Decoding androgen receptor inhibitor resistance in prostate cancer

Abstract

In prostate cancer, aberrant androgen receptor (AR) function leads to the deregulation of AR target gene expression, driving prostate tumorigenesis. AR-directed therapies are transiently effective, and resistance to these drugs inevitably occurs. Therefore, there is a pressing need to better understand therapeutic resistance to AR inhibitors.

In this study, we identified transcription factor TLE3 as a novel regulator of enzalutamide and apalutamide resistance using a genome-wide CRISPR-Cas9 resistance screen in LNCaP cells. At the molecular level, TLE3 loss rescued expression of a subset of AR target genes in enzalutamide-treated cells. The second-most upregulated gene in enzalutamide- treated TLE3KO cells was the glucocorticoid receptor (GR), which was previously shown to substitute AR in prostate cancer cells thereby conferring enzalutamide resistance. We found that TLE3 and AR bind at the GR locus, consistent with upregulation of GR in this context. Genetic and pharmacological perturbation of GR rescued the resistance phenotype. Moreover, GR binding at TLE3/AR-shared genes further supports the role of GR in enzalutamide resistance in TLE3KO cells. Analysis of patient samples revealed an association between TLE3 and GR expression in prostate tumors that are in line with our in vitro findings, of which the clinical relevance is yet to be established.

Furthermore, we reveal aberrant MAPK signaling as a determinant of enzalutamide sensitivity in CWR-R1 prostate cancer cells. Using a kinome-centered CRISPR-Cas9 screen, we identified BRAF as a critical node required for cell growth in enzalutamide-treated CWR-R1 cells. Inhibition of downstream MAPK components MEK or ERK in conjunction with enzalutamide yielded similar results, showing strong synergistic inhibition of cell proliferation. Characterization of the BRAF gene revealed a mutation in the activating kinase domain of BRAF. The lack of response to enzalutamide in two patients harboring mutations in the activating kinase domain of BRAF is consistent with our findings in CWR-R1 cells in vitro. Combined, our findings suggest that co-targeting of the AR and MAPK pathway may be effective in patients with an activated MAPK pathway, particularly those harboring tumors with oncogenic BRAF mutations.

Finally, we explored molecular data with the aim to reveal genetic determinants associated with organ-specific metastasis in prostate cancer. To this end, we set out to characterize genetic features associated with metastasis site through molecular profiling of 326 prostate cancer metastases. We found RB1 and PIK3CA alterations to be enriched in metastases of the lymph nodes and liver respectively when compared to other sites. Aggregated pathway alteration data showed a trend for enrichment of PI3K and DNA repair pathway alterations in lymph node compared to bone metastases. Furthermore, analysis of TMB revealed it was increased in visceral and liver metastases, which was associated with an MMR deficiency signature. In line with these findings, a significant proportion of high-TMB liver metastases showed alterations in MSH6, MLH1 and POLD3, while high-TMB visceral metastases were characterized by MSH2 and POLD1 alterations. Together, our data show differential enrichment of TMB/MMR-deficiency, and alterations affecting prostate cancer- related oncogenic drivers, at distinct metastatic sites, which may potentially impact therapy response and disease progression.