine learning technique with leave-pair-out cross validation (LPOCV)). The performance of classifier was measured using receiver operating characteristic (ROC) analysis with area under the curve (AUC) as the primary evaluation metric. The functional significance of miR-28-3p was studied by its overexpression and knockdown in PCa cell lines followed by functional assays. Results: Unsupervised analysis of sequencing data by principal component analyses (PCA) revealed distinct clustering of the CRPC-NE tumors from CRPC-Adenocarcinomas based on miRNA profiles suggesting that miRNA profiles can be used to stratify these tumor types. Our analyses showed that a set of 5 miRNAs of the classifier are important in distinguishing between CRPC-Adeno vs CRPC-NE with an AUC=0.8318. Further, miR 28-3p potentially has a biphasic role in PCa, where it is oncogenic in early stages and acts as a tumor suppressor in late stage PCa. Conclusions: A ‘5- miRNA’ classifier was validated to be of significance in two independent validation cohorts. We propose this miRNA classifier as an important tool for diagnosing NED in CRPC patients. Citation Format: Amritha Sreekumar, Sharanjot Saini. MicroRNA regulators of neuroendocrine differentiation of prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B025.

Abstract B007: New generation mTOR blocker sensitizes prostate cancer models to AR-targeting therapy

Abstract Advanced stage prostate cancers (PCa) can develop bone metastasis, a lethal feature of castration-resistant prostate cancer (CRPC) for which no curative options are currently available. Androgen deprivation therapy (ADT) is the main therapeutic strategy for treating advanced PCa; patients with bone metastasis, however, are frequently developing resistance to ADT, progressing to CRPC. Dysregulation of the mTOR pathway has been linked with AR reactivation and with progression to CRPC, and mutations in targets of the PI3K/Akt/mTOR pathway are frequently found in CRPC patients. This study investigates the effect of synergistic AR and mTOR pathways blockade on therapy resistance in advanced PCa in vitro and in vivo. LNCaP and LAPC4 cells were treated in vitro with AR blockers (apalutamide, enzalutamide), mTOR blockers (rapamycin, everolimus, MLN0128 or Rapalink-1) or a combination of both blockers. Viability was measured in a timecourse from 1 to 10 days after drug administration using CellTiter Acqueous 96. To mimick ADT in vitro, cells were cultured for 7 days in charcoal-stripped serum in medium without dihydroxytestosterone or phenol red before the drug screening. Two already established PDX models of bone-metastatic PCa, BM18 and LAPC9 respectively androgen-dependent and -independent, were used to assess the effect of combined ADT and mTOR-targeting therapy in vivo. Both LNCaP and LAPC4 models showed a higher sensitivity to MLN0128 and Rapalink-1 compared to rapalogs, LAPC4 cells in particular were more sensitive than LNCaP cells to mTOR inhibitors. AR blockers were moderately effective and only at concentrations between 10 and 100 µM, when used alone. However, treatment with sublethal doses of Rapalink-1 (0.001-0.01 µM) sensitized both cell lines to 2- to 3-log fold lower concentrations of apalutamide. In vivo, treatment with Rapalink-1 alone significantly reduced the growth rate of LAPC9 PDX, while co-treatment with ADT and Rapalink-1 resulted in regression in this CRPC model, a result not achievable by either treatment alone. Treatment with Rapalink-1, while effectively reducing cell viability in vitro already when used alone, could sensitize the tested models to AR blockers, at clinically relevant concentrations. In vivo treatment with Rapalink-1 was well-tolerated and sufficient to control the growth of the BM18 PDX model. When used in combination with ADT, it could induce regression of the CRPC model LAPC9. Citation Format: Shuang Li, Federico La Manna, Panagiotis Chouvardas, George Thalmann, Marianna Kruithof-de Julio. New generation mTOR blocker sensitizes prostate cancer models to AR-targeting therapy [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B007.

