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Gene therapy rescues disease phenotype in a spinal muscular atrophy with respiratory distress type 1 (SMARD1) mouse model.

Nizzardo M, Simone C, Rizzo F, Salani S, Dametti S, Rinchetti P, Del Bo R, Foust K, Kaspar BK, Bresolin N, Comi GP, Corti S - Sci Adv (2015)

Bottom Line: It is caused by mutations in the IGHMBP2 gene (11q13) and presently has no cure.AAV9-IGHMBP2 administration restored protein levels and rescued motor function, neuromuscular physiology, and life span (450% increase), ameliorating pathological features in the central nervous system, muscles, and heart.To test this strategy in a human model, we transferred wild-type IGHMBP2 into human SMARD1-induced pluripotent stem cell-derived motor neurons; these cells exhibited increased survival and axonal length in long-term culture.

View Article: PubMed Central - PubMed

Affiliation: Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), University of Milan, and Neurology Unit, IRCCS Foundation Ca' Granda, Ospedale Maggiore Policlinico, 20122 Milan, Italy.

ABSTRACT
Spinal muscular atrophy with respiratory distress type 1 (SMARD1) is an autosomal recessive motor neuron disease affecting children. It is caused by mutations in the IGHMBP2 gene (11q13) and presently has no cure. Recently, adeno-associated virus serotype 9 (AAV9)-mediated gene therapy has been shown to rescue the phenotype of animal models of another lower motor neuron disorder, spinal muscular atrophy 5q, and a clinical trial with this strategy is ongoing. We report rescue of the disease phenotype in a SMARD1 mouse model after therapeutic delivery via systemic injection of an AAV9 construct encoding the wild-type IGHMBP2 to replace the defective gene. AAV9-IGHMBP2 administration restored protein levels and rescued motor function, neuromuscular physiology, and life span (450% increase), ameliorating pathological features in the central nervous system, muscles, and heart. To test this strategy in a human model, we transferred wild-type IGHMBP2 into human SMARD1-induced pluripotent stem cell-derived motor neurons; these cells exhibited increased survival and axonal length in long-term culture. Our data support the translational potential of AAV-mediated gene therapies for SMARD1, opening the door for AAV9-mediated therapy in human clinical trials.

No MeSH data available.


Related in: MedlinePlus

AAV9-IGHMBP2 treatment protects human iPSC-derived spinal motor neurons from SMARD1-induced degeneration.(A) Schematic of the experimental strategy used to differentiate motor neurons from iPSCs. (B) Compared to WT (left), significantly fewer SMARD1 motor neurons were observed after long-term culture (right). Genetic correction with IGHMBP2 increased the number of SMARD1 motor neurons (TR-SMARD1, middle panel). SMI32, green; ChAT, red; DAPI, blue. Scale bar, 75 μm. (C) Genetic correction (green bar) protected motor neurons in long-term culture, increasing their survival versus SMARD1 cells (yellow bar) (8 weeks, *P < 0.001). (D) At 8 weeks, SMARD1 motor neurons (yellow) presented shorter axons than did WT cells (black). TR-SMARD1 (green) had longer axons than did SMARD1 motor neurons (P < 0.001). (E) IGHMBP2 staining (green) of WT motor neurons (left), SMARD1 motor neurons (right), and TR-SMARD1 (middle). Nuclei were counterstained with DAPI (blue). Scale bar, 70 μm. (F) Western blot of genetically corrected SMARD1 iPSCs (TR-SMARD1) indicated significantly increased expression of IGHMBP2 with respect toSMARD1 cells (SMARD1). (G) Quantification of IGHMBP2 levels in transfected SMARD1 iPSCs (TR-SMARD1) versus untransfected SMARD1 cells (SMARD1). Values are means ± SEM (*P < 0.01).
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Figure 5: AAV9-IGHMBP2 treatment protects human iPSC-derived spinal motor neurons from SMARD1-induced degeneration.(A) Schematic of the experimental strategy used to differentiate motor neurons from iPSCs. (B) Compared to WT (left), significantly fewer SMARD1 motor neurons were observed after long-term culture (right). Genetic correction with IGHMBP2 increased the number of SMARD1 motor neurons (TR-SMARD1, middle panel). SMI32, green; ChAT, red; DAPI, blue. Scale bar, 75 μm. (C) Genetic correction (green bar) protected motor neurons in long-term culture, increasing their survival versus SMARD1 cells (yellow bar) (8 weeks, *P < 0.001). (D) At 8 weeks, SMARD1 motor neurons (yellow) presented shorter axons than did WT cells (black). TR-SMARD1 (green) had longer axons than did SMARD1 motor neurons (P < 0.001). (E) IGHMBP2 staining (green) of WT motor neurons (left), SMARD1 motor neurons (right), and TR-SMARD1 (middle). Nuclei were counterstained with DAPI (blue). Scale bar, 70 μm. (F) Western blot of genetically corrected SMARD1 iPSCs (TR-SMARD1) indicated significantly increased expression of IGHMBP2 with respect toSMARD1 cells (SMARD1). (G) Quantification of IGHMBP2 levels in transfected SMARD1 iPSCs (TR-SMARD1) versus untransfected SMARD1 cells (SMARD1). Values are means ± SEM (*P < 0.01).

