Background: The androgen receptor axis inhibitors (ARi) (e.g, enzalutamide, abiraterone acetate) are administered in daily practice for men with metastatic castration-resistant prostate cancer (mCRPC). However, not all patients respond, and mechanisms of both primary and acquired resistance remain largely unknown. Methods: In a prospective trial MATCH-R (NCT02517892), 55 mCRPC patients underwent whole exome sequencing (WES) (n=45) and RNA-sequencing (RNA-seq) (n=52) of metastatic biopsies before starting ARi. Also, 16 mCRPC patients underwent biopsy at time of resistance (WES=14, RNA-seq = 14). The objectives were to identify genomic alterations associated with resistance to ARi as well as to describe clonal evolution. Primary resistance was determined at 4 months of treatment using composite criteria for progression that included serum prostate specific antigen measurements, bone scan, CT imaging and symptom assessments. Acquired resistance was defined by occurrence of progressive disease after initial response or stable disease. Associations of genomic and transcriptomic alterations with primary resistance were determined using Wilcoxon and Fisher's exact tests. Results: At 4 months, 22/55 patients in the cohort had disease progression (primary resistance). No genomic alterations from WES analysis were significantly associated with primary resistance. Analysis of sequential biopsies suggests that mCRPC follows mainly a parallel evolution model and involve DNA-repair related mutational processes. At time of acquired resistance to ARi, most tumors acquired new drivers affecting AR pathway (e.g, AR, NCOR1/2) or lineage switching (e.g, RB1, PTEN, TP53). Using computational methods, we measured AR transcriptional function and performed gene set enrichment analysis to identify pathways whose activity state correlated with resistance. AR gene alterations and AR expression were similar between responding and non-responding patients. Transcriptional analysis demonstrated that multiple specific gene sets — including those linked to low AR transcriptional activity, stemness program, RB loss and homologous repair deficiency — were activated in both primary and acquired resistance. Conclusion: Resistance to AR axis inhibitors results from multiple transcriptional programs already activated in pre-treatment samples. Clonal evolution analysis along with RNA-seq data indicate the role of genomic instability and lineage switching in driving acquired resistance Citation Format: Naoual Menssouri, Loic Poiraudeau, Carole Helissey, Ludovic Bigot, Jonathan Sabio, Tony Ibrahim, Claudio Nicotra, Maud Ngocamus, Lambros Tselikas, Thierry De Baere, Etienne Rouleau, Ludovic Lacroix, Anne Chaucherau, Luc Friboulet, Ronan Flippot, Giulia Baciarello, Laurence Albiges, Emeline Colomba, Pernelle Lavaud, Stefan Michiels, Aline Maillard, Antoine Italiano, Fabrice Barlesi, Jean-Charles Soria, Jean-Yves Scoazec, christophe Massard, Benjamin Besse, Fabrice André, Karim Fizazi, Daniel Gautheret, Yohann Loriot. A prospective study of prostate cancer metastases identifies an androgen receptor activity-low, stemness program associated with resistance to androgen receptor axis inhibitors and unveils mechanisms of clonal evolution [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 358.

