Friday, May 17, 2024
5/17/2024
Project Summary: Despite the remarkable success in the treatment of some hematological cancers, the risk of serious adverse events and loss of potency remain major problems that threaten to curtail widespread application of chimeric antigen receptor (CAR) T-cell therapies to other cancer types including solid tumors. These problems, all of which are influenced by the binding affinity of the CAR targeting domain, result from the propensity for unpredictable hyperactivation that can lead to potentially fatal systemic toxicities such as cytokine release syndrome (CRS), tumor lysis syndrome (TLS) and neurotoxicity. Furthermore, on-target off-tumor toxicities can result from the indiscriminate binding of constitutively expressed CARs to tumor antigens on healthy tissues, whilst disease relapses can occur due to antigen escape and/or T cell exhaustion. Current and emerging CAR- T cell strategies and designs to address these problems are often too slow in terms of response times and onset of action. Furthermore, they prematurely and permanently eliminate the “high value” cells and often require the expression of additional proteins which may constrain the DNA payload capacity of current lenti- and retro- virus vectors. There is thus an urgent unmet need for in-vivo regulatable CAR-T cell platforms that are rapid, reversible and don’t exert a substantial DNA burden on the CAR delivery vector. The objective of this proposal is to assess the feasibility to regulate the activation, effector functions and cytokine release of CAR T-cells using an exogenously administered ligand that modulates the affinity of the CAR T-cell targeting domain. To this end we will utilize our proprietary universal allosteric-linker and switch module for antibodies (UNASMA) technology to incorporate an ON/OFF affinity switch into a well characterized antibody single chain variable fragment (scFv) against a clinically validated hematological cancer target which will subsequently be formatted as a switchable affinity chimeric antigen receptor (SaefCAR). We postulate that the resulting SaefCAR following transduction into a T-cell, will enable the in-vivo regulation of the SaefCAR T-cells’ activity via exogenous administration of a small molecule or peptide ligand, potentially solving several of the problems of current CAR-T cell therapies by: 1) Ameliorating intrinsic toxicities via enablement of a slow start to T-cell activation, 2) Preventing on-target off- tumor toxicities by switching off the CAR-T cell without eliminating it 3) Preventing exhaustion of tumor-specific CAR T cells caused by tonic signaling and other mechanisms via intermittent ON/OFF switching of the CAR. Specifically in Aim 1 we will develop switchable forms of the scFv which will subsequently be formatted into a CAR and transduced into a T-cell in Aim 2. In Aim 3 we will validate proof-of-concept of the regulated SaefCAR T-cells in in-vitro and in-vivo studies. We aim to demonstrate that a ligand-controllable affinity switch in the binding domain of SaefCAR T-cells can regulate it’s activation and effector functions against a tumor. Importantly due to the universal character of our approach, it provides a blueprint to improve any scFv-based CAR T-cell therapy in the future by adding a capability to regulate the CAR affinity for the tumor antigen.
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