Buthionine sulfoximine and chemoresistance in cancer treatments: a systematic review with meta-analysis of preclinical studies.

Buthionine sulfoximine (BSO) is a synthetic amino acid that blocks the biosynthesis of reduced glutathione (GSH), an endogenous antioxidant cellular component present in tumor cells. GSH levels have been associated with tumor cell resistance to chemotherapeutic drugs and platinum compounds. Consequently, by depleting GSH, BSO enhances the cytotoxicity of chemotherapeutic agents in drug-resistant tumors. Therefore, the aim of this study was to conduct a systematic review with meta-analysis of preclinical studies utilizing BSO in cancer treatments. The systematic search was carried out using the following databases: PubMed, Web of Science, Scopus, and EMBASE up until March 20, 2023, in order to collect preclinical studies that evaluated BSO, alone or in association, as a strategy for antineoplastic therapy. One hundred nine investigations were found to assess the cytotoxic potential of BSO alone or in combination with other compounds. Twenty-one of these met the criteria for performing the meta-analysis. The evidence gathered indicated that BSO alone exhibits cytotoxic activity. However, this compound is generally used in combination with other antineoplastic strategies, mainly chemotherapy ones, to improve cytotoxicity to carcinogenic cells and treatment efficacy. Finally, this review provides important considerations regarding BSO use in cancer treatment conditions, which might optimize future studies as a potential adjuvant antineoplastic therapeutic tool.

Amplification of Tumor Oxidative Stresses with Liposomal Fenton Catalyst and Glutathione Inhibitor for Enhanced Cancer Chemotherapy and Radiotherapy.

Amplification of intracellular oxidative stress has been found to be an effective strategy to induce cancer cell death. To this end, we prepare a unique type of ultrasmall gallic acid-ferrous (GA-Fe(II)) nanocomplexes as the catalyst of Fenton reaction to enable persistent conversion of H2O2 to highly cytotoxic hydroxyl radicals (•OH). Then, both GA-Fe(II) and l-buthionine sulfoximine (BSO), an inhibitor of glutathione (GSH) synthesis, are coencapsulated within a stealth liposomal nanocarrier. Interestingly, the obtained BSO/GA-Fe(II)@liposome is able to efficiently amplify intracellular oxidative stress via increasing •OH generation and reducing GSH biosynthesis. After chelating with 99mTc4+ radioisotope, such BSO/GA-Fe(II)@liposome could be tracked under in vivo single-photon-emission-computed-tomography (SPECT) imaging, which illustrates the time-dependent tumor homing of such liposomal nanoparticles after intravenous injection. With GA-Fe(II)-mediated •OH production and BSO-mediated GSH depletion, treatment with such BSO/GA-Fe(II)@liposome would lead to dramatically enhanced intratumoral oxidative stresses, which then result in remarkably improved therapeutic efficacies of concurrently applied chemotherapy or radiotherapy. This work thus presents the concise fabrication of biocompatible BSO/GA-Fe(II)@liposome as an effective adjuvant nanomedicine to promote clinically used conventional cancer chemotherapy and radiotherapy, by greatly amplifying the intratumoral oxidative stress.

A Phase I New Approaches to Neuroblastoma Therapy Study of Buthionine Sulfoximine and Melphalan With Autologous Stem Cells for Recurrent/Refractory High-Risk Neuroblastoma.

BACKGROUND: Myeloablative therapy for high-risk neuroblastoma commonly includes melphalan. Increased cellular glutathione (GSH) can mediate melphalan resistance. Buthionine sulfoximine (BSO), a GSH synthesis inhibitor, enhances melphalan activity against neuroblastoma cell lines, providing the rationale for a Phase 1 trial of BSO-melphalan. PROCEDURES: Patients with recurrent/resistant high-risk neuroblastoma received BSO (3 gram/m(2) bolus, then 24 grams/m(2) /day infusion days -4 to -2), with escalating doses of intravenous melphalan (20-125 mg/m(2) ) days -3 and -2, and autologous stem cells day 0 using 3 + 3 dose escalation. RESULTS: Among 28 patients evaluable for dose escalation, one dose-limiting toxicity occurred at 20 mg/m(2) melphalan (grade 3 aspartate aminotransferase/alanine aminotransferase) and one at 80 mg/m(2) (streptococcal bacteremia, grade 4 hypotension/pulmonary/hypocalcemia) without sequelae. Among 25 patients evaluable for response, there was one partial response (PR) and two mixed responses (MRs) among eight patients with prior melphalan exposure; one PR and three MRs among 16 patients without prior melphalan; one stable disease with unknown melphalan history. Melphalan pharmacokinetics with BSO were similar to reports for melphalan alone. Melphalan Cmax for most patients was below the 10 µM concentration that showed neuroblastoma preclinical activity with BSO. CONCLUSIONS: BSO (75 gram/m(2) ) with melphalan (125 mg/m(2) ) is tolerable with stem cell support and active in recurrent/refractory neuroblastoma. Further dose escalation is feasible and may increase responses.

