m6A epitranscriptome analysis reveals differentially methylated transcripts that drive early chemoresistance in bladder cancer.

N ⁶-Methyladenosine (m⁶A) RNA modifications dynamically regulate messenger RNA processing, differentiation and cell fate. Given these functions, we hypothesized that m⁶A modifications play a role in the transition to chemoresistance. To test this, we took an agnostic discovery approach anchored directly to chemoresistance rather than to any particular m⁶A effector protein. Specifically, we used methyl-RNA immunoprecipitation followed by sequencing (MeRIP-seq) in parallel with RNA sequencing to identify gene transcripts that were both differentially methylated and differentially expressed between cisplatin-sensitive and cisplatin-resistant bladder cancer (BC) cells. We filtered and prioritized these genes using clinical and functional database tools, and then validated several of the top candidates via targeted quantitative polymerase chain reaction (qPCR) and MeRIP-PCR. In cisplatin-resistant cells, SLC7A11 transcripts had decreased methylation associated with decreased m⁶A reader YTHDF3 binding, prolonged RNA stability, and increased RNA and protein levels, leading to reduced ferroptosis and increased survival. Consistent with this, cisplatin-sensitive BC cell lines and patient-derived organoids exposed to cisplatin for as little as 48 h exhibited similar mechanisms of SLC7A11 upregulation and chemoresistance, trends that were also reflected in public cancer survival databases. Collectively, these findings highlight epitranscriptomic plasticity as a mechanism of rapid chemoresistance and a potential therapeutic target.