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Title: Highly efficient gene transfer using a retroviral vector into murine T cells for preclinical chimeric antigen receptor-expressing T cell therapy

Abstract

Adoptive immunotherapy using chimeric antigen receptor-expressing T (CAR-T) cells has attracted attention as an efficacious strategy for cancer treatment. To prove the efficacy and safety of CAR-T cell therapy, the elucidation of immunological mechanisms underlying it in mice is required. Although a retroviral vector (Rv) is mainly used for the introduction of CAR to murine T cells, gene transduction efficiency is generally less than 50%. The low transduction efficiency causes poor precision in the functional analysis of CAR-T cells. We attempted to improve the Rv gene transduction protocol to more efficiently generate functional CAR-T cells by optimizing the period of pre-cultivation and antibody stimulation. In the improved protocol, gene transduction efficiency to murine T cells was more than 90%. In addition, almost all of the prepared murine T cells expressed CAR after puromycin selection. These CAR-T cells had antigen-specific cytotoxic activity and secreted multiple cytokines by antigen stimulation. We believe that our optimized gene transduction protocol for murine T cells contributes to the advancement of T cell biology and development of immunotherapy using genetically engineered T cells. - Highlights: • We established highly efficient gene transduction protocols for murine T cells. • CD8{sup +} CAR-T cells had antigen-specific cytotoxic activity.more » • CD4{sup +} CAR-T cells secreted multiple cytokines by antigen stimulation. • This finding can contribute to the development of T-cell biology and immunotherapy.« less

Authors:
; ; ; ; ;
Publication Date:
OSTI Identifier:
22596340
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemical and Biophysical Research Communications; Journal Volume: 473; Journal Issue: 1; Other Information: Copyright (c) 2016 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ACCURACY; ANTIGENS; CATTLE; DISEASE VECTORS; ENZYMES; FLUORESCENCE; FUNCTIONAL ANALYSIS; GENES; IMMUNOTHERAPY; INTERFERON; LYMPHOCYTES; MICE; MONOCLONAL ANTIBODIES; NEOPLASMS; POLYMERASE CHAIN REACTION; PUROMYCIN; RECEPTORS; RIBOSOMES; STIMULATION

Citation Formats

Kusabuka, Hotaka, Fujiwara, Kento, Tokunaga, Yusuke, Hirobe, Sachiko, Nakagawa, Shinsaku, E-mail: nakagawa@phs.osaka-u.ac.jp, and Okada, Naoki, E-mail: okada@phs.osaka-u.ac.jp. Highly efficient gene transfer using a retroviral vector into murine T cells for preclinical chimeric antigen receptor-expressing T cell therapy. United States: N. p., 2016. Web. doi:10.1016/J.BBRC.2016.03.054.
Kusabuka, Hotaka, Fujiwara, Kento, Tokunaga, Yusuke, Hirobe, Sachiko, Nakagawa, Shinsaku, E-mail: nakagawa@phs.osaka-u.ac.jp, & Okada, Naoki, E-mail: okada@phs.osaka-u.ac.jp. Highly efficient gene transfer using a retroviral vector into murine T cells for preclinical chimeric antigen receptor-expressing T cell therapy. United States. doi:10.1016/J.BBRC.2016.03.054.
Kusabuka, Hotaka, Fujiwara, Kento, Tokunaga, Yusuke, Hirobe, Sachiko, Nakagawa, Shinsaku, E-mail: nakagawa@phs.osaka-u.ac.jp, and Okada, Naoki, E-mail: okada@phs.osaka-u.ac.jp. Fri . "Highly efficient gene transfer using a retroviral vector into murine T cells for preclinical chimeric antigen receptor-expressing T cell therapy". United States. doi:10.1016/J.BBRC.2016.03.054.
@article{osti_22596340,
title = {Highly efficient gene transfer using a retroviral vector into murine T cells for preclinical chimeric antigen receptor-expressing T cell therapy},
author = {Kusabuka, Hotaka and Fujiwara, Kento and Tokunaga, Yusuke and Hirobe, Sachiko and Nakagawa, Shinsaku, E-mail: nakagawa@phs.osaka-u.ac.jp and Okada, Naoki, E-mail: okada@phs.osaka-u.ac.jp},
abstractNote = {Adoptive immunotherapy using chimeric antigen receptor-expressing T (CAR-T) cells has attracted attention as an efficacious strategy for cancer treatment. To prove the efficacy and safety of CAR-T cell therapy, the elucidation of immunological mechanisms underlying it in mice is required. Although a retroviral vector (Rv) is mainly used for the introduction of CAR to murine T cells, gene transduction efficiency is generally less than 50%. The low transduction efficiency causes poor precision in the functional analysis of CAR-T cells. We attempted to improve the Rv gene transduction protocol to more efficiently generate functional CAR-T cells by optimizing the period of pre-cultivation and antibody stimulation. In the improved protocol, gene transduction efficiency to murine T cells was more than 90%. In addition, almost all of the prepared murine T cells expressed CAR after puromycin selection. These CAR-T cells had antigen-specific cytotoxic activity and secreted multiple cytokines by antigen stimulation. We believe that our optimized gene transduction protocol for murine T cells contributes to the advancement of T cell biology and development of immunotherapy using genetically engineered T cells. - Highlights: • We established highly efficient gene transduction protocols for murine T cells. • CD8{sup +} CAR-T cells had antigen-specific cytotoxic activity. • CD4{sup +} CAR-T cells secreted multiple cytokines by antigen stimulation. • This finding can contribute to the development of T-cell biology and immunotherapy.},
doi = {10.1016/J.BBRC.2016.03.054},
journal = {Biochemical and Biophysical Research Communications},
number = 1,
volume = 473,
place = {United States},
year = {Fri Apr 22 00:00:00 EDT 2016},
month = {Fri Apr 22 00:00:00 EDT 2016}
}