Abstract 2176: Using a PSMA-specific low-molecular-weight compound for prostate cancer treatment with rapidly switchable universal CAR-T cells: Overcoming the challenges of cellular immunotherapies in solid tumors

CAR-T cell therapy holds great promise for treating a wide range of malignancies. Nevertheless, the CAR-T approach faces multiple challenges, including lack of suitable targets, insufficient tumor penetration and a microenvironment hostile to CAR-T cells. Here we provide pre-clinical evidence for using a low-molecular-weight, chemically synthesized compound to re-target CAR-T cells against solid tumors in conjunction with the CD28 co-stimulatory domain to address several of these challenges. PSMA is a validated target for treatment of advanced and metastatic prostate cancer (PCa), but also broadly expressed on tumor neo-vasculature beyond PCa. PSMA is also known to be expressed on a number of normal tissues, albeit to a much lower extent, which necessitates additional safety mechanisms for CAR-T therapy. Therefore, a rapidly switchable universal CAR-T platform (UniCAR) was developed. In this system, antigen-specificity is provided by soluble adaptors termed targeting modules (TM), which redirect T-cells engineered to express a universal CAR in an antigen-specific manner against tumors. The TM explored in the present study incorporates a clinically well-characterized radiotracer peptide motif binding to the enzymatic groove of PSMA. The small-sized TM has favorable pharmacokinetic and pharmacodynamics properties (short half-life of < 30 min; rapid internalization in < 1h), which allows a fast silencing and excellent controllability of UniCAR-T reactivity (< 4 h). Furthermore, the small molecule TM efficiently penetrates and rapidly accumulates within solid tumors as clinically demonstrated by related radiotracers. Results from our pre-clinical in vivo model demonstrate potent recruitment of UniCAR-T into tumor tissue by the TM and an efficient anti-tumor response. Of note, it was recently shown that the chosen PCa model suppresses CAR-T response by secreting high levels of transforming growth factor beta (TGF-β), creating an immunosuppressive milieu. We could overcome immunosuppression utilizing CD28 compared to 4-1-BB in the intracellular signaling domain of the UniCAR. Administered CD28/zeta UniCAR efficiently suppressed tumor growth in our in vivo model and could be re-activated by consecutive cycles of TM administration even after several weeks. In contrast to CARs with fixed binding moieties, which continuously signal and thereby induce T- cell exhaustion, the discontinuous activation of UniCAR keeps a balanced mix of T-effector and T-memory cells. In