In vivo and ex vivo lentiviral and stem cell therapy approaches for progressive systemic skeletal dysplasia: mucopolysaccharidosis IVA
| Author(s) | Celik, Betul | |
| Date Accessioned | 2025-04-14T16:53:38Z | |
| Date Available | 2025-04-14T16:53:38Z | |
| Publication Date | 2025 | |
| SWORD Update | 2025-04-14T04:01:38Z | |
| Abstract | Mucopolysaccharidosis IVA (MPS IVA) is caused by a mutation in the N-acetylgalactosamine-6-sulfate sulfatase (GALNS) gene, leading to glycosaminoglycan (GAG) accumulation in multiple tissues, resulting in progressive skeletal dysplasia and poor quality of life. There is no effective treatment for this skeletal disease at present. This study proposes an extensive evaluation of novel lentiviral (LV) and hematopoietic stem cell (HSC) gene therapy (GT) that produce and secrete the active GALNS enzyme at supraphysiologic levels within the cells. LVs carrying the native GALNS encoding sequence (cDNA) were made under three different promoters: CBh, COL2A1, and CD11b. Moreover, we designed LVs carrying the native GALNS cDNA tagged with D8 octapeptide under CD11b promoter and a human codon-optimized GALNS cDNA under CBh promoter, respectively. We first confirmed the potential impact of LVs in vitro and translated our research into an MPS IVA animal model. Translational research was performed under both ex vivo LV-mediated HSC transplantation into myeloablated recipient MPS IVA mice and in vivo direct infusion of LVs into bloodstream MPS IVA mice. Following tissue collections, enzyme activity, GAG levels, vector copy numbers (VCNs), histopathological evaluations, and bone morphometric analysis were performed at both cellular levels and in treated mice. LVs under the COL2A1 promoter produced the highest enzyme activity and normalized the storage materials in vitro, followed by the CBh promoter. Both LVs effectively modified donor MPS IVA HSCs that were further transplanted into busulfan-myeloablated MPS IVA recipients. This ex vivo LVGT, although the expression of hGALNS was lower, provided full correction of heart tissue and slight correction of bone pathology. We also investigated the potential effects of direct infusion of LVs. In this part of our research, we evaluated age-dependency (newborn vs. older mice), therapeutic doses (low-moderate-high doses), administration routes (intravenous, intramuscular, and intravenous), promoters (CBh, COL2A1, and CD11b) with high potential to drive gene expression in vivo and codon optimization of hGALNS enzyme in vivo. Our data confirmed the efficiency of systemic delivery rather than local administration under high doses of LVs into newborn MPS IVA mice. Parallel to in vitro and ex vivo assessment, we found that intravenous administration of LV-CBh or LV-COL2A1 viral vectors drove the hGALNS transgene expression at high levels. However, most LVs were integrated in the liver and the rest cleared from the body, which resulted in a slight correction of bone pathology. Local administrations, however, did not improve the course of disease biochemically or pathologically. Additionally, we noted an increase in anti-hGALNS antibodies, suggesting an immune response to the therapy. Overall, these findings of a broad range of therapeutic strategies evaluated here indicate that LVGT could be a promising approach for treating MPS IVA and similar skeletal disorders when combined into HSCs, offering hope for future advancements in the field. | |
| Advisor | Tomatsu, Shunji | |
| Degree | Ph.D. | |
| Department | University of Delaware, Department of Biological Sciences | |
| Unique Identifier | 1515021897 | |
| URL | https://udspace.udel.edu/handle/19716/36032 | |
| Language | en | |
| Publisher | University of Delaware | |
| URI | https://www.proquest.com/pqdtlocal1006271/dissertations-theses/em-vivo-ex-lentiviral-stem-cell-therapy/docview/3190112894/sem-2?accountid=10457 | |
| Keywords | Hematopoietic stem cell | |
| Keywords | Enzyme activity | |
| Keywords | Glycosaminoglycan | |
| Title | In vivo and ex vivo lentiviral and stem cell therapy approaches for progressive systemic skeletal dysplasia: mucopolysaccharidosis IVA | |
| Type | Thesis |