Abstract B023: Targeting noncanonical Wnt5a signaling suppresses the neural lineage network and overcomes enzalutamide resistance in advanced prostate cancer

Abstract INTRODUCTION AND OBJECTIVES Androgen receptor (AR) blockade using antiandrogens is a mainstay for the treatment of castration-resistant prostate cancer (CRPC). However, resistance to antiandrogens occurs unavoidably due to the escaping mechanisms of tumor cells that are not completely understood. Upregulation of non-canonical Wnt ligand Wnt5a is identified from circulating tumor cells from CRPC patients after being treated with enzalutamide. FZD2, the cognate frizzled receptor for Wnt5a, is the most commonly co-upregulated non-canonical Wnt signaling molecule in prostate cancer. In this study, we determined the role of non-canonical Wnt5a/FZD2 to enzalutamide resistance and neural lineage plasticity, and explore the potential of targeting Wnt5a/FZD2 to overcome resistance in advanced prostate cancer. METHODS Wnt5a and FZD2 expression was determined in enzalutamide-resistant C4-2B MDVR cells. Wnt5a and FZD2 expressions were modulated using specific siRNAs. Cell growth, colony formation, and migration were studied in vitro. Transcriptomic analysis was performed on C4-2B MDVR cells treated with FZD2 knocked down; gene program of non-canonical Wnt signaling, hallmark androgen response and AR-V7 associated genes, and neural lineage pathways were analyzed. A novel tRNA bioengineered Wnt5a siRNA was developed to target Wnt5a/FZD2 signaling. The effect of tRNA-siWnt5a on tumor growth, sensitivity to enzalutamide treatment, and neural lineage plasticity was evaluated in vitro and in vivo. RESULTS Wnt5a and FZD2 are highly upregulated in castration-resistant prostate cancer patients, which is verified overexpression in enzalutamide-resistant C4-2B MDVR cells. Knocking down Wnt5a and FZD2 diminishes the enrichment of the non-canonical Wnt signaling pathway. Downregulating Wnt5a and FZD2 expression by specific siRNAs suppresses prostate cancer cell proliferation and resensitized C4-2B MDVR cells to enzalutamide treatment. Suppressing Wnt5a and FZD2 abrogated the enrichment of gene programs regulating cancer cell survival/proliferation, and neural lineage pathways including neuron differentiation and embryonic stem cell markers. Using the bioengineered tRNA-Wnt5a siRNA effectively inhibited the growth of enzalutamide-resistant prostate cancer cells and resensitized tumor cells to enzalutamide treatment in vitro, and resistant CRPC PDX LuCaP35CR tumor growth in vivo. Conclusions Our results suggest that activation of noncanonical Wnt5a/FZD2 signaling confers to enzalutamide resistance and neural lineage plasticity. Targeting the Wnt5a signal could provide benefits for CRPC patients with tumors expressing a high level of Wnt5a, FZD2, and lineage markers, not only enhancing the anti-tumor effects of enzalutamide but suppressing the potential of neuroendocrine differentiation in advanced prostate cancer patients. Citation Format: Shu Ning, Wei Lou, Chengfei Liu, Alan Paul Lombard, Leandro S. D' Abronzo, Neelu Batra, Aiming Yu, Christopher P. Evans, Allen Gao. Targeting noncanonical Wnt5a signaling suppresses the neural lineage network and overcomes enzalutamide resistance in advanced prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B023.

Abstract PR013: Unlocking the proteome of metastatic prostate cancer circulating tumor cells

