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Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene

Chimeric antigen receptor (CAR)-T cell immunotherapy is at the forefront of innovative cancer therapeutics. However, lack of standardization of cellular products within the same clinical trial and lack of harmonization between different trials have hindered the clear identification of efficacy and s...

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Autores principales: Casucci, Monica, Falcone, Laura, Camisa, Barbara, Norelli, Margherita, Porcellini, Simona, Stornaiuolo, Anna, Ciceri, Fabio, Traversari, Catia, Bordignon, Claudio, Bonini, Chiara, Bondanza, Attilio
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5871667/
https://www.ncbi.nlm.nih.gov/pubmed/29619024
http://dx.doi.org/10.3389/fimmu.2018.00507
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author Casucci, Monica
Falcone, Laura
Camisa, Barbara
Norelli, Margherita
Porcellini, Simona
Stornaiuolo, Anna
Ciceri, Fabio
Traversari, Catia
Bordignon, Claudio
Bonini, Chiara
Bondanza, Attilio
author_facet Casucci, Monica
Falcone, Laura
Camisa, Barbara
Norelli, Margherita
Porcellini, Simona
Stornaiuolo, Anna
Ciceri, Fabio
Traversari, Catia
Bordignon, Claudio
Bonini, Chiara
Bondanza, Attilio
author_sort Casucci, Monica
collection PubMed
description Chimeric antigen receptor (CAR)-T cell immunotherapy is at the forefront of innovative cancer therapeutics. However, lack of standardization of cellular products within the same clinical trial and lack of harmonization between different trials have hindered the clear identification of efficacy and safety determinants that should be unveiled in order to advance the field. With the aim of facilitating the isolation and in vivo tracking of CAR-T cells, we here propose the inclusion within the CAR molecule of a novel extracellular spacer based on the low-affinity nerve-growth-factor receptor (NGFR). We screened four different spacer designs using as target antigen the CD44 isoform variant 6 (CD44v6). We successfully generated NGFR-spaced CD44v6 CAR-T cells that could be efficiently enriched with clinical-grade immuno-magnetic beads without negative consequences on subsequent expansion, immuno-phenotype, in vitro antitumor reactivity, and conditional ablation when co-expressing a suicide gene. Most importantly, these cells could be tracked with anti-NGFR monoclonal antibodies in NSG mice, where they expanded, persisted, and exerted potent antitumor effects against both high leukemia and myeloma burdens. Similar results were obtained with NGFR-enriched CAR-T cells specific for CD19 or CEA, suggesting the universality of this strategy. In conclusion, we have demonstrated that the incorporation of the NGFR marker gene within the CAR sequence allows for a single molecule to simultaneously work as a therapeutic and selection/tracking gene. Looking ahead, NGFR spacer enrichment might allow good manufacturing procedures-manufacturing of standardized CAR-T cell products with high therapeutic potential, which could be harmonized in different clinical trials and used in combination with a suicide gene for future application in the allogeneic setting.
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spelling pubmed-58716672018-04-04 Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene Casucci, Monica Falcone, Laura Camisa, Barbara Norelli, Margherita Porcellini, Simona Stornaiuolo, Anna Ciceri, Fabio Traversari, Catia Bordignon, Claudio Bonini, Chiara Bondanza, Attilio Front Immunol Immunology Chimeric antigen receptor (CAR)-T cell immunotherapy is at the forefront of innovative cancer therapeutics. However, lack of standardization of cellular products within the same clinical trial and lack of harmonization between different trials have hindered the clear identification of efficacy and safety determinants that should be unveiled in order to advance the field. With the aim of facilitating the isolation and in vivo tracking of CAR-T cells, we here propose the inclusion within the CAR molecule of a novel extracellular spacer based on the low-affinity nerve-growth-factor receptor (NGFR). We screened four different spacer designs using as target antigen the CD44 isoform variant 6 (CD44v6). We successfully generated NGFR-spaced CD44v6 CAR-T cells that could be efficiently enriched with clinical-grade immuno-magnetic beads without negative consequences on subsequent expansion, immuno-phenotype, in vitro antitumor reactivity, and conditional ablation when co-expressing a suicide gene. Most importantly, these cells could be tracked with anti-NGFR monoclonal antibodies in NSG mice, where they expanded, persisted, and exerted potent antitumor effects against both high leukemia and myeloma burdens. Similar results were obtained with NGFR-enriched CAR-T cells specific for CD19 or CEA, suggesting the universality of this strategy. In conclusion, we have demonstrated that the incorporation of the NGFR marker gene within the CAR sequence allows for a single molecule to simultaneously work as a therapeutic and selection/tracking gene. Looking ahead, NGFR spacer enrichment might allow good manufacturing procedures-manufacturing of standardized CAR-T cell products with high therapeutic potential, which could be harmonized in different clinical trials and used in combination with a suicide gene for future application in the allogeneic setting. Frontiers Media S.A. 2018-03-21 /pmc/articles/PMC5871667/ /pubmed/29619024 http://dx.doi.org/10.3389/fimmu.2018.00507 Text en Copyright © 2018 Casucci, Falcone, Camisa, Norelli, Porcellini, Stornaiuolo, Ciceri, Traversari, Bordignon, Bonini and Bondanza. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Immunology
Casucci, Monica
Falcone, Laura
Camisa, Barbara
Norelli, Margherita
Porcellini, Simona
Stornaiuolo, Anna
Ciceri, Fabio
Traversari, Catia
Bordignon, Claudio
Bonini, Chiara
Bondanza, Attilio
Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene
title Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene
title_full Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene
title_fullStr Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene
title_full_unstemmed Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene
title_short Extracellular NGFR Spacers Allow Efficient Tracking and Enrichment of Fully Functional CAR-T Cells Co-Expressing a Suicide Gene
title_sort extracellular ngfr spacers allow efficient tracking and enrichment of fully functional car-t cells co-expressing a suicide gene
topic Immunology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5871667/
https://www.ncbi.nlm.nih.gov/pubmed/29619024
http://dx.doi.org/10.3389/fimmu.2018.00507
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