Inhibition of the glutamate-cysteine ligase catalytic subunit with buthionine sulfoximine enhances the cytotoxic effect of doxorubicin and cyclophosphamide in Burkitt lymphoma cells.

Burkitt lymphoma (BL) is a highly aggressive lymphoma that mainly affects children and young adults. Chemotherapy is effective in young BL patients but the outcome in adults is less satisfactory. Therefore, there is a need to enhance the cytotoxic effect of drugs used in BL treatment. Glutathione (GSH) is an important antioxidant involved in processes such as regulation of oxidative stress and drug detoxification. Elevated GSH levels have been observed in many cancers and were associated with chemoresistance. We previously identified GCLC, encoding an enzyme involved in GSH biosynthesis, as an essential gene in BL. We now confirm that knockout of GCLC decreases viability of BL cells and that the GCLC protein is overexpressed in BL tissues. Moreover, we demonstrate that buthionine sulfoximine (BSO), a known inhibitor of GCLC, decreases growth of BL cells but does not affect control B cells. Furthermore, we show for the first time that BSO enhances the cytotoxicity of compounds commonly used in BL treatment, doxorubicin, and cyclophosphamide. Given the fact that BSO itself was not toxic to control cells and well-tolerated in clinical trials, combination of chemotherapy with BSO may allow reduction of the doses of cytotoxic drugs required to obtain effective responses in BL patients.

Acquired Radiation Resistance Induces Thiol-dependent Cisplatin Cross-resistance.

Resistance to radiation remains a significant clinical challenge in non-small cell lung carcinoma (NSCLC). It is therefore important to identify the underlying molecular and cellular features that drive acquired resistance. We generated genetically matched NSCLC cell lines to investigate characteristics of acquired resistance. Murine Lewis lung carcinoma (LLC) and human A549 cells acquired an approximate 1.5-2.5-fold increase in radiation resistance as compared to their parental match, which each had unique intrinsic radio-sensitivities. The radiation resistance (RR) was reflected in higher levels of DNA damage and repair marker γH2AX and reduced apoptosis induction after radiation. Morphologically, we found that radiation resistance A549 (A549-RR) cells exhibited a greater nucleus-to-cytosol (N/C) ratio as compared to its parental counterpart. Since the N/C ratio is linked to the differentiation state, we next investigated the epithelial-to-mesenchymal transition (EMT) phenotype and cellular plasticity. We found that A549 cells had a greater radiation-induced plasticity, as measured by E-cadherin, vimentin and double-positive (DP) modulation, as compared to LLC. Additionally, migration was suppressed in A549-RR cells, as compared to A549 cells. Subsequently, we confirmed in vivo that the LLC-RR and A549-RR cells are also more resistance to radiation than their isogenic-matched counterpart. Moreover, we found that the acquired radiation resistance also induced resistance to cisplatin, but not carboplatin or oxaliplatin. This cross-resistance was attributed to induced elevation of thiol levels. Gamma-glutamylcysteine synthetase inhibitor buthionine sulfoximine (BSO) sensitized the resistant cells to cisplatin by decreasing the amount of thiols to levels prior to obtaining acquired radiation resistance. By generating radiation-resistance genetically matched NSCLC we were able to identify and overcome cisplatin cross-resistance. This is an important finding arguing for combinatorial treatment regimens including glutathione pathway disruptors in patients with the potential of improving clinical outcomes in the future.

