Glutathione S-transferases and S-glutathionylation in cancer

CN04-02 The glutathione-S-transferase (GST) super family comprises multiple isozymes (Alpha, Mu, Pi, Omega, Theta, and Zeta) with evidence of functional polymorphic variations. Correlative studies have shown that genetic differences of GST isozyme expression may contribute to cancer susceptibility and treatment. Over the last two decades, a body of data has accumulated linking aberrant expression of GST isozymes with the development and expression of resistance to cancer drugs. In particular, GST Pi (GSTP) is over-expressed in a number of different tumors compared to normal tissues (including ovarian, non-small cell lung, breast, colon, liver, pancreas and lymphoma). Moreover, a significant range of anticancer drugs can cause an increased expression of GSTP in drug resistant selected cell lines (and drug treated patients). From a functional viewpoint, not all drugs used to select for resistance are substrates for catalysis by GSTP. In fact, catalytic constants for GSTP conjugation reactions with virtually all standard cancer drugs are poor. So why is GSTP so frequently over-expressed in these situations? There is recent evidence that GST isozymes, and GSTP in particular, have multiple functions in cells, many unrelated to thioether bond catalysis with chemical moieties. These include: (1) ligand biding and transport of heme, bilirubin, nitric oxide; (2) protein: protein interactions with possible chaperone like functions; (3) regulation of mitogen activated protein kinases, particularly c-jun NH2 terminal kinase (JNK); (4) mediation of the forward reaction of the post-translational process of S-glutathionylation. Of these, aberrant kinase signaling pathways and altered protein S-glutathionylation patterns are both characteristic of the malignant phenotype. What happens in the absence of GSTP? GSTP null animals have essentially normal development and life spans. Mouse embryo fibroblast (MEF) cells isolated from wild type or GSTP null animals differ in a number of characteristics related to signaling and growth. For example, the doubling time for wild type cells is 33.6 h compared to 26.2 h for GSTP null. Both early passage and immortalized MEF cells from GSTP null animals express significantly elevated activities of extracellular regulated kinases (ERK1/ERK2). Null animals had constitutively elevated c-jun NH2-terminal kinase (JNK) activity compared to wild type and this is correlated with altered regulation of genes downstream of JNK. As a whole, the genetic absence of GSTP influences the capacity of stress kinases to regulate gene expression and this can have an impact on cell proliferation pathways. The non-lethality of the deletion points to possible functional redundancy and implies that other GST (or other redox proteins) may substitute for the absence of GSTP. This conclusion would seem to be supported by the data suggesting general redundancy of function amongst, and within, this protein cluster. Cysteine residues in proteins provide a nucleophilic site for a number of disparate post-translational modifications. For example, disulfide bonds between vicinal thiols have major implications for three dimensional protein structures. Contingent upon the steric properties and local environment of the cysteine, lipidation of these residues may occur through S-isoprenylation, S-farnesylation, S-geranylgeranylation or S-palmitoylation. The direct addition of GSH to cysteines with low pKa’s creates an S-glutathionylated residue, resulting in an increase in both molecular weight (of 605) and negative charge (from the glutamic acid residue). The scheme in the Figure summarizes some components of this reaction. GSTP can facilitate the forward reaction and glutaredoxin, sulfiredoxin or thioredoxin can contribute to the reverse reaction. The reversible nature of this pathway is important in facilitating a sulfur based regulatory pathway that can expedite response to stress conditions. Importantly, a number of phosphatases can be regulated by S-glutathionylation and this provides a link and conduit with phosphate based signaling pathways. Because the structure, function and cell distribution of proteins can be affected by S-glutathionylation, the importance of GSTP in mediating this reaction could have significant consequences and may be a contributory factor in the high expression levels of GSTP in many tumors. These multiple functionalities contribute to the recent rational efforts to target GSTs with novel small molecule therapeutics. While the ultimate success of these attempts remains to be shown in the clinic, at least three drugs are in late-stage clinical testing. Two of these (NOV-002 and Telintra) are being tested therapeutically as small molecule myeloproliferative agents. As the field progresses, the concept of designing new drugs that might interfere with protein:protein interactions between GSTs and regulatory kinases provides a novel approach to identify new targets in the search for cancer therapeutics. Relevant References 1. Tew KD. Redox in Redux: Emergent Roles for Glutathione S-Transferase P (GSTP) in Regulation of Cell Signaling and S-glutathionylation. Biochemical Pharmacology 2007;73:1257-69. 2. McIlwain CC, Townsend DM, Tew KD. Glutathione S-Transferase Polymorphisms and Cancer Incidence and Therapy. Oncogene 2006;25: 1639-48.