Background <p>Current treatment strategies for high-grade gliomas have a limited overall patient benefit. Here, Methylated RNA immunoprecipitation sequencing (MeRIP-seq) of glioma was conducted to screen the lncRNA regulated by m6A modification in glioma, and provide an innovative chemotherapeutic strategy to improved the therapeutic efficacy.</p> Methods <p>To elucidate the impact and the molecular mechanism of ALKBH5/SH3RF3-AS1/SH3RF3 regulation on glioma malignancy, proliferation phenotypic assays, MeRIP-qPCR, RIP-qPCR and western blotting were used. Expressions and clinical significances of ALKBH5, SH3RF3-AS1, and SH3RF3 were assessed in tissue chip. Furthermore, we designed a nanodrug (Nano^PCPT/siRNA@U251) for the targeted co-delivery of CPT (targeting type Ⅰ topoisomerase) and si-SH3RF3-AS1 (inhibiting SH3RF3, a Rac1-JNK activator) in glioma treatment.</p> Results <p>SH3RF3-AS1 was demethylated and overexpressed in glioma. We elucidated the mechanistic link between ALKBH5-mediated N6-methyladenosine (m6A) modification and SH3RF3-AS1 expression, and the cis-regulation of SH3RF3-AS1 to SH3RF3. Clinically, the ALKBH5, SH3RF3-AS1, and SH3RF3 expression increased with increasing glioma grade, high SH3RF3-AS1 expression correlated with unfavorable 5-year overall survival and progression-free survival in a cohort of 148 glioma patients. SH3RF3-AS1 drived glioma progression and promoted chemoresistance to camptothecin (CPT). We designed a nanodrug (Nano^PCPT/siRNA@U251) for the targeted co-delivery of CPT (targeting type Ⅰ topoisomerase) and si-SH3RF3-AS1 (inhibiting SH3RF3, a Rac1-JNK activator) in glioma treatment, and it effectively suppressed glioma cell growth both in vivo and in vitro models.</p> Conclusion <p>Our findings provide valuable insights into the roles of m6A demethylation-mediated upregulation of SH3RF3-AS1 in supporting glioma proliferation and demonstrate the potential of Nano^PCPT/siRNA@U251 as an innovative chemotherapeutic strategy for glioma.</p>