How Sung Lee

A Small Molecule Inhibitor of Isoprenylcysteine Carboxymethyltransferase Induces Autophagic Cell Death in PC3 Prostate Cancer Cells

A number of proteins involved in cell growth control, including members of the Ras family of GTPases, are modified at their C terminus by a three-step posttranslational process termed prenylation. The enzyme isoprenylcysteine carboxylmethyl-transferase (Icmt) catalyzes the last step in this process, and genetic and pharmacological suppression of Icmt activity significantly impacts on cell growth and oncogenesis. Screening of a diverse chemical library led to the identification of a specific small molecule inhibitor of Icmt, cysmethynil, that inhibited growth factor signaling and tumorigenesis in an in vitro cancer cell model (Winter-Vann, A. M., Baron, R. A., Wong, W., dela Cruz, J., York, J. D., Gooden, D. M., Bergo, M. O., Young, S. G., Toone, E. J., and Casey, P. J. (2005) Proc. Natl. Acad. Sci. U. S. A. 102, 4336–4341). To further evaluate the mechanisms through which this Icmt inhibitor impacts on cancer cells, we developed both in vitro and in vivo models utilizing PC3 prostate cancer cells. Treatment of these cells with cysmethynil resulted in both an accumulation of cells in the G1 phase and cell death. Treatment of mice harboring PC3 cell-derived xenograft tumors with cysmethynil resulted in markedly reduced