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Novel Genome-Editing Tools to Model and Correct Primary Immunodeficiencies.

Ott de Bruin LM, Volpi S, Musunuru K - Front Immunol (2015)

Bottom Line: With this treatment, severe complications may result due to integration within oncogenes.With these genome-editing tools a correct copy can be inserted in a precisely targeted "safe harbor." They can also be used to correct pathogenic mutations in situ and to develop cellular or animal models needed to study the pathogenic effects of specific genetic defects found in immunodeficient patients.This review discusses the advantages and disadvantages of these endonucleases in gene correction and modeling with an emphasis on CRISPR/Cas9, which offers the most promise due to its efficacy and versatility.

View Article: PubMed Central - PubMed

Affiliation: Division of Immunology, Boston Children's Hospital, Harvard Medical School , Boston, MA , USA ; Department of Pediatric Immunology, Wilhelmina Children's Hospital, University Medical Center Utrecht , Utrecht , Netherlands.

ABSTRACT
Severe combined immunodeficiency (SCID) and other severe non-SCID primary immunodeficiencies (non-SCID PID) can be treated by allogeneic hematopoietic stem cell (HSC) transplantation, but when histocompatibility leukocyte antigen-matched donors are lacking, this can be a high-risk procedure. Correcting the patient's own HSCs with gene therapy offers an attractive alternative. Gene therapies currently being used in clinical settings insert a functional copy of the entire gene by means of a viral vector. With this treatment, severe complications may result due to integration within oncogenes. A promising alternative is the use of endonucleases such as ZFNs, TALENs, and CRISPR/Cas9 to introduce a double-stranded break in the DNA and thus induce homology-directed repair. With these genome-editing tools a correct copy can be inserted in a precisely targeted "safe harbor." They can also be used to correct pathogenic mutations in situ and to develop cellular or animal models needed to study the pathogenic effects of specific genetic defects found in immunodeficient patients. This review discusses the advantages and disadvantages of these endonucleases in gene correction and modeling with an emphasis on CRISPR/Cas9, which offers the most promise due to its efficacy and versatility.

No MeSH data available.


Related in: MedlinePlus

In vitro modeling. (A) Induced pluripotent stem cells (iPSCs) are reprogrammed from a patient(s) and from a healthy control(s). The iPSCs are differentiated into a cell type of interest, and the phenotypes of the patient-derived cells are compared to the phenotypes of the healthy control cells. The cells that are compared do not have the exact same genetic background (genetically and epigenetically unmatched). This can lead to confounding. (B) Using genome editing with engineered nucleases like ZFNs, TALENs, and CRISPR/Cas9, a pathogenetic mutation can be corrected in patient-derived cells or introduced into healthy control cells, and isogenic cell lines (i.e., identical genetic background) can be compared for relevant phenotypes.
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Figure 3: In vitro modeling. (A) Induced pluripotent stem cells (iPSCs) are reprogrammed from a patient(s) and from a healthy control(s). The iPSCs are differentiated into a cell type of interest, and the phenotypes of the patient-derived cells are compared to the phenotypes of the healthy control cells. The cells that are compared do not have the exact same genetic background (genetically and epigenetically unmatched). This can lead to confounding. (B) Using genome editing with engineered nucleases like ZFNs, TALENs, and CRISPR/Cas9, a pathogenetic mutation can be corrected in patient-derived cells or introduced into healthy control cells, and isogenic cell lines (i.e., identical genetic background) can be compared for relevant phenotypes.

Mentions: The advent of next-generation sequencing has stimulated a new wave of discovery of novel inborn errors of immunity (102). The ability to correct patient-specific iPSCs, or conversely, to introduce patient-specific mutations into a wild-type iPSC line using endonucleases represents an invaluable tool to prove the pathogenicity of newly discovered mutations and to gain insight into disease mechanisms in different cell types, depending on patients’ phenotypes. This approach also makes it possible to study the contribution of genetic background to the phenotypes arising from specific mutations by comparing patient-derived iPSCs with wild-type iPSCs into which the same mutations are introduced (Figure 3). In a recent study, iPSCs were generated from patients with Parkinson disease caused by the G2019S mutation of the LRRK2 gene and from healthy controls. When comparing the whole-genome gene expression patterns, the investigators found a high degree of heterogeneity among the different iPSCs lines. However, when they used ZFNs to correct the mutation in three of the patient-derived iPSC lines and compared these lines to the original lines, and when they introduced the mutation into a healthy control line and compared this line to the original line, the lines were much more closely matched with respect to gene expression (83). This shows the importance of comparing isogenic lines, as confounding due to differences in genetic background is minimized.


