Role of glutathione and glutathione S-transferase in chlorambucil resistance.

A chlorambucil (CLB)-resistant cell line, N50-4, was developed from the established mouse fibroblast cell line NIH 3T3, by multistep drug selection. The mutant cells exhibited greater than 10-fold resistance to CLB. Alterations in GSH and glutathione S-transferase (GST) were found in CLB-resistant variants. A 7-10-fold increase in cellular GSH content and a 3-fold increase in GST activity were detected in N50-4 cells, compared with parental cells, as determined by enzymatic assays. An increase in steady state levels of the GST-alpha isozyme mRNA was found in the CLB-resistant cells, as analyzed by Northern blotting. No GST gene amplification or rearrangement was shown by Southern blot analysis. To test the relative roles of GSH and GST in CLB resistance, a number of GSH- and GST-blocking agents were used. The CLB toxicity was significantly enhanced in N50-4 cells by administration of either the GSH-depleting agent buthionine sulfoximine or the GST inhibitors ethacrynic acid or indomethacin. The resistance to CLB cytotoxicity in N50-4 cells, however, was still significantly higher than that of parental cells. The resistance of N50-4 cells to CLB was almost completely abolished by combination pretreatment yielding both GSH depletion and GST inhibition. The results indicate that both increased cellular GSH content and increased GST activity play major roles in CLB resistance in N50-4 mutant cells.     

Metabolism of nitroglycerin by smooth muscle cells. Involvement of glutathione and glutathione S-transferase.

Metabolism of nitroglycerin (GTN) in the vascular smooth muscle is required for the drug to be effective in the treatment of angina pectoris and congestive heart failure. The usefulness of GTN is limited by the development of tolerance to the drug. The metabolism of GTN was studied in its target tissue, vascular smooth muscle. Inorganic nitrite was produced by cultured smooth muscle cells when GTN was added to the culture dish. Nitrite production increased with increasing GTN concentration and with incubation time. The enzymatic nature of GTN metabolism to nitrite was assessed by enzyme inhibition studies. Indocyanine green, a non-substrate inhibitor of glutathione S-transferase, inhibited GTN metabolism by smooth muscle cells. Cellular glutathione is also involved in GTN metabolism by the smooth muscle cell. Pretreatment with phorone, a glutathione S-transferase substrate, depleted cellular glutathione and decreased nitrite production from GTN. Pretreatment with buthionine sulfoximine, inhibitor of gamma-glutamylcysteine synthetase, decreased intracellular glutathione and caused decreased GTN metabolism in smooth muscle cells. Removal of cysteine from the smooth muscle cell incubation medium in combination with buthionine sulfoximine pretreatment decreased GTN metabolism to a lower level than buthionine sulfoximine pretreatment alone. This study shows that glutathione S-transferase and glutathione are involved in GTN metabolism by cultured smooth muscle cells.     

Heterogeneity of glutathione S-transferase enzyme and gene expression in ovarian carcinoma.

Expression of the three major cytosolic classes of glutathione S-transferases (GST; Pi, Alpha and Mu) was examined by 2D gel analysis and Western blotting of biopsies from 26 patients diagnosed with ovarian carcinoma. In contrast to other tissues, at least one 'constitutive' subunit from each of the three major cytosolic GST classes was expressed. In most cases, pi appeared to be the major form present, although levels of alpha and mu subunit expression were approximately equal to pi in some patients. There was no detectable effect of prior chemotherapy on enzyme activity. Mean transferase activity for primary carcinoma was 79.9 +/- 11.9 (mean +/- SEM; nmol min-1 mg-1), with three pair-matched normal tissues showing minor decreases in transferase activity. One sample, in which a 32% increase in tumour enzyme activity was noted, was from a patient with primary disease and was associated with marked overexpression of a relatively basic form of alpha which was absent from the matching normal tissue, but present in 20% of all tumours examined. RFLP analysis of genomic tumour DNA using a human mu class cDNA probe indicated that at least two of the three mu forms (the 'constitutive' form and one other) observed in ovarian tissue were allelic variants, as a one-to-one correlation was observed between the presence of two Hind III fragments at 13.1 and 2.2 kb and expression of a second, more basic, variable form. This latter form was positively identified as the mu class subunit mu based on Southern analysis and was seen to be present in 40% of the samples examined. However, in the absence of mu expression, at least one other mu class subunit probably corresponding to GST psi, was seen to be present. Thus, at least in ovarian tissues, absence of the mu subunit does not necessarily imply a lack of ability to metabolize mu substrates, as psi has similar catalytic activity. A third mu subunit, probably corresponding to GST phi based on its relatively acidic pI, was also noted in 72% of samples examined, but has unknown substrate specificity. Increased expression of both alpha and mu forms may be of relevance to disease diagnosis and drug response.     

