Description

Structure guided DMF5 TCR mutations can alter antigen specificity, off target cross reactivity, and polyfunctional phenotypes of T cells

Kendra Foley¹, Lance Hellman², Timothy Spear¹, David Murray¹, Brian Baker², Michael I. Nishimura¹

¹Loyola University, Department of Surgery, Cardinal Bernardin Cancer Center, Maywood, IL, USA

²University of Notre Dame, Department of Chemistry & Biochemistry, Notre Dame, IN, USA

TCR gene modified T cells in adoptive cell transfer have been shown to mediate objective clinical responses in treating melanoma and other malignancies. Despite this success, there remains the potential for cross reactivity and unanticipated off-target reactivity which could lead to autoimmunity. One approach to improve specific antigen reactivity is to modify the TCR in positive and negative structure guided mutations. This strategy includes mutations that will alter the TCR/pMHC to enhance antigen specificity, while additional mutations will alter the TCR/pMHC contact residues to reduce TCR binding with the MHC. To study these TCR mutations, our lab uses the HLA-A2 restricted DMF5 TCR which is reactive against the melanoma antigen MART-1. Five structure guided mutant DMF5 variants were constructed based on the crystal structure of the pMHC/TCR interaction. To determine the effects of these mutations on T cell function, a panel of naturally occurring MART-1 homolog peptides were generated and used to measure lysis and multi-cytokine production of transduced T cells. Our results revealed that these mutations can alter polyfunctionality and cytokine production patterns to not only the MART-1 peptide, but can also alter cross reactivity against the MART-1 homologs. These data can help us understand how the structure and affinity of the TCR/pMHC interaction can influence T cell function and cross reactivity to result in better TCRs to be used in engineered T cells for adoptive cell transfer.