A cancer-specific activatable theranostic nanodrug for enhanced therapeutic efficacy via amplification of oxidative stress.

Rationale: Despite considerable advances, the reactive oxygen species (ROS)-mediated cancer treatment suffers from the problems of up-regulation of adaptive antioxidants in cancer cells as well as side effects to normal cells. Therefore, development of a new generation of cancer-specific nanomedicine capable of amplifying oxidative stress would be of great interest for accurate and effective cancer treatment. Methods: Herein, transferrin (Tf)-decorated, dihydroartemisinin (DHA), L-buthionine-sulfoximine (BSO), and CellROX-loaded liposomal nanoparticles (Tf-DBC NPs) were developed for precise cancer theranositcs. Tf-DBC NPs could specifically recognize cancer cells via Tf-Tf receptor binding and be uptaken into the lysosomes of cancer cells, where Tf-DBC NPs were activated to release Fe(II), DHA, and BSO. ROS was generated by DHA in the presence of Fe(II), and GSH was depleted by BSO to disrupt the redox balance in cancer cells. Furthermore, CellROX, as a fluorescent probe for imaging of intracellular oxidative stress, was used to monitor the therapeutic efficacy. Results: The integration of Tf, DHA, and BSO into the acidic pH-responsive liposomes selectively and effectively killed cancer cells and prevented the oxidative injury to normal cells. The high oxidative state was visualized at the tumor site and the amplification of oxidative stress enabled tumor eradication by Tf-DBC NPs, demonstrating the successful implementation of this novel strategy in vivo. Conclusion: Our study provides a new paradigm for the design of ROS-mediated therapeutics and offers a promising perspective for precise cancer treatment.

Synergistic effect of buthionine sulfoximine on the chlorin e6-based photodynamic treatment of cancer cells.

In this study, we investigated the synergistic effect of L-buthionine sulfoximine (BSO) on the chlorin e6 (Ce6)-based photodynamic therapy (PDT) of cancer cells. Among various cancer cells, HCT116 cells have highest intracellular L-glutathione (GSH) level and SNU478 cells showed the lowest GSH level. BSO alone showed negligible intrinsic cytotoxicity against CCD986sk cells. Since HCT116 and SNU478 cells showed the highest and the lowest intracellular GSH levels, respectively, those were used to test synergistic effect on the Ce6-based PDT. In the absence of light, BSO and Ce6 combination did not practically increase reactive oxygen species (ROS) in either of HCT116 or SNU478 cells, while light irradiation increased ROS level dose-dependently. 10 µM BSO treatment significantly depleted total GSH level in cancer cells, i.e. total GSH level decreased to one-fourth of the control in HCT116 cells while it decreased to two-fifth of the control treatment at SNU478 cell. BSO showed synergistic effect on the ROS production in HCT116 cells while it has practically no benefits in ROS production of SNU478 cells. No synergistic effect was observed in viability of SNU478 cells because BSO itself was cytotoxic to SNU478 cells. However, BSO had negligible cytotoxicity against HCT116 cells and showed synergistic anticancer effect in combination with Ce6-based PDT. Furthermore, the addition of glutathione reduced ethyl ester (GSH-OEt), recovered intracellular GSH level, and cell viability with reduced the intracellular ROS level. We suggest that synergistic effect of BSO in the Ce6-based PDT should be considered with intrinsic intracellular GSH level of cancer cells.

Buthionine sulfoximine has anti-Trypanosoma cruzi activity in a murine model of acute Chagas' disease and enhances the efficacy of nifurtimox.

L-buthionine (S,R)-sulfoximine (BSO) at a dose of 220 mg/kg of body weight/day showed an anti-Trypanosoma cruzi effect in infected mice, increasing their survival rate and decreasing the parasitemias and parasite burden in the hearts. Treatment with BSO plus nifurtimox caused an increase in the survival rate in comparison to the rates with treatment with each drug alone.

GSH loss per se does not affect neuroblastoma survival and is not genotoxic.

