Biology subjects

Izraelson, M.

Publications and source records attributed to Izraelson, M..

5 recordsLinked to original sources

bioRxiv2025

Molecule template estimation using Validator Barcodes in multiplex PCR for adaptive immune repertoire profiling

TCR- and BCR-sequencing (TCR/BCR-seq) are two important technologies in studying the immune repertoire of samples such as PBMCs or tumors. In their most common form, these assays combine multiplex PCR of the repertoire using primers targeting regions of the V(D)J and the constant region with next-generation sequencing (NGS). The data produced by this assay provide a slew of information regarding immune repertoire(s) including the presence critical clonotypes, repertoire diversity, variable (V) gene usage, analysis of public clonotypes, etc. One issue that can arise during generation of the TCR/BCR-seq data is sequence bias during the PCR or NGS steps. To combat this, unique molecular identifiers (UMIs) have been used to identify and eliminate sequence bias. However, UMI fragments can be long and very diverse, resulting in the UMI sequences interfering with any of the multitude of primers during multiplex PCR. Here, we introduce Validator Barcodes (VBCs), a set of eight short barcodes (6-9 nucleotides in length). This compact set of barcodes improves PCR efficiency and facilitates PCR primer designs. Also, like UMIs, the VBCs may be used to estimate the number of template molecules (RNA or DNA). Using VBC-labeled primers for TCR and BCR repertoire profiling from PBMCs produces highly comparable results and similarly template values to those obtained through UMI-based assay counts. Overall, VBCs are a useful and simpler alternative to UMIs in assaying TCR and BCR repertoires.

immunology↗

bioRxiv2024

Tracking antigen-specific T cell response to cancer immunotherapy

In each human or model animal, T cell receptor (TCR) of each effector/memory T cell clone recognizes from one to several cognate peptide-MHC complexes (pMHC). Limited knowledge on TCR repertoire specificities restricts our capacity to rationally interpret this information, both diagnostically and in preclinical research. Here we 1) develop cost-efficient wet lab and computational pipeline to identify mouse TCRs specific to particular peptides, 2) produce dataset of helper T cells (Th) TCR beta CDR3s specific to B16 melanoma neoantigens in the I-Ab pMHCII context, available in VDJdb, and 3) apply this dataset to track tumor-specific T cell response to the CTLA4 blocking immunotherapy on orthotopic B16 melanoma model. We show that tumor promotes clonally independent Tregs carrying tumor-specific TCRs characteristic for Th cells. We further show that CTLA4 blockade promotes Th clonal expansion and induces general, non-tumor-specific Th-to-Treg plasticity. Altogether, we provide a universal pipeline for the investigation of mouse T cell responses at the antigen-specific level, facilitating development and validation of immunotherapeutic and vaccination approaches.

immunology↗

bioRxiv2023

Robust detection of SARS-CoV-2 exposure in population using T-cell repertoire profiling

The COVID-19 pandemic offers a powerful opportunity to develop methods for monitoring the spread of infectious diseases based on their signatures in population immunity. Adaptive immune receptor repertoire sequencing (AIRR-seq) has become the method of choice for identifying T cell receptor (TCR) biomarkers encoding pathogen specificity and immunological memory. AIRR-seq can detect imprints of past and ongoing infections and facilitate the study of individual responses to SARS-CoV-2, as shown in many recent studies. Here, we have applied a machine learning approach to two large AIRR-seq datasets with more than 1,200 high-quality repertoires from healthy and COVID-19-convalescent donors to infer TCR repertoire features that were induced by SARS-CoV-2 exposure. The new batch effect correction method allowed us to use data from different batches together, as well as combine the analysis for data obtained using different protocols. Proper standardization of AIRR-seq batches, access to human leukocyte antigen (HLA) typing, and the use of both - and {beta}-chain sequences of TCRs resulted in a high-quality biomarker database and a robust and highly accurate classifier for COVID-19 exposure. This classifier is applicable to individual TCR repertoires obtained using different protocols, paving the way to AIRR-seq-based immune status assessment in large cohorts of donors.

immunology↗

bioRxiv2023

Ultrasensitive allele inference from immune repertoire sequencing data with MiXCR

Allelic variability in the adaptive immune receptor loci, which harbor the gene segments that encode B cell and T cell receptors (BCR/TCR), has been shown to be of critical importance for immune responses to pathogens and vaccines. In recent years, B cell and T cell receptor repertoire sequencing (Rep-Seq) has become widespread in immunology research making it the most readily available source of information about allelic diversity in immunoglobulin (IG) and T cell receptor (TR) loci in different populations. Here we present a novel algorithm for extra-sensitive and specific variable (V) and joining (J) gene allele inference and genotyping allowing reconstruction of individual high-quality gene segment libraries. The approach can be applied for inferring allelic variants from peripheral blood lymphocyte BCR and TCR repertoire sequencing data, including hypermutated isotype-switched BCR sequences, thus allowing high-throughput genotyping and novel allele discovery from a wide variety of existing datasets. The developed algorithm is a part of the MiXCR software (https://mixcr.com) and can be incorporated into any pipeline utilizing upstream processing with MiXCR. We demonstrate the accuracy of this approach using Rep-Seq paired with long-read genomic sequencing data, comparing it to a widely used algorithm, TIgGER. We applied the algorithm to a large set of IG heavy chain (IGH) Rep-Seq data from 450 donors of ancestrally diverse population groups, and to the largest reported full-length TCR alpha and beta chain (TRA; TRB) Rep-Seq dataset, representing 134 individuals. This allowed us to assess the genetic diversity of genes within the IGH, TRA and TRB loci in different populations and demonstrate the connection between antibody repertoire gene usage and the number of allelic variants present in the population. Finally we established a database of allelic variants of V and J genes inferred from Rep-Seq data and their population frequencies with free public access at https://vdj.online.

immunology↗

bioRxiv2018

T cell immunity does not age in a long-lived rodent species

Numerous studies have demonstrated that the percentage of naive T cells and diversity of T cell receptor (TCR) repertoire decrease with age, with some findings likewise suggesting that increased repertoire diversity may be associated with longer lifespan and healthy aging. In this work, we have analyzed peripheral TCR diversity from humans, mice, and blind mole-rats (Spalax spp.)--long-lived, hypoxia- and cancer-tolerant rodents. We employed a quantitative approach to TCR repertoire profiling based on 5RACE with unique molecular identifiers (UMI) to achieve accurate comparison of repertoire diversity, which also required development of specific wet lab protocol and TCR gene reference for Spalax. Our direct comparison reveals a striking phenomenon. Whereas TCR diversity of mice and humans decreases with age, resulting primarily from the shrinkage of the naive T cell pool, Spalax TCR diversity remains stable even for the animals that reach extreme old age (15-17 years). This indicates that T cell immunity does not meaningfully age in long-lived rodents, at least in terms of the classical understanding of immunosenescence, which is associated with the accumulation of large numbers of memory clones. We suggest that the extraordinary longevity of Spalax may be attributable at least in part to the distinctive organization of their T cell immunity. Our findings should therefore encourage a close re-examination of the contribution of immunosenescence to life span in mammals.

immunology↗