Abstract In recent years, liquid biopsies have emerged to provide improved insights on the current disease state of a patient, but they generally rely on a small number of protein markers from immunofluorescent staining or cell-free DNA, which provides limited information especially around drug targets. As more targeted therapies and immunotherapies enter the market, selecting the optimal drug target could improve patient outcomes drastically. One way to achieve this goal is to generate multi-omic datasets from the cancer cells in circulation, known as circulating tumor cells (CTCs). In this study, we collaborated with Astrin Biosciences and used their patented isolation technology to enrich CTCs with high purity from the blood of 15 metastatic castration resistant prostate cancer (mCRPC) patients. The enriched CTCs and a patient matched control sample consisting primarily of white blood cells were divided such that RNA and protein could be interrogated from the same patient. Label free unbiased mass spectrometry-based proteomics was used to identify the CTC-enriched proteins and marks the first time a global proteomic assessment has been performed on CTCs from cancer patients. Initial analysis identified over 1,500 peptides corresponding to greater than 500 proteins from each patients’ CTCs; including increased epithelial cell markers in the CTC-enriched sample and increased WBC markers in the control sample. Hallmarks of androgen response and detection of the androgen receptor were also identified in the CTC-enriched proteome. Overall, this study was designed to show the feasibility of obtaining large scale proteomic data from a liquid biopsy and future studies will provide more insight into the clinical utility of such data with a focus on cancer signaling mechanisms and biomarkers. Importantly, as we analyze more patients, we plan to compare the proteomic data with future RNA signatures across mCRPC patients to eventually be able to correlate them with drug response with the overall goal to improve treatment selection and patient outcomes. Citation Format: Justin M. Drake, Zoi Sychev, Alec Horrmann, Kaylee Judith Smith, Catalina Galeano-Garces, Ali Arafa, Nicholas Heller, Mahdi Ahmadi, Jiarong Hong, Megan Ludwig, Justin Hwang, Emmanuel S. Antonarakis, Jayant Parthasarathy. Unlocking the proteome of metastatic prostate cancer circulating tumor cells [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr PR013.

Abstract B024: Nuclear size and nuclear shape instability in prostate cancer epithelial cells during metastatic progression

Abstract Very small nuclear (vsn) circulating tumor cells (CTCs) are detected in a subgroup of metastatic, castration resistant prostate cancer (mCRPC) patients who develop visceral metastasis following intensive androgen receptor suppression. In addition, we have previously observed that nuclear shape instability (characterized by the loss and/or mis-localization of the nuclear envelop protein emerin) is present in human prostate cancer tissues, and in CTCs from patients with metastatic disease. Herein, we investigated the biological and clinical implications of the vsn phenotype in prostate CTCs. Patients with vsnCTCs had poor overall survival and low PSA response to androgen receptor suppression compared with non-vsnCTC patients. The vsnCTCs exhibited a loss/downregulation of emerin (EMD) and we found a direct correlation between emerin loss and nuclear size reduction in CTC from mCRPC patients. Further, in vitro studies revealed that the vsn phenotype was characterized by high invasive capacity, enhanced cellular plasticity, and resistance to AR target therapy. Finally, depletion of EMD in prostate tumor cells resulted in the vsn phenotype. Together, these findings demonstrate that the vsnCTC phenotype is characterized by deregulation of EMD expression, and the presence of these cells result in poor clinical outcomes in prostate cancer. Citation Format: Karen Cavassani, Edwin Posadas, Pai-Chi Teng, Sadaf Pustchi Pustchi, Minhyung Kim, Neil Bhowmick, Sungyong You, Michael Freeman, Leigh Ellis, Dolores Di Vizio, Hsian-Rong Tseng, Leland W. Chung, Susan Yao, Catherine Grasso. Nuclear size and nuclear shape instability in prostate cancer epithelial cells during metastatic progression [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B024.

Abstract A005: Androgen receptor variants mRNA absolute quantification in prostate cancer cell models

