Joanna Cyrta

Cancer cell–autonomous contribution of type I interferon signaling to the efficacy of chemotherapy

Some of the anti-neoplastic effects of anthracyclines in mice originate from the induction of innate and T cell–mediated anticancer immune responses. Here we demonstrate that anthracyclines stimulate the rapid production of type I interferons (IFNs) by malignant cells after activation of the endosomal pattern recognition receptor Toll-like receptor 3 (TLR3). By binding to IFN-α and IFN-β receptors (IFNARs) on neoplastic cells, type I IFNs trigger autocrine and paracrine circuitries that result in the release of chemokine (C-X-C motif) ligand 10 (CXCL10). Tumors lacking Tlr3 or Ifnar failed to respond to chemotherapy unless type I IFN or Cxcl10, respectively, was artificially supplied. Moreover, a type I IFN–related signature predicted clinical responses to anthracycline-based chemotherapy in several independent cohorts of patients with breast carcinoma characterized by poor prognosis. Our data suggest that anthracycline-mediated immune responses mimic those induced by viral pathogens. We surmise that such viral mimicry constitutes a hallmark of successful chemotherapy.

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, in which tumor cells demonstrate low to absent AR expression and often have neuroendocrine features. The etiology and molecular basis for this alternative treatment-resistant cell state remain incompletely understood. Here, by analyzing whole-exome sequencing data of metastatic biopsies from patients, we observed substantial genomic overlap between castration-resistant tumors that were histologically characterized as prostate adenocarcinomas (CRPC-Adeno) and neuroendocrine prostate cancer (CRPC-NE); analysis of biopsy samples from the same individuals over time points to a model most consistent with divergent clonal evolution. Genome-wide DNA methylation analysis revealed marked epigenetic differences between CRPC-NE tumors and CRPC-Adeno, and also designated samples 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.

N-Myc Induces an EZH2-Mediated Transcriptional Program Driving Neuroendocrine Prostate Cancer

The transition from castration-resistant prostate adenocarcinoma (CRPC) to neuroendocrine prostate cancerxa0(NEPC) has emerged as an important mechanism of treatment resistance. NEPC is associated with overexpression and gene amplification of MYCN (encoding N-Myc). N-Myc is an established oncogene in several rare pediatric tumors, but its role in prostate cancer progression is not well established. Integrating a genetically engineered mouse model and human prostate cancer transcriptome data, we show that N-Myc overexpression leads to the development of poorly differentiated, invasive prostate cancer that is molecularly similar to human NEPC. This includes an abrogation of androgen receptor signaling and induction of Polycomb Repressive Complex 2 signaling. Altogether, our data establishes N-Myc as an oncogenic driver of NEPC.

Genomic landscape of adenoid cystic carcinoma of the breast.

Adenoid cystic carcinoma (AdCC) is a rare type of triple‐negative breast cancer (TNBC) characterized by the presence of the MYB–NFIB fusion gene. The molecular underpinning of breast AdCCs other than the MYB–NFIB fusion gene remains largely unexplored. Here we sought to define the repertoire of somatic genetic alterations of breast AdCCs. We performed whole‐exome sequencing, followed by orthogonal validation, of 12 breast AdCCs to determine the landscape of somatic mutations and gene copy number alterations. Fluorescence in situ hybridization and reverse‐transcription PCR were used to define the presence of MYB gene rearrangements and MYB–NFIB chimeric transcripts. Unlike common forms of TNBC, we found that AdCCs have a low mutation rate (0.27 non‐silent mutations/Mb), lack mutations in TP53 and PIK3CA and display a heterogeneous constellation of known cancer genes affected by somatic mutations, including MYB, BRAF, FBXW7, SMARCA5, SF3B1 and FGFR2. MYB and TLN2 were affected by somatic mutations in two cases each. Akin to salivary gland AdCCs, breast AdCCs were found to harbour mutations targeting chromatin remodelling, cell adhesion, RNA biology, ubiquitination and canonical signalling pathway genes. We observed that, although breast AdCCs had rather simple genomes, they likely display intra‐tumour genetic heterogeneity at diagnosis. Taken together, these findings demonstrate that the mutational burden and mutational repertoire of breast AdCCs are more similar to those of salivary gland AdCCs than to those of other types of TNBCs, emphasizing the importance of histological subtyping of TNBCs. Furthermore, our data provide direct evidence that AdCCs harbour a distinctive mutational landscape and genomic structure, irrespective of the disease site of origin. Copyright

Development and validation of a whole-exome sequencing test for simultaneous detection of point mutations, indels and copy-number alterations for precision cancer care

We describe Exome Cancer Test v1.0 (EXaCT-1), the first New York State-Department of Health-approved whole-exome sequencing (WES)-based test for precision cancer care. EXaCT-1 uses HaloPlex (Agilent) target enrichment followed by next-generation sequencing (Illumina) of tumour and matched constitutional control DNA. We present a detailed clinical development and validation pipeline suitable for simultaneous detection of somatic point/indel mutations and copy-number alterations (CNAs). A computational framework for data analysis, reporting and sign-out is also presented. For the validation, we tested EXaCT-1 on 57 tumours covering five distinct clinically relevant mutations. Results demonstrated elevated and uniform coverage compatible with clinical testing as well as complete concordance in variant quality metrics between formalin-fixed paraffin embedded and fresh-frozen tumours. Extensive sensitivity studies identified limits of detection threshold for point/indel mutations and CNAs. Prospective analysis of 337 cancer cases revealed mutations in clinically relevant genes in 82% of tumours, demonstrating that EXaCT-1 is an accurate and sensitive method for identifying actionable mutations, with reasonable costs and time, greatly expanding its utility for advanced cancer care.

