NF-κB-driven immune checkpoint knockdown and cytokine expression in cancer cells for tumor immunotherapy.

Cancer immunotherapy has made significant advancements, as immune checkpoint inhibitors and cytokines are widely applied in the clinic. Nonetheless, these therapies are often challenged by severe side effects arising from inappropriate activation of the immune system or systemic toxicity. Herein, we constructed a tumor-selective, safe therapeutic gene cassette specifically activated by NF-κB to produce cytokine IL-15 and microRNAs silencing two key immune checkpoints, PD-L1 and CD47. High NF-κB activity in cancer cells drove expression of the cassette to produce IL-15 and suppress PD-L1 and CD47, while low basal NF-κB rendered the cassette silent in most normal cells. This design simultaneously activated and enhanced adaptive and innate immunity. A recombinant adeno-associated virus (AAV2) delivering the cassette showed significant antitumor effects, favorable tumor selectivity, and biosafety across multiple murine solid tumor models. Consequently, this treatment may offer a potentially more effective and safer immunotherapy against cancers in the future.