Abstract Background: Patients treated with Androgen Deprivation Therapy (ADT) have been shown to increase the expression of constitutively active Androgen Receptor (AR) splice variants (AR-Vs). These splice variants lack the ligand binding domain at the C-terminus of AR, required for binding to antiandrogen therapies. AR-V7 has also been correlated to the development and progression of castration resistant prostate cancer (CRPC). As not all circulating tumor cells (CTCs) contain AR-V7, we hypothesized that other AR-Vs are involved in the mechanism of resistance to ADT commonly seen in CRPC. To test this, we developed an mRNA quantitative assay targeting AR-Vs and then correlated our results with a targeted mass spectrometry (MS) based protein assay. This work will provide a feasible quantification of AR-V transcripts and that may provide a stratification for predicted responses to ADT treatment in patients that could be useful to clinicians. Methods: The absolute copy number of total AR, AR-V2, AR-V7, AR-V12, and AR-V23 were determined in eight different PCa cell lines and 48 patient derived xenografts (PDXs) using qRT-PCR assays. We performed siRNA targeting of these variants to evaluate primer specificity by evaluating knockdown efficiencies. These results will be correlated to targeted MS quantification of AR-Vs in each cell model. Results: We found that the most highly expressed variants in PCa cell lines were AR-V7 and AR-V12. In the LuCaP PDX model, AR-V12 was shown to have the highest expression of the AR-Vs tested. Specificity of AR-V knockdown was tested using AD-1 (which only expresses the full- length AR transcript) and R1-D567 (which only expresses AR-V567es, an AR-V12-like variant) cell lines. Conclusions: We were able to determine the landscape of AR-V mRNA expression in eight PCa cell lines and in 48 PDX tumor samples. This data will be correlated to protein expression currently being analyzed using a targeted MS method developed in our lab. Our work will be useful to help clinicians stratify PCa patients based on their AR-V expression profiles. This will establish a prognostic biomarker program that measures AR-V proteins in real time from clinical biopsy tissues, circulating tumor cells, or exosomes and informs the clinician on which course of treatment may be effective for each patient. Establishing a CRPC biomarker other than AR-V7 that can help explain the AR-V7 negative CRPC patients could eventually improve clinical application and predictive treatment outcomes. Citation Format: Gabrianne Larson, Zoi Sychev, Stephen Plymate, Eva Corey, Justin M. Drake. Androgen receptor variants mRNA absolute quantification in prostate cancer cell models [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr A005.

Abstract A078: Identifying and targeting the genetic determinants of immune suppression and immunotherapy failure in prostate cancer

Abstract The advent of immunotherapies such as immune checkpoint blockade (ICB) has greatly expanded the therapeutic options available to patients with treatment-refractory solid tumors. However, patients with castration-resistant prostate cancer (CRPC) generally remain unresponsive to ICB therapy due to an immunologically “cold” prostate tumor microenvironment (TME) with limited accessibility to cytotoxic lymphocytes such as Natural Killer (NK) and T cells that can mediate potent anti-tumor immunity. During the course of tumor evolution, CRPCs acquire many genomic alterations and copy number changes that can influence therapy outcomes. Yet the impact of such genetic alterations on the immune TME and response to ICB remains largely unknown, in part due to the lack of models to functionally study them in an immune competent setting. To this end, we previously developed a novel electroporation-based genetically engineered mouse model (EPO-GEMM) to rapidly generate genetically defined prostate tumors de novo within an intact prostate TME (Leibold*, Ruscetti* et al., Cancer Discovery 2020). Here, we applied the EPO-GEMM approach to characterize the immune landscapes of distinct genetic subtypes of CRPC and subsequently identify and target tumor intrinsic mediators of immune suppression for prostate cancer immunotherapy. Using transposon systems to overexpress the MYC oncogene and CRISPR/Cas9 vectors to inactivate tumor suppressor genes (TSGs) commonly altered in CRPC (Pten, P53, Rb1, Apc), we generated a suite of genetically distinct prostate cancer-bearing EPO-GEMMs. Histological analysis revealed genotype-specific immune infiltrates, with MYC-driven tumors generally lacking cytotoxic CD8+ T cells and harboring increased numbers of regulatory T cells (Tregs). A similar correlation was also observed between MYC levels and cytotoxic lymphocyte suppression in CRPC patient samples. In particular, tumors with compound MYC overexpression and p53 loss (MP) displayed drastic NK cell suppression. Cytokine and RNA-seq profiling of primary EPO-GEMM CRPCs, tumor-derived cell lines, and isogenic Myc-CaP cells engineered with TSG losses revealed that MP alterations led to not only inhibition of inflammatory cytokine and interferon signaling and antigen presentation/processing gene expression, but also induction of VEGF signaling and increased angiogenesis in the prostate TME. Treatment of MP tumors with a VEGFR2 antibody to block VEGF signaling significantly reduced tumor growth and increased NK and CD8+T cell infiltration and activation, indicating a phenotypic switch from a traditionally “cold” TME. Our results demonstrate that the genetic configuration of prostate cancer shapes its surrounding TME. Additionally, targeting VEGF signaling may be a mechanism to overcome ICB resistance in prostate cancer. More broadly, we believe that by understanding the tumor intrinsic mechanisms driving immune suppression, we can identify rational immunotherapy combinations for CRPC based on the genetic fingerprint of a patient’s tumor for precision medicine. Citation Format: Katherine C. Murphy, Kelly DeMarco, Yvette Lopez-Diaz, Lin Zhou, Marcus Ruscetti. Identifying and targeting the genetic determinants of immune suppression and immunotherapy failure in prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr A078.

