Simultaneously Targeting Lineage Plasticity and Therapy Resistance Drivers Using a Novel Dual-PROTAC Strategy

PROJECT SUMMARY Lineage plasticity has been recognized as a key mechanism that enables cancer cells to evade targeted therapies. Prostate cancer (PCa), especially its most aggressive form, metastatic castration-resistant prostate cancer (mCRPC), exemplifies lineage plasticity-based resistance to Androgen Receptor (AR) targeted therapies. This resistance significantly constrains patient clinical outcomes, rendering mCRPC incurable, and underscores the critical demand for elucidating the mechanisms of resistance and developing novel therapeutic approach to overcome resistance. Although various lineage plasticity and resistance drivers have been identified recently, including GR and JAK-STAT, a notable characteristic of this plasticity-derived therapy resistance is the capacity of the cancer cells to switch their lineage swiftly and reversibly, relying on multiple dysregulated lineage survival factors. This remarkable adaptability of cancer cells underscores the necessity of simultaneously inhibiting alternative resistance drivers and the lineages they drive to effectively overcome therapy resistance. Although PROTAC protein degraders have emerged as a significant pharmacological advancement, they lack the ability to simultaneously target multiple oncogenic drivers, which greatly limits the available clinical approaches to overcome resistance. To address this gap, we propose to test the hypothesis that the simultaneous inhibition of both the original and alternative tumor driver proteins is necessary for effectively suppressing therapy resistance driven by lineage plasticity. Leveraging our newly developed high-throughput PROTAC-type degrader synthesis platform, the overall goal of this study is to develop an innovative dual-target PROTAC-based protein degrader targeting lineage plasticity drivers in mCRPC. This system will provide an effective method to concurrently target two alternative oncogenic drivers essential for the survival of therapy-resistant mCRPC cells. In Aim 1, we will first synthesize a series of mono-PROTAC degraders targeting GR and JAK1/JAK2, utilizing our high-throughput PROTAC platform. Building on the successful development of GR mono-PROTACs, we will then evolve a multicomponent coupling strategy to synthesize dual-PROTACs, GAP-1, that simultaneously target both GR and AR. Finally, we will synthesize a novel dual-PROTAC degrader, JAP-1, to simultaneously target ectopic JAK- STAT and AR signaling. In Aim 2, we will examine the efficacy of the dual-PROTAC degraders in vitro and in vivo, using a collection of xenograft-derived cell lines, patient-derived organoids (PDO), and patient-derived explants (PDE). Then, using scRNA-seq and spatial transcriptomics, we will comprehensively assess the impact of our dual-PROTAC degraders on temporal and spatial intratumor heterogeneity and lineage plasticity at single- cell resolution. The successful completion of the proposed study will open a novel path to mitigate resistance in numerous advanced cancers dependent on multiple oncogenic drivers and have significant immediate and profound impacts on the clinical outcomes of patients with lethal mCRPC.