Cargando…

Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources

BACKGROUND: Adoptive cell therapy with chimeric antigen receptor T cells (CAR-T) has become a standard treatment for patients with certain aggressive B cell malignancies and holds promise to improve the care of patients suffering from numerous other cancers in the future. However, the high manufactu...

Descripción completa

Detalles Bibliográficos
Autores principales: Schmidts, Andrea, Marsh, Leah C, Srivastava, Ambike A, Bouffard, Amanda A, Boroughs, Angela C, Scarfò, Irene, Larson, Rebecca C, Bedoya, Felipe, Choi, Bryan D, Frigault, Matthew J, Bailey, Stefanie R, Leick, Mark B, Vatsa, Sonika, Kann, Michael C, Prew, Michelle S, Kleinstiver, Benjamin P, Joung, J Keith, Maus, Marcela V
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BMJ Publishing Group 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7477986/
https://www.ncbi.nlm.nih.gov/pubmed/32900862
http://dx.doi.org/10.1136/jitc-2020-000990
_version_ 1783579983713665024
author Schmidts, Andrea
Marsh, Leah C
Srivastava, Ambike A
Bouffard, Amanda A
Boroughs, Angela C
Scarfò, Irene
Larson, Rebecca C
Bedoya, Felipe
Choi, Bryan D
Frigault, Matthew J
Bailey, Stefanie R
Leick, Mark B
Vatsa, Sonika
Kann, Michael C
Prew, Michelle S
Kleinstiver, Benjamin P
Joung, J Keith
Maus, Marcela V
author_facet Schmidts, Andrea
Marsh, Leah C
Srivastava, Ambike A
Bouffard, Amanda A
Boroughs, Angela C
Scarfò, Irene
Larson, Rebecca C
Bedoya, Felipe
Choi, Bryan D
Frigault, Matthew J
Bailey, Stefanie R
Leick, Mark B
Vatsa, Sonika
Kann, Michael C
Prew, Michelle S
Kleinstiver, Benjamin P
Joung, J Keith
Maus, Marcela V
author_sort Schmidts, Andrea
collection PubMed
description BACKGROUND: Adoptive cell therapy with chimeric antigen receptor T cells (CAR-T) has become a standard treatment for patients with certain aggressive B cell malignancies and holds promise to improve the care of patients suffering from numerous other cancers in the future. However, the high manufacturing cost of CAR-T cell therapies poses a major barrier to their broader clinical application. Among the key cost drivers of CAR-T production are single-use reagents for T cell activation and clinical-grade viral vector. The presence of variable amounts of contaminating monocytes in the starting material poses an additional challenge to CAR-T manufacturing, since they can impede T cell stimulation and transduction, resulting in manufacturing failure. METHODS: We created K562-based artificial antigen-presenting cells (aAPC) with genetically encoded T cell stimulation and costimulation that represent an inexhaustible source for T cell activation. We additionally disrupted endogenous expression of the low-density lipoprotein receptor (LDLR) on these aAPC (aAPC-ΔLDLR) using CRISPR-Cas9 gene editing nucleases to prevent inadvertent lentiviral transduction and avoid the sink effect on viral vector during transduction. Using various T cell sources, we produced CD19-directed CAR-T cells via aAPC-ΔLDLR-based activation and tested their in vitro and in vivo antitumor potency against B cell malignancies. RESULTS: We found that lack of LDLR expression on our aAPC-ΔLDLR conferred resistance to lentiviral transduction during CAR-T production. Using aAPC-ΔLDLR, we achieved efficient expansion of CAR-T cells even from unpurified starting material like peripheral blood mononuclear cells or unmanipulated leukapheresis product, containing substantial proportions of monocytes. CD19-directed CAR-T cells that we produced via aAPC-ΔLDLR-based expansion demonstrated potent antitumor responses in preclinical models of acute lymphoblastic leukemia and B-cell lymphoma. CONCLUSIONS: Our aAPC-ΔLDLR represent an attractive approach for manufacturing of lentivirally transduced T cells that may be simpler and more cost efficient than currently available methods.
format Online
Article
Text
id pubmed-7477986
institution National Center for Biotechnology Information
language English
publishDate 2020
publisher BMJ Publishing Group
record_format MEDLINE/PubMed
spelling pubmed-74779862020-09-21 Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources Schmidts, Andrea Marsh, Leah C Srivastava, Ambike A Bouffard, Amanda A Boroughs, Angela C Scarfò, Irene Larson, Rebecca C Bedoya, Felipe Choi, Bryan D Frigault, Matthew J Bailey, Stefanie R Leick, Mark B Vatsa, Sonika Kann, Michael C Prew, Michelle S Kleinstiver, Benjamin P Joung, J Keith Maus, Marcela V J Immunother Cancer Immune Cell Therapies and Immune Cell Engineering BACKGROUND: Adoptive cell therapy with chimeric antigen receptor T cells (CAR-T) has become a standard treatment for patients with certain aggressive B cell malignancies and holds promise to improve the care of patients suffering from numerous other cancers in the future. However, the