Novel Genome-Editing Tools to Model and Correct Primary Immunodeficiencies.

Ott de Bruin LM, Volpi S, Musunuru K - Front Immunol (2015)

In vitro modeling. (A) Induced pluripotent stem cells (iPSCs) are reprogrammed from a patient(s) and from a healthy control(s). The iPSCs are differentiated into a cell type of interest, and the phenotypes of the patient-derived cells are compared to the phenotypes of the healthy control cells. The cells that are compared do not have the exact same genetic background (genetically and epigenetically unmatched). This can lead to confounding. (B) Using genome editing with engineered nucleases like ZFNs, TALENs, and CRISPR/Cas9, a pathogenetic mutation can be corrected in patient-derived cells or introduced into healthy control cells, and isogenic cell lines (i.e., identical genetic background) can be compared for relevant phenotypes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4440404&req=5

Figure 3: In vitro modeling. (A) Induced pluripotent stem cells (iPSCs) are reprogrammed from a patient(s) and from a healthy control(s). The iPSCs are differentiated into a cell type of interest, and the phenotypes of the patient-derived cells are compared to the phenotypes of the healthy control cells. The cells that are compared do not have the exact same genetic background (genetically and epigenetically unmatched). This can lead to confounding. (B) Using genome editing with engineered nucleases like ZFNs, TALENs, and CRISPR/Cas9, a pathogenetic mutation can be corrected in patient-derived cells or introduced into healthy control cells, and isogenic cell lines (i.e., identical genetic background) can be compared for relevant phenotypes.
Mentions: The advent of next-generation sequencing has stimulated a new wave of discovery of novel inborn errors of immunity (102). The ability to correct patient-specific iPSCs, or conversely, to introduce patient-specific mutations into a wild-type iPSC line using endonucleases represents an invaluable tool to prove the pathogenicity of newly discovered mutations and to gain insight into disease mechanisms in different cell types, depending on patients’ phenotypes. This approach also makes it possible to study the contribution of genetic background to the phenotypes arising from specific mutations by comparing patient-derived iPSCs with wild-type iPSCs into which the same mutations are introduced (Figure 3). In a recent study, iPSCs were generated from patients with Parkinson disease caused by the G2019S mutation of the LRRK2 gene and from healthy controls. When comparing the whole-genome gene expression patterns, the investigators found a high degree of heterogeneity among the different iPSCs lines. However, when they used ZFNs to correct the mutation in three of the patient-derived iPSC lines and compared these lines to the original lines, and when they introduced the mutation into a healthy control line and compared this line to the original line, the lines were much more closely matched with respect to gene expression (83). This shows the importance of comparing isogenic lines, as confounding due to differences in genetic background is minimized.

Bottom Line: With this treatment, severe complications may result due to integration within oncogenes.With these genome-editing tools a correct copy can be inserted in a precisely targeted "safe harbor." They can also be used to correct pathogenic mutations in situ and to develop cellular or animal models needed to study the pathogenic effects of specific genetic defects found in immunodeficient patients.This review discusses the advantages and disadvantages of these endonucleases in gene correction and modeling with an emphasis on CRISPR/Cas9, which offers the most promise due to its efficacy and versatility.

View Article: PubMed Central - PubMed

Affiliation: Division of Immunology, Boston Children's Hospital, Harvard Medical School , Boston, MA , USA ; Department of Pediatric Immunology, Wilhelmina Children's Hospital, University Medical Center Utrecht , Utrecht , Netherlands.

ABSTRACT
Severe combined immunodeficiency (SCID) and other severe non-SCID primary immunodeficiencies (non-SCID PID) can be treated by allogeneic hematopoietic stem cell (HSC) transplantation, but when histocompatibility leukocyte antigen-matched donors are lacking, this can be a high-risk procedure. Correcting the patient's own HSCs with gene therapy offers an attractive alternative. Gene therapies currently being used in clinical settings insert a functional copy of the entire gene by means of a viral vector. With this treatment, severe complications may result due to integration within oncogenes. A promising alternative is the use of endonucleases such as ZFNs, TALENs, and CRISPR/Cas9 to introduce a double-stranded break in the DNA and thus induce homology-directed repair. With these genome-editing tools a correct copy can be inserted in a precisely targeted "safe harbor." They can also be used to correct pathogenic mutations in situ and to develop cellular or animal models needed to study the pathogenic effects of specific genetic defects found in immunodeficient patients. This review discusses the advantages and disadvantages of these endonucleases in gene correction and modeling with an emphasis on CRISPR/Cas9, which offers the most promise due to its efficacy and versatility.

No MeSH data available.


Related in: MedlinePlus