HSV1-specific thymidylate kinase activity in infected cells

Area of Science:

• Virology
• Molecular Biology
• Antiviral Drug Development

Background:

• Herpes simplex virus type 1 (HSV1) is a significant human pathogen.
• Antiviral drugs targeting viral enzymes are crucial for managing HSV1 infections.
• Understanding drug resistance mechanisms is vital for developing effective therapies.

Purpose of the Study:

• To classify 5-methoxymethyldeoxyuridine (MMdU)-resistant HSV1 mutants.
• To investigate the relationship between drug resistance and viral enzyme activity.
• To determine the impact of enzyme alterations on sensitivity to various antiviral drugs.

Main Methods:

• Generation and isolation of MMdU-resistant HSV1 mutants.
• Sensitivity testing of viral mutants against deoxythymidine kinase (dTK)-dependent drugs (acyclovir, araT, BVdU) and dTK-independent drug (PAA).
• Enzymatic assays to assess the phosphorylation activity of viral dTK in mutant strains.

Main Results:

• Five dTK-negative (dTK-) mutants exhibited high resistance to BVdU and MMdU (≥500-fold), moderate resistance to acyclovir (ACV) and araT (10- to 100-fold), and no resistance to PAA.
• The MMdUr-20 mutant (dTK+) showed reduced dTMP phosphorylation by its dTK, without altered deoxythymidine affinity.
• Other drug-resistant mutants (S1, B3) also displayed diminished dTMP phosphorylation.

Conclusions:

• Drug resistance in HSV1 can arise from mutations affecting deoxythymidine kinase (dTK) activity.
• Alterations in thymidylate kinase activity may also contribute to antiviral drug resistance.
• These findings provide insights into the molecular basis of HSV1 drug resistance and inform future therapeutic strategies.