Investigation of n-terminus mll complexes on the reversion of taxane resistance in castration-resistant prostate cancer

Prostate cancer (PC) typically relies on androgen for abnormal growth, and androgen deprivation therapy (ADT) is preferred as the primary treatment. However, patients frequently progress to a castration-resistant (CR) stage, where tumor growth becomes unresponsive to ADT. Despite the common use of conventional chemotherapeutics like Taxanes (Docetaxel-Dtx, Cabazitaxel-Cbz), either alone or in conjunction with hormone therapies, certain tumors may experience recurrence. This work focuses on the epigenetic regulations conferring Dtx resistance in CRPC. Firstly, the generation of Dtx-resistant cells was conducted using the dose increment method, and subsequently, our model was confirmed via in vitro and in vivo models. Epigenetic drug screening identified MLL-Menin and MLL-WDR5 inhibitors as hit molecules that effectively reverse drug resistance through G2/M arrest and apoptosis induction. N- and C-terminal binding partners of MLL were individually knocked out via CRISPR-cas9. Our analysis led to the discovery of sensitivity on DtxR cells upon Menin depletion, while no effect was observed on parental counterparts. On the other hand, parental cells lacking Menin expression showed reduced capacity to develop drug resistance, suggesting an indispensable role of Menin for the drug refractory phenotype. Restoring several Menin mutants led to the identification of another effective factor, LEDGF, which specifically diminishes colony growth on the DtxR model. RNA-seq analysis was conducted on parental and DtxR cells, upon Menin and LEDGF ablation. GSEA analysis revealed positively enriched mTOR signaling, E2F targets, and G2M checkpoints gene sets in DtxR cells. Interestingly, Menin and LEDGF depletion significantly reversed the enrichment profile of the interested gene sets. Initially, we revealed the essentiality of the mTOR pathway in DtxR maintenance and cell growth. Furthermore, mTOR expression was significantly reduced in Menin knockout cells. On the other hand, Menin depletion triggered a significant synergy with Torin (mTOR inhibitor) and Dtx in our resistant model. Recovering Menin also induced mTOR expression in our DtxR model and abolished the observed synergy. Menin and mTOR correlation was also increased in metastatic CRPC in patient-derived clinical data. Furthermore, our ChIP-qPCR experiments demonstrated Menin enrichment on mTOR promoter region in DtxR CRPC cells. Competition experiments exhibited significant domination by control cells; indicating Menin depletion results in a slower rate of cell division. A higher proportion of Menin knockout cells accumulated in the G1 cell cycle state. Furthermore, two important factors promoting G1-S transition, Cyclin D1 and CDK20 were significantly lower in Menin-depleted cells. Restoring Menin also rescued expression patterns of these targets; furthermore, Menin occupied the promoter region of Cyclin D1 and CDK20. This slow growth rate observed in Menin ablated cells also provided a slight resistance against CDK4/6 inhibitors. Overall, Menin appears as a key regulator that confers drug resistance through mTOR upregulation and controls G1-S progression via Cyclin D1 and CDK20 in our DtxR model. Menin and LEDGF both contribute to essentiality in DtxR cells, while Menin shows enrichment on the promoter regions of specific targets. Our study provides a detailed analysis of Dtx resistance in CRPC, through epigenetic regulation.