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4094 Structural Determinants of Immunogenicity for Peptide-Based Immunotherapy
- Jason Devlin, Jesus Alonso, Grant Keller, Sara Bobisse, Alexandre Harari, Brian Baker
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- Journal:
- Journal of Clinical and Translational Science / Volume 4 / Issue s1 / June 2020
- Published online by Cambridge University Press:
- 29 July 2020, p. 16
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OBJECTIVES/GOALS: Neoantigen vaccine immunotherapies have shown promise in clinical trials, but identifying which peptides to include in a vaccine remains a challenge. We aim to establish that molecular structural features can help predict which neoantigens to target to achieve tumor regression. METHODS/STUDY POPULATION: Proteins were prepared by recombinant expression in E. coli followed by in vitro refolding. Correctly folded proteins were purified by chromatography. Affinities of protein-protein interactions were measured by surface plasmon resonance (SPR) and thermal stabilities of proteins were determined by differential scanning fluorimetry. All experiments were performed at least in triplicate. Protein crystals were obtained by hanging drop vapor diffusion. The protein crystal structures were solved by molecular replacement and underwent several rounds of automated refinement. Molecular dynamics simulations were performed using the AMBER molecular dynamics package. RESULTS/ANTICIPATED RESULTS: A T cell receptor (TCR) expressed by tumor-infiltrating T cells exhibited a 20-fold stronger binding affinity to the neoantigen peptide compared to the self-peptide. X-ray crystal structures of the peptides with the major histocompatibility complex (MHC) protein demonstrated that a non-mutated residue in the peptide samples different positions with the mutation. The difference in conformations of the non-mutated residue was supported by molecular dynamics simulations. Crystal structures of the TCR engaging both peptide/MHCs suggested that the conformation favored by the mutant peptide was crucial for TCR binding. The TCR bound the neoantigen/MHC with faster binding kinetics. DISCUSSION/SIGNIFICANCE OF IMPACT: Our results suggest that the mutation impacts the conformation of another residue in the peptide, and this alteration allows for more favorable T cell receptor binding to the neoantigen. This highlights the potential of non-mutated residues in contributing to neoantigen recognition.
3348 Structural Determinants of Neoantigen Immunogenicity for Cancer Therapy
- Jason Devlin, Sara Bobisse, Alexandre Harari, Brian Baker
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- Journal:
- Journal of Clinical and Translational Science / Volume 3 / Issue s1 / March 2019
- Published online by Cambridge University Press:
- 26 March 2019, p. 22
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OBJECTIVES/SPECIFIC AIMS: We are exploring the structure of the interaction between an immunogenic neoantigen and a T cell receptor (TCR) that recognizes the neoantigen while tolerating the counterpart self antigen. No structural example exists to date of how a TCR can discriminate between a neoantigen and the self antigen. We aim to determine the structural and biophysical features that underlie the immunogenicity for this neoantigen, and the features we determine are likely to be present in other immunogenic neoantigens. Algorithms to predict the immunogenicity of neoantigens are available, but do not incorporate structural or biophysical factors. We aim to improve these methods for immunogenic neoantigen prediction by determining structural and biophysical factors that result in recognition by the immune system. METHODS/STUDY POPULATION: Recombinant protein expression, production, and purification. Protein x-ray crystallography. Biophysical protein-protein binding experiments RESULTS/ANTICIPATED RESULTS: The T cell receptor (TCR) bound to the neoantigen with an affinity 15-fold higher than the self antigen. The leucine to phenylalanine mutation occurs at position 8 of a 9-amino acid long peptide antigen. This position is typically in the interface bound by the T cell receptor. The structures of the unbound neoantigen and self antigen showed that the mutated residue was in the TCR interface. Additionally we noted a change in the side chain position of a proximal tryptophan, potentially due to clashes with the larger phenylalanine residue. The structure of the TCR bound to the neoantigen showed that the TCR interacted with the tryptophan in the mutation-induced conformation and with the phenylalanine residue. Thus the mutation may be altering TCR binding affinity by interactions of the residue itself with the TCR, and by locking the proximal tryptophan residue in an optimal position to interact with the TCR. We are testing the contributions of each of these factors to the overall affinity change. Hydrophobicity has been linked to immunogenicity, so mutations that increase hydrophobicity compared to the self antigen are likely to be immunogenic. However, leucine and phenylalanine are similar on hydrophobicity scales. On the other hand, a side chain rotation is unlikely to represent a large energy barrier. Therefore, we hypothesize that another property of the phenylalanine, such as size or aromaticity, is driving the affinity difference. DISCUSSION/SIGNIFICANCE OF IMPACT: Traditional forms of cancer therapy do not specifically target cancer cells, and their toxicity to healthy cells limits their effectiveness. Immunotherapy, which involves orchestrating a specific anti-cancer immune response, is now an established cancer therapy. Several forms of immunotherapy target “neoantigens,” which are derived from mutated proteins in cancer, and are therefore are cancer-specific. Neoantigens represent a foothold that can allow the immune system to distinguish between cancer cells and healthy cells, and thus specifically target cancer cells for destruction while imparting no activity toward healthy cells that lack the neoantigen. Most cancer mutations that result in neoantigens arise from random passenger mutations in cancer and will be different among patients. Neoantigen-based cancer therapies are thus a precision medicine technique. The quality of neoantigens to induce an immune response (immunogenicity), which relates to how likely they are to be presented to the immune system and recognized as foreign, has been shown to be a critical factor in predicting the outcome of immunotherapy treatment. We are investigating, on a structural and biophysical level, features that may increase the likelihood of a neoantigen being recognized as foreign by the immune system. The structural insight we gain can be incorporated into algorithms that predict neoantigens from cancer exome sequencing for patient-specific identification of immunogenic neoantigens for immunotherapeutic intervention.
Yehuda Elkana (1934–2012)
- Leo Corry, Moritz Epple, Orna Harari, Alexandre Métraux, Jürgen Renn
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- Journal:
- Science in Context / Volume 26 / Issue 1 / March 2013
- Published online by Cambridge University Press:
- 11 February 2013, pp. 1-2
- Print publication:
- March 2013
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We mourn the loss of Yehuda Elkana, founding editor of this journal. Setting science in context was a mission of his life. For him this did not mean to relativize and historicize science to the point where it is no longer distinguishable as central to the human quest for knowledge. Rather, an understanding of science as being rooted in social, material, and cultural contexts was for him the key to its central role for solving the problems of humanity with which he was so deeply concerned.