Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selection
Abstract Objective Adoptive immunotherapy with ex vivo expanded tumor‐specific T cells has potential as anticancer therapy. Preferentially expressed antigen in melanoma (PRAME) is an attractive target overexpressed in several cancers including melanoma and acute myeloid leukaemia (AML), with low exp...
| Main Authors: | , , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Wiley
2020-01-01
|
| Series: | Clinical & Translational Immunology |
| Subjects: | |
| Online Access: | https://doi.org/10.1002/cti2.1200 |
| _version_ | 1818300180945960960 |
|---|---|
| author | Koon H Lee Kavitha Gowrishankar Janine Street Helen M McGuire Fabio Luciani Brendan Hughes Mandeep Singh Leighton E Clancy David J Gottlieb Kenneth P Micklethwaite Emily Blyth |
| author_facet | Koon H Lee Kavitha Gowrishankar Janine Street Helen M McGuire Fabio Luciani Brendan Hughes Mandeep Singh Leighton E Clancy David J Gottlieb Kenneth P Micklethwaite Emily Blyth |
| author_sort | Koon H Lee |
| collection | DOAJ |
| description | Abstract Objective Adoptive immunotherapy with ex vivo expanded tumor‐specific T cells has potential as anticancer therapy. Preferentially expressed antigen in melanoma (PRAME) is an attractive target overexpressed in several cancers including melanoma and acute myeloid leukaemia (AML), with low expression in normal tissue outside the gonads. We developed a GMP‐compliant manufacturing method for PRAME‐specific T cells from healthy donors for adoptive immunotherapy. Methods Mononuclear cells were pulsed with PRAME 15‐mer overlapping peptide mix. After 16 h, activated cells expressing CD137 were isolated with immunomagnetic beads and cocultured with irradiated CD137neg fraction in medium supplemented with interleukin (IL)‐2, IL‐7 and IL‐15. Cultured T cells were restimulated with antigen‐pulsed autologous cells after 10 days. Cellular phenotype and cytokine response following antigen re‐exposure were assessed with flow cytometry, enzyme‐linked immunospot (ELISPOT) and supernatant cytokine detection. Detailed phenotypic and functional analysis with mass cytometry and T‐cell receptor (TCR) beta clonality studies were performed on selected cultures. Results PRAME‐stimulated cultures (n = 10) had mean expansion of 2500‐fold at day 18. Mean CD3+ percentage was 96% with CD4:CD8 ratio of 4:1. Re‐exposure to PRAME peptide mixture showed enrichment of CD4 cells expressing interferon (IFN)‐γ (mean: 12.2%) and TNF‐α (mean: 19.7%). Central and effector memory cells were 23% and 72%, respectively, with 24% T cells expressing PD1. Mass cytometry showed predominance of Th1 phenotype (CXCR3+/CCR4neg/CCR6neg/Tbet+, mean: 73%) and cytokine production including IL‐2, IL‐4, IL‐8, IL‐13 and GM‐CSF (2%, 6%, 8%, 4% and 11%, respectively). Conclusion PRAME‐specific T cells for adoptive immunotherapy were enriched from healthy donor mononuclear cells. The products were oligoclonal, exhibited Th1 phenotype and produced multiple cytokines. |
| first_indexed | 2024-12-13T05:03:02Z |
| format | Article |
| id | doaj.art-c5ccad7e20d64a35b9c22b710d0f04af |
| institution | Directory Open Access Journal |
| issn | 2050-0068 |
| language | English |
| last_indexed | 2024-12-13T05:03:02Z |
| publishDate | 2020-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Clinical & Translational Immunology |
| spelling | doaj.art-c5ccad7e20d64a35b9c22b710d0f04af2022-12-21T23:58:44ZengWileyClinical & Translational Immunology2050-00682020-01-01910n/an/a10.1002/cti2.1200Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selectionKoon H Lee0Kavitha Gowrishankar1Janine Street2Helen M McGuire3Fabio Luciani4Brendan Hughes5Mandeep Singh6Leighton E Clancy7David J Gottlieb8Kenneth P Micklethwaite9Emily Blyth10Westmead Institute for Medical Research Westmead NSW AustraliaWestmead Institute for Medical Research Westmead NSW AustraliaWestmead Institute for Medical Research Westmead NSW AustraliaFaculty of Medicine and Health Sydney Medical School Sydney NSW AustraliaThe Kirby Institute University of New South Wales Darlinghurst NSW AustraliaThe Kirby Institute University of New South Wales Darlinghurst NSW AustraliaThe Garvan Institute of Medical Research Darlinghurst NSW AustraliaWestmead Institute for Medical Research