Aberrant Activation of a Gastrointestinal Transcriptional Circuit in Prostate Cancer Mediates Castration Resistance

Prostate cancer exhibits a lineage-specific dependence on androgen signaling. Castration resistance involves reactivation of androgen signaling or activation of alternative lineage programs to bypass androgen requirement. We describe an aberrant gastrointestinal-lineage transcriptome expressed in ∼5% of primary prostate cancer that is characterized by abbreviated response to androgen-deprivation therapy and in ∼30% of castration-resistant prostate cancer. This program is governed by a transcriptional circuit consisting of HNF4G and HNF1A. Cistrome and chromatin analyses revealed that HNF4G is a pioneer factor that generates and maintains enhancer landscape at gastrointestinal-lineage genes, independent of androgen-receptor signaling. In HNF4G/HNF1A-double-negative prostate cancer, exogenous expression of HNF4G at physiologic levels recapitulates the gastrointestinal transcriptome, chromatin landscape, and leads to relative castration resistance.

Patient derived organoids to model rare prostate cancer phenotypes

A major hurdle in the study of rare tumors is a lack of existing preclinical models. Neuroendocrine prostate cancer is an uncommon and aggressive histologic variant of prostate cancer that may arise de novo or as a mechanism of treatment resistance in patients with pre-existing castration-resistant prostate cancer. There are few available models to study neuroendocrine prostate cancer. Here, we report the generation and characterization of tumor organoids derived from needle biopsies of metastatic lesions from four patients. We demonstrate genomic, transcriptomic, and epigenomic concordance between organoids and their corresponding patient tumors. We utilize these organoids to understand the biologic role of the epigenetic modifier EZH2 in driving molecular programs associated with neuroendocrine prostate cancer progression. High-throughput organoid drug screening nominated single agents and drug combinations suggesting repurposing opportunities. This proof of principle study represents a strategy for the study of rare cancer phenotypes.There are few available models to study neuroendocrine prostate cancer. Here they develop and characterize patient derived organoids from metastatic lesions, use these models to show the role of EZH2 in driving neuroendocrine phenotype, and perform high throughput organoid screening to identify therapeutic drug combinations.

Bone biopsy protocol for advanced prostate cancer in the era of precision medicine.

Metastatic biopsies are increasingly being performed in patients with advanced prostate cancer to search for actionable targets and/or to identify emerging resistance mechanisms. Due to a predominance of bone metastases and their sclerotic nature, obtaining sufficient tissue for clinical and genomic studies is challenging.

Abstract IA19: Phenotype plasticity—a novel mechanism of targeted therapy resistance

Prostate cancer (PCa) is the most commonly diagnosed cancer and the third highest cause of cancer-related death in men in Europe, where it is responsible for over 90,000 deaths a year. The mainstay of treatment for metastatic PCa is androgen-deprivation therapy (ADT). Although initially effective, the treatment ultimately fails and progression to castration-resistant prostate cancer (CRPC) occurs. Given that CRPC is still driven by hormonal signaling through aberrant activation of the androgen receptor (AR), improved, more potent AR-targeting therapies have been developed for CRPC patients. However, resistance to these therapies ultimately occurs as well. One form of resistance identified by my group results in a phenotypic switch leading to androgen receptor indifference and progression to neuroendocrine prostate cancer (NEPC), which shows a distinct histomorphology and expresses neural-like markers. Unlike more commonly recognized mechanisms of ADT resistance due to AR mutations or amplification, NEPC no longer responds to AR-targeting therapy and has a mean survival of 7 months. There is mounting evidence supporting the role of epigenetic events as a mechanism for transdifferentiation of PCa to an androgen signaling-indifferent state under specific genomic conditions, involving but not limited to TP53, RB1, and PTEN loss. However, the epigenetic regulators at work and the specific changes in the epigenetic landscape are unknown. The SWI/SNF complex is a major epigenetic player, both in regulating normal cell differentiation and in cancer biology. This presentation will focus on novel data supporting the role of alterations in this complex that could contribute to PCa phenotype plasticity. Citation Format: Joanna Cyrta, David Wilkes, Sung Suk Chae, Matteo Benelli, Rohan Bareja, Davide Prandi, Paola Maria Giovanna Cavaliere, Himisha Beltran, Andrea Sboner, Francesca Demichelis, Mark A. Rubin. Phenotype plasticity—a novel mechanism of targeted therapy resistance [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 IA19.

Bone biopsy protocol for advanced prostate cancer in the era of precision medicine: Bone Biopsy Protocol for NGS

Metastatic biopsies are increasingly being performed in patients with advanced prostate cancer to search for actionable targets and/or to identify emerging resistance mechanisms. Due to a predominance of bone metastases and their sclerotic nature, obtaining sufficient tissue for clinical and genomic studies is challenging.