Abstract A022: Alternative polyadenylation as a therapeutic vulnerability in prostate cancer

Abstract Prostate cancer is the second leading cause of male cancer death in the United States. While localized disease can be cured by radiation or surgery, metastatic prostate cancer presents a clinical challenge. Metastatic prostate cancer can initially be controlled by endocrine therapies that target the androgen receptor (AR), however, these tumors will inevitably develop resistance. This stage of the disease, termed castration-resistant prostate cancer (CRPC), is responsible for the majority of prostate cancer-specific deaths. Current treatment options for CRPC patients are not curable and highlight the need to develop novel therapeutic options that will more effectively treat the disease. Spliced mRNA molecules undergo cleavage and polyadenylation ~25 base pair downstream from a canonical 5’AAUAAA poly(A) site located within the 3’ untranslated region (UTR). Shortening of mRNA 3’UTRs through alternative polyadenylation has been found in numerous cancers leading to downregulation of tumor suppressor genes and upregulation of oncogenes. To investigate the function of alternative polyadenylation in CRPC, we conducted a loss-of-function screen of mRNA polyadenylation factors in hormone-sensitive LNCaP cells and hormone-insensitive LNCaP95 and 22Rv1 cells. We found that knockdown of the cleavage and polyadenylation specificity factor (CPSF) component, CPSF1, inhibited growth of all cell lines. To define pathways that are regulated by CPSF1 in prostate cancer, we identified CPSF1-dependent gene expression using RNA-seq and CPSF1-dependent poly(A) sites using poly(A)-ClickSeq (PAC-Seq). Gene set enrichment analysis of RNA-seq data revealed the Glycolysis Hallmark and Hypoxia Hallmark gene sets were positively regulated by CPSF1. Analysis of PAC-seq data revealed global induction of poly(A) sites distal to 3’ UTRs upon CPSF1 knockdown. By integrating these RNA-seq and PAC-seq datasets, we have identified AR-independent pathways that may be attractive for therapeutic targeting in prostate cancer. This study has revealed post-transcriptional gene regulation by CPSF1 as an important pathway used in CRPC. Citation Format: Kiel T. Tietz, Braedan M. McClukey, Sarah A. Munro, Conor R. Miller, Scott M. Dehm. Alternative polyadenylation as a therapeutic vulnerability in prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr A022.

Use of poly (adp-ribose) polymerase inhibitors (PARPi) in metastatic castration resistant prostate cancer (mCRPC) patients stratified by homologous recombination repair mutations.

e17053 Background: Approval of the PARPi in May 2020 caused a paradigm shift in the treatment of mCRPC patients with homologous recombination repair (HRR) mutations. Gaps remain in the optimal prescribing of PARPi in eligible patients, with negative consequences on clinical outcomes. The goal of this study was to look at PARPi prescribing practices in patients with HRR mutations overall and by sub-mutation. Methods: The Integra Connect PQ-deidentified real-world database was used, comprising over 1 million cancer patients across 275 sites of care, of which 80% were community practices and 20% academic settings. We performed medical chart reviews and identified 261 patients with HRRm mCRPC that received therapy between 5/1/2020 to 1/24/2023. We segmented patients into BRCA-positive with or without other HRR mutations, ATM positive without BRCA mutations and with or without other HRR mutations, and other HRR mutations without BRCA or ATM mutations. We compared utilization of PARPi in HRRm patients by sub-group. Descriptive statistics were used to assess proportions of utilization of PARPi treatment among HRRm patients. Proportions were compared using the two-proportion z-test. Results: Among HRRm mCRPC patients with prior next-generation hormonal agent (NHA) treatment, the use of PARPi is 61% (158 of 261). Among HRRm mCRPC patients who received PARPi after prior NHA treatment, 76% (78 of 103) were in the BRCA positive group, 62% (48 of 78) in the ATM positive group and 40% (32 of 80) in the other HRR mutations positive group. Patients in the BRCA positive group were more likely to receive a PARPi than those in either the ATM or other HRR mutations group (p &lt; 0.01). Patients harboring ATM mutations were more likely to receive a PARPi than patients with non-BRCA HRR mutations (p &lt; 0.01). Conclusions: This real-world study shows an opportunity to increase rates of PARPi treatment in patients with HRRm mCRPC across all HRR gene mutations. mCRPC patients with BRCA and ATM mutations were more likely to receive PARPi than those with other HRR mutations.