Depletion of glutathione (GSH) by buthionine sulfoximine (BSO) has been reported to be toxic against some cancer cells and to sensitize many tumours including neuroblastoma (NB) to anticancer drugs. The balance between the production rate of reactive oxygen species (ROS) and the function of GSH affects the intracellular reduction-oxidation status, which is crucial for the regulation of several cellular physiological functions. To assess the role of glutathione in neuroblastoma therapy, the effect of sublethal concentrations of BSO was studied in a panel of neuroblastoma cell lines characterized by different MYCN status. We found that GSH depletion per se not accompanied by ROS overproduction, does not affect cell survival, and is not genotoxic but induces HO-1 expression in GI-ME-N cell line, a representative example of MYCN non-amplified NB cells, having the highest basal levels of GSH among the tested NB lines. These observations might open a novel therapeutic window based on the possibility of modulating the cellular 'activity' of GSH.

Cyanidin-3-O-glucoside ameliorates diabetic nephropathy through regulation of glutathione pool.

Diabetic nephropathy (DN) is a common complication of diabetes and the major cause of chronic kidney disease. Cyanidin 3-glucoside (C3G) is the most widespread anthocyanin in nature. In the present study, we aimed to investigate the possible effects of C3G on DN in db/db mice. We found that body weights and high levels of fasting blood glucose, serum insulin, C-peptide, glycosylated hemoglobin A1c, and systolic blood pressure in diabetic mice were significantly reduced by C3G. C3G also reduced the ratio of kidney to body weight and the levels of blood urea nitrogen (BUN), serum creatinine, urinary albumin content and albumin/creatinine ratio (ACR), ameliorated the pathological changes of kidneys, reduced the surface area of Bowman's capsule, glomerular tuft, Bowman's space, and decreased renal expression of collagen IV, fibronectin, transforming growth factor ß 1 (TGFß1), matrix metalloprotein 9 (MMP9) and α-smooth muscle actin (α-SMA) in db/db mice. The Lee's index, perirenal white adipose tissue weight, and high levels of blood and renal triglyceride and cholesterol were decreased by C3G. Moreover, C3G reduced systemic levels and renal expression of tumor necrosis factor ɑ (TNFɑ), IL-1ɑ, and monocyte chemotactic protein-1 (MCP-1), indicating the inhibition of inflammation. Furthermore, C3G increased glutathione (GSH) level and decreased GSSG level in kidneys of diabetic mice. The renal mRNA expression of glutamate-cysteine ligase catalytic subunit (GCLC) and glutamate-cysteine ligase modifier subunit (GCLM) was increased by C3G in diabetic mice. Buthionine sulphoximine (BSO), an inhibitor of GSH synthesis, inhibited the effects of C3G on glucose metabolic dysfunction and DN. The data demonstrates that enhancement of GSH pool is involved in the renal-protective effects of C3G. Overall, C3G could be a promising therapeutic option for attenuation of diabetes and DN.

Lack of mitochondrial depolarization by oxidative stress is associated with resistance to buthionine sulfoximine in acute lymphoblastic leukemia cells.

Raised intracellular glutathione is one characteristics of high-risk childhood acute lymphoblastic leukemia (ALL). Depletion of glutathione by buthionine sulfoximine (BSO) has been reported to be toxic against some cancer cells. To assess the role of glutathione in ALL, the toxicity of BSO was studied in B-precursor ALL cell lines. BSO increased oxidative stress equally in all cell lines; however mitochondrial depolarization was observed only in BSO-sensitive cells. BSO up-regulated Bcl-2 protein, and antagonized the anti-ALL effect of prednisolone in BSO-resistant cells. A lack of mitochondrial death-signal activation by oxidative stress seemed to be associated with BSO-resistance in ALL.

3-bromopyruvate and buthionine sulfoximine effectively kill anoikis-resistant hepatocellular carcinoma cells.

