Single-Cell TCR Repertoire Profiling for Paired-Chain Identification
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Solution Overview
Problem
Existing methods fail to provide personalized immunotherapy for cancer patients by matching subject-specific T cell receptors with tumor neoantigens, necessitating a method to profile and utilize T cell receptor repertoires for targeted treatments.
Innovation Solution
A method involving single-cell bar-coded droplets (SCBD) technology to profile T cell receptor repertoires, identify subject-specific TCRs, and match them with tumor neoantigens, followed by cloning and expressing these receptors in T cells for personalized immunotherapy.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional TCR sequencing methods are used, then TCR sequences can be obtained, but paired TCR chains from single cells cannot be determined
Solution Approach 1:
The patent segments the TCR sequencing problem into single-cell isolation, single-cell genomic DNA extraction, and paired-chain amplification steps. By isolating individual T cells and processing their DNA separately, the method enables determination of paired TCR chains from the same cell, resolving the limitation of conventional bulk sequencing methods that cannot associate alpha and beta chains from the same T cell.
Solution Approach 2:
The patent uses PCR-based amplification with specific primers as an intermediary to amplify and identify paired TCR chains from single-cell genomic DNA. The TCR-SCAN method employs primers that specifically amplify TCR alpha and beta chain genes from single-cell DNA, enabling the determination of which chains are paired together in the same T cell receptor complex.
2Productivity
If bulk TCR sequencing is performed, then large numbers of TCR sequences can be obtained, but subject-specific paired TCR chains cannot be identified
Solution Approach 1:
The patent segments the TCR analysis into bulk sequencing for high throughput and single-cell analysis for paired-chain identification. By processing single-cell genomic DNA through PCR amplification with TCR-specific primers, the method identifies which TCR chains are paired together while maintaining the ability to analyze multiple samples.
Solution Approach 2:
The patent performs preliminary isolation and characterization of TCR sequences from single-cell genomic DNA before applying these sequences to identify subject-specific T cells. The TCR-SCAN method pre-identifies the paired TCR chains from single cells, which then can be used to guide subsequent enrichment and therapy development for subject-specific treatments.
3Adaptability or versatility
If personalized TCR therapy is developed, then subject-specific cancer treatment can be provided, but the method to identify and match TCRs with neoantigens is not established
Solution Approach 1:
The patent performs preliminary identification and characterization of subject-specific TCR sequences from single-cell genomic DNA before matching them with tumor neoantigens. The TCR-SCAN method pre-identifies the TCR repertoire of the subject, including paired alpha and beta chains, which then can be screened for reactivity against subject-specific tumor antigens to develop personalized therapies.
Solution Approach 2:
The patent employs a feedback loop where identified TCR sequences are screened for reactivity against tumor cells or neoantigens, and this reactivity information feeds back into the selection and optimization of TCR candidates for therapy. The method iteratively refines the TCR selection based on functional assays that test recognition of subject-specific tumor antigens.
Data Source
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AI summary
The present disclosure relates to methods for profiling subject specific and personalized T cell receptor (TCR) repertoires using a single-cell sequencing method. More particularly, disclosed are methods for determining binding of T cell receptors to subject specific neoantigens. In addition, the techniques herein may identify the antigenic targets of T cell receptors in the context of tumor neoantigens. Moreover, the present disclosure enables the discovery of T cell targets in numerous diseases, with implications for understanding the basic mechanisms of the mammalian immune response and for developing antigen-specific diagnostic markers and therapies. Finally, cloned TCRs can be used to formulate personalized immunotherapies for those inflicted with a disease, such as cancer.