Castration-resistant Neuroendocrine Prostate Cancer Research Articles

Abstract 6913: A preclinical platform of prostate cancer PDX and derived cellular models as a tool for pharmacological screening and functional studies

Abstract Prostate Cancer (PCa) is the most frequently diagnosed cancer in men and remains one of the leading causes of cancer death worldwide. Despite undeniable progress in understanding the biology and genetics of this highly heterogenous and complex disease, the development of effective therapies is hampered by the lack of relevant experimental models recapitulating its diversity. To tackle this issue, over the last years, we have generated and characterized at the molecular and pharmacology level a collection of prostate cancer patient-derived xenografts (PDX) that accurately reproduce PCa histological and molecular heterogeneity. Although the development of these PCa PDX has been challenging, we are now able to offer a preclinical platform of 11 fully characterized PCa PDX models and in vitro cellular counterparts for preclinical evaluation of novel treatment modalities. Our platform consists of a collection of 4 androgen receptor high (ARPC), 2 androgen receptor low (ARLPC), 1 amphicrine prostate carcinoma (AMPC) and 4 castration-resistant neuroendocrine prostate cancer (CRPC-NE) PDX models and 6 cellular models derived from these PDX (PDXDC). Most of these PDX models were obtained by transplantation of post-surgery tumor specimens either by grafting of tumor fragments in the interscapular region of nude mice or in the subrenal capsule of NOD scid gamma mice. One model is a CTC-derived eXplant established using circulating tumor cells (CTCs) obtained by diagnostic leukapheresis injected into the fat pad of NOD scid gamma mice. PDXDC were obtained from dissociated PDX tumors cultured under various media and matrix conditions. They were characterized by comparison with the parental PDX by Short Tandem Repeat (STR) profiling before performing a master bank. All models (PDX and PDXDC) were characterized at molecular level by whole exome and RNA sequencing. PDX were characterized pharmacologically for response to standards of care, physical castration and response to the androgen receptor inhibitor enzalutamide. In vitro drug sensitivity of PDXDC was compared with their parental PDX in vivo drug response. This unique PDX panel reflects the molecular heterogeneity of prostate cancer and reproduces the molecular and drug response profile of human tumors. Associated to in vitro cell derivatives, our collection provides a powerful preclinical platform for translational research to identify mechanisms underlying acquired resistance to current therapies and develop novel treatment strategies against PCa and CRPC. Citation Format: Emilie Indersie, Léa Sinayen, Ludovic Bigot, Katell Mevel, Aurore Gorse, Marie Tavernier, Enora Le Ven, Luc Friboulet, Françoise Farace, Ester Morgado, Laurent Pouyet, Yohann Loriot, François Romagné, Olivier Déas. A preclinical platform of prostate cancer PDX and derived cellular models as a tool for pharmacological screening and functional studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6913.

RNA Splicing Factors SRRM3 and SRRM4 Distinguish Molecular Phenotypes of Castration-Resistant Neuroendocrine Prostate Cancer.

Neuroendocrine (NE) differentiation in metastatic castration-resistant prostate cancer (mCRPC) is an increasingly common clinical feature arising from cellular plasticity. We recently characterized two mCRPC phenotypes with NE features: androgen receptor (AR)-positive NE-positive amphicrine prostate cancer (AMPC) and AR-negative small cell or neuroendocrine prostate cancer (SCNPC). Here, we interrogated the regulation of RE1-silencing transcription factor (REST), a transcriptional repressor of neuronal genes, and elucidated molecular programs driving AMPC and SCNPC biology. Analysis of prostate cancer cell lines, mCRPC specimens, and LuCaP patient-derived xenograft models detected alternative splicing of REST to REST4 and attenuated REST repressor activity in AMPC and SCNPC. The REST locus was also hypermethylated and REST expression was reduced in SCNPC. While serine/arginine repetitive matrix protein 4 (SRRM4) was previously implicated in alternative splicing of REST in mCRPC, we detected SRRM3 expression in REST4-positive, SRRM4-negative AMPC, and SCNPC. In CRPC cell lines, SRRM3 induced alternative splicing of REST to REST4 and exacerbated the expression of REST-repressed genes. Furthermore, SRRM3 and SRRM4 expression defined molecular subsets of AMPC and SCNPC across species and tumor types. Two AMPC phenotypes and three SCNPC phenotypes were characterized, denoted either by REST attenuation and ASCL1 activity or by progressive activation of neuronal transcription factor programs, respectively. These results nominate SRRM3 as the principal REST splicing factor expressed in early NE differentiation and provide a framework to molecularly classify diverse NE phenotypes in mCRPC. SIGNIFICANCE: This study identifies SRRM3 as a key inducer of cellular plasticity in prostate cancer with neuroendocrine features and delineates distinct neuroendocrine phenotypes to inform therapeutic development and precision medicine applications.