Prevalence of intra-patient inter-metastatic heterogeneity in mCRPC patients based on triple-tracer PET imaging: The 3TMPO study.

5033 Background: Intra-patient inter-metastatic heterogeneity (IIH) has been demonstrated in metastatic castration resistant prostate cancer (mCRPC) patients based on genomic and imaging studies, most often after several lines of therapies. IIH is of utmost importance for PSMA radioligand therapy (RLT) eligibility, biopsy-based precision medicine and/or treatment intensification decision-making. The Triple-Tracer strategy against Metastatic PrOstate cancer (3TMPO) study (NCT04000776) is a prospective multicenter PET imaging study that was designed to determine the prevalence of IIH in mCRPC patients and to determine candidacy for RLT. Methods: 3TMPO is a PET-imaging trial including mCRPC patients showing at least 3 metastases on conventional imaging with evidence of biochemical or radiographic progression, at least 3 months after initiation of the last systemic therapy. 68Ga-PSMA-617 and 18F-FDG PET/CT scans were performed within 10 days and analyzed quantitatively. A third scan with 68Ga-Octreotate was done when a PSMA-/FDG+ lesion was found. For all tracers, positivity of a lesion was defined as its SUVpeak being 1.5 times higher than the SUVmean of the liver. Lesions smaller than 1 cc and likely benign foci of uptake were excluded. IIH prevalence was the primary outcome, defined as the percentage of patients having at least two lesions with discordant features on PET imaging. Results: We included 98 patients in the final analysis. Patients had a mean age of 69 years and a median PSA of 51.1 ng/mL. Number of metastases were &lt;5 in 46.9% and ≥10 in 33.7% of patients and 10.2, 20.4, 18.4 and 51.0% had received 0, 1, 2 or &gt;2 lines of systemic therapies for mCRPC, respectively. Prevalence of IIH was 83.7% based on pre-specified PET criteria. Overall, seven different combinations of lesion phenotypes were found among patients based on FDG and PSMA PETs, and at least one PSMA-/FDG+ lesion was found in 46 patients (46.9%). Of the 44 patients who underwent Octreotate PET, six (13.6%) had at least one Octreotate-positive lesion. In this FDG/PSMA/Octreotate-imaged subgroup, 11 different combinations of phenotypes were observed between metastases of individual patients. Overall, 52.0% (IC95: 41.7-62.2) of patients were found to be candidate for PSMA RLT, but none for Octreotate RLT. Conclusions: The majority of mCRPC patients showed IIH. Based on a multi-tracer approach, up to 11 lesion phenotype combinations were found amongst patients. Correlation of these imaging phenotypes with genomics and treatment response will be highly relevant for optimized precision medicine, especially with respect to PSMA-RLT. Clinical trial information: NCT04000776 . [Table: see text]

Presence of somatic/germline homologous recombination repair (HRR) mutations and outcomes in metastatic castration-resistant prostate cancer (mCRPC) patients (pts) receiving first-line (1L) treatment stratified by BRCA status.