high manufacturing cost of CAR-T cell therapies poses a major barrier to their broader clinical application. Among the key cost drivers of CAR-T production are single-use reagents for T cell activation and clinical-grade viral vector. The presence of variable amounts of contaminating monocytes in the starting material poses an additional challenge to CAR-T manufacturing, since they can impede T cell stimulation and transduction, resulting in manufacturing failure. METHODS: We created K562-based artificial antigen-presenting cells (aAPC) with genetically encoded T cell stimulation and costimulation that represent an inexhaustible source for T cell activation. We additionally disrupted endogenous expression of the low-density lipoprotein receptor (LDLR) on these aAPC (aAPC-ΔLDLR) using CRISPR-Cas9 gene editing nucleases to prevent inadvertent lentiviral transduction and avoid the sink effect on viral vector during transduction. Using various T cell sources, we produced CD19-directed CAR-T cells via aAPC-ΔLDLR-based activation and tested their in vitro and in vivo antitumor potency against B cell malignancies. RESULTS: We found that lack of LDLR expression on our aAPC-ΔLDLR conferred resistance to lentiviral transduction during CAR-T production. Using aAPC-ΔLDLR, we achieved efficient expansion of CAR-T cells even from unpurified starting material like peripheral blood mononuclear cells or unmanipulated leukapheresis product, containing substantial proportions of monocytes. CD19-directed CAR-T cells that we produced via aAPC-ΔLDLR-based expansion demonstrated potent antitumor responses in preclinical models of acute lymphoblastic leukemia and B-cell lymphoma. CONCLUSIONS: Our aAPC-ΔLDLR represent an attractive approach for manufacturing of lentivirally transduced T cells that may be simpler and more cost efficient than currently available methods. BMJ Publishing Group 2020-09-07 /pmc/articles/PMC7477986/ /pubmed/32900862 http://dx.doi.org/10.1136/jitc-2020-000990 Text en © Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ. http://creativecommons.org/licenses/by-nc/4.0/ http://creativecommons.org/licenses/by-nc/4.0/This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See http://creativecommons.org/licenses/by-nc/4.0/.
spellingShingle Immune Cell Therapies and Immune Cell Engineering
Schmidts, Andrea
Marsh, Leah C
Srivastava, Ambike A
Bouffard, Amanda A
Boroughs, Angela C
Scarfò, Irene
Larson, Rebecca C
Bedoya, Felipe
Choi, Bryan D
Frigault, Matthew J
Bailey, Stefanie R
Leick, Mark B
Vatsa, Sonika
Kann, Michael C
Prew, Michelle S
Kleinstiver, Benjamin P
Joung, J Keith
Maus, Marcela V
Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources
title Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources
title_full Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources
title_fullStr Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources
title_full_unstemmed Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources
title_short Cell-based artificial APC resistant to lentiviral transduction for efficient generation of CAR-T cells from various cell sources
title_sort cell-based artificial apc resistant to lentiviral transduction for efficient generation of car-t cells from various cell sources
topic Immune Cell Therapies and Immune Cell Engineering
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7477986/
https://www.ncbi.nlm.nih.gov/pubmed/32900862
http://dx.doi.org/10.1136/jitc-2020-000990
work_keys_str_mv AT schmidtsandrea cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT marshleahc cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT srivastavaambikea cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT bouffardamandaa cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT boroughsangelac cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT scarfoirene cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT larsonrebeccac cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT bedoyafelipe cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT choibryand cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT frigaultmatthewj cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT baileystefanier cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT leickmarkb cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT vatsasonika cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT kannmichaelc cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT prewmichelles cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT kleinstiverbenjaminp cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT joungjkeith cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources
AT mausmarcelav cellbasedartificialapcresistanttolentiviraltransductionforefficientgenerationofcartcellsfromvariouscellsources