谷胱甘肽-S转移酶-中期因子融合蛋白原核表达及多克隆抗体的制备

目的 构建谷胱甘肽-S-转移酶(GST)和中期因子(MK)融合蛋白的原核表达质粒,并表达和纯化蛋白,制备多克隆抗体.方法 通过RT-PCR技术从人胃癌组织中扩增入MK编码序列,克隆入表达载体pGEX-1λT中,获得表达质粒pGEX-MK,并在大肠杆菌BL21 (DE3)中经IPTG诱导表达,通过亲和层析纯化表达的GST-MK融合蛋白,并以重组蛋白免疫兔子.结果 成功构建了GST-MK融合蛋白的原核表达载体,经诱导表达纯化得到GST-MK融合蛋白.免疫兔子后取多抗血清以间接ELISA检测效价达1∶64 000,Western blotting分析显示多克隆抗血清对MK蛋白特异结合.结论 MK在大肠杆菌中成功表达及其多克隆抗体的获得,为研究MK生物功能奠定了基础.     

Role of glutathione S-transferase 8-8 in allylamine resistance of vascular smooth muscle cells in vitro.

Allylamine (AA) is a cardiovascular toxin that causes lesions resembling atherosclerosis in several mammalian species. AA's toxic effects are thought to be exerted through its conversion to acrolein (AC), a potent electrophilic alkylating agent and atherogen. Semicarbazide sensitive amine oxidase (SSAO) catalyzes the oxidation of AA to AC. Glutathione S-transferases (GST) can catalyze the first step of detoxification of AC to mercapturic acid. Our previous studies suggest that the isozyme rGST8-8 is a principal defense against electrophilic stress exerted by alpha,beta-unsaturated carbonyls such as AC. In the present studies, we use cultured rat vascular smooth muscle cells (VSMC) to examine the relative roles of SSAO and rGST8-8 in the cytotoxic effects of the atherogens, AA and AC. Exposure derived AA-resistant cells (VSMC-AA) were 3.5-fold more resistant to AA when compared to VSMC and 1.8-fold more resistant to acrolein. SSAO activity was 2-fold higher in VSMC-AA than in VSMC. Consistent with the role of SSAO in biotransformation of AA, the SSAO inhibitor semicarbazide (SC; 100 microM) provided nearly complete protection from AA to both VSMC-AA and VSMC. As expected, SC did not affect the cytotoxicity of AC. Pretreatment with 100 microM sulfasalazine (SS), a GST inhibitor, potentiated AA and AC toxicity in both VSMC-AA and VSMC, indicating a protective role of GST. Catalytic efficiency (K(cat)/K(m)) of GSTs was higher toward 4-hydroxynonenal (4-HNE) (0.65 mM(-1) s(-1)) than toward 1-chloro-2, 4-dinitrobenzene (CDNB) (0.14 mM(-1) s(-1)) for VSMC. In VSMC-AA, K(cat)/K(m) was increased 4.1-fold toward CDNB (0.58 mM(-1) s(-1)) and 6-fold toward 4HNE (3.9 mM(-1) s(-1)) when compared to VSMC, indicating a preferential increase in VSMC-AA of GST isozymes which utilize alpha,beta-unsaturated carbonyls. Western blots confirmed induction of rGST8-8 in VSMC-AA. Expression of recombinant mGSTA4 (the mouse homolog of rGST8-8) in VSMC caused a 1.6-fold increase in resistance to AA and AC. This resistance was fully reversed by 50 microM SS. Our results demonstrate that GSTs are an important defense against electrophilic atherogens and that isozymes with high activity toward alpha,beta-unsaturated carbonyls are particularly important in the vascular wall.     

Genotoxicity of 4-hydroxy-2-nonenal in human colon tumor cells is associated with cellular levels of glutathione and the modulation of glutathione S-transferase A4 expression by butyrate.

