Project Summary:
Adoptive cellular immunotherapy utilizing T cells engineered with chimeric antigen receptor (CAR) has shown
durable clinical responses in hematologic malignancies. However, relapse remains a challenge, urging for better
CAR designs. CAR, analogous to the T cell receptor (TCR), is specifically engineered to redirect T cell specificity
towards tumor antigens. Optimal regulation on TCR dynamics is crucial for proper TCR function. Altered
dynamics of TCR leads to disfunction or even immune disorders. While the importance of control TCR dynamics
has been well established, there is a notable lack of comprehensive studies focusing specifically on CAR
dynamics, including endocytosis, recycling/degradation, and its impact on CAR-T function. My previous work
demonstrated that modulating molecular dynamics of CAR greatly improves CAR-T function. By utilizing the
endocytic feature of the cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) cytoplasmic tail (CCT), I
reprogrammed CAR dynamics and substantially enhanced CAR-T efficacy in vivo. To control CAR dynamics
more precisely and elucidate its impact on CAR-T function, further modulation on CCT fusion and investigation
on cell-intrinsic regulators to control CAR dynamics are needed. In Aim 1 of this K99/R00 application, I propose
to identify the optimal position and functional motif of CCT fusion. Completion of this aim will not only develop a
novel CAR engineering approach but also provide a deeper understanding of how altering CAR dynamics
contributes to improved CAR-T function. In Aim 2, I plan to uncover cell-intrinsic regulator of CAR dynamics
using CRISPR-based gene perturbation. This aim will serve to fill the gap in our understanding of the improved
anti-tumor efficacy of CAR-CCT but also uncover novel regulators on CTLA-4 dynamics. In Aim 3, I propose to
develop self-regulated CAR with CCT fusion in response to antigen stimulation. This novel engineer approach
will enable precise control over the timing and duration of CAR signaling, which is crucial for maintaining CAR-
T efficacy with limited toxicity. All aims center on engineering CAR dynamics with CCT fusion to develop novel
immunotherapies. The innovative insights and versatile engineering approach proposed in this study will also
contribute significantly to addressing the existing knowledge gap in CAR engineering. This grant will facilitate my
career development during both the postdoctoral training phase and transition phase as an independent faculty
and group leader.