Mentions: Our next question was whether the transfer of IGHMBP2 also protected human SMARD1 motor neurons against degeneration induced by the loss of IGHMBP2. For this purpose, we used iPSCs that we generated from SMARD1 patients and healthy subjects (Fig. 5A) (17). We differentiated these cells into motor neurons, monitoring their phenotype by immunocytochemistry for motor neuronal markers. Human motor neurons extended processes and expressed SMI32 and choline acetyltransferase (ChAT) (Fig. 5B). For further evaluation, we examined several features of motor neurons that are likely relevant to SMARD1 pathogenesis, including cell survival, axonal elongation, and growth cone formation. At 8 weeks, SMARD1 iPSC–derived motor neurons had significantly reduced cell numbers and axon length compared to motor neurons derived from wild-type iPSCs (P < 0.001). However, motor neurons derived from genetically corrected SMARD1 cells (TR-SMARD1) presented an ameliorated phenotype, with increased cell survival at 8 weeks of culture (P < 0.001; Fig. 5C). These improved features were linked to increased IGHMBP2 expression; TR-SMARD1 motor neurons also presented increased neurite lengths compared to untreated neurons (P < 0.001; Fig. 5D). TR-SMARD1 motor neurons and their parental iPSCs expressed IGHMBP2 at higher levels than did untreated cells, as revealed by immunocytochemistry and Western blotting (Fig. 5, E to G). Thus, transferring wild-type IGHMBP2 can protect human motor neurons from SMARD1-induced degeneration.


Gene therapy rescues disease phenotype in a spinal muscular atrophy with respiratory distress type 1 (SMARD1) mouse model.

Nizzardo M, Simone C, Rizzo F, Salani S, Dametti S, Rinchetti P, Del Bo R, Foust K, Kaspar BK, Bresolin N, Comi GP, Corti S - Sci Adv (2015)

AAV9-IGHMBP2 treatment protects human iPSC-derived spinal motor neurons from SMARD1-induced degeneration.(A) Schematic of the experimental strategy used to differentiate motor neurons from iPSCs. (B) Compared to WT (left), significantly fewer SMARD1 motor neurons were observed after long-term culture (right). Genetic correction with IGHMBP2 increased the number of SMARD1 motor neurons (TR-SMARD1, middle panel). SMI32, green; ChAT, red; DAPI, blue. Scale bar, 75 μm. (C) Genetic correction (green bar) protected motor neurons in long-term culture, increasing their survival versus SMARD1 cells (yellow bar) (8 weeks, *P < 0.001). (D) At 8 weeks, SMARD1 motor neurons (yellow) presented shorter axons than did WT cells (black). TR-SMARD1 (green) had longer axons than did SMARD1 motor neurons (P < 0.001). (E) IGHMBP2 staining (green) of WT motor neurons (left), SMARD1 motor neurons (right), and TR-SMARD1 (middle). Nuclei were counterstained with DAPI (blue). Scale bar, 70 μm. (F) Western blot of genetically corrected SMARD1 iPSCs (TR-SMARD1) indicated significantly increased expression of IGHMBP2 with respect toSMARD1 cells (SMARD1). (G) Quantification of IGHMBP2 levels in transfected SMARD1 iPSCs (TR-SMARD1) versus untransfected SMARD1 cells (SMARD1). Values are means ± SEM (*P < 0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4643829&req=5

