258 Enhancing tumor infiltration of CAR T cells in solid tumors

Background

CD8⁺ naive T cells undergo clonal expansion upon antigen encounter, generating effector cells that eliminate infected and malignant cells. Following clearance, most effector cells terminally differentiate, while a subset of memory precursor effector cells survive contraction, giving rise to circulating memory T cells providing long-term protection. Some precursor cells infiltrate tissues, becoming tissue-resident memory T (T_RM) cells. T_RM cells persist in peripheral tissues, preventing reinfection and tumor growth. This project focuses on understanding the molecular networks controlling T_RM tissue infiltration and long-term persistence with the final aim to optimize and improve adoptive cell transfer immunotherapy in solid cancers. Despite the success of chimeric antigen receptor (CAR) T cells in liquid tumors, their efficacy in solid tumors is hindered by immunosuppression, metabolic factors, and physical barriers, compromising functionality and tumor infiltration. Indeed, anti-tumor T cell responses resembling T_RM correlate with improved prognosis.^{1 2} However, the molecular networks regulating adaptation to persistent antigen stimulation and tumor infiltration in CAR T cell therapy remain unknown. Thus, targeting solid tumors is still particularly challenging.

Methods

To study the infiltration of CAR T cells into solid tumors, we are conducting co-transductions of CD8 T cells using a CAR specific to the CD19 molecule (CAR19), along with genes encoding proteins of interest. The modified cells are then tested both in vitro and in vivo using tumor models, such as B16 mouse melanoma tumors that overexpress the CD19 molecule (B16.CD19).

Results

Our hypothesis is that providing tissue residency transcriptional programs to CAR T cells can enhance their ability to control solid tumors. As a proof of concept, we have simultaneously overexpressed the CAR and the transcription factor RUNX3, master regulator of tissue residency. This approach aims to promote tumor infiltration and T cell accumulation within the tumor. In vitro coculture experiments using the modified T cells and the B16.CD19 cell line have confirmed the specific killing activity of the co-transduced CAR T cells, regardless the overexpression of RUNX3. In vivo, tumor monitoring has revealed that RUNX3⁺CAR19⁺ CD8 T cells present superior control of tumor growth over time and higher accumulation within the tumor compared to RUNX3^–CAR19⁺ cells. Notably, the RUNX3⁺CAR19⁺ T cells display increased expression of T-bet and CD44, suggesting a heightened effector potential, and of PD-1 and TCF-1, a phenotype associated with favorable prognosis in cancer patients.

Conclusions

These results highlight the importance of dissecting tissue-resident memory cells and their regulatory circuits to treat solid tumors.

References

Ethics Approval

All animal studies were approved by the Institutional Animal Care and Use Committees of the University of California, San Diego (UCSD, protocol #S04105) and performed in accordance with UC guidelines.