BACKGROUND & AIMS: Acquisition of anoikis resistance is a prerequisite for metastasis in hepatocellular carcinoma (HCC). However, little is known about how energy metabolism and antioxidant systems are altered in anoikis-resistant (AR) HCC cells. We evaluated anti-tumor effects of a combination treatment of 3-bromopyruvate (3-BP) and buthionine sulfoximine (BSO) in AR HCC cells. METHODS: We compared glycolysis, reactive oxygen species (ROS) production, and chemoresistance among Huh-BAT, HepG2 HCC cells, and the corresponding AR cells. Expression of hexokinase II, gamma-glutamylcysteine synthetase (rGCS), and epithelial-mesenchymal transition (EMT) markers in AR cells was assessed. Anti-tumor effects of a combination treatment of 3-BP and BSO were evaluated in AR cells and an HCC xenograft mouse model. RESULTS: AR HCC cells showed significantly higher chemoresistance, glycolysis and lower ROS production than attached cells. Expression of hexokinase II, rGCS, and EMT markers was higher in AR HCC cells than attached cells. A combination treatment of 3-BP/BSO effectively suppressed proliferation of AR HCC cells through apoptosis by blocking glycolysis and enhancing ROS levels. In xenograft mouse models, tumor growth derived from AR HCC cells was significantly suppressed in the group treated with 3-BP/BSO compared to the group treated with 3-BP or sorafenib. CONCLUSIONS: These results demonstrated that a combination treatment of 3-BP/BSO had a synergistic anti-tumor effect in an AR HCC model. This strategy may be an effective adjuvant therapy for patients with sorafenib-resistant HCC.

Neuroprotective effects of a glutathione depletor in rat post-ischemic reperfusion brain damage.

The induction of heme oxygenase (HO), the rate-limiting enzyme in heme degradation, occurs as an adaptative response to oxidative stress and is consequent to decrease in cellular glutathione levels. Our previous studies demonstrated significant increase in survival rates of rats treated with glutathione depletors and submitted to transient cerebral ischemia. The aim of the present research was to test the effects of L-Buthionine sulfoximine (BSO), a glutathione depletor, during cerebral post-ischemic reperfusion. Cerebral ischemia was induced by bilateral clamping of common carotid arteries for 20 min. Each sample was used for glutathione ad lipid peroxidation level dosage and for evaluating the expression of heme oxygenase both after a single subcutaneous administration of BSO and without treatment. In the same experimental conditions, endothelial, inducible and neuronal Nitric Oxide Synthase (eNOS, iNOS and nNOS) and Dimethylarginine Dimethyl amine Hydrolases (DDAH-1 and DDAH-2) were also evaluated. Results obtained in the present study suggested that HO-1 over-expression may be implicated in the protective effect of BSO in post-ischemic reperfusion brain damage, although the involvement of other important stress mediators cannot be ruled out.

Pharmacological modulation of reactive oxygen species in cancer treatment.

Aerobic metabolism of mammalian cells leads to the generation of reactive oxygen species (ROS). To cope with this toxicity, evolution provided cells with effective antioxidant systems like glutathione. Current anticancer therapies focus on the cancer dependence on oncogenes and non-oncogenes. Tumors trigger mechanisms to circumvent the oncogenic stress and to escape cell death. In this context we have studied 2-phenylethinesulfoxamine (PES), which disables the cell protective mechanisms to confront the proteotoxicity of damaged and unfolded proteins. Proteotoxic stress is increased in tumor cells, thus providing an explanation for the anticancer selectivity of PES. In addition, we have found that PES induces a severe oxidative stress and the activation of p53. The reduction of the cell content in glutathione by means of L-buthionine-sulfoximine (BSO) synergizes with PES. In conclusion, we have found that ROS constitutes a central element in a series of positive feed-back loops in the cell. ROS, p53, proteotoxicity, autophagy and mitochondrial dynamics are interconnected with the mechanisms leading to cell death, either apoptotic or necrotic. This network of interactions provides multiple targets for drug discovery and development in cancer.

Importance of dose intensity in neuro-oncology clinical trials: summary report of the Sixth Annual Meeting of the Blood-Brain Barrier Disruption Consortium.

