Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers

Abstract Background Adoptive transfer of chimeric antigen receptor (CAR)-engineered T cells combined with checkpoint inhibition may prevent T cell exhaustion and improve clinical outcomes. However, the approach is limited by cumulative costs and toxicities. Methods To overcome this drawback, we crea...

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Autores principales:	Zhifen Yang, Lingyu Li, Ahu Turkoz, Pohan Chen, Rona Harari-Steinfeld, Maggie Bobbin, Ofir Stefanson, Hana Choi, Violena Pietrobon, Bennett Alphson, Angshumala Goswami, Vitaly Balan, Alper Kearney, Dharmesh Patel, Jin Yang, Damla Inel, Veena Vinod, Alessandra Cesano, Bing Wang, Kyung-Ho Roh, Lei S. Qi, Francesco M. Marincola
Formato:	article
Lenguaje:	EN
Publicado:	BMC 2021
Materias:	Medicine R
Acceso en línea:	https://doaj.org/article/fb0228a28866446a85e61c3402715418
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spelling	oai:doaj.org-article:fb0228a28866446a85e61c34027154182021-11-14T12:08:16ZContextual reprogramming of CAR-T cells for treatment of HER2+ cancers10.1186/s12967-021-03132-61479-5876https://doaj.org/article/fb0228a28866446a85e61c34027154182021-11-01T00:00:00Zhttps://doi.org/10.1186/s12967-021-03132-6https://doaj.org/toc/1479-5876Abstract Background Adoptive transfer of chimeric antigen receptor (CAR)-engineered T cells combined with checkpoint inhibition may prevent T cell exhaustion and improve clinical outcomes. However, the approach is limited by cumulative costs and toxicities. Methods To overcome this drawback, we created a CAR-T (RB-340-1) that unites in one product the two modalities: a CRISPR interference-(CRISPRi) circuit prevents programmed cell death protein 1 (PD-1) expression upon antigen-encounter. RB-340-1 is engineered to express an anti-human epidermal growth factor receptor 2 (HER2) CAR single chain variable fragment (scFv), with CD28 and CD3ζ co-stimulatory domains linked to the tobacco etch virus (TEV) protease and a single guide RNA (sgRNA) targeting the PD-1 transcription start site (TSS). A second constructs includes linker for activation of T cells (LAT) fused to nuclease-deactivated spCas9 (dCas9)-Kruppel-associated box (KRAB) via a TEV-cleavable sequence (TCS). Upon antigen encounter, the LAT-dCas9-KRAB (LdCK) complex is cleaved by TEV allowing targeting of dCas9-KRAB to the PD-1 gene TSS. Results Here, we show that RB-340-1 consistently demonstrated higher production of homeostatic cytokines, enhanced expansion of CAR-T cells in vitro, prolonged in vivo persistence and more efficient suppression of HER2+ FaDu oropharyngeal cancer growth compared to the respective conventional CAR-T cell product. Conclusions As the first application of CRISPRi toward a clinically relevant product, RB-340-1 with the conditional, non-gene editing and reversible suppression promotes CAR-T cells resilience to checkpoint inhibition, and their persistence and effectiveness against HER2-expressing cancer xenografts.Zhifen YangLingyu LiAhu TurkozPohan ChenRona Harari-SteinfeldMaggie BobbinOfir StefansonHana ChoiViolena PietrobonBennett AlphsonAngshumala GoswamiVitaly BalanAlper KearneyDharmesh PatelJin YangDamla InelVeena VinodAlessandra CesanoBing WangKyung-Ho RohLei S. QiFrancesco M. MarincolaBMCarticleMedicineRENJournal of Translational Medicine, Vol 19, Iss 1, Pp 1-18 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R
spellingShingle	Medicine R Zhifen Yang Lingyu Li Ahu Turkoz Pohan Chen Rona Harari-Steinfeld Maggie Bobbin Ofir Stefanson Hana Choi Violena Pietrobon Bennett Alphson Angshumala Goswami Vitaly Balan Alper Kearney Dharmesh Patel Jin Yang Damla Inel Veena Vinod Alessandra Cesano Bing Wang Kyung-Ho Roh Lei S. Qi Francesco M. Marincola Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers
description	Abstract Background Adoptive transfer of chimeric antigen receptor (CAR)-engineered T cells combined with checkpoint inhibition may prevent T cell exhaustion and improve clinical outcomes. However, the approach is limited by cumulative costs and toxicities. Methods To overcome this drawback, we created a CAR-T (RB-340-1) that unites in one product the two modalities: a CRISPR interference-(CRISPRi) circuit prevents programmed cell death protein 1 (PD-1) expression upon antigen-encounter. RB-340-1 is engineered to express an anti-human epidermal growth factor receptor 2 (HER2) CAR single chain variable fragment (scFv), with CD28 and CD3ζ co-stimulatory domains linked to the tobacco etch virus (TEV) protease and a single guide RNA (sgRNA) targeting the PD-1 transcription start site (TSS). A second constructs includes linker for activation of T cells (LAT) fused to nuclease-deactivated spCas9 (dCas9)-Kruppel-associated box (KRAB) via a TEV-cleavable sequence (TCS). Upon antigen encounter, the LAT-dCas9-KRAB (LdCK) complex is cleaved by TEV allowing targeting of dCas9-KRAB to the PD-1 gene TSS. Results Here, we show that RB-340-1 consistently demonstrated higher production of homeostatic cytokines, enhanced expansion of CAR-T cells in vitro, prolonged in vivo persistence and more efficient suppression of HER2+ FaDu oropharyngeal cancer growth compared to the respective conventional CAR-T cell product. Conclusions As the first application of CRISPRi toward a clinically relevant product, RB-340-1 with the conditional, non-gene editing and reversible suppression promotes CAR-T cells resilience to checkpoint inhibition, and their persistence and effectiveness against HER2-expressing cancer xenografts.
format	article
author	Zhifen Yang Lingyu Li Ahu Turkoz Pohan Chen Rona Harari-Steinfeld Maggie Bobbin Ofir Stefanson Hana Choi Violena Pietrobon Bennett Alphson Angshumala Goswami Vitaly Balan Alper Kearney Dharmesh Patel Jin Yang Damla Inel Veena Vinod Alessandra Cesano Bing Wang Kyung-Ho Roh Lei S. Qi Francesco M. Marincola
author_facet	Zhifen Yang Lingyu Li Ahu Turkoz Pohan Chen Rona Harari-Steinfeld Maggie Bobbin Ofir Stefanson Hana Choi Violena Pietrobon Bennett Alphson Angshumala Goswami Vitaly Balan Alper Kearney Dharmesh Patel Jin Yang Damla Inel Veena Vinod Alessandra Cesano Bing Wang Kyung-Ho Roh Lei S. Qi Francesco M. Marincola
author_sort	Zhifen Yang
title	Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers
title_short	Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers
title_full	Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers
title_fullStr	Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers
title_full_unstemmed	Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers
title_sort	contextual reprogramming of car-t cells for treatment of her2+ cancers
publisher	BMC
publishDate	2021
url	https://doaj.org/article/fb0228a28866446a85e61c3402715418
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Contextual reprogramming of CAR-T cells for treatment of HER2+ cancers

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