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Preclinical Evaluation of a Novel TALEN Targeting CCR5 Confirms Efficacy and Safety in Conferring Resistance to HIV-1 Infection


Romito, Marianna; Juillerat, Alexandre; Kok, Yik Lim; Hildenbeutel, Markus; Rhiel, Manuel; Andrieux, Geoffroy; Geiger, Johannes; Rudolph, Carsten; Mussolino, Claudio; Duclert, Aymeric; Metzner, Karin J; Duchateau, Philippe; Cathomen, Toni; Cornu, Tatjana I (2021). Preclinical Evaluation of a Novel TALEN Targeting CCR5 Confirms Efficacy and Safety in Conferring Resistance to HIV-1 Infection. Biotechnology Journal, 16(1):2000023.

Abstract

Therapies to treat patients infected with human immunodeficiency virus (HIV) aim at preventing viral replication but fail to eliminate the virus. Although transplantation of allogeneic CCR5Δ32 homozygous stem cell grafts provided a cure for a few patients, this approach is not considered a general therapeutic strategy because of potential side effects. Conversely, gene editing to disrupt the C-C chemokine receptor type 5 (CCR5) locus, which encodes the major HIV coreceptor, has shown to confer resistance to CCR5-tropic HIV strains. Here, an engineered transcription activator-like effector nuclease (TALEN) that enables efficient CCR5 editing in hematopoietic cells is presented. After transferring TALEN-encoding mRNA into primary CD4+ T cells, up to 89% of CCR5 alleles are disrupted. Genotyping confirms the genetic stability of the CCR5-edited cells, and genome-wide off-target analyses established the absence of relevant mutagenic events. When challenging the edited T cells with CCR5-tropic HIV, protection in a dose-dependent manner is observed. Functional assessments reveal no significant differences between edited and control cells in terms of proliferation and their ability to secrete cytokines upon exogenous stimuli. In conclusion, a highly active and specific TALEN to disrupt CCR5 is successfully engineered, paving the way for its clinical application in hematopoietic stem cell grafts.

Abstract

Therapies to treat patients infected with human immunodeficiency virus (HIV) aim at preventing viral replication but fail to eliminate the virus. Although transplantation of allogeneic CCR5Δ32 homozygous stem cell grafts provided a cure for a few patients, this approach is not considered a general therapeutic strategy because of potential side effects. Conversely, gene editing to disrupt the C-C chemokine receptor type 5 (CCR5) locus, which encodes the major HIV coreceptor, has shown to confer resistance to CCR5-tropic HIV strains. Here, an engineered transcription activator-like effector nuclease (TALEN) that enables efficient CCR5 editing in hematopoietic cells is presented. After transferring TALEN-encoding mRNA into primary CD4+ T cells, up to 89% of CCR5 alleles are disrupted. Genotyping confirms the genetic stability of the CCR5-edited cells, and genome-wide off-target analyses established the absence of relevant mutagenic events. When challenging the edited T cells with CCR5-tropic HIV, protection in a dose-dependent manner is observed. Functional assessments reveal no significant differences between edited and control cells in terms of proliferation and their ability to secrete cytokines upon exogenous stimuli. In conclusion, a highly active and specific TALEN to disrupt CCR5 is successfully engineered, paving the way for its clinical application in hematopoietic stem cell grafts.

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Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Institute of Medical Virology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Applied Microbiology and Biotechnology
Life Sciences > Molecular Medicine
Language:English
Date:1 January 2021
Deposited On:19 Jan 2021 15:58
Last Modified:21 Jan 2021 11:22
Publisher:Wiley-VCH Verlag
ISSN:1860-6768
OA Status:Hybrid
Publisher DOI:https://doi.org/10.1002/biot.202000023
PubMed ID:33103367

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