Therapeutic options for the treatment of malignant brain tumors have been limited, in part, because of the presence of the blood-brain barrier. For this reason, the Sixth Annual Meeting of the Blood-Brain Barrier Disruption Consortium, the focus of which was the "Importance of Dose Intensity in Neuro-Oncology Clinical Trials," was convened in April 2000, at Government Camp, Mount Hood, Oregon. This meeting, which was supported by the National Cancer Institute, the National Institute of Neurological Disorders and Stroke, and the National Institute of Deafness and Other Communication Disorders, brought together clinicians and basic scientists from across the U.S. to discuss the role of dose intensity and enhanced chemotherapy delivery in the treatment of malignant brain tumors and to design multicenter clinical trials. Optimizing chemotherapy delivery to the CNS is crucial, particularly in view of recent progress identifying certain brain tumors as chemosensitive. The discovery that specific constellations of genetic alterations can predict which tumors are chemoresponsive, and can therefore more accurately predict prognosis, has important implications for delivery of intensive, effective chemotherapy regimens with acceptable toxicities. This report summarizes the discussions, future directions, and key questions regarding dose-intensive treatment of primary CNS lymphoma, CNS relapse of systemic non-Hodgkin's lymphoma, anaplastic oligodendroglioma, high-grade glioma, and metastatic cancer of the brain. The promising role of cytoenhancers and chemoprotectants as part of dose-intensive regimens for chemosensitive brain tumors and development of improved gene therapies for malignant gliomas are discussed.

Retroviral transfer of MRP1 and gamma-glutamyl cysteine synthetase modulates cell sensitivity to L-buthionine-S,R-sulphoximine (BSO): new rationale for the use of BSO in cancer therapy.

MRP1 (multidrug resistance protein 1) co-exports glutathione (GSH) and drug(s) and exports GSH, glucuronide, and sulphate-conjugated drugs. Human Fly-eco fibrosarcoma cells producing the MRP1-expressing retrovirus SF91MRP (Fly-eco MRP1), as well as 3T3 cells transduced with SF91MRP (3T3/MRP1), presented a decrease in intracellular GSH levels, as measured by two different methods. The enhanced export of GSH caused by the overexpression of MRP1 was partially counterbalanced by an increased rate of GSH synthesis. Fly-eco MRP1 and 3T3/MRP1 were hypersensitive to the GSH-depleting and cytotoxic activities of L-buthionine-S,R-sulphoximine (BSO), compared with their parental counterparts. In addition, the potentiation by BSO of the cytotoxic activity of chlorambucil and doxorubicin in Fly-eco MRP1 cells was greater than in parental Fly-eco cells. Although the turnover time of GSH, i.e. the theoretical time in which the entire GSH pool is resynthesised, was approximately 50% faster in Fly-eco MRP1 cells than in parental cells, this was not sufficient to fully restore the intracellular GSH level. In addition, mrp1 (-/-) mice were resistant to the GSH-depleting activity of intraperitoneally (i.p.) injected BSO, compared with mrp1 (+/+) mice. Co-transfer of the cDNAs for MRP1 and the heavy subunit of gamma-glutamyl cysteine synthetase (GCS) resulted in increased intracellular GSH levels and in high-level resistance to the GSH-depleting and cytotoxic activities of BSO. These data, and in particular the elevated single-agent cytotoxicity of BSO, provide a new rationale for the use of BSO in the treatment of MRP1-overexpressing tumours.

The emerging role of reactive oxygen species in cancer therapy.

The generation of reactive oxygen species (ROS) can be exploited therapeutically in the treatment of cancer. One of the first drugs to be developed that generates ROS was procarbazine. It is oxidised readily in an oxic environment to its azo derivative, generating ROS. Forty years ago, Berneis reported a synergistic effect in DNA degradation when procarbazine was combined with radiation; this was confirmed in preclinical in vivo modes. Early uncontrolled clinical trials suggested an enhancement of the radiation effect with procarbazine, but two randomised trials failed to confirm this. The role of ROS in cancer treatments and in the development of resistance to chemotherapy is now better understood. The possibility of exploiting ROS as a cancer treatment is re-emerging as a promising therapeutic option with the development of agents such as buthionine sulfoximine and motexafin gadolinium.

Enhancement of lipid peroxidation and of the antitumor effect of hyperthermia upon combination with oral eicosapentaenoic acid.

