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Abstract As the most common modification of RNA, N6-methyladenosin (m6A) has been confirmed to be involved in the occurrence and development of various cancers. However, the relationship between m6A and castration resistance prostate cancer (CRPC), has not been fully studied. By m6A-sequencing of patient cancer tissues, we identified that the overall level of m6A in CRPC was up-regulated than castration sensitive prostate cancer (CSPC). Based on the analysis of m6A-sequencing data, we found m6A modification level of HRas proto-oncogene, GTPase (HRAS) and mitogen-activated protein kinase kinase 2 (MEK2 or MAP2K2) were enhanced in CRPC. Specifically, tissue microarray analysis and molecular biology experiments confirmed that METTL3, an m6A “writer” up-regulated after castration, activated the ERK pathway to contribute to malignant phenotype including ADT resistance, cell proliferation and invasion. We revealed that METTL3-mediated ERK phosphorylation by stabilizing the transcription of HRAS and positively regulating the translation of MEK2. In the Enzalutamide-resistant (Enz-R) C4-2 and LNCap cell line (C4-2R, LNCapR) established in the current study, the ERK pathway was confirmed to be regulated by METTL3. We also found that applying antisense oligonucleotides (ASOs) to target the METTL3/ERK axis can restore Enzalutamide resistance in vitro and in vivo. In conclusion, METTL3 activated the ERK pathway and induced the resistance to Enzalutamide by regulating the m6A level of critical gene transcription in the ERK pathway.

Abstract Enabled resistance or innate insensitiveness to antiandrogen are lethal for castration-resistant prostate cancer (CRPC). Unfortunately, there seems to be little can be done to overcome the antiandrogen resistance because of the largely unknown mechanisms. In prospective cohort study, we found that HOXB3 protein level was an independent risk factor of PSA progression and death in patients with metastatic CRPC. In vivo, upregulated HOXB3 contributed to CRPC xenografts progression and abiraterone resistance. To uncover the mechanism of HOXB3 driving tumor progression, we performed RNA-sequencing in HOXB3 negative (HOXB3-) and HOXB3 high (HOXB3 + ) staining CRPC tumors and determined that HOXB3 activation was associated with the expression of WNT3A and enriched WNT pathway genes. Furthermore, extra WNT3A and APC deficiency led HOXB3 to be isolated from destruction-complex, translocated to nuclei, and then transcriptionally regulated multiple WNT pathway genes. What’s more, we also observed that the suppression of HOXB3 could reduce cell proliferation in APC-downregulated CRPC cells and sensitize APC-deficient CRPC xenografts to abiraterone again. Together, our data indicated that HOXB3 served as a downstream transcription factor of WNT pathway and defined a subgroup of CRPC resistant to antiandrogen which would benefit from HOXB3-targeted therapy.

Abstract Androgen receptor (AR) plays an important role in the progression of prostate cancer and has been targeted by castration or AR-antagonists. The emergence of castration-resistant prostate cancer (CRPC) after androgen deprivation therapy (ADT) is inevitable. However, it is not entirely clear how ADT fails or how it causes resistance. Through analysis of RNA-seq data, we nominate ARHGEF2 as a pivotal androgen-repressed gene. We show that ARHGEF2 is directly suppressed by androgen/AR. AR occupies the enhancer and communicates with the promoter region of ARHGEF2. Functionally, ARHGEF2 is important for the growth, lethal phenotype, and survival of CRPC cells and tumor xenografts. Correspondingly, AR inhibition or AR antagonist treatment can restore ARHGEF2 expression, thereby allowing prostate cancer cells to induce treatment resistance and tolerance. Overall, our findings provide an explanation for the contradictory clinical results that ADT resistance may be caused by the up-regulation of ARHGEF2 and provide a novel target.