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Integrating T Cell Receptor Beta Chain Profiling Enhances Neoantigen Prediction and Prioritization in Colorectal Cancer

Core Concepts
Integrating T cell receptor beta chain (TCRβ) profiling data with peptide-HLA binding prediction improves the identification and prioritization of immunogenic neoantigens for personalized cancer immunotherapy.
The study aimed to develop a novel workflow that integrates T cell receptor beta chain (TCRβ) sequencing data with peptide-HLA (pHLA) binding prediction to enhance the identification and prioritization of immunogenic neoantigens for personalized cancer immunotherapy. Key highlights: TCRβ sequencing of tumor-infiltrating lymphocytes (TILs) from 27 colorectal cancer (CRC) patients revealed substantial intra-tumor and inter-patient heterogeneity in the TCRβ repertoire, likely due to the stochastic utilization of V and J segments in response to neoantigens. Analysis of publicly available datasets showed that both pHLA binding and pHLA-TCR binding strength are important determinants of neoantigen immunogenicity. Integrating these two features in a combined machine learning model improved the sensitivity and specificity for identifying immunogenic neoantigens compared to using either feature alone. Experimental validation using ELISpot assays on peripheral blood mononuclear cells (PBMCs) from 4 CRC patients confirmed that the combined model outperformed the conventional pHLA-based approach in selecting immunogenic neoantigen candidates. The study identified several novel neoantigen candidates with confirmed immunogenicity that were not previously reported in public databases, highlighting the potential of this integrated approach to uncover effective targets for personalized cancer immunotherapy.
The TCRβ sequencing analysis yielded an average of 3,066,197 productive TCR reads per sample, with a range between 256,035 and 10,888,726. Across the 27 patients, the number of TCRβ-CDR3 clonotypes ranged from 433 to 27,749, indicating substantial intra-tumor heterogeneity. MSI-H tumors displayed a significantly lower number of TCRβ clonotypes compared to MSS tumors (2185 versus 6070, p=0.047).
"The recognition of the neoantigen-HLA complex through TCR is of paramount importance for T cell activation and eliciting an immune response." "Sequencing the T cell receptors (TCRs) of TILs or lymphocytes found in peripheral blood provides crucial insights into the T-cell repertoire and their responses against neoantigens associated with tumors." "The integration of both pHLA binding and pHLA-TCR binding strength features in our approach exhibited superior performance in neoantigen selection and prioritization when compared to the single-feature method."

Deeper Inquiries

How can the insights from TCRβ profiling be leveraged to develop personalized TCR-based therapies for CRC patients

The insights gained from TCRβ profiling in CRC patients can be instrumental in developing personalized TCR-based therapies. By analyzing the TCR repertoire of TILs, researchers can identify specific TCR sequences that are reactive to neoantigens present in the tumor. These TCR sequences can then be isolated, engineered, and utilized in TCR-based therapies such as TCR-engineered T cell therapy. One approach is to isolate the TCR sequences that show reactivity to neoantigens and introduce them into patient-derived T cells. These engineered T cells can then be expanded in vitro and reintroduced into the patient to target and destroy cancer cells expressing the specific neoantigens. This personalized approach ensures that the T cells used in therapy are specifically tailored to recognize and attack the patient's unique tumor antigens, potentially leading to more effective and targeted treatment outcomes. Furthermore, the TCRβ profiling data can also be used to identify common TCR sequences that are shared among patients with similar neoantigen profiles. This information can guide the development of off-the-shelf TCR-based therapies that target commonly expressed neoantigens in a subset of CRC patients, providing a more standardized treatment option for a specific patient population.

What are the potential limitations of using PBMCs instead of TILs for the experimental validation of neoantigen candidates, and how can this be addressed in future studies

Using PBMCs instead of TILs for experimental validation of neoantigen candidates may have limitations due to differences in the TCR repertoire and activation status between these cell types. TILs are T cells that have infiltrated the tumor microenvironment and are likely to have been exposed to tumor antigens, leading to a more specific and activated TCR repertoire compared to peripheral blood T cells. One potential limitation is that PBMCs may not accurately represent the TCR diversity and reactivity present in TILs, leading to discrepancies in the validation of neoantigen candidates. To address this limitation in future studies, researchers can consider isolating TILs directly from tumor tissues to validate neoantigens. This approach ensures that the T cells used in the validation process closely resemble the T cells that are actively engaging with tumor antigens in the tumor microenvironment. Additionally, researchers can explore methods to activate and expand TILs ex vivo to enhance their reactivity and specificity before using them in experimental validation assays. By priming TILs to recognize specific neoantigens and boosting their activation status, researchers can improve the accuracy and reliability of neoantigen validation using TILs.

Given the observed differences in TCRβ repertoire between MSI-H and MSS CRC subtypes, how can this information be utilized to design more effective immunotherapeutic strategies for each subtype

The differences observed in the TCRβ repertoire between MSI-H and MSS CRC subtypes provide valuable insights that can be leveraged to design more effective immunotherapeutic strategies for each subtype. For MSI-H CRC, which is characterized by high mutation and neoantigen burdens, the reduced TCR clonotypes and diversity observed in these patients suggest a potential enrichment of neoantigen-reactive TCR clonotypes. To design more effective immunotherapies for MSI-H CRC, researchers can focus on identifying and targeting these enriched TCR clonotypes that are specific to neoantigens present in MSI-H tumors. By developing personalized TCR-based therapies that harness the unique TCR repertoire of MSI-H patients, it may be possible to enhance the anti-tumor immune response and improve treatment outcomes. For MSS CRC, which may have lower mutation and neoantigen burdens, the diverse TCR repertoire observed indicates a broader range of TCR clonotypes that may not be specifically reactive to tumor antigens. In this case, researchers can explore strategies to enhance TCR reactivity and specificity towards tumor antigens in MSS CRC patients. This could involve TCR engineering or combination therapies that boost the immune response against tumor cells in MSS CRC patients, potentially overcoming the challenges posed by lower neoantigen burdens in this subtype.