Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cells
Background: Recent advances in single-cell technologies and an improved understanding of tumor antigens have empowered researchers to investigate tumor antigen-specific CD8+ T cells at the single-cell level. Peptide-MHC I tetramers are often utilized to enrich antigen-specific CD8+ T cells, which ho...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-01-01
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| Series: | Translational Oncology |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S1936523322002182 |
| _version_ | 1811295033027985408 |
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| author | Kamalakannan Rajasekaran Xiangnan Guan Alireza Tafazzol Habib Hamidi Martine Darwish Mahesh Yadav |
| author_facet | Kamalakannan Rajasekaran Xiangnan Guan Alireza Tafazzol Habib Hamidi Martine Darwish Mahesh Yadav |
| author_sort | Kamalakannan Rajasekaran |
| collection | DOAJ |
| description | Background: Recent advances in single-cell technologies and an improved understanding of tumor antigens have empowered researchers to investigate tumor antigen-specific CD8+ T cells at the single-cell level. Peptide-MHC I tetramers are often utilized to enrich antigen-specific CD8+ T cells, which however, introduces the undesired risk of altering their clonal distribution or their transcriptional state. This study addresses the feasibility of utilizing tetramers to enrich antigen-specific CD8+ T cells for single-cell analysis. Methods: HLA-A*02:01-restricted human cytomegalovirus (CMV) pp65 peptide-specific CD8+ T cells were used as a model for analyzing antigen-specific CD8+ T cells. Single-cell RNA sequencing and TCR sequencing were performed to compare the frequency and gene expression profile of pp65-specific TCR clones between tetramer-sorted, unstimulated- and tetramer-stimulated total CD8+ T cells. Results: The relative frequency of pp65-specific TCR clones and their transcriptional profile remained largely unchanged following tetramer-based sorting. In contrast, tetramer-mediated stimulation of CD8+ T cells resulted in significant gene expression changes in pp65-specific CD8+ T cells. An Antigen-Specific Response (ASR) gene signature was derived from tetramer-stimulated pp65-specific CD8+ T cells. The ASR signature had a predictive value and was significantly associated with progression free survival in lung cancer patients treated with anti-PD-L1, anti-VEGF, chemotherapy combination (NCT02366143). The predictive power of the ASR signature was independent of the conventional CD8 effector signature. Conclusions: Our findings validate the approach of enriching antigen-specific CD8+ T cells through tetramer-aided Fluorescence-Activated Cell Sorting (FACS) sorting for single-cell analysis and also identifies an ASR gene signature that has value in predicting response to cancer immunotherapy. |
| first_indexed | 2024-04-13T05:26:27Z |
| format | Article |
| id | doaj.art-2c46d971ad4c4375b7ee0e590ee03894 |
| institution | Directory Open Access Journal |
| issn | 1936-5233 |
| language | English |
| last_indexed | 2024-04-13T05:26:27Z |
| publishDate | 2023-01-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Translational Oncology |
| spelling | doaj.art-2c46d971ad4c4375b7ee0e590ee038942022-12-22T03:00:34ZengElsevierTranslational Oncology1936-52332023-01-0127101559Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cellsKamalakannan Rajasekaran0Xiangnan Guan1Alireza Tafazzol2Habib Hamidi3Martine Darwish4Mahesh Yadav5Corresponding authors.; Genentech, 1 DNA way, South San Francisco, CA 94080, USAGenentech, 1 DNA way, South San Francisco, CA 94080, USAGenentech, 1 DNA way, South San Francisco, CA 94080, USAGenentech, 1 DNA way, South San Francisco, CA 94080, USAGenentech, 1 DNA way, South San Francisco, CA 94080, USACorresponding authors.; Genentech, 1 DNA way, South San Francisco, CA 94080, USABackground: Recent advances in single-cell technologies and an improved understanding of tumor antigens have empowered researchers to investigate tumor antigen-specific CD8+ T cells at the single-cell level. Peptide-MHC I tetramers are often utilized to enrich antigen-specific CD8+ T cells, which however, introduces the undesired risk of altering their clonal distribution or their transcriptional state. This study addresses the feasibility of utilizing tetramers to enrich antigen-specific CD8+ T cells for single-cell analysis. Methods: HLA-A*02:01-restricted human cytomegalovirus (CMV) pp65 peptide-specific CD8+ T cells were used as a model for analyzing antigen-specific CD8+ T cells. Single-cell RNA sequencing and TCR sequencing were performed to compare the frequency and gene expression profile of pp65-specific TCR clones between tetramer-sorted, unstimulated- and tetramer-stimulated total CD8+ T cells. Results: The relative frequency of pp65-specific TCR clones and their transcriptional profile remained largely unchanged following tetramer-based sorting. In contrast, tetramer-mediated stimulation of CD8+ T cells resulted in significant gene expression changes in pp65-specific CD8+ T cells. An Antigen-Specific Response (ASR) gene signature was derived from tetramer-stimulated pp65-specific CD8+ T cells. The ASR signature had a predictive value and was significantly associated with progression free survival in lung cancer patients treated with anti-PD-L1, anti-VEGF, chemotherapy combination (NCT02366143). The predictive power of the ASR signature was independent of the conventional CD8 effector signature. Conclusions: Our findings validate the approach of enriching antigen-specific CD8+ T cells through tetramer-aided Fluorescence-Activated Cell Sorting (FACS) sorting for single-cell analysis and also identifies an ASR gene signature that has value in predicting response to cancer immunotherapy.http://www.sciencedirect.com/science/article/pii/S1936523322002182CMV pp65-specific CD8+ T cellspMHC TetramersFACS sortingSingle-cell RNA sequencingTCR analysisAntigen-specific response gene signature |
| spellingShingle | Kamalakannan Rajasekaran Xiangnan Guan Alireza Tafazzol Habib Hamidi Martine Darwish Mahesh Yadav Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cells Translational Oncology CMV pp65-specific CD8+ T cells pMHC Tetramers FACS sorting Single-cell RNA sequencing TCR analysis Antigen-specific response gene signature |
| title | Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cells |
| title_full | Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cells |
| title_fullStr | Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cells |
| title_full_unstemmed | Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cells |
| title_short | Tetramer-aided sorting and single-cell RNA sequencing facilitate transcriptional profiling of antigen-specific CD8+ T cells |
| title_sort | tetramer aided sorting and single cell rna sequencing facilitate transcriptional profiling of antigen specific cd8 t cells |
| topic | CMV pp65-specific CD8+ T cells pMHC Tetramers FACS sorting Single-cell RNA sequencing TCR analysis Antigen-specific response gene signature |
| url | http://www.sciencedirect.com/science/article/pii/S1936523322002182 |
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