An ALKBH3 Inhibitor HUHS199 Suppresses Proliferation of Glioblastoma Cells by Targeting 1-Methyladenosine on GADD45A mRNA.

Glioblastoma (GBM) is a highly aggressive, poor-prognosis brain tumor classified as WHO grade IV, for which effective treatments remain limited. Although AlkB homolog 3 (ALKBH3), a demethylase for 1-methyladenosine (m1A) and 3-methylcytidine (m3C) in DNA and RNA, has been implicated in cancer cell proliferation, its functional role in GBM remains unclear. We aimed to characterize the role of ALKBH3 in GBM and develop and evaluate a patented ALKBH3 inhibitor, HUHS199, to identify its molecular targets and therapeutic potential. We found that ALKBH3 was highly expressed in clinical GBM specimens; its knockdown inhibited GBM cell proliferation. HUHS199 inhibited GBM cell proliferation in a dose-dependent manner, induced G1 phase cell cycle arrest, and increased m1A levels in RNA. Furthermore, RNA immunoprecipitation-microarray analysis using an anti-m1A antibody identified growth arrest and DNA-damage-inducible protein GADD45 alpha (GADD45A) mRNA as a target of ALKBH3-mediated demethylation. These findings suggest that ALKBH3 promotes GBM cell proliferation via m1A demethylation of GADD45A mRNA. Collectively, this study highlights the therapeutic potential of ALKBH3 inhibition and presents HUHS199 as a first-in-class candidate for GBM treatment.