AAV-Mediated CAR-T therapy eradicate HIV reservoirs with augmented safety and persistence

Abstract HIV-1 persists in latent reservoirs within CD4 cells, posing a major barrier to cure. Chimeric antigen receptor (CAR) T-cell therapy has shown promise against HIV but is limited by several key challenges, including persistent and protecting CAR-T cells expression, particularly those expressing CD4-based receptors from HIV infection, preventing immune evasion through viral mutation, and effectively targeting transcriptionally silent reservoirs that lack antigen expression and thus remain undetectable to CAR-based therapies. AAV-based CAR-T therapy addresses these challenges by providing stable and safe CAR delivery with low integration risk, enhancing persistence while reducing insertional mutagenesis concerns. AAV-CD4CAR transduced primary human T cells at high efficiency and viability; the resulting CAR T cells selectively recognized and killed HIV-1–infected CD4+ T-cells, significantly reducing viral p24 levels in infected PBMC cultures. In a humanized mouse model engrafted with HIV, AAV-CD4CAR T cells led to sustained CAR expression on circulating T cells. Treated mice showed marked suppression of plasma viremia and preservation of CD4+ T-cell counts compared to controls, without evidence of systemic toxicity. The CAR T cells exhibited effector cytokine production and memory markers, indicating functional activation and persistence. Moreover, we found that the AAV-based CAR is safe and biocompatible and does not affect the physiological function of mice challenged with HIV. Taken together, these results verify that AAV-CD4CAR is effective in clearing viral load from ex vivo human samples and in mice models and is safe and biocompatible. Our findings provide proof of concept that an AAV-CD4CAR approach can induce durable anti-HIV immunity, suggesting a new avenue for curative therapy.