Advances in CAR optimization strategies based on CD28

Sijin Li ^1,2,3 Yusi Zhou ^1,2,3 Hairong Wang ^1,2,3 Gexi Qu ^1,2,3 Xuan Zhao ^1,2,3 Xu Wang ^1,2,3 Rui Hou ⁴ Zhangchun Guan ^1,2,3 Dan Liu ^1,2,3* Junnian Zheng ^2,3* Ming Shi ^1,2,3*

• ¹ Cancer Institute, Xuzhou Medical University, Xuzhou, China
• ² Center of Clinical Oncology, Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu Province, China
• ³ Jiangsu Center for the Collaboration and Innovation of Cancer Biotherapy, Xuzhou, Jiangsu Province, China
• ⁴ College of Pharmacy, Xuzhou Medical University, Xuzhou, Jiangsu Province, China

Chimeric antigen receptor (CAR)-T cell therapy, which utilizes genetic engineering techniques to modify T-cells to achieve specific targeting of cancer cells, has made significant breakthroughs in cancer treatment in recent years. All marketed CAR-T products are second-generation CAR-T cells containing co-stimulatory structural domains, and co-stimulatory molecules are critical for CAR-T cell activation and function. Although CD28-based co-stimulatory molecules have demonstrated potent cytotoxicity in the clinical application of CAR-T cells, they still suffer from high post-treatment relapse rates, poor efficacy durability, and accompanying severe adverse reactions. In recent years, researchers have achieved specific results in enhancing the anti-tumor function of CD28 by mutating its signaling motifs, combining the co-stimulatory structural domains, and modifying other CAR components besides co-stimulation. This paper reviewed the characteristics and roles of CD28 in CAR-T cell-mediated anti-tumor signaling and activation. We explored potential strategies to enhance CAR-T cell efficacy and reduce side effects by optimizing CD28 motifs and CAR structures, aiming to provide a theoretical basis for further clinical CAR-T cell therapy development.