Westmead NSW AustraliaWestmead Institute for Medical Research Westmead NSW AustraliaWestmead Institute for Medical Research Westmead NSW AustraliaWestmead Institute for Medical Research Westmead NSW AustraliaAbstract Objective Adoptive immunotherapy with ex vivo expanded tumor‐specific T cells has potential as anticancer therapy. Preferentially expressed antigen in melanoma (PRAME) is an attractive target overexpressed in several cancers including melanoma and acute myeloid leukaemia (AML), with low expression in normal tissue outside the gonads. We developed a GMP‐compliant manufacturing method for PRAME‐specific T cells from healthy donors for adoptive immunotherapy. Methods Mononuclear cells were pulsed with PRAME 15‐mer overlapping peptide mix. After 16 h, activated cells expressing CD137 were isolated with immunomagnetic beads and cocultured with irradiated CD137neg fraction in medium supplemented with interleukin (IL)‐2, IL‐7 and IL‐15. Cultured T cells were restimulated with antigen‐pulsed autologous cells after 10 days. Cellular phenotype and cytokine response following antigen re‐exposure were assessed with flow cytometry, enzyme‐linked immunospot (ELISPOT) and supernatant cytokine detection. Detailed phenotypic and functional analysis with mass cytometry and T‐cell receptor (TCR) beta clonality studies were performed on selected cultures. Results PRAME‐stimulated cultures (n = 10) had mean expansion of 2500‐fold at day 18. Mean CD3+ percentage was 96% with CD4:CD8 ratio of 4:1. Re‐exposure to PRAME peptide mixture showed enrichment of CD4 cells expressing interferon (IFN)‐γ (mean: 12.2%) and TNF‐α (mean: 19.7%). Central and effector memory cells were 23% and 72%, respectively, with 24% T cells expressing PD1. Mass cytometry showed predominance of Th1 phenotype (CXCR3+/CCR4neg/CCR6neg/Tbet+, mean: 73%) and cytokine production including IL‐2, IL‐4, IL‐8, IL‐13 and GM‐CSF (2%, 6%, 8%, 4% and 11%, respectively). Conclusion PRAME‐specific T cells for adoptive immunotherapy were enriched from healthy donor mononuclear cells. The products were oligoclonal, exhibited Th1 phenotype and produced multiple cytokines.https://doi.org/10.1002/cti2.1200adoptive immunotherapyantigen‐specific T cellsPRAMEpreferential expressed antigen in melanoma |
| spellingShingle | Koon H Lee Kavitha Gowrishankar Janine Street Helen M McGuire Fabio Luciani Brendan Hughes Mandeep Singh Leighton E Clancy David J Gottlieb Kenneth P Micklethwaite Emily Blyth Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selection Clinical & Translational Immunology adoptive immunotherapy antigen‐specific T cells PRAME preferential expressed antigen in melanoma |
| title | Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selection |
| title_full | Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selection |
| title_fullStr | Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selection |
| title_full_unstemmed | Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selection |
| title_short | Ex vivo enrichment of PRAME antigen‐specific T cells for adoptive immunotherapy using CD137 activation marker selection |
| title_sort | ex vivo enrichment of prame antigen specific t cells for adoptive immunotherapy using cd137 activation marker selection |
| topic | adoptive immunotherapy antigen‐specific T cells PRAME preferential expressed antigen in melanoma |
| url | https://doi.org/10.1002/cti2.1200 |
| work_keys_str_mv | AT koonhlee exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT kavithagowrishankar exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT janinestreet exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT helenmmcguire exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT fabioluciani exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT brendanhughes exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT mandeepsingh exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT leightoneclancy exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT davidjgottlieb exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT kennethpmicklethwaite exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection AT emilyblyth exvivoenrichmentofprameantigenspecifictcellsforadoptiveimmunotherapyusingcd137activationmarkerselection |