H3K36me3‐Guided m ⁶ A Modification of Oncogenic L1CAM‐AS1 Drives Macrophage Polarization and Immunotherapy Resistance in Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is one of the most lethal malignancies and epigenetic modifiers play a key role in HCC progression. Histone H3 trimethylation at lysine-36 (H3K36me3) determines deposition of mRNA de novo N ⁶-methyladenosine (m⁶A) modification. However, it remains largely elusive how long noncoding RNAs (lncRNAs) are selected for proper m⁶A methylation. The current study provides evidence for L1CAM-AS1 as a novel H3K36me3-guided, m⁶A-modified lncRNA through integration of genome-wide H3K36me3 profiles and transcriptome-wide m⁶A profiles of HCC cells. The crucial m⁶A-modification site in L1CAM-AS1 exon 3 is recognized by IGF2BP1, leading to increased lncRNA stability. Oncogenic L1CAM-AS1 shows higher expression in HCC tissues than in normal specimens, and its elevated expression is associated with shorten patient survival. Mechanistically, L1CAM-AS1 interrupts binding of RAN to the E3 ligase OSTM1, suppresses RAN ubiquitination at Lys152 and Lys167, stabilizes RAN protein, enhances nuclear import of RELA (p65), and activates the NF-κB signaling, leading to up-regulated CCL2 expression. L1CAM-AS1-induced CCL2 secretion from HCC cells enhances M2 polarization of tumor-associated macrophages (TAMs). Meanwhile, immunosuppressive M2 macrophages-released CCL5 augments RELA nuclear import in HCC cells, which in turn activates the NF-κB signaling. Given the critical role of macrophages in anti-tumor immunity, inhibition of the L1CAM-AS1-RAN axis promotes the efficacy of PD-1 blockade via TAM reprogramming in HCC mouse models. In conclusion, this study provides novel insights into how epigenetic alternations are involved in antitumor immunity modulation and illustrates promising potentials of L1CAM-AS1 in immune-checkpoint inhibitor treatments for HCC.