5003 Background: Understanding of the association between HRR mutations and outcomes in mCRPC pts is limited. This analysis investigated the prevalence and outcomes of pts with/out HRR mutations (somatic and/or germline), stratified as BRCA, non-BRCA, or HRR non-BRCA, who initiated 1L mCRPC treatment with novel hormonal therapy (NHT) or taxane. Methods: Eligible pts from PROREPAIR-B (NCT03075735), PROSENZA (NCT02922218), PROSTAC (NCT02362620), and PROSABI (NCT02787837) studies underwent paired somatic/germline DNA analyses using a custom NGS panel that included ATM, BRCA1, BRCA2, BRIP1, CDK12, CHEK2, FANCA, HDAC2, PALB2, RAD51B, and RAD54L. Those with pathogenic (likely) mutations in ≥1 allele of ≥1 gene were considered deficient (HRR). rPFS, PFS2, and OS were reported for BRCA, non-BRCA, and HRR non-BRCA subgroups; associations between mutations and outcomes were assessed using inverse probability weighted Cox models, which balanced the baseline (BL) characteristics between subgroups. Hazard ratios (HRs) with 95% confidence intervals (CIs) and p values are presented. Results: Of 729 pts, 223 (30.6%) were HRR, including 96 (13.2%) BRCA. 60.4% of pts were treated with NHT and 39.6% with taxane in 1L; 80.7% received at least second-line treatment. Median age at BL was 72.2 years, 63.5% had Gleason &gt;7, 53.1% had ECOG ≥1, and 13.3% presented with visceral metastases. BL characteristics after adjustment were similar (standard mean difference: &lt;0.1 in HRR and maximum 0.11 for all pts). BRCA pts had significantly worse rPFS, PFS2, and OS than non-BRCA pts; BRCA pts also had significantly worse PFS2 and OS than HRR non-BRCA pts. There were no significant differences between the outcomes of somatic and germline BRCA pts. Conclusions: BRCA pts, regardless of somatic/germline origin, had significantly worse rPFS, PFS2, and OS than the total non-BRCA population and significantly worse PFS2 and OS than non-BRCA pts in the HRR subgroup. It is crucial to screen early for HRR mutations, particularly in BRCA1/2, to begin timely, targeted mCRPC treatment and improve prognosis.[Table: see text]

Development and initial clinical testing of a multiplex assay evaluating histone modifier p300 acetylation in prostate cancer circulating tumor cells.

e17020 Background: Histone modifying enzymes (HMEs) hold promise as targets for drug discovery in cancer. Histone acetyltransferases and histone deacetylases dictate the presence of acetyl marks, which regulate chromatin and transcriptional activation. We previously developed a novel ‘histone array on a chip’ to measure HME dysregulation associated with the development of castration resistant prostate cancer (CRPC) (ACS Chem Biol 2017). This approach identified decreased SIRT2 expression and increased p300 activity leads to a concerted mechanism of hyperacetylation at specific histone lysine sites (H3K9, H3K14, and H3K18) in CRPC xenografts. These alterations were subsequently validated in castration-resistant tissues (BMC Cancer 2017). The goal of the current study was to determine if circulating tumor cells (CTCs) could be used as a therapeutic biomarker to identify patients with altered p300 acetylation activity. Methods: Blood was collected from 11 CRPC and 3 hormone-sensitive patients with advanced PC under an IRB-approved protocol. PBMCs were isolated with Ficoll-paque PLUS, and lymphocytes were depleted using anti-CD45 magnetic beads. CTCs were then captured with EpCAM labeled coated magnetic beads. Bound cells were isolated with Exclusion-based Sample Preparation (ESP) technology. Bound cells were moved through staining and washing wells prior to imaging at 20x on a fluorescent microscope. Images were analyzed with NIS Elements software. Results: We initially optimized the staining of acetyl-p300, total p300, H3K18ac, and SIRT2 in single cell prostate cancer cell cultures. Exposure to A-485, a selective p300/CBP catalytic inhibitor, reduced levels of p300 and H3K18 acetylation confirming assay specificity. CTCs from patient samples were then captured by EpCAM antibody binding and target staining revealed clear expression of acetyl-p300, acetyl-H3K18, SIRT2, and CK (cytokeratin). No expression of CD45+ cells (lymphocyte exclusion) is noted. Acetyl-p300 displayed a strong positive correlation with its target acetylated H3k18 (Pearson’s r = 0.61), and SIRT2 expression showed robust negative correlation with H3k18 (r = -0.60). A distinct group of 6/11 (55%) of CRPC patients demonstrated altered expression. In one subject with an untreated CSPC sample and subsequent CRPC sample ( &gt; 12mo) a loss of SIRT2 expression, increased p300 activity, and histone H3 hyperacetylation are associated with the transition from hormone-sensitive to CRPC. Conclusions: Acetyl-p300, H3K18ac, and SIRT2 increase in a subset of CTC from patients with CRPC. These biomarkers are associated with increased CBP/p300 activity. This biomarker suite could potentially be used for noninvasive monitoring of outcomes and predicting tumor responses to CBP/p300 acetylation inhibitors in clinical development.