Purpose: The androgen receptor axis inhibitors (ARPI) (e.g, enzalutamide, abiraterone acetate) are administered in daily practice for men with metastatic castration-resistant prostate cancer (mCRPC). However, not all patients respond, and mechanisms of both primary and acquired resistance remain largely unknown Experimental Design: In a prospective trial MATCH-R (NCT02517892), 59 mCRPC patients underwent whole exome sequencing (WES) and/or RNA-sequencing (RNA-seq) of samples collected before starting ARPI. Also, 18 mCRPC patients underwent biopsy at time of resistance. The objectives were to identify genomic alterations associated with resistance to ARPIs as well as to describe clonal evolution. Associations of genomic and transcriptomic alterations with primary resistance were determined using Wilcoxon and Fisher's exact tests. Results: WES analysis indicated that no single-gene genomic alterations was strongly associated with primary resistance. RNA-seq analysis showed that AR gene alterations and expression levels were similar between responders and non-responders. RNA-based pathway analysis found that patients with primary resistance had a higher Hedgehog pathway score, a lower AR pathway score and a lower NOTCH pathway score than patients with a response. Subclonal evolution and acquisition of new alterations in AR-related genes or neuroendocrine differentiation are associated with acquired resistance. ARPIs do not induce significant changes in the tumor transcriptome of most patients; however, programs associated with cell proliferation are enriched in resistant samples. Conclusions:Low AR activity, activation of stemness programs, and Hedgehog pathway were associated with primary ARPIs’ resistance, whereas most acquired resistance was associated with subclonal evolution, AR-related events, and neuroendocrine differentiation

Background: Recent advances in the biology of prostate cancer have identified aggressive variants of metastatic castration-resistant prostate cancer [mCRPC] (e.g, CRPC with neuroendocrine (NE) features, or microsatellite instability [MSI], or BRCA2 mutations). However, few preclinical models have been successfully established to study these aggressive mCRPC variants owing to their historically low establishment success and to the difficulty in accessing such clinical samples Methods: Fresh tumor biopsy specimens were obtained prospectively from patients with mCRPC through a prospective single-institution clinical trial (MATCH-R, NCT02517892). Patient-derived xenografts (PDX) in NOD Scid Gamma mice were developed and characterized as well as PDX-derived organoids derived from the same PDX (PDXO). Whole-exome sequencing, RNA sequencing and immunohistochemistry were performed on human samples and their corresponding PDX and PDXO. The primary aim was to successfully derive PDX and PDXO models reproducing the clinical features of mCRPC aggressive variants. Results: As of November 2019, 83 tumor biopsies were obtained from 61 mCRPC patients, 16 biopsies from 13 patients were successfully engrafted with an overall success rate of 26% (16/61). In addition, 16 PDXO were developed with a success rate of 100%. Overall, we developed 4 PDX and 4 PDXO from 2 patients with germline BRCA2 mutation, 2 PDX and 2 PDXO from 1 patient with MSI-high CPRC, 10 PDX and 6 PDXO from 10 patients with NE mCPRC. Molecular profiling revealed a high concordance between PDX, PDXO and human tumor samples for histological phenotype, driver mutations and transcriptomic phenotypes. The two models harbouring BRCA2 mutation were highly sensitive to both carboplatin and olaparib (in PDX and PDXO) reflecting the clinical scenario observed in the patients. BRCA2-mutated PDX models were treated in vivo to derive olaparib-resistant mCRPC models. Genetic analysis identified secondary mutations restoring the reading-frame of the gene which reversed the sensitivity of the PDX to carboplatin and olaparib. Preclinical models derived from mCRPC with NE features (including the case of small cell carcinoma) globally reproduced the clinical patterns seen in human samples. Results from high-throughput organoid drug screening suggest good concordance with patients clinical response Conclusion: The study demonstrated the feasibility of establishing preclinical models (PDX and PDXO) derived from aggressive mCRPC variants. Overall, our models reproduce the phenotypic, molecular and pharmacological characteristics of their initial human samples and may represent a unique preclinical platform modeling BRCA2 mutated, MSI and neuroendocrine prostate cancers. Citation Format: Ludovic Bigot, Jonathan Sabio, Tony Ibrahim, Naoual Menssouri, Loic poiraudeau, Carole Helissey, Jean-Yves Scoazec, Zahira Merabet, Thierry De Baere, Frederic Deschamps, Maud Ngocamus, Claudio Nicotra, Etienne Rouleau, Ludovic Lacroix, Olivier Deas, Luc Friboulet, Gilles Vassal, Eric Solary, Jean-Charles Soria, Karim Fizazi, Fabrice André, Christophe Massard, Benjamin Besse, Yohann Loriot. Novel preclinical models of aggressive variants of castration-resistant prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1114.