Optimizing TCR avidity with Somatic Hypermutation

Introduction:

Adoptive transfer of T cells that have been genetically modified to express an anti-tumor TCR is a potent immunotherapy, but only if TCR avidity is sufficiently high. Endogenous TCRs specific to shared (self) tumor associated antigens (TAAs) have low affinity due to central tolerance. Therefore, for effective therapy, anti-TAA TCRs with higher and optimal avidity must be generated. We have developed a new in-vitro TCR avidity maturation system using somatic hypermutation (SHM) driven by Activation Induced cytidine Deaminase (AID), a mechanism used in nature by B cells for antibody optimization.

Materials and Methods:

Host cells, such as 293TREx cells, are transfected with a TCR, CD3, Tet-repressor, and AID (Tet-regulated). SHM is initiated by induction of AID expression, and cells with improved TCR avidity are selected by FACS. Mutant TCRs are transduced into primary T cells are analyzed with in-vitro and in-vivo T cell assays.

Results and discussion:

We identified 44 point mutations to the Pmel-1 TCR, specific for the H2-D^b-gp100_25-33 melanoma antigen. Primary T cells transduced with TCRs containing two or three of these mutations had enhanced activity in-vitro. Furthermore, the triple-mutant TCR improved in-vivo therapy of tumor-bearing mice, which exhibited improved survival, smaller tumors, and delayed or no relapse. We are now working on a human anti-tumor TCR using a refined system with promising preliminary results.

Conclusion:

TCR avidity matuation by SHM may be an effective strategy to improve cancer immunotherapy.