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YJC-10592, a new chemokine receptor 2 (CCR-2) antagonist, was newly synthesized for the treatment of asthma and atopic dermatitis. To evaluate the pharmacokinetic characteristics of YJC-10592, a high-performance liquid chromatographic (HPLC) method was developed for the determination of YJC-10592 in rat plasma, urine, and tissue homogenates, using YJC-10450 as an internal standard. The mobile phase was a gradient of acetate buffer (50 mM, pH 3.0) in acetonitrile at a flow rate of 1.0 mL/min. Chromatograms were monitored by an ultraviolet detector at 230 nm. The retention times for YJC-10592 and the internal standard were 8.0 and 6.3 min, respectively, and the lower limits of quantification (LLOQ) for YJC-10592 in rat plasma and urine were 0.05 and 0.2 μg/mL, respectively. The intra- and inter-assay precisions (coefficients of variation, CVs) were generally low (below 11.4 %) for rat plasma and urine samples. The accuracies (relative errors) ranged from 94.2 to 108 %. No interferences from endogenous substances were found. YJC-10592 was stable in rat whole blood but unstable in the liver homogenate. An initial pharmacokinetic study was performed in Sprague–Dawley rats and demonstrated that YJC-10592 has a half-life of 86.8 min and wide tissue distribution. In summary, this HPLC method can be applied to future preclinical and clinical evaluation of YJC-10592.

YJC-10592, a novel chemokine receptor 2 (CCR-2) antagonist, was developed for treating asthma and atopic dermatitis. We studied the pharmacokinetic characteristics of YJC-10592 after intravenous (5, 10 and 20 mg/kg) and oral (100 and 200 mg/kg) administration of the drug to rats. Tissue distribution of YJC-10592 was also evaluated after intravenous administration of YJC-10592, 10 mg/kg, to rats. The pharmacokinetics of YJC-10592 was dose-dependent from 20 mg/kg after intravenous administration to rats. The values of the area under the plasma concentration–time curve from time zero to infinity (AUC) of YJC-10592 were dose-dependent from 20 mg/kg and the time-averaged total body (CL) and nonrenal (CLNR) clearances of YJC-10592 were significantly lower at dose of 20 mg/kg, suggesting that saturable metabolism may be involved. The absolute bioavailability (F) of YJC-10592 was generally low (

Protein N-acetylglucosamine modification (O-GlcNAcylation) plays a critical role in cell-cycle regulation, apoptosis and signal transduction. Thr-58 of c-Myc, a mutational hot spot in lymphomas, is a site for both phosphorylation (primed by Ser-62 phosphorylation) and O-GlcNAcylation. Whereas Thr-58 O-GlcNAcylation induces ubiquitin-dependent c-Myc degradation, Thr-58/Ser-62 phosphorylation increases c-Myc stability and thus induces invasiveness and tumorigenesis of MCF-7 human breast cancer cells. Using form-specific antibodies for Thr-58-O-GlcNAcylated c-Myc, Western blot analysis was carried out and found that Thr-58-O-GlcNAcylated c-Myc proteins were detected primarily in the nuclear fraction as 68 kDa proteins. MCF-7 cells were treated for 4 hr with 1% DMSO or 2 mM ketoconazole [β-N-acetylglucosaminidase (O-GlcNAcase) inhibitor, dissolved in DMSO]. Western blot analysis of nuclear lysates of MCF-7 cells revealed that the levels of Thr-58-O-GlcNAcylated c-Myc dramatically increased after ketoconazole treatment. Thr-58-O-GlcNAcylated c-Myc levels also increased in the nuclear lysates of Mia Paca-2 and SW480 cells treated with 50 µg/ml for 48 hours. The ketoconazole treatment increased cell death in MCF-7 cells 84% higher than in MCF10A non-cancerous cells by lactate dehydrogenase (LDH) cytotoxicity assays. Treatment with 50 µg/ml ketoconazole for 72 hr inhibited cell proliferation of colon cancer cells, SW480 and HT29, by 63.3% and 67.4%, respectively, lung cancer cells, H1437 and A549, by 44.9% and 51.3%, respectively, and a pancreatic cancer cell, Mia Paca-2, by 60.2% in a dose-dependent manner. These results demonstrated that treatment of ketoconazole inhibited the proliferation of MCF-7 breast cancer cells as well as colon, lung and pancreatic cancer cells. Wound healing assays were carried out to verify the inhibition of cell migration. Whereas 55.3% of Mia Paca-2 cells migrated toward a scratched field after 24-hr treatment, only 11.6% of the cells migrated after treatment with 100 µg/ml ketoconazole. PANC-1 cells, which failed to increase Thr-58-O-GlcNAcylated c-Myc expression and cell proliferation after ketoconazole treatment, showed no significant difference of cell migration after 24 hr with 100 µg/ml ketoconazole treatment. Surprisingly, colon cancer cells, SW480 and HT29, showed an inhibition of invasiveness as evidenced by the inhibition of cell proliferation and cell migration after treatment with 100 µg/ml ketoconazole. Collectively, our results demonstrated that treatment of ketoconazole, an O-GlcNAcase inhibitor, increased Thr-58-O-GlcNAcylated c-Myc level and inhibited cell proliferation in MCF-7, SW480, HT29, H1437, A549 and Mia Paca-2 cells, which may decrease invasiveness and migration of the breast, colon, lung and pancreatic cancer cells. Supported by NCI SBIR Phase II Contract No. HHSN261201300058C. Citation Format: So Hee Kim, Eun-Sin Du, Aby Joiakim, Sung-Su Park, David Kaplan, David Putt, Hyesook Kim. Ketoconazole-induced O-GlcNAcylated c-Myc expression and inhibition of cell proliferation in cancers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3513. doi:10.1158/1538-7445.AM2014-3513