Investigating RUNX2 KO in B16 melanoma cells as a potential strategy to enhance in vivo tumor response to therapeutic approaches

This work investigates the role of RUNX2 in Cutaneous Melanoma (CM) resistance. Bioinformatic analysis (TIMER2.0, TISIDB) shows RUNX2 correlates with immune inhibitors and therapy non-responsiveness. We describe the generation of a RUNX2 knockout (KO) in B16 melanoma cells using the CRISPR/Cas9 system. Successful gene editing in five clones was confirmed by Sanger sequencing and Western blot, showing an in-frame deletion in the Runt domain and loss of protein expression. These engineered B16 cells provide a crucial tool for dissecting RUNX2-mediated immune evasion in syngeneic mouse models.