The cellular production of 4-hydroxy-2-nonenal (HNE), a product of endogenous lipid peroxidation, constitutes a genotoxic risk factor for carcinogenesis. Our previous studies have shown that human HT29 colon cells developed resistance toward HNE injury after treatment with butyrate, a diet-associated gut fermentation product. This resistance was attributed to the induction of certain glutathione S-transferases (hGSTP1-1, hGSTM2-2, and hGSTA1-1) and also for the tripeptide glutathione (GSH) synthesizing enzymes. In the present study, we have investigated in HT29 cells whether hGSTA4-4, which has a high substrate specificity for HNE, was also inducible by butyrate and, thus, could contribute to the previously observed chemoresistance. In addition, we investigated if cellular depletion of GSH by L-buthionine-S,R-sulfoximine (BSO) enhances chemosensitivity to HNE injury in HT29 cells. Incubation of HT29 cells with butyrate (2-4 mM) significantly elicited a 1.8 to 3-fold upregulation of steady state hGSTA4 mRNA over 8-24 h after treatment. Moreover, 4 mM butyrate tended to increase hGSTA4-4 protein concentrations. Incubation with 100 microM BSO decreased cellular GSH levels by 77% without significant changes in cell viability. Associated with this was a 2-fold higher level of HNE-induced DNA damage as measured by the comet assay. Collectively, the results of this study and our previous work indicate that the genotoxicity of HNE is highly dependent on cellular GSH status and those GSTs that contribute toward HNE conjugation, including hGSTA4-4. Since HNE contributes to colon carcinogenesis, the favorable modulation of the GSH/GST system by butyrate may contribute to chemoprevention and reduction of the risks.     

Glutathione and glutathione S-transferase activities of mammalian cells in culture

Continuous cell cultures derived from various tissues of rat, mouse, hamster and man were assayed for their glutathione (GSH) content and glutathione S-transferase activities. GSH S-transferase activities were monitored toward the substrates 1-chloro-2,4-dinitro-benzene (CDNB), 1,2-dichloro-4-nitrobenzene (DCNB) and 1,2-epoxy-3-(p-nitrophenoxy)propane (PO). All cell lines tested contained appreciable amounts of GSH ranging from 10 to 65 mol/mg cellular protein. Likewise, all cell lines expressed GSH S-transferase activities. However, the various cell lines differed considerably in their relative transferase activities exhibiting some degree of species-specificity.     

催化合成谷胱甘肽的基因工程菌的构建及表达

目的利用基因工程菌合成谷胱甘肽。方法克隆γ-谷氨酰转肽酶,构建重组质粒pET30a-GGT,乳糖诱导融合蛋白的表达,利用pET30a-GGT/E.coliBL21和pET32a-GSH-II/E.coliBL21分两步法合成γ-谷氨酰半胱氨酸与谷胱甘肽。结果工程菌的最佳乳糖诱导条件为:浓度3mmoL.L-1、时间6h、温度28℃。工程菌经最佳条件诱导后,γ-谷氨酰转肽酶的酶活大约是出发菌株E.coliDH5α的21倍,酶活力得到明显提高,谷胱甘肽的产量达到0.524g.L-1。结论成功地为今后利用基因工程菌催化合成谷胱甘肽寻找到一种新的方法 。     

Induction of glutathione S-transferase activity and protein expression in brown bullhead (Ameiurus nebulosus) liver by ethoxyquin.

The inducibility of hepatic cytosolic glutathione S-transferases (GSTs) was examined in brown bullheads, a freshwater fish that is highly susceptible to hepatic neoplasia following exposure to carcinogen-contaminated sediments. Juvenile bullheads were fed a semi-purified antioxidant-free diet supplemented with ethoxyquin (0.5% w/w dissolved in 3% corn oil), a prototypical rodent GST-inducing agent, twice daily for 14 days. Control bullheads received the antioxidant-free diet supplemented with corn oil (3% w/w). A significant increase (1.6-fold, p < or = 0.01) in hepatic cytosolic GST activity toward 1-chloro-2,4-dinitrobenzene (CDNB) was observed in the ethoxyquin-treated bullheads relative to control fish. A trend toward increased GST-NBC activity was observed in the ethoxyquin-treated fish (1.2-fold, p = 0.06), whereas no treatment-related effects were observed on GST activities toward ethacrynic acid (ECA). In contrast, GST activity toward (+/-)-anti-benzo[a]pyrene-trans-7,8-dihydrodiol-9,10-epoxide (BPDE) was repressed in affinity-purified cytosolic fractions prepared from ethoxyquin-treated bullheads relative to control bullheads. Silver staining and densitometric analysis of isoelectric-focused, affinity-purified GST proteins revealed increased expression of two basic GST-like isoforms in ethoxyquin-treated fish. In summary, exposure to ethoxyquin increases brown bullhead GST-CDNB catalytic activity and hepatic cationic GST protein expression. However, the increase in overall GST-CDNB activity by ethoxyquin is associated with repression of GST-BPDE activity, suggesting differential effects on hepatic bullhead GST isoforms by ethoxyquin. The potential repression of bullhead GST isoforms that conjugate the carcinogenic metabolites of PAH metabolism under conditions of environmental chemical exposure could be a contributing factor in the sensitivity of bullheads to pollutant-associated neoplasia.     

