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Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells
Dominant negative mutations in the STAT3 gene account for autosomal dominant hyper-IgE syndrome (AD-HIES). Patients typically present high IgE serum levels, recurrent infections, and soft tissue abnormalities. While current therapies focus on alleviating the symptoms, hematopoietic stem cell transpl...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9601366/ https://www.ncbi.nlm.nih.gov/pubmed/36292796 http://dx.doi.org/10.3390/genes13101912 |
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author | König, Saskia Fliegauf, Manfred Rhiel, Manuel Grimbacher, Bodo Cornu, Tatjana I. Cathomen, Toni Mussolino, Claudio |
author_facet | König, Saskia Fliegauf, Manfred Rhiel, Manuel Grimbacher, Bodo Cornu, Tatjana I. Cathomen, Toni Mussolino, Claudio |
author_sort | König, Saskia |
collection | PubMed |
description | Dominant negative mutations in the STAT3 gene account for autosomal dominant hyper-IgE syndrome (AD-HIES). Patients typically present high IgE serum levels, recurrent infections, and soft tissue abnormalities. While current therapies focus on alleviating the symptoms, hematopoietic stem cell transplantation (HSCT) has recently been proposed as a strategy to treat the immunological defect and stabilize the disease, especially in cases with severe lung infections. However, because of the potentially severe side effects associated with allogeneic HSCT, this has been considered only for a few patients. Autologous HSCT represents a safer alternative but it requires the removal of the dominant negative mutation in the patients’ cells prior to transplantation. Here, we developed allele-specific CRISPR-Cas9 nucleases to selectively disrupt five of the most common STAT3 dominant negative alleles. When tested ex vivo in patient-derived hematopoietic cells, allele-specific disruption frequencies varied in an allele-dependent fashion and reached up to 62% of alleles harboring the V637M mutation without detectable alterations in the healthy STAT3 allele. However, assessment of the gene expression profiles of the STAT3 downstream target genes revealed that, upon activation of those edited patient cells, mono-allelic STAT3 expression (functional haploinsufficiency) is not able to sufficiently restore STAT3-dependent signaling in edited T cells cultured in vitro. Moreover, the stochastic mutagenesis induced by the repair of the nuclease-induced DNA break could further contribute to dominant negative effects. In summary, our results advocate for precise genome editing strategies rather than allele-specific gene disruption to correct the underlying mutations in AD-HIES. |
format | Online Article Text |
id | pubmed-9601366 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-96013662022-10-27 Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells König, Saskia Fliegauf, Manfred Rhiel, Manuel Grimbacher, Bodo Cornu, Tatjana I. Cathomen, Toni Mussolino, Claudio Genes (Basel) Article Dominant negative mutations in the STAT3 gene account for autosomal dominant hyper-IgE syndrome (AD-HIES). Patients typically present high IgE serum levels, recurrent infections, and soft tissue abnormalities. While current therapies focus on alleviating the symptoms, hematopoietic stem cell transplantation (HSCT) has recently been proposed as a strategy to treat the immunological defect and stabilize the disease, especially in cases with severe lung infections. However, because of the potentially severe side effects associated with allogeneic HSCT, this has been considered only for a few patients. Autologous HSCT represents a safer alternative but it requires the removal of the dominant negative mutation in the patients’ cells prior to transplantation. Here, we developed allele-specific CRISPR-Cas9 nucleases to selectively disrupt five of the most common STAT3 dominant negative alleles. When tested ex vivo in patient-derived hematopoietic cells, allele-specific disruption frequencies varied in an allele-dependent fashion and reached up to 62% of alleles harboring the V637M mutation without detectable alterations in the healthy STAT3 allele. However, assessment of the gene expression profiles of the STAT3 downstream target genes revealed that, upon activation of those edited patient cells, mono-allelic STAT3 expression (functional haploinsufficiency) is not able to sufficiently restore STAT3-dependent signaling in edited T cells cultured in vitro. Moreover, the stochastic mutagenesis induced by the repair of the nuclease-induced DNA break could further contribute to dominant negative effects. In summary, our results advocate for precise genome editing strategies rather than allele-specific gene disruption to correct the underlying mutations in AD-HIES. MDPI 2022-10-20 /pmc/articles/PMC9601366/ /pubmed/36292796 http://dx.doi.org/10.3390/genes13101912 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article König, Saskia Fliegauf, Manfred Rhiel, Manuel Grimbacher, Bodo Cornu, Tatjana I. Cathomen, Toni Mussolino, Claudio Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells |
title | Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells |
title_full | Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells |
title_fullStr | Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells |
title_full_unstemmed | Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells |
title_short | Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells |
title_sort | allele-specific disruption of a common stat3 autosomal dominant allele is not sufficient to restore downstream signaling in patient-derived t cells |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9601366/ https://www.ncbi.nlm.nih.gov/pubmed/36292796 http://dx.doi.org/10.3390/genes13101912 |
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