Circulating tumor DNA (ctDNA) to detect neuroendocrine prostate cancer genomic and DNA methylation changes.

8 Background: Loss of androgen receptor signaling dependence occurs in approximately 20% of treatment resistant prostate cancers, and this may manifest as transformation to castration resistant neuroendocrine prostate cancer (CRPC-NE). The diagnosis of CRPC-NE relies on a metastatic tumor biopsy, which is invasive for patients. Here we study ctDNA as an approach to identify CRPC-NE-associated genomic and epigenomic changes. We also use this as a tool to understand tumor heterogeneity and clonal dynamics that occurs during CRPC-NE progression. Methods: 64 patients with metastatic prostate cancer (10 hormone naive, 35 castration resistant adenocarcinoma (CRPC-Adeno), 17 CRPC-NE) were prospectively enrolled for matched metastatic biopsy and blood collection. Twenty-four had multiple time-points, for total of 69 plasma and 98 metastatic tissues. Whole exome sequencing (WES) and whole genome bisulfite sequencing(of subset) of ctDNA,germline DNA, and metastatic biopsies were performed. Results: Overall there was high concordance of alterations between ctDNA and patient-matched metastasis, though this was highest in CRPC-NE. There was also less heterogeneity across patients with CRPC-NE. Alteration frequencies were consistent with published tissue-based studies with AR alterations enriched in CRPC-Adeno and TP53and RB1loss enriched in CRPC-NE ctDNA. The prognostic significance of these alterations differed based on histologic subtype. Allele-specific copy number analysis and serial sampling allowed for the detection and tracking of clonal and subclonal tumor cell populations. cfDNA methylation was reflective of methylation patterns in biopsies and detected CRPC-NE-associated epigenetic changes.A targeted set combining genomic ( TP53, RB1, CYLD, AR) and epigenomic(20 differential DNA methylation sites) was capable of identifying patients with CRPC-NE. Conclusions: WES and whole genome methylation of ctDNA is feasible, concordant with biopsy tissues, and identifies the spectrum and frequency of CRPC-NE genomic and epigenomic changes. A targeted panel extended and validated in larger cohorts could potentially improve the detection of CRPC-NE using non-invasive ctDNA profiling.

Abstract CN06-01: Lineage plasticity and the neuroendocrine phenotype as a resistance mechanism in prostate cancer

