Darshit Shah

Title: HLA-peptide discovery to develop tumor-specific immunotherapies for the treatment of cancer

Affiliations: Regeneron Pharmaceuticals, 777 Old Saw Mill River Road, Tarrytown, NY 10591, USA

Abstract: The efficacy of T cell immune checkpoint inhibitors in patients provide proof-of-concept that T cells can recognize melanoma-associated antigens to kill tumor cells. However, many tumors do not respond to checkpoint inhibitors, and targeted immunotherapies such as CD3 bispecifics, CAR T or TCR T cells represent promising modalities to specifically target tumors for destruction. Peptides presented by human leukocyte antigen (HLA) molecules on the cell surface play a crucial role in adaptive immunity, mediating the communication between T cells and antigen-presenting cells. Such tumor-specific HLA-peptide complexes can potentially serve as antigens for cancer immunotherapy, especially when there are no cell surface protein targets available. In our study, we utilized the advanced capabilities of the Bruker TimsTOF Ultra mass spectrometer and optimized our Immunopeptidomics workflow utilizing a multiple myeloma cell line (IM-9) to conduct an in-depth proteogenomic profiling of HLA-presented peptides. We have detected > 6000 9mer peptides from 5E+6 cells in-depth mass spectrometry analysis. We have also successfully applied this sensitive workflow to smaller melanoma tissues of 0.1mg or lower. Our comprehensive study of melanoma tissues has led to the detection of numerous peptides derived from tumor-specific cancer testis (CT) antigen-like genes. Notably, some of these peptides have been previously identified in genes such as MAGE-A3 and MAGE-A4. Through an enhanced proteogenomic analysis of patient samples, we have further successfully identified ‘Dark Antigens’ that remained undetected using conventional open-source databases. Our optimized pipeline also facilitated the identification of peptides originating from non-coding regions, mutants, HERVs, and transposons. This discovery of ‘Dark Antigens’ paves the way for the potential development of personalized and/or shared immunotherapies for melanoma, and potentially other cancer types.

In summary, our research underscores the efficacy of comprehensive proteogenomic profiling, particularly when utilizing the high sensitivity and resolution of the TimsTOF Ultra mass spectrometer, in validating and uncovering novel antigens for cancer immunotherapy from smaller tissues sizes. This could potentially lead to the development of more personalized and effective therapies for melanoma and potentially other cancer types utilizing biopsy-sized material.

Bio: Darshit Shah is a seasoned professional in the field of Biotechnology, currently serving as a Senior Scientist at Regeneron Pharmaceutical Inc. for the past 12 years. He holds a Master’s degree from The University of Texas at San Antonio. Darshit has a strong background in establishing, optimizing, and performing HLA peptidome analysis related to various cancers and viral targets. His expertise lies in using both discovery and targeted approaches to reveal candidates for peptide-based immunotherapy.

At Regeneron, Darshit has developed quantitative assays for immunopeptidomes using Mass Spectrometry and has created workflows to analyze immunopeptidomes for cell lines, xenografts, and patient tumors. He is also proficient in designing proteomics experiments to analyze genomics profiling studies (RNA-seq) to support drug target and biomarker identification.

His innovative approach led him to develop a novel, efficient high-throughput biomarker discovery method utilizing Mass Spectrometry. He has also designed proteomics experiments to compare and contrast genomics profiling studies (RNA-seq) and has performed affinity-purification coupled to mass spectrometry (AP-MS) for ligand fishing experiments.

Darshit’s work in quantitative proteomics has been instrumental in identifying biomarkers in serum. He has also identified HLA presented peptides to design patient-based therapy. His extensive knowledge and innovative approach make him a valuable asset in the field of Biotechnology and a key contributor to the success of Regeneron Pharmaceutical Inc