Persistent androgen receptor (AR) transcriptional activity underlies resistance to AR-targeted therapy and progression to lethal castration resistant prostate cancer (CRPC). Recent success in re-targeting persistent AR activity with next-generation androgen/AR axis inhibitors such as enzalutamide (MDV3100) has validated AR as a master regulator during all stages of disease progression. However, resistance to next-generation AR inhibitors limits therapeutic efficacy for many patients. One emerging mechanism of CRPC progression is AR gene rearrangement, promoting synthesis of constitutively-active truncated AR splice variants (AR-Vs) that lack the AR ligand binding domain. In this study, we demonstrate that cells with AR gene rearrangements expressing both full-length and AR-Vs are androgen-independent and enzalutamide-resistant. However, selective knock-down of AR-V expression inhibited androgen-independent growth and restored responsiveness to androgens and antiandrogens. In heterogeneous cell populations, AR gene rearrangements marked individual AR-V-dependent cells that were resistant to enzalutamide. Gene expression profiling following knock-down of full-length AR or AR-Vs demonstrated that AR-Vs drive resistance to AR-targeted therapy by functioning as constitutive and independent effectors of the androgen/AR transcriptional program. Further, mitotic genes deemed previously to be unique AR-V targets were found to be biphasic targets associated with a proliferative level of signaling output from either AR-Vs or androgen-stimulated AR. Overall, these studies highlight AR-Vs as key mediators of persistent AR signaling and resistance to the current arsenal of conventional and next-generation AR-directed therapies, advancing the concept of AR-Vs as therapeutic targets in advanced disease.

Plant organ development and polarity establishment is mediated by the action of several transcription factors. Among these, the KANADI (KAN) subclade of the GARP protein family plays important roles in polarity-associated processes during embryo, shoot and root patterning. In this study, we have identified a set of potential direct target genes of KAN1 through a combination of chromatin immunoprecipitation/DNA sequencing (ChIP-Seq) and genome-wide transcriptional profiling using tiling arrays. Target genes are over-represented for genes involved in the regulation of organ development as well as in the response to auxin. KAN1 affects directly the expression of several genes previously shown to be important in the establishment of polarity during lateral organ and vascular tissue development. We also show that KAN1 controls through its target genes auxin effects on organ development at different levels: transport and its regulation, and signaling. In addition, KAN1 regulates genes involved in the response to abscisic acid, jasmonic acid, brassinosteroids, ethylene, cytokinins and gibberellins. The role of KAN1 in organ polarity is antagonized by HD-ZIPIII transcription factors, including REVOLUTA (REV). A comparison of their target genes reveals that the REV/KAN1 module acts in organ patterning through opposite regulation of shared targets. Evidence of mutual repression between closely related family members is also shown.

Loss of cell-cycle control is a hallmark of cancer, which can be targeted with agents, including cyclin-dependent kinase-4/6 (CDK4/6) kinase inhibitors that impinge upon the G-S cell-cycle checkpoint via maintaining activity of the retinoblastoma tumor suppressor (RB). This class of drugs is under clinical investigation for various solid tumor types and has recently been FDA-approved for treatment of breast cancer. However, development of therapeutic resistance is not uncommon. In this study, palbociclib (a CDK4/6 inhibitor) resistance was established in models of early stage, RB-positive cancer. This study demonstrates that acquired palbociclib resistance renders cancer cells broadly resistant to CDK4/6 inhibitors. Acquired resistance was associated with aggressive and phenotypes, including proliferation, migration, and invasion. Integration of RNA sequencing analysis and phosphoproteomics profiling revealed rewiring of the kinome, with a strong enrichment for enhanced MAPK signaling across all resistance models, which resulted in aggressive and phenotypes and prometastatic signaling. However, CDK4/6 inhibitor-resistant models were sensitized to MEK inhibitors, revealing reliance on active MAPK signaling to promote tumor cell growth and invasion. In sum, these studies identify MAPK reliance in acquired CDK4/6 inhibitor resistance that promotes aggressive disease, while nominating MEK inhibition as putative novel therapeutic strategy to treat or prevent CDK4/6 inhibitor resistance in cancer. .

The androgen receptor (AR) is a driver of prostate cancer (PCa) cell growth and disease progression. Therapies for advanced PCa exploit AR dependence by blocking the production or action of androgens, but these interventions inevitably fail via multiple mechanisms including mutation or deletion of the AR ligand binding domain (LBD). Thus, the development of new inhibitors which act through non-LBD interfaces is an unmet clinical need. EPI-001 is a bisphenol A-derived compound shown to bind covalently and inhibit the AR NH2-terminal domain (NTD). Here, we demonstrate that EPI-001 has general thiol alkylating activity, resulting in multilevel inhibitory effects on AR in PCa cell lines and tissues. At least one secondary mechanism of action associated with AR inhibition was found to be selective modulation of peroxisome proliferator activated receptor-gamma (PPARγ). These multi-level effects of EPI-001 resulted in inhibition of transcriptional activation units (TAUs) 1 and 5 of the AR NTD, and reduced AR expression. EPI-001 inhibited growth of AR-positive and AR-negative PCa cell lines, with the highest sensitivity observed in LNCaP cells. Overall, this study provides new mechanistic insights to the chemical biology of EPI-001, and raises key issues regarding the use of covalent inhibitors of the intrinsically unstructured AR NTD.