Figure 5: AAV9-IGHMBP2 treatment protects human iPSC-derived spinal motor neurons from SMARD1-induced degeneration.(A) Schematic of the experimental strategy used to differentiate motor neurons from iPSCs. (B) Compared to WT (left), significantly fewer SMARD1 motor neurons were observed after long-term culture (right). Genetic correction with IGHMBP2 increased the number of SMARD1 motor neurons (TR-SMARD1, middle panel). SMI32, green; ChAT, red; DAPI, blue. Scale bar, 75 μm. (C) Genetic correction (green bar) protected motor neurons in long-term culture, increasing their survival versus SMARD1 cells (yellow bar) (8 weeks, *P < 0.001). (D) At 8 weeks, SMARD1 motor neurons (yellow) presented shorter axons than did WT cells (black). TR-SMARD1 (green) had longer axons than did SMARD1 motor neurons (P < 0.001). (E) IGHMBP2 staining (green) of WT motor neurons (left), SMARD1 motor neurons (right), and TR-SMARD1 (middle). Nuclei were counterstained with DAPI (blue). Scale bar, 70 μm. (F) Western blot of genetically corrected SMARD1 iPSCs (TR-SMARD1) indicated significantly increased expression of IGHMBP2 with respect toSMARD1 cells (SMARD1). (G) Quantification of IGHMBP2 levels in transfected SMARD1 iPSCs (TR-SMARD1) versus untransfected SMARD1 cells (SMARD1). Values are means ± SEM (*P < 0.01).
Mentions: Our next question was whether the transfer of IGHMBP2 also protected human SMARD1 motor neurons against degeneration induced by the loss of IGHMBP2. For this purpose, we used iPSCs that we generated from SMARD1 patients and healthy subjects (Fig. 5A) (17). We differentiated these cells into motor neurons, monitoring their phenotype by immunocytochemistry for motor neuronal markers. Human motor neurons extended processes and expressed SMI32 and choline acetyltransferase (ChAT) (Fig. 5B). For further evaluation, we examined several features of motor neurons that are likely relevant to SMARD1 pathogenesis, including cell survival, axonal elongation, and growth cone formation. At 8 weeks, SMARD1 iPSC–derived motor neurons had significantly reduced cell numbers and axon length compared to motor neurons derived from wild-type iPSCs (P < 0.001). However, motor neurons derived from genetically corrected SMARD1 cells (TR-SMARD1) presented an ameliorated phenotype, with increased cell survival at 8 weeks of culture (P < 0.001; Fig. 5C). These improved features were linked to increased IGHMBP2 expression; TR-SMARD1 motor neurons also presented increased neurite lengths compared to untreated neurons (P < 0.001; Fig. 5D). TR-SMARD1 motor neurons and their parental iPSCs expressed IGHMBP2 at higher levels than did untreated cells, as revealed by immunocytochemistry and Western blotting (Fig. 5, E to G). Thus, transferring wild-type IGHMBP2 can protect human motor neurons from SMARD1-induced degeneration.

Bottom Line: It is caused by mutations in the IGHMBP2 gene (11q13) and presently has no cure.AAV9-IGHMBP2 administration restored protein levels and rescued motor function, neuromuscular physiology, and life span (450% increase), ameliorating pathological features in the central nervous system, muscles, and heart.To test this strategy in a human model, we transferred wild-type IGHMBP2 into human SMARD1-induced pluripotent stem cell-derived motor neurons; these cells exhibited increased survival and axonal length in long-term culture.

View Article: PubMed Central - PubMed

Affiliation: Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), University of Milan, and Neurology Unit, IRCCS Foundation Ca' Granda, Ospedale Maggiore Policlinico, 20122 Milan, Italy.

ABSTRACT
Spinal muscular atrophy with respiratory distress type 1 (SMARD1) is an autosomal recessive motor neuron disease affecting children. It is caused by mutations in the IGHMBP2 gene (11q13) and presently has no cure. Recently, adeno-associated virus serotype 9 (AAV9)-mediated gene therapy has been shown to rescue the phenotype of animal models of another lower motor neuron disorder, spinal muscular atrophy 5q, and a clinical trial with this strategy is ongoing. We report rescue of the disease phenotype in a SMARD1 mouse model after therapeutic delivery via systemic injection of an AAV9 construct encoding the wild-type IGHMBP2 to replace the defective gene. AAV9-IGHMBP2 administration restored protein levels and rescued motor function, neuromuscular physiology, and life span (450% increase), ameliorating pathological features in the central nervous system, muscles, and heart. To test this strategy in a human model, we transferred wild-type IGHMBP2 into human SMARD1-induced pluripotent stem cell-derived motor neurons; these cells exhibited increased survival and axonal length in long-term culture. Our data support the translational potential of AAV-mediated gene therapies for SMARD1, opening the door for AAV9-mediated therapy in human clinical trials.

No MeSH data available.


Related in: MedlinePlus