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Differential ganciclovir-mediated cell killing by glutamine 125 mutants of herpes simplex virus type 1 thymidine

R R Drake1, T N Wilbert, T A Hinds

  • 1Department of Biochemistry, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA. drakerickr@exchange.uams.edu

The Journal of Biological Chemistry
|December 22, 1999
PubMed

Insights

Herpes simplex virus type 1 thymidine kinase (HSV-1 TK) gene therapy with ganciclovir (GCV) shows promise for cancer treatment. Mutants of HSV-1 TK influence GCV metabolism and cell death mechanisms, offering insights into optimizing this cancer therapy.

Area of Science:

  • Molecular Biology
  • Cancer Therapy
  • Enzyme Engineering

Background:

  • The herpes simplex virus type 1 thymidine kinase (HSV-1 TK) gene combined with ganciclovir (GCV) is a promising cancer therapy.
  • GCV is phosphorylated by HSV-1 TK to a toxic metabolite that induces tumor cell death.
  • The precise mechanisms of GCV metabolite-induced cell death are not fully understood.

Purpose of the Study:

  • To investigate the cellular and pharmacological mechanisms of GCV-induced cell death.
  • To characterize the function of mutated HSV-1 TK enzymes at residue Gln-125.
  • To explore how distinct TK mutations affect GCV metabolism and cancer cell responses.

Main Methods:

  • Expression of wild-type and mutated HSV-1 TK (Asn-125, Glu-125) in mammalian cell lines (NIH3T3, HCT-116).
  • Assessment of GCV metabolism, sensitivity, bystander effect, and apoptosis induction.
  • Utilized cell cycle analysis, DAPI staining, and caspase 3 activation assays.

Main Results:

  • The Asn-125 HSV-1 TK mutant showed efficacy comparable to wild-type TK in GCV metabolism, cell killing, and apoptosis, but lacked deoxypyrimidine metabolism.
  • The Glu-125 HSV-1 TK mutant exhibited attenuated GCV metabolism, with diminished but still effective cell sensitivity and bystander killing at higher GCV concentrations.
  • Distinct cell death responses were observed for Glu-125 TK-expressing cells compared to wild-type or Asn-125 TK cells, linked to GCV-triphosphate levels.

Conclusions:

  • Mutations in HSV-1 TK at Gln-125 significantly alter GCV metabolism and downstream cell death pathways.
  • The Asn-125 mutant retains therapeutic potential with altered metabolic profiles.
  • The Glu-125 mutant provides insights into differential GCV-induced apoptosis and bystander effects, guiding future therapeutic optimization.

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