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Role of GRP58 in mitomycin C-induced DNA cross-linking

Claudia M Celli1, Anil K Jaiswal

  • 1Department of Pharmacology, Baylor College of Medicine, Houston, Texas 77030, USA.

Cancer Research
|October 3, 2003
PubMed

Insights

Researchers identified glucose-regulatory protein 58 (GRP58) as a key enzyme in activating the anticancer drug Mitomycin C (MMC). GRP58 enhances MMC-induced DNA cross-linking and cytotoxicity, offering a new target for cancer therapy.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Cancer Research

Background:

  • Mitomycin C (MMC) is an anticancer drug requiring reductive activation for its cytotoxic effects.
  • DNA cross-linking by MMC is a primary mechanism for cell death, with NQO1 and microsomal enzymes implicated in its activation.
  • NQO1's minor role suggests other cytosolic factors contribute significantly to MMC-induced DNA cross-linking.

Purpose of the Study:

  • To identify and characterize the unique cytosolic activity responsible for Mitomycin C (MMC)-induced DNA cross-linking in NQO1-null mice.
  • To elucidate the role of Glucose-Regulatory Protein 58 (GRP58) in MMC activation and its potential as a therapeutic target.

Main Methods:

  • Purification and peptide sequencing of the unique cytosolic activity from NQO1-null mouse liver cytosol.
  • Immunodepletion assays using anti-GRP58 antibodies to assess its role in MMC-induced DNA cross-linking.
  • Overexpression studies in Chinese hamster ovary (CHO) cells and functional assays with bacterially expressed GRP58.
  • Tissue array analysis to determine GRP58 expression in mouse tissues and human tumors.

Main Results:

  • A unique, cofactor-independent, and dicoumarol-insensitive cytosolic activity was identified and purified.
  • Peptide sequencing identified this activity as Glucose-Regulatory Protein 58 (GRP58).
  • Immunodepletion of GRP58 abolished MMC-induced DNA cross-linking, confirming its essential role.
  • GRP58 overexpression in CHO cells increased MMC-induced DNA cross-linking and cytotoxicity.
  • GRP58 is ubiquitously expressed in mouse tissues and upregulated in human breast, uterus, lung, and stomach tumors.

Conclusions:

  • Glucose-Regulatory Protein 58 (GRP58) is a critical enzyme mediating reductive activation of Mitomycin C (MMC).
  • GRP58 significantly contributes to MMC-induced DNA cross-linking and cytotoxicity.
  • GRP58's upregulation in various human tumors suggests its potential as a predictive biomarker and therapeutic target in cancer treatment.

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