Mapping and removal of T-cell epitopes in the PE38 portion of immunotoxins

Monday, October 24, 2011 — Poster Session I
Noon – 2:00 p.m.	Natcher Conference Center	NCI	CANCER-7

Authors

• J Eberle
• R Mazor
• A Vassall
• R Beers
• I Pastan

Abstract

Recombinant immunotoxins (RITs) are genetically engineered proteins that target cancer cells. They contain the variable region of a Mab that reacts with a cancer cell fused to a bacterial toxin. We make RITs that employ a 38kDa fragment of Pseudomonas exotoxin A (PE38), which eventually elicits an immune response in patients due to its bacterial origin, thereby limiting treatment cycles. To improve the usefulness of RITs we aim to eliminate immunodominant T cell epitopes for increased efficacy. We developed an IL2 ELISpot assay to identify T cell epitopes in which mononuclear cells are first incubated with immunotoxin and then stimulated with peptides derived from PE38. We identified one dominant T cell epitope and used alanine-scanning mutagenisis to identify key residues in the peptide. We then prepared a mutant immunotoxin, which retains full cytotoxic activity, but no longer stimulates a T cell response. We are now testing this mutant to see if its ability to induce antibodies is also diminished. We conclude that the PE38 RITs can be made less immunogenic, and therefore more efficacious, through single amino acid mutations. This method of screening toxins to identify and remove immunogenic regions should make the clinical use of RITs more successful.

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