Summary The lack of in vitro prostate cancer models that recapitulate the diversity of human prostate cancer has hampered progress in understanding disease pathogenesis and therapy response. Using a 3D “organoid” system, we report success in long-term culture of prostate cancer from biopsy specimens and circulating tumor cells. The first seven fully characterized organoid lines recapitulate the molecular diversity of prostate cancer subtypes, including TMPRSS2-ERG fusion, SPOP mutation, SPINK1 overexpression and CHD1 loss. Whole exome sequencing shows a low mutational burden, consistent with genomics studies, but with mutations in FOXA1 and PIK3R1, as well as of DNA repair and chromatin modifier pathways that have been reported in advanced disease. Loss of p53 and RB tumor suppressor pathway function are the most common feature shared across the organoid lines. The methodology described here should enable the generation of a large repertoire of patient-derived prostate cancer lines amenable to genetic and pharmacologic studies.

Some cancers evade targeted therapies through a mechanism known as lineage plasticity, whereby tumor cells acquire phenotypic characteristics of a cell lineage whose survival no longer depends on the drug target. We use in vitro and in vivo human prostate cancer models to show that these * Corresponding author. sawyersc@mskcc.org. SUPPLEMENTARY MATERIALS

Malignant Peripheral Nerve Sheath Tumors (MPNSTs) represent a group of highly aggressive soft tissue sarcomas that may occur sporadically, in association with neurofibromatosis type I (NF1-), or after radiotherapy1–3. Using comprehensive genomic approaches, we identified loss-of-function (LOF) somatic alterations of the Polycomb repressive complex 2 (PRC2) core components, EED or SUZ12, in 92% of sporadic, 70% of NF1-associated and 90% of radiotherapy-associated MPNSTs. MPNSTs with PRC2 loss showed complete loss of H3K27me3 and aberrant transcriptional activation of multiple PRC2-repressed homeobox master regulators and their regulated developmental pathways. Introduction of the PRC2 component in a PRC2-deficient MPNST cell line restored H3K27me3 and decreased cell growth. Additionally, we identified frequent somatic alterations of CDKN2A (81% of all MPNSTs) and NF1 (72% of non-NF1-associated MPNSTs), and they significantly co-occur with PRC2 alterations. The highly recurrent and specific inactivation of PRC2, NF1, CDKN2A posits their critical and potentially cooperative roles in MPNST pathogenesis.

Activation of oncogenes by mechanisms other than genetic aberrations such as mutations, translocations, or amplifications is largely undefined. Here we report a novel isoform of the anaplastic lymphoma kinase (ALK) that is expressed in ~ 11% of melanomas and sporadically in other human cancer types, but not in normal tissues. The novel ALK transcript initiates from a de novo alternative transcription initiation (ATI) site in ALK intron 19, and was termed ALKATI. In ALKATI-expressing tumours, the ATI site is enriched for H3K4me3 and RNA polymerase II, chromatin marks characteristic of active transcription initiation sites1. ALKATI is expressed from both ALK alleles, and no recurrent genetic aberrations are found at the ALK locus, indicating that the transcriptional activation is independent of genetic aberrations at the ALK locus. The ALKATI transcript encodes three proteins with molecular weights of 61.1, 60.8 and 58.7 kilodaltons, consisting primarily of the intracellular tyrosine kinase domain. ALKATI stimulates multiple oncogenic signalling pathways, drives growth-factor-independent cell proliferation in vitro, and promotes tumorigenesis in vivo in mouse models. ALK inhibitors can suppress the kinase activity of ALKATI, suggesting that patients with ALKATI-expressing tumours may benefit from ALK inhibitors. Our findings suggest a novel mechanism of oncogene activation in cancer through de novo alternative transcription initiation.

Supplementary data are available at Bioinformatics online.

Gastrointestinal stromal tumour (GIST), originating from the interstitial cells of Cajal (ICCs), is characterized by frequent activating mutations of the KIT receptor tyrosine kinase. Despite the clinical success of imatinib that targets KIT, most advanced GIST patients develop resistance and eventually die of the disease. The ETS family transcription factor, ETV1, is a master regulator of the ICC lineage. Using mouse models of Kit activation and Etv1 ablation, we demonstrate that Etv1 is required for GIST initiation and proliferation in vivo, validating it as a therapeutic target. We further uncover a positive feedback circuit where MAP kinase activation downstream of KIT stabilizes the ETV1 protein and ETV1 positively regulates KIT expression. Combined targeting of ETV1 stability by imatinib and MEK162 resulted in increased growth suppression in vitro and complete tumour regression in vivo. The combination strategy to target ETV1 may provide an effective therapeutic strategy in GIST clinical management.

Summary Metastatic prostate cancer is characterized by recurrent genomic copy number alterations that are presumed to contribute to resistance to hormone therapy. We identified CHD1 loss as a cause of antiandrogen resistance in an in vivo small hairpin RNA (shRNA) screen of 730 genes deleted in prostate cancer. ATAC-seq and RNA-seq analyses showed that CHD1 loss resulted in global changes in open and closed chromatin with associated transcriptomic changes. Integrative analysis of this data, together with CRISPR-based functional screening, identified four transcription factors (NR3C1, POU3F2, NR2F1, and TBX2) that contribute to antiandrogen resistance, with associated activation of non-luminal lineage programs. Thus, CHD1 loss results in chromatin dysregulation, thereby establishing a state of transcriptional plasticity that enables the emergence of antiandrogen resistance through heterogeneous mechanisms.

Summary 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.