Seguin, Sandlin P
(2011)
Identification of Small Molecule Inhibitors of Polyomavirus Replication.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
Polyomaviruses (PyVs) are ubiquitous DNA viruses that are not generally associated with pathogenicity. However, in immunosuppressed populations, PyVs can cause diseases, including BK Virus associated nephropathy, hemorrhagic cystitis and progressive multifocal leukoencephalopathy. There is currently no PyV specific inhibitor for these diseases. Because all PyVs express a conserved large T antigen (TAg) that is essential for viral replication, I hypothesized that inhibitors of the model TAg from Simian Virus 40 (SV40) would inhibit the replication of PyVs in general. TAg has multiple essential activities, but my work focused on the ATPase activity of TAg, which provides the helicase activity during viral replication. Two high throughput screens were performed on purified TAg to identify inhibitors of TAg ATPase activity. In the first screen, a quinaldine red screen of the MS2000 library of commercially available compounds identified the FDA approved compounds bithionol and hexachlorophene as inhibitors of the ATPase activity of TAg. The structure activity relationship of these bisphenols was refined, and the results suggested that inhibition of TAg was unique to these compounds, and is not a general feature of bisphenols. The compounds also inhibited SV40 DNA replication and infection, and are the most potent SV40 inhibitors reported. Surprisingly, bithionol and hexachlorophene inhibit replication of the clinically relevant PyV BK Virus (BKV), but not to the same extent.In the second screen, which employed the ADP Hunter assay and the NIH Molecular Libraries Probe Centers Network (MLPCN) compound library, three scaffolds of interest were identified. However, all three had limited potency when characterized in other in vitro assays. Chemical refinement of these scaffolds identified another bisphenol that inhibits TAg activity and TAg dependent DNA replication in vitro. Unfortunately, the known cytotoxicity of this compound limits its use as a therapeutic. Although I have not yet identified a PyV specific inhibitor that would be a suitable therapeutic, this work supports my overall hypothesis: small molecules that inhibit the ATPase activity of TAg reduce viral DNA replication, and viral infection. My work has also provided valuable insights to design future screens to identify inhibitors of PyV replication.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
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| ETD Committee: |
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| Date: |
30 September 2011 |
| Date Type: |
Completion |
| Defense Date: |
29 June 2011 |
| Approval Date: |
30 September 2011 |
| Submission Date: |
15 July 2011 |
| Access Restriction: |
5 year -- Restrict access to University of Pittsburgh for a period of 5 years. |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Dietrich School of Arts and Sciences > Biological Sciences |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
drug discovery; molecular virology; transplant science |
| Other ID: |
http://etd.library.pitt.edu/ETD/available/etd-07152011-155444/, etd-07152011-155444 |
| Date Deposited: |
10 Nov 2011 19:51 |
| Last Modified: |
15 Nov 2016 13:46 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/8396 |
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