Abstract Loss of androgen receptor (AR) signaling dependence occurs in approximately 15-20% of treatment resistant prostate cancers, and this may manifest clinically as transformation from a prostate adenocarcinoma histology to a castration resistant neuroendocrine prostate cancer (CRPC-NE). The diagnosis of CRPC-NE currently relies on a metastatic tumor biopsy, which is invasive for patients and often challenging to diagnose due to tumor heterogeneity. By studying whole exome sequencing and whole genome bisulfite sequencing of metastatic tumor biopsies and matched cell free DNA (cfDNA), we identified distinct genomic and epigenomic features of CRPC-NE and patterns of tumor evolution that occur during clinical progression and treatment resistance. Loss of RB1 and TP53 are enriched in CRPC-NE compared with castration resistant prostate adenocarcinoma, as are significant changes in DNA methylation, which are detectable by cfDNA. There was significantly higher concordance between cfDNA and biopsy tissue genomic alterations in CRPC-NE patients compared to castration resistant adenocarcinoma, supporting greater intra-individual genomic consistency across metastases. cfDNA and serial tumor biopsies allowed for the tracking of dynamic clonal and subclonal tumor cell populations as patients progressed and identified CRPC-NE alterations sometimes prior to the development of clinical features. CRPC-NE appears to arise clonally from a prostate adenocarcinoma precursor likely through a dynamic process of clonal selection and trans-differentiation that occurs during resistance to AR-directed therapies. In addition to loss of AR expression and canonical AR signaling, we identified a dysregulation of key pathways in CRPC-NE including loss of Notch signaling, reactivation of developmental programs, and gain of neuronal/neuroendocrine programs including critical lineage determining transcription factors (LDTFs) such as ASCL1, MYCN, BRN2, pointing to novel biomarkers and potential targets for CRPC-NE. Patient-derived organoids and xenografts were used to interrogate the timing by which genomic and epigenomic changes and LDTFs contribute to lineage plasticity and the development of CRPC-NE as a resistance mechanism in prostate cancer. Citation Format: Himisha Beltran, Sheng-Yu Ku, Vincenza Conteduca, Alessandro Romanel, Loredana Puca, Michael Sigouros, Juan Miguel Mosquera, Scott T. Tagawa, Andrea Sboner, Olivier Elemento, David Goodrich, David Rickman, Amina Zoubeidi, Francesca Demichelis. Lineage plasticity and the neuroendocrine phenotype as a resistance mechanism in prostate cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr CN06-01. doi:10.1158/1535-7163.TARG-19-CN06-01

Abstract IA05: Emerging therapeutic strategies for androgen indifferent prostate cancer

Abstract The development of castration resistance in prostate cancer most commonly involves reactivation of androgen receptor (AR) signaling; this knowledge has informed the clinical development of potent AR-targeted agents now commonly used in the clinic. However, a subset of prostate cancers may evolve under therapeutic pressure of the AR towards non-AR driven, or “androgen indifferent,” disease. This has been associated with aggressive clinical features with repeat biopsies enriched with TP53 and RB1 genomic loss, overexpression of MYCN, epigenetic changes, and activation of alternative lineage programs such as neuronal and neuroendocrine pathways. One extreme manifestation is complete transformation to a small cell/neuroendocrine carcinoma with loss of AR expression. Although castration-resistant neuroendocrine prostate cancer evolves clonally from prostate adenocarcinoma likely through a trans-differentiation process, it acquires molecular alterations seen in other poorly differentiated neuroendocrine cancers. This talk will focus on emerging biomarker-driven therapeutic strategies for androgen-indifferent disease, including updates on targeting the N-myc/Aurora kinase A complex, the potential role for epigenetic therapies, and new data to inform ADC and immunotherapy development for this subset. Citation Format: Himisha Beltran. Emerging therapeutic strategies for androgen indifferent prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr IA05.

Abstract 1947: Rovalpituzumab tesirine as a therapeutic agent for neuroendocrine prostate cancer