Factors associated with thrombocytopenia in patients treated with pegfilgrastim.

Pegfilgrastim is a long-acting, granulocyte colony-stimulating factor approved in Japan for the prevention of neutropenia caused by antineoplastic agents. Severe thrombocytopenia was reported with pegfilgrastim, however, the factors associated with thrombocytopenia are unclear. This study aimed to explore the factors associated with thrombocytopenia in patients with metastatic castration-resistant prostate cancer treated with pegfilgrastim for primary prophylaxis of febrile neutropenia (FN) with cabazitaxel. This study included metastatic castration-resistant prostate cancer patients who received pegfilgrastim for primary prophylaxis of FN with cabazitaxel. The timing and severity of thrombocytopenia and factors associated with the reduction rate of platelets were examined in patients who received pegfilgrastim for the primary prevention of FN during the first course of cabazitaxel and by multiple regression analysis. Thrombocytopenia was most common within 7 days of pegfilgrastim administration, with 32 cases of grade 1 and 6 cases of grade 2 as per the Common Terminology Criteria for Adverse Events version 5.0. Multiple regression analysis revealed that the reduction rate of platelets after pegfilgrastim administration was significantly positively correlated with monocytes. In contrast, the presence of liver metastases and neutrophils was significantly negatively correlated with the reduction rate of platelets. Thrombocytopenia due to pegfilgrastim administered as primary prophylaxis for FN with cabazitaxel was most likely to occur within one week after pegfilgrastim administration, suggesting that monocytes, neutrophils, and liver metastases were associated with a reduction in platelets.

The utility of the [-2]pro-prostate-specific antigen level as a prognostic marker in patients with castration-resistant prostate cancer treated with enzalutamide.

The prostate health index (phi) derived using [-2]pro-prostate-specific antigen (p2PSA), a precursor of PSA, has been shown to predict cancer in the gray zone. However, the utility of p2PSA in predicting outcomes for castration-resistant prostate cancer (CRPC) patients remains unknown. Therefore, in this study, we aimed to evaluate the usefulness of p2PSA in predicting the efficacy and prognosis of enzalutamide treatment in CRPC patients. We conducted a prospective study of CRPC patients treated with enzalutamide at our institution, measuring p2PSA levels in 98 pre-treatment serum samples. All patients were divided into two groups based on the median values of each parameter. The PSA progression-free survival (PSA-PFS) and overall survival (OS) were compared using the Kaplan-Meier method. This study was approved by the Institutional Review Board of Gunma University Hospital (IRB No. 2021-092, 1983). The median PSA level before enzalutamide treatment was 25.59 ng/mL, the median p2PSA level was 208.75 pg/mL, and the median phi was 187.95. PSA, p2PSA, and phi were not all predictors of PSA-PFS. However, the OS was significantly better in the low-value groups (log-rank p-values of PSA, p2PSA, and phi were 0.024, 0.034, and 0.018, respectively). In the docetaxel (DOC)-naive group (n = 58), PSA was not a predictor of OS, but p2PSA and phi were significantly associated with better OS in the low group. This relationship was not observed in the DOC-treated group. Our study elucidates the usefulness of p2PSA in predicting outcomes for CRPC patients treated with enzalutamide. It suggests that p2PSA and phi may be prognostic markers after enzalutamide administration in CRPC patients.