The present study was designed to determine the effect of eicosapentaenoic acid (EPA) on the susceptibility of tumor cells to treatments that kill the cells by lipid peroxidation. Using AH109A carcinoma, a rat liver cancer, we measured EPA content, levels of antioxidants, and degree of lipid peroxidation in tumor tissue and normal liver tissue after oral administration of EPA. In the control group treated with distilled water, EPA in tumor tissue was lower than in normal liver tissue, suggesting that its content of polyunsaturated fatty acids (the substrates for lipid peroxidation) was inherently low. Levels of antioxidants also tended to be lower in tumor tissue. EPA level increased in both tumor and normal tissues after oral administration of EPA. At the same time, glutathione peroxidase (GSH-Px) increased in normal tissue, whereas tumor tissue displayed no increase in antioxidants; instead GSH decreased. The EPA-induced change in balance between substrates for lipid peroxidation and antioxidants suggested that tumor tissue might become more susceptible to lipid peroxidation than normal liver tissue. In fact, hyperthermia treatment did enhance lipid peroxidation and antitumor action. Our results indicate that oral EPA specifically increases the susceptibility of liver tumor tissue to lipid peroxidation, and hence enhance the antitumor effect of hyperthermia and prolongs survival.

Potentiation of treosulfan toxicity by the glutathione-depleting agent buthionine sulfoximine in human malignant glioma cells: the role of bcl-2.

Median survival of human malignant glioma patients is less than one year even with cytoreductive surgery and postoperative radiotherapy. Adjuvant chemotherapy has been rather ineffective. Here, we studied the potentiation by L-buthionine-[S,R]-sulfoximine (BSO), a glutathione-depleting agent, of anticancer drug actions on two human malignant glioma cell lines, LN-229 and T98G. LN-229 has wild-type p53 status, T98G is mutant for p53. Glutathione levels were depleted by BSO with similar kinetics in both cell lines. Only LN-229 cells were growth-inhibited by BSO. BSO had minor effects on the toxicity of doxorubicin, ACNU (1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-3-(2-chloroethyl)-3-nitrosou rea, nimustine) and vincristine. BSO failed to alter teniposide or cytarabine toxicity. BSO induced prominent sensitization to the alkylating agent, treosulfan, in both cell lines, as assessed by viability assays, in situ DNA end labeling and quantitative DNA fragmentation. Treosulfan is thought to mediate toxicity via formation of reactive epoxides. In the absence of BSO, treosulfan had little acute cytotoxic and moderate antiproliferative effects. Synergistic glioma cell cytotoxicity induced by treosulfan and BSO was not associated with reactive oxygen species formation. Ectopic expression of bcl-2 did not alter basal glutathione levels but attenuated glutathione depletion induced by BSO. Bcl-2 provided only moderate protection from synergistic induction of glioma cell death by treosulfan and BSO. Glutathione depletion may play a role in BSO-mediated chemosensitization, but other mechanisms are probably involved as well. BSO may be a useful agent for glioma cell sensitization to specific chemotherapeutic drugs such as treosulfan.

Role of glutathione in protection against noise-induced hearing loss.

A potential mechanism of hearing loss due to acoustic overstimulation is the generation of reactive oxygen species (ROS). ROS not removed by antioxidant defenses could be expected to cause significant damage to the sensory cells of the cochlea. We studied the influence of the antioxidant glutathione (GSH) on noise-induced hearing loss by using l-buthionine-[S,R]-sulfoximine (BSO), an inhibitor of GSH synthesis, and 2-oxothiazolidine-4-carboxylate (OTC), a cysteine prodrug, which promotes rapid restoration of GSH when GSH is acutely depleted. Pigmented female guinea pigs were exposed to broadband noise (102 dB SPL, 3 h/day, 5 days) while receiving daily injections of BSO, OTC, or saline. By weeks 2 and 3 after noise exposure, BSO-treated animals showed significantly greater threshold shifts above 12 kHz than saline-treated subjects, whereas OTC-treated animals showed significantly smaller threshold shifts at 12 kHz than controls. Histologically assessed noise-induced damage to the organ of Corti, predominantly basal turn row 1 outer hair cells, was most pronounced in BSO-treated animals. High performance liquid chromatographic analysis showed that OTC significantly increased cysteine levels, but not GSH levels, in the cochlea. These findings show that GSH inhibition increases the susceptibility of the cochlea to noise-induced damage and that replenishing GSH, presumably by enhancing availability of cysteine, attenuates noise-induced cochlear damage.