Expression of glutathione S-transferase (GST) genes in the marine copepod Tigriopus japonicus exposed to trace metals

The intertidal copepod, Tigriopus japonicus has been recognized as a potential model species for marine pollution toxicity testing. Toxicity ranges of several biocides, endocrine-disrupting chemicals (EDCs), and trace metals are known in T. japonicus. A large number of expressed sequence tags (ESTs) and genomic DNA are also sequenced from T. japonicus. In this study, expression of ten glutathione S-transferase (GST) genes was studied in the copepods exposed to trace metals. Expression of these genes was also studied against exposure to hydrogen peroxide (H(2)O(2)) used as a positive control with prooxidant activity. Of all genes, expression of GST-Sigma (GSTS) was highly upregulated in H(2)O(2) as well as trace metal-exposed copepods. In the time-course study, expression of GSTS mRNA was more consistent compared to other GSTs such as GST-Omega, GST-Delta1, GST-Theta3 or microsomal GST1 (mGST1). GSTS is predominantly reported from the insects. Coupled with the previous study of the in vitro antioxidant role of T. japonicus GSTS, these findings imply an antioxidant role for GSTS and highlight its importance as a biomarker of exposure to trace metals in T. japonicus. However, further validation and field trials would be necessary to propose GSTS gene expression as biomarker of exposure to trace metals, as for some trace metals such as silver the response was not consistent in concentration and time-series exposure experiments.     

Constitutive mRNA expression of various glutathione S-transferase isoforms in different tissues of mice.

Glutathione S-transferase (Gst) enzymes are instrumental in protecting cellular macromolecules against electrophiles and products of oxidative stress. Of interest primarily to pharmacologists and toxicologists is the ability of these enzymes to metabolize cancer chemotherapeutic drugs, insecticides, herbicides, and carcinogens. Thus, constitutive expression of Gsts might determine a tissue's ability to handle certain forms of chemical stress. In the present study, the constitutive mRNA expression of 19 different Gst enzymes was investigated in 14 different tissues in mice. The information obtained from the present study could be distilled into a few generalized principles: in all tissues examined, multiple isoforms of Gst were constitutively expressed; several isoforms, such as Gstk1, Gstm1, Gstm4, Gstm6, and Gstt1, were expressed in most of the tissues studied; at least five Gst isoforms were highly expressed in the gonads, about three in heart, and at least one in brain (Gstm5). Gender differences in the expression of various Gst isoforms were pronounced. With a few exceptions, most of the Gst isoforms expressed in kidney showed higher expression in females than males; the same trend was observed for heart and gonads. At least eight Gst isoforms showed very high expression in stomach. This was a unique finding in the current study because drug-metabolizing enzymes that are highly expressed in the gastrointestinal (GI) tract tend to have the highest expression in small intestine with low or no expression in the stomach. In summary, most Gst isoforms are most highly expressed in the GI tract and liver, which strongly suggests an important role of many Gst isoforms in detoxification of ingested xenobiotics.     

Detoxification of 1-chloro-2,4-dinitrobenzene in MCF7 breast cancer cells expressing glutathione S-transferase P1-1 and/or multidrug resistance protein 1.

