Alkylating agents, such as temozolomide, are among the most effective cytotoxic agents used for malignant gliomas, but responses remain very poor. The DNA repair protein O 6 -methylguanine-DNA methyltransferase (MGMT) plays an important role in cellular resistance to alkylating agents. IFN-B can act as a drug sensitizer, enhancing toxicity against a variety of neoplasias, and is widely used in combination with other antitumor agents such as nitrosoureas. Here, we show that IFN-B sensitizes glioma cells that harbor the unmethylated MGMT promoter and are resistant to temozolomide. By means of oligonucleotide microarray and RNA interference, we reveal that the sensitizing effect of IFN-B was possibly due to attenuation of MGMT expression via induction of the protein p53. Our study suggests that clinical efficacy of temozolomide might be improved by combination with IFN-B using appropriate doses and schedules of administration. (Cancer Res 2005; 65(17): 7573-9)

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Objective-When the availability of tetrahydrobiopterin (BH4) is deficient, endothelial nitric oxide synthase (eNOS) produces superoxide rather than NO (uncoupled eNOS). We have shown that the atherosclerotic lesion size was augmented in apolipoprotein E-deficient (ApoE-KO) mice overexpressing eNOS because of the enhanced superoxide production. In this study, we addressed the specific importance of uncoupled eNOS in atherosclerosis, and the potential mechanistic role for specific versus nonspecific antioxidant strategies in restoring eNOS coupling. Methods and Results-We crossed mice overexpressing eNOS in the endothelium (eNOS-Tg) with mice overexpressing GTP-cyclohydrolase I (GCH), the rate-limiting enzyme in BH4 synthesis, to generate ApoE-KO/eNOS-Tg/GCH-Tg mice. As a comparison, ApoE-KO/eNOS-Tg mice were treated with vitamin C. Atherosclerotic lesion formation was increased in ApoE-KO/eNOS-Tg mice compared with ApoE-KO mice. GCH overexpression in ApoE-KO/eNOS-Tg/ GCH-Tg mice increased vascular BH4 levels and reduced plaque area. This reduction was associated with decreased superoxide production from uncoupled eNOS. Vitamin C treatment failed to reduce atherosclerotic lesion size in ApoE-KO/eNOS-Tg mice, despite reducing overall vascular superoxide production. Conclusion-In contrast to vitamin C treatment, augmenting BH4 levels in the endothelium by GCH overexpression reduced the accelerated atherosclerotic lesion formation in ApoE-KO/eNOS-Tg mice, associated with a reduction of superoxide production from uncoupled eNOS. Key Words: eNOS uncoupling Ⅲ tetrahydrobiopterin Ⅲ vitamin C Ⅲ atherosclerosis Ⅲ apolipoprotein E-deficient mice N itric oxide (NO) derived from endothelial NO synthase (eNOS) is a critical signaling molecule in the vasculature and has a range of antiatherogenic effects. 1 In eNOS/ apolipoprotein E (ApoE) double-knockout mice, atherosclerosis is increased, suggesting a protective effect of eNOSderived NO. 2,3 However, certain vascular diseased states are associated with an increase rather than a decrease in the expression of eNOS. 4,5 We have shown that endotheliumtargeted overexpression of eNOS in ApoE-KO (ApoE-KO/ eNOS-Tg) mice surprisingly resulted in decreased endothelial NO availability, increased vascular superoxide production, and accelerated atherosclerosis. 6 See page 1493These observations could be explained by a relative deficiency of the cofactor tetrahydrobiopterin (BH4) causing eNOS uncoupling, 7 where the enzymatic reduction of molecular oxygen by eNOS is no longer coupled to L-arginine oxidation, resulting in production of superoxide rather than NO. In ApoE-KO/eNOS-Tg mice, dietary BH4 supplementation reduced superoxide production and increased NO availability, although it was unclear whether this was a general antioxidant effect of BH4 or a specific effect on eNOS coupling. 6 Indeed, dietary supplementation with the antioxidant vitamin C can also reduce vascular oxidative stress, increase BH4 levels, and was sufficient to improve the depressed endothelium-dependent relaxatio...