A large number of human genetic diseases result from mutations that cause premature termination of the synthesis of proteins encoded by mutant genes. Currently, hundreds of such nonsense mutations are known, and several where shown to account for certain cases of fatal diseases, including cystic fibrosis (CF), Duchenne muscular dystrophy (DMD), Tay-Sachs, and more. For many of those diseases there is presently no effective treatment. In the last several years, it was shown that some aminoglycoside antibiotics (including gentamicin) have the ability to allow the mammalian ribosome to selectively read past a false-stop signal, but not a normal termination signal, and generate full-length functional proteins. However, high toxicity of these drugs in humans limits their therapeutic use. The main objective of this research is to develop novel aminoglycosides that will have efficient termination suppression activity, and at the same time will have reduced toxicity against mammalian cells. To date, the clinical application of aminoglycosides is limited to their use as antibacterial drugs, and no efforts have been made to optimize their activity as stop codon read-through inducers. Toward these ends, a collaborative effort is under way between several different laboratories and combines expertise of number of complementary disciplines to synthesize and elucidate the structure-activity- toxicity relationships of the designed drugs. The hypothesis behind the proposal is to separate the elements of the aminoglycosides structures that cause toxicity from those that are required for inducing nonsense suppression: designed structures exhibiting extensive specificity and selectivity for the cytoplasmic rRNA A site can decrease the functional dosing ranges and subsequently decrease the anticipated toxicity, including deleterious effects on mitochondrial protein synthesis machinery, making them potential drugs for the treatments of human genetic disorders. Our hypothesis is based on a series of recent observations in which we have shown that by reducing the specificity to prokaryotic ribosome and as such wiping away from aminoglycosides their """"""""natural"""""""" antibacterial activity we reduce their action on eukaryotic mitochondrial protein synthesis machinery and as such significantly reduce their toxic effects on humans. Substantial therapeutic and economic benefits are anticipated from this study. The designed structures presented here are simple for preparation and the preliminary tests already discovered some variants with lower toxicity and greater read-through efficacy to restore functional CFTR protein from the mutant gene both in vitro and in vivo, including models closely predictive of results with human CF subjects, than those of gentamicin. Fulfillment of the goals of this project will provide the necessary knowledge and tools for uncovering new structures, which may act as novel drugs.

Public Health Relevance

Many human genetic diseases including cystic fibrosis, Duchenne muscular dystrophy, Usher syndrome, Hurler syndrome and numerous types of cancer are caused by nonsense mutations. According to the Human Gene Mutation Database, nonsense mutations represent about 12% of all mutations reported, and for many of those diseases there is presently no effective treatment. The proposed research aims to develop potent aminoglycosides derivatives with high potential for immediate therapeutic application for treatment of genetic diseases caused by nonsense mutations.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM094792-04
Application #
8636483
Study Section
Therapeutic Approaches to Genetic Diseases (TAG)
Program Officer
Marino, Pamela
Project Start
2011-04-01
Project End
2015-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
4
Fiscal Year
2014
Total Cost
$346,427
Indirect Cost
$25,661
Name
Technion-Israel Institute of Technology
Department
Type
DUNS #
600133854
City
Haifa
State
Country
Israel
Zip Code
32000
Shalev, Moran; Rozenberg, Haim; Smolkin, Boris et al. (2015) Structural basis for selective targeting of leishmanial ribosomes: aminoglycoside derivatives as promising therapeutics. Nucleic Acids Res 43:8601-13
Gunn, Gwen; Dai, Yanying; Du, Ming et al. (2014) Long-term nonsense suppression therapy moderates MPS I-H disease progression. Mol Genet Metab 111:374-381
Shulman, Eli; Belakhov, Valery; Wei, Gao et al. (2014) Designer aminoglycosides that selectively inhibit cytoplasmic rather than mitochondrial ribosomes show decreased ototoxicity: a strategy for the treatment of genetic diseases. J Biol Chem 289:2318-30
Shalev, Moran; Baasov, Timor (2014) When Proteins Start to Make Sense: Fine-tuning Aminoglycosides for PTC Suppression Therapy. Medchemcomm 5:1092-1105
Xue, Xiaojiao; Mutyam, Venkateshwar; Tang, Liping et al. (2014) Synthetic aminoglycosides efficiently suppress cystic fibrosis transmembrane conductance regulator nonsense mutations and are enhanced by ivacaftor. Am J Respir Cell Mol Biol 50:805-16
Shalev, Moran; Kondo, Jiro; Kopelyanskiy, Dmitry et al. (2013) Identification of the molecular attributes required for aminoglycoside activity against Leishmania. Proc Natl Acad Sci U S A 110:13333-8
Keeling, Kim M; Wang, Dan; Dai, Yanying et al. (2013) Attenuation of nonsense-mediated mRNA decay enhances in vivo nonsense suppression. PLoS One 8:e60478
Shalev, Moran; Kandasamy, Jeyakumar; Skalka, Nir et al. (2013) Development of generic immunoassay for the detection of a series of aminoglycosides with 6'-OH group for the treatment of genetic diseases in biological samples. J Pharm Biomed Anal 75:33-40
Keeling, Kim M; Wang, Dan; Conard, Sara E et al. (2012) Suppression of premature termination codons as a therapeutic approach. Crit Rev Biochem Mol Biol 47:444-63
Kandasamy, Jeyakumar; Atia-Glikin, Dana; Shulman, Eli et al. (2012) Increased selectivity toward cytoplasmic versus mitochondrial ribosome confers improved efficiency of synthetic aminoglycosides in fixing damaged genes: a strategy for treatment of genetic diseases caused by nonsense mutations. J Med Chem 55:10630-43

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