All ETDs from UAB

Advisory Committee Chair

David M Bedwell

Advisory Committee Members

Kim M Keeling

Robert A Kesterson

Trenton R Schoeb

Jphn J Shacka

Document Type

Dissertation

Date of Award

2017

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Nonsense mutations introduce a premature termination codon (PTC) into the open reading frame of an mRNA resulting in premature translation termination, loss of functional protein, and rapid degradation of the mutant mRNA. Approximately 11% of human genetic disorders are attributable to a nonsense mutation. Several small molecules have been identified as potential nonsense suppression compounds. These compounds increase the frequency of PTC recognition by near-cognate aminoacyl tRNAs resulting in PTC “readthrough” and restored protein production. Mucopolysaccharidosis type I-Hurler (MPS IH) is the severe form of the autosomal recessive lysosomal storage disorder caused by mutations in the iduronidase (IDUA) gene. 60-80% of the MPS IH patient population have a nonsense mutation in at least one allele of the gene. Preliminary studies in our laboratory indicated that the designer aminoglycoside NB84 could restore functional enzyme and alleviate GAG accumulation both in vitro and in vivo in a nonsense model of MPS IH. Long-term, early intervention treatment with NB84 in the W402X mouse model resulted in sustained restoration of enzyme activity, reduced GAG accumulation, slowed disease progression in multiple tissues, and no evidence of toxicity. These results demonstrate that nonsense suppression therapy can effectively ameliorate disease progression and that designer aminoglycosides represent a safe and more effective alternative to traditional aminoglycosides. The non-aminoglycoside compounds PTC124, 414, and 415 were identified by high throughput screening as candidates for nonsense suppression therapy. Extensive evidence demonstrates that PTC124 is orally bioavailable and induces nonsense mutation readthrough in a variety of disease contexts. We evaluated drug mechanism, safety and efficacy of these three compounds using heterologous reporter assays, in vitro, and in vivo studies in the context of MPS IH. We found that all three compounds resulted in decreased GAG accumulation in vitro. PTC124 produced modest but statistically significant improvements in vitro after both short- and long-term administration. Our results demonstrate that nonsense suppression therapy is a viable approach for the treatment of genetic disorders but that, due to its modest efficacy, it may be best used in combination with existing or new therapies.

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