Date of Award:

5-1-1981

Document Type:

Dissertation

Degree Name:

Doctor of Philosophy (PhD)

Department:

Biology

Department name when degree awarded

Life Sciences:Biology

Committee Chair(s)

Robert W. Sidwell

Committee

Robert W. Sidwell

Committee

Rex S. Spendlove

Committee

Bill B. Barnett

Committee

Rhagubir P. Sharma

Committee

Thomas F. Emery

Abstract

Known RNA virus inhibitors were evaluated in vitro and in vivo to determine if any had specific inhibitory effects on reo-, rota- and orbivirus infections. Test compounds included 1-ß-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (ribavirin), 3-deazaguanine (3-DG), 3-deazauridine (3-DU), and 9-(S)-(2,3-dihydroxypropyl) adenine ([S]-DHPA). Certain selected studies were also run with the following ribavirin derivatives: ribavirin 2' ,3' ,5'-triacetate, ribavirin 5'-monophosphate and ribavirin 5'-triphosphate. Viruses included types 1, 2, and 3 reovirus; bovine, porcine and simian rotaviruses; and bluetongue and Colorado tick fever orbiviruses. Results of the in vitro antiviral studies indicated ribavirin to be the most potent anti-reovirus agent, and 3-DG the strongest rota- and orbivirus inhibitor. Ribavirin, 3-DG, and 3-DU possessed in vitro activity against all viruses, whereas (S)-DHPA was inactive against Colorado tick fever virus and only weakly active against reoviruses. Cytopathic effect inhibition, immunofluorescent cell count reduction, and plaque reduction assays were equally useful to determine antiviral activities of the test compounds. The agents also inhibited the production of infectious simian rotavirus particles in MA-104 cell cultures. Actinomycin D reversed the anti-rotavirus activity of all compounds except (S)-DHPA, and anti-reovirus activities were reversed by the appropriate natural nucleoside (either guanosine, uridine, or adenosine). All drugs reduced the levels of dense (precursor) and light (complete) simian rotavirus particle yields but did not alter the dense to light particle ratio, suggesting the agents did not interfere with virus assembly. Only ribavirin and 3-DG inhibited rotavirus polypeptide synthesis, and none of the agents nor phosphorylated derivatives of ribavirin inhibited simian rotavirus polymerase activity. Radiolabeled precursor uptake studies indicated each compound to inhibit 3H-thymidine, 3H-uridine, and/or 3H-leucine or 3H-amino acid incorporation into acid soluble and acid insoluble fractions of MDBK and MA-104 cells. Minimum anti-cellular effects occurred at doses approximating minimum antiviral concentrations, but the results suggest that viral and cellular functions may be inhibited by different mechanisms. In mouse studies, ribavirin triacetate increased the number of surviving mice infected intracerebrally with Colorado tick fever virus. 3-DG increased mean survival time, and ribavirin produced no beneficial effect against the infection. Ribavirin triacetate may be able to cross the blood-brain barrier in concentrations high enough to be virus-inhibitory, based upon an increase in survivors in intraperitoneally-treated, infected mice. Against murine rotavirus infections, (S)-DHPA and ribavirin triacetate treatment increased mean survival time. Infected mice treated with (S)-DHPA and 3-DG gained weight at a considerably faster rate than rotavirus-infected controls, suggesting these agents may have lessened the severity of the disease in survivors. This study has served to establish methodologies and techniques for evaluating antiviral substances against infections caused by viruses of the Reoviridae family. The antiviral activities of four known RNA virus inhibitors have been defined, possible mechanisms of drug action were determined, and the usefulness of treating Reoviridae-induced animal diseases with the compounds was examined. The methods employed in this report will be applicable to evaluate other, possibly more specific anti-Reoviridae compounds.

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