Abstract Background: Delta-like protein 3 (DLL3) is expressed on the surface of small cell lung cancer (SCLC) tumor-initiating cells, and the DLL3 targeted antibody-drug conjugate, Rovalpituzumab tesirine (Rova-T™; SC16LD6.5), has shown promise for patients with SCLC. Neuroendocrine prostate cancer (NEPC) is an emerging late stage subtype of castration resistant prostate cancer with limited therapeutic options. Based on clinical and molecular similarities with SCLC, we investigated expression of DLL3 and the use of Rovalpituzumab tesirine in NEPC xenografts. Methods: We evaluated mRNA and/or protein expression of DLL3 in a cohort of 361 patients (552 samples) ranging from benign prostate (BEN), localized prostate adenocarcinoma (PCA), castration resistant adenocarcinoma (CRPC), and castration resistant NEPC and correlated with pathologic and genomic features. mRNA was assessed by RNAseq. Protein was assessed by immunohistochemistry (DLL3 SP347 antibody). Prostate cancer cell lines were engrafted in mice and treated with SC16LD6.5 in vivo. Results: DLL3 was expressed at the mRNA and/or protein level in 0/132 Benign (0%), 1/254 (0.4%) PCA, 10/90 (11.1%) CRPC and 47/76 (61.8%) NEPC samples. DLL3 IHC was of higher intensity in NEPC and co-localized with classical neuroendocrine (NE) markers. DLL3 was amongst the most differentially expressed genes by RNA-seq in NEPC versus CRPC (p=<0.0001, fold change=71), and positively correlated with ASCL1 expression (r= 0.88) and RB1 genomic loss (83%), and inversely with AR expression. siRNA knockdown of DLL3 did not alter AR signaling or NE score. In vivo treatment of the NEPC, NCI-H660, xenografts with vehicle, IgG1LD6.5, or SC16LD6.5 (0.3mg/Kg, intraperitoneal single dose) showed complete responses to SC16LD6.5 with no recurring tumors after 100 days; while the tumor developed from the CRPC line, DU145, which do not express DLL3, continued growing despite the treatment and were sacrified when tumors reached the maximum size allowed by our IACUC protocol. Conclusions: DLL3 is expressed on the surface of the majority of NEPC cases evaluated and is not expressed in primary prostate cancer or benign tissues. Modulation of DLL3 expression does not appear to affect AR or NEPC signaling in cell lines. SC16LD6.5 (Rova-T) demonstrates preferential preclinical activity in NEPC that express DLL3 compared to CRPC-adenocarcinoma that are DLL3 negative, in vivo. A Phase 1 Basket trial investigating Rova-T is now open with a dedicated NEPC arm (NCT02709889). Citation Format: Loredana Puca, Verena Sailor, Katie Gavyert, Etienne Dardenne, Kumiko Isse, Michael Sigouros, David M. Nanus, Scott T. Tagawa, Juan Miguel Mosquera, Laura Saunders, Himisha Beltran. Rovalpituzumab tesirine as a therapeutic agent for neuroendocrine prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1947.

Abstract 3357: Genetic regulation of RUNX2 induce apoptotic cell death through regulating the expression of SOX2 in CRPC-NE cells

Abstract Although runt-related transcription factor 2 (RUNX2) is well known for its bone-specific transcriptional regulator in highly metastatic prostate cancer, there is little knowledge about its role of RUNX2 in castration-resistant neuroendocrine prostate cancer (it is named CRPC-NE). It is well known that expression of Androgen Receptor (AR) in CRPC-NE cases is lower than other types and AR suppresses RUNX2 expression by binding it and abrogating its recruitment to DNA. This fact suggests that in CRPC-NE, low expression of AR, the addiction of RUNX2 is relatively higher than that of other prostate cancer types. Extensive analysis of the clinical datasets revealed that RUNX2 is one of the most consistently up-regulated genes in CRPC-NE and the overexpression of SOX2 confers an accelerated disease progression and shortened overall survival periods to CRPC-NE patients. We also found that the silencing RUNX2 induced apoptotic cell death in CRPC-NE cells independent on p53, but through transcriptionally down-regulating SOX2 expressions. Mechanistically, ChIP-qPCR assay and luciferase reporter experiments confirmed that RUNX2 positively regulates SOX2 expression by binding to the consensus RUNX2 binding sequence (5'-TGTGGT-3') located in the proximal promoter region of SOX2. Short hairpin RNA (shRNA)-mediated knockdown of RUNX2 in the CRPC-NE cell line PC-3 induced the decreased expression of SOX2 and the increased expression of cleaved form of caspase-3 and PARP in a p53 independent manner. Silencing of SOX2 in PC-3 cells suppressed the proliferation of these cancer cells and induced apoptosis, which phenotypes were rescued by restoring SOX2 in the cells. These data indicates that inhibition of SOX2-apoptosis axis via RUNX2 could be a better therapeutic choice in CRPC-NE. Lastly, we examined the efficacy of our novel molecule Chlorambucil-conjugated Pyrrole Imidazole Polyamide (we named it as Chb-M'), which specifically binds to the consensus RUNX2 binding sequence and inhibits RUNX2 target gene. Surprisingly, Chb-M' had tremendous inhibitory effect on PC-3 cells (IC50 value at 620 nM) through effective SOX2 inhibition. Additionally, it was exceptionally well-tolerated in mice and exerted excellent efficacy against xenograft mice models of CRPC-NE. Taken together, our work identified a novel interaction of RUNX2 and SOX2-apoptosis axis, offering a new strategy for the management of poor-prognostic advanced stage CRPC-NE cancer patients. Citation Format: Yuki Noguchi, Natsuki Wariishi, Shiina Iwai, Gengo Kashiwazaki, Junichi Taniguchi, Toshikazu Bando, Masaya Baba, Souichi Adachi, Hiroshi Sugiyama, Yasuhiko Kamikubo. Genetic regulation of RUNX2 induce apoptotic cell death through regulating the expression of SOX2 in CRPC-NE cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3357.

Neuroendocrine Differentiation in Prostate Cancer: Emerging Biology, Models, and Therapies.

Although a de novo clinical presentation of small cell neuroendocrine carcinoma of the prostate is rare, a subset of patients previously diagnosed with prostate adenocarcinoma may develop neuroendocrine features in later stages of castration-resistant prostate cancer (CRPC) progression as a result of treatment resistance. Despite sharing clinical, histologic, and some molecular features with other neuroendocrine carcinomas, including small cell lung cancer, castration-resistant neuroendocrine prostate cancer (CRPC-NE) is clonally derived from prostate adenocarcinoma. CRPC-NE therefore retains early prostate cancer genomic alterations and acquires new molecular changes making them resistant to traditional CRPC therapies. This review focuses on recent advances in our understanding of CRPC-NE biology, the transdifferentiation/plasticity process, and development and characterization of relevant CRPC-NE preclinical models.

Development of Neuroendocrine Prostate Cancers by the Ser/Arg Repetitive Matrix 4-Mediated RNA Splicing Network.

While the use of next-generation androgen receptor pathway inhibition (ARPI) therapy has significantly increased the survival of patients with metastatic prostate adenocarcinoma (AdPC), several groups have reported a treatment-resistant mechanism, whereby cancer cells can become androgen receptor (AR) indifferent and gain a neuroendocrine (NE)-like phenotype. This subtype of castration-resistant prostate cancer has been termed “treatment-induced castration-resistant neuroendocrine prostate cancer” (CRPC-NE). Recent reports indicate that the overall genomic landscapes of castration-resistant tumors with AdPC phenotypes and CRPC-NE are not significantly altered. However, CRPC-NE tumors have been found to contain a NE-specific pattern throughout their epigenome and splicing transcriptome, which are significantly modified. The molecular mechanisms by which CRPC-NE develops remain unclear, but several factors have been implicated in the progression of the disease. Recently, Ser/Arg repetitive matrix 4 (SRRM4), a neuronal-specific RNA splicing factor that is upregulated in CRPC-NE tumors, has been shown to establish a CRPC-NE-unique splicing transcriptome, to induce a NE-like morphology in AdPC cells, and, most importantly, to transform AdPC cells into CRPC-NE xenografts under ARPI. Moreover, the SRRM4-targeted splicing genes are highly enriched in various neuronal processes, suggesting their roles in facilitating a CRPC-NE program. This article will address the importance of SRRM4-mediated alternative RNA splicing in reprogramming translated proteins to facilitate NE differentiation, survival, and proliferation of cells to establish CRPC-NE tumors. In addition, we will discuss the potential roles of SRRM4 in conjunction with other known pathways and factors important for CRPC-NE development, such as the AR pathway, TP53 and RB1 genes, the FOXA family of proteins, and environmental factors. This study aims to explore the multifaceted functions of SRRM4 and SRRM4-mediated splicing in driving a CRPC-NE program as a coping mechanism for therapy resistance, as well as define future SRRM4-targeted therapeutic approaches for treating CRPC-NE or mitigating its development.

Transcriptional Regulation in Prostate Cancer.

Prostate cancer development involves corruption of the normal prostate transcriptional network, following deregulated expression or mutation of key transcription factors. Here, we provide an overview of the transcription factors that are important in normal prostate homeostasis (NKX3-1, p63, androgen receptor [AR]), primary prostate cancer (ETS family members, c-MYC), castration-resistant prostate cancer (AR, FOXA1), and AR-independent castration-resistant neuroendocrine prostate cancer (RB1, p53, N-MYC). We use functional (in vitro and in vivo) as well as clinical data to discuss evidence that unveils their roles in the initiation and progression of prostate cancer, with an emphasis on results of chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq).

Sensitization of neuroendocrine prostate cancer by RRx-001.

280 Background: RRx-001 is a first-in-class dinitroazetidine nontoxic anticancer agent. Its actions include upregulation of oxidative stress, depletion of GSH and NADPH, anti-angiogenesis, epigenetic modulation and induction of viral mimicry. In a phase II clinical study (NCT02489903), RRx-001 reversed resistance to reintroduced platinum doublets in small cell lung cancer (SCLC) and other tumor types. RRx001 demonstrated clinical activity with a complete metabolic response of metastatic castration-resistant neuroendocrine prostate cancer (NEPC) after treatment with RRx-001 and reintroduced platinum doublets. Here, we used a preclinical model to further investigate RRx-001 sensitization of NEPC to platinum therapy. Methods: NCI-H660 spheroids were pre-treated with 1 mM RRx-001 or vehicle prior to treatment with carboplatin or docetaxel and cell proliferation was assessed. Cells were also pre-treated with both 1 mM RRx-001 and 10 mM N-acetyl cysteine (NAC) or 1 mM Buthionine-S,R-sulfoximine (BSO) to investigate the role of reactive oxygen species (ROS) in RRx-001 activity. NCI-H660 xenografts were pre-treated with RRx-001 (mimicking clinical activity) or vehicle prior to treatment with carboplatin, and tumor volume and body weight were monitored. Results: RRx-001 pre-treatment significantly decreased the IC50 of carboplatin from ~86 mM to 36 mM in vitro, whereas no significant effect was observed on the IC50 of docetaxel. The ROS inhibitor NAC reversed > 70% of RRx-001 sensitization to carboplatin, while co-pre-treatment with the ROS generator BSO significantly enhanced RRx-001 activity. In vivo, pre-treatment with RRx-001 prior to carboplatin significantly decreased tumor volume compared to vehicle, with ~50% greater reduction in mean tumor volume at 30 days. Body weight was not affected by RRx-001. Conclusions: This preclinical study supports the concept that “priming” with RRx-001 may be an effective therapeutic strategy to resensitize NEPC to initially highly effective platinum doublets. A phase 3 trial is planned, and studies with additional preclinical NEPC models will identify the mechanism of action of RRx-001.

Abstract LB-122: Tumor heterogeneity in castration resistant neuroendocrine prostate cancer from whole exome sequencing of circulating tumor DNA

Abstract Background: We recently identified mechanisms underlying the clonal evolution of castration resistant prostate adenocarcinoma (CRPC-Adeno) to a neuroendocrine resistance phenotype (Beltran et al, Nature Med 2016). In the current study, we aimed to develop a non-invasive approach to identify patients that are developing neuroendocrine prostate cancer (NEPC). Methods: We performed whole exome sequencing (WES) of matched circulating tumor DNA (ctDNA), germline DNA, and metastatic biopsies from patients with CRPC-Adeno and NEPC using minimum 50ng DNA, NimbleGen library prep, Illumina platform (ctDNA >300X). After applying ad hoc partial duplication filtering, we used FACETS segmentation followed by extended CLONET to calculate the fraction of tumor DNA in the circulation and assess the clonality of genomic lesions. Results: 64 CRPC patients were prospectively enrolled. The spectrum of alterations captured by WES ctDNA was consistent with those commonly observed in CRPC validating the feasibility of the approach. The SCNA similarity between tumor tissue and ctDNA was higher in NEPC compared to CRPC-Adeno (p=0.0001) suggesting less heterogeneity in NEPC. There was enrichment of RB1 and TP53 loss in NEPC ctDNA and AR gains in CRPC-Adeno. The overall fraction of mutations shared by ctDNA and tumoral tissue was ~80%. We compared three different tumor biopsy time-points of patient PM161—CRPC-Adeno (lymph node), CRPC-Adeno (bone), and NEPC (liver). Unexpectedly the baseline ctDNA profile (at time of CRPC-Adeno) displayed genomic features most similar to the NEPC liver biopsy. These data suggest that NEPC alterations are detectable in the circulation, potentially prior to the development of NEPC clinical features. We compared the ctDNA of another patient PM0 with his 6 sites of NEPC metastases obtained 6 days later at autopsy; the relative contribution of tumor alterations in ctDNA was highest for the liver metastasis (similarity 0.59) compared with other sites suggesting differential contribution of sites of metastases in the circulation, with implications for the interpretation of single site clinical biopsies. Conclusions: This is the first study to show that WES of ctDNA is feasible in CRPC and can help elucidate intra-patient heterogeneity and identify the spectrum and frequency of NEPC-specific genomic changes. We have incorporated our WES ctDNA discoveries into a targeted panel for larger scale validation using prospective cohorts. Our goal is to improve the earlier detection of patients transforming towards NEPC. Other data to be presented are targeted methylation of ctDNA and integrative molecular analysis of rapid autopsies (n=5) to better understand the role of intra-patient heterogeneity in NEPC. Citation Format: Himisha Beltran, Alssandro Romanel, Nicola Casiraghi, Michael Sigouros, Matteo Benelli, Jenny Xiang, Francesca Demichelis. Tumor heterogeneity in castration resistant neuroendocrine prostate cancer from whole exome sequencing of circulating tumor DNA [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 LB-122. doi:10.1158/1538-7445.AM2017-LB-122

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Genomic Aberrations Drive Clonal Evolution of Neuroendocrine Tumors

Molecular features of castration-resistant neuroendocrine prostate cancer (CRPC-NE) are not well characterized. A recent study that investigated genomic aberrations of CRPC-NE tumors suggests their clonal evolution from CRPC adenocarcinoma. Furthermore, the existence of a distinct DNA methylation profile in CRPC-NE implicates a critical role for epigenetic modification in the development of CRPC-NE.

Divergent clonal evolution of castration-resistant neuroendocrine prostate cancer.

An increasingly recognized resistance mechanism to androgen receptor (AR)-directed therapy in prostate cancer involves epithelial plasticity, wherein tumor cells demonstrate low to absent AR expression and often neuroendocrine features. The etiology and molecular basis for these “alternative” treatment-resistant cell states remain incompletely understood. Here, by analyzing whole exome sequencing data of metastatic biopsies from patients, we observed significant genomic overlap between castration resistant adenocarcinoma (CRPC-Adeno) and neuroendocrine histologies (CRPC-NE); analysis of serial progression samples points to a model most consistent with divergent clonal evolution. Genome-wide DNA methylation revealed marked epigenetic differences between CRPC-NE and CRPC-Adeno that also designated cases of CRPC-Adeno with clinical features of AR-independence as CRPC-NE, suggesting that epigenetic modifiers may play a role in the induction and/or maintenance of this treatment-resistant state. This study supports the emergence of an alternative, “AR-indifferent” cell state through divergent clonal evolution as a mechanism of treatment resistance in advanced prostate cancer.