We examined the roles of glutathione S-transferase (GST) P1-1 and the glutathione S-conjugate (GS-X) transporter, multidrug resistance protein 1 (MRP1), singly or in combination, in the detoxification of 1-chloro-2,4-dinitrobenzene (CDNB). Derivatives of MCF7 breast carcinoma cells expressing GST P1-1 and MRP1 alone or in combination were developed. Detoxification was measured in cells as formation of the glutathione conjugate of CDNB, S-(2,4-dinitrophenyl)-glutathione (DNP-SG), efflux of DNP-SG, and ultimately protection from CDNB cytotoxicity. MRP1 expression in the absence of GST P1-1 confers a three- to fourfold resistance to CDNB, which is associated with a >10-fold increase in the maximum rate of DNP-SG efflux. DNP-SG efflux in MRP1-expressing MCF7 cells was ATP-dependent and exhibited an apparent Km for DNP-SG of 95 microM. MRP1 expression alone, however, had no effect on DNP-SG formation. Combined expression of GST P1-1 and MRP1 increased the rates of DNP-SG formation when cells were exposed to 10 microM CDNB. Moreover, combined expression of GSTP1-1 with MRP1 moderately augmented MRP1-mediated resistance to CDNB but only during short term (10 min) exposures to CDNB where IC50 values were in the 8-10 microM range. In contrast, expression of GST P1-1 in the absence of MRP1 slightly sensitized cells to the toxicity of CDNB (10 min exposures), despite increasing rates of DNP-SG formation. The sensitization to CDNB in cells expressing GST P1-1 alone was associated with increased intracellular accumulation of DNP-SG, indicating that DNP-SG may contribute to CDNB toxicity. The potential toxicity of DNP-SG is also suggested by the finding that inhibition of DNP-SG formation by prior glutathione depletion confers resistance to CDNB cytotoxicity in MRP1-poor MCF7 cells. Altogether, our results demonstrate that glutathione conjugation and MRP1-mediated conjugate efflux can operate together to confer resistance to CDNB. The data indicate that MRP1-mediated conjugate efflux is required for cytoprotection from CDNB because its conjugate (DNP-SG), when present at high intracellular levels, may also be toxic to cells.     

Anthracycline resistance in murine leukemic P388 cells. Role of drug efflux and glutathione related enzymes

Energy-dependent drug efflux is a major factor in cellular resistance of P388/R84 mouse leukemic cells to anthracyclines such as doxorubicin (DOX), and blocking of efflux increases sensitivity. However, efflux does not play a significant role in resistance to N-trifluoroacetyladriamycin-14-valerate (AD 32), a DOX analog. Since drug efflux alone cannot account for resistance to anthracyclines, we have, in the present study, measured cellular glutathione (GSH) content and activity of GSH cycle related enzymes to determine their role in resistance. Cellular GSH content was similar in DOX-sensitive and -resistant mouse leukemic cells (P388 and P388/R84). GSH peroxidase, glucose-6-phosphate dehydrogenase and glutathione reductase activities were 1.36-, 1.58- and 1.14-fold higher in P388/R84 cells. Incubation of P388/R84 cells with 100 microM buthionine-S,R-sulfoximine (BSO) for 24 hr reduced cellular GSH content to 6% of control and reduced their resistance to DOX [dose modification factor (DMF) 3.9]. GSH depletion had no significant effect on the cytotoxicity of AD 32 (DMF 1.5). Exposure of P388/R84 cells to BSO (for GSH depletion) and trifluoperazine (for efflux blocking) further reduced their resistance to DOX (DMF 14). These results indicate that DOX resistance in P388/R84 cells is multifactorial and that changes in GSH cycle related enzymes such as GSH peroxidase may also contribute to their resistance.     

Effect of monensin on liver glutathione, glutathione S-transferase and monooxygenases in rats.

Monensin administered ip to male rats at a dosage of 2.5 mg/kg/d for 3 consecutive days did not change the liver levels of glutathione, but depressed significantly the amount of cytochrome P-450 and the activities of aniline hydroxylase and a cytosolic CDNB-specific glutathione S-transferase. There was a marked decrease in the aminopyrine N-demethylase activity and a significant increase in the pentobarbital sleeping time in rats treated with monensin. In contrast, no change in these parameters was found 2 h after a single ip dose (7.5 mg/kg) of monensin. The results suggest that monensin-induced inhibition of the liver cytosolic glutathione S-transferase and microsomal monooxygenases is non-specific.     

Inhibition of glutathione S-transferase M1 by new gabosine analogues is essential for overcoming cisplatin resistance in lung cancer cells

A new class of human GST inhibitors has been identified via rational design approach; we report their discovery, synthesis, inhibitory activity, and synergetic effect in combination with cisplatin against A549 lung cancer cell line. The results of this effort show that the lead 4-O-decyl-gabosine D (24) has optimum synergetic effect in A549 human lung adenocarcinoma epithelial cell and that this activity involves inhibition of glutathione S-transferase M1, apparently consistent with siRNA-mediated knockdown of GSTM1 gene.     

Effects of lead on glutathione S-transferase expression in rat kidney: a dose-response study.

Glutathione S-transferases (GST, EC 2.5.1.18) are a family of phase II detoxification enzymes involved in the conjugation of glutathione to a highly diverse group of compounds. The purpose of this study was to evaluate the dose-response effects of lead acetate administration on the expression of rat kidney GST. Sprague-Dawley rats were injected with doses of lead acetate ranging from 0.11 to 114 mg/kg (0.3 to 300 mumol/kg) for three consecutive days and sacrificed 24 h later. Kidney GST activity, GST isoform HPLC profiles, blood lead analysis, and electron microscopy were performed. A dose of 1.1 mg/kg lead acetate resulted in a blood lead level of 26 micrograms/dl and produced a significant increase in GST activity which continued to increase with dose up to 38 mg/kg. Morphological changes were detected at 3.8 mg/kg and increasing severity of cellular damage paralleled dose, blood lead levels, and changes in body weight. Individual GST isoforms exhibited different thresholds and maxima; rGSTP1 and rGSTM1 had thresholds of 1.1 and 3.8 mg/kg, respectively, very similar rates of increase with dose, and a maximum yield that was 450% above control at a dose of 38 mg/kg for both enzymes. rGSTA1 and rGSTA3 showed similar thresholds (1.1 mg/kg) and maximal fold increase (275%) but varied in the relative response to each dose. These results indicate that renal GST increases occur at lead levels which are environmentally significant, that these changes precede cellular damage, and suggest that GST may serve as a tissue biomarker of lead exposure.     

Expression of arylhydrocarbon hydroxylase, epoxide hydrolases, glutathione S-transferase and UDP-glucuronosyltransferases in H5-6 hepatoma cells

1. The presence of arylhydrocarbon hydroxylase (cytochrome P-450 IA1 dependent), glutathione S-transferase, two distinct forms of epoxide hydrolases and UDP-glucuronosyltransferases was detected in H5-6 hepatoma cell homogenates using model substrates, selective inhibitors and specific antibodies. 2. The activity of arylhydrocarbon hydroxylase decreased strongly at the first days after plating and remained at a minimal value (1.5 pmol/min per mg) after 5 days of culture. 3. The hydratation of trans-stilbene oxide catalyzed by the soluble form of epoxide hydrolase was very low (11.0 pmol/min per mg), whereas the hepatoma cells contained appreciable amounts of the membrane-bound epoxide hydrolase and glutathione S-transferase measured with cis-stilbene oxide as substrate (maximal specific activity: 1.46 and 2.73 nmol/min per mg, respectively). 4. These cells also glucuronidated 1-naphthol efficiently (6 nmol/min per mg) and, at a lower extent, bilirubin (12 pmol/min per mg). 5. Addition of fenofibrate (70 microM) into the culture medium for 1-3 days failed to significantly stimulate the activity of cytosolic epoxide hydrolase. Only bilirubin glucuronidation increased 2-fold after 2 days of presence of the drug.     

Expression of gamma-glutamyltransferase in cancer cells and its significance in drug resistance

The expression of gamma-glutamyltransferase (GGT), a cell surface enzyme involved in cellular glutathione homeostasis, is often significantly increased in human tumors, and its role in tumor progression, invasion and drug resistance has been repeatedly suggested. As GGT participates in the metabolism of cellular glutathione, its activity has been mostly regarded as a factor in reconsitution of cellular antioxidant/antitoxic defences. On this basis, an involvement of GGT expression in resistance of cancer cells to cytotoxic drugs (in particular, cisplatin and other electrophilic agents) has been envisaged. Mechanistic aspects of GGT involvement in antitumor pharmacology deserve however further investigations. Recent evidence points to a more complex role of GGT in modulation of redox equilibria, with effects acting both intracellularly and in the extracellular microenvironment. Indications exist that the protective effects of GGT may be independent of intracellular glutathione, and derive rather from processes taking place at extracellular level and involving reactions of electrophilic drugs with thiol metabolites originating from GGT-mediated cleavage of extracellular glutathione. Although expression of GGT cannot be regarded as a general mechanism of resistance, the involvement of this enzyme in modulation of redox metabolism is expected to have impact in cellular response to several cytotoxic agents. The present commentary is a survey of data concerning the role of GGT in tumor cell biology and the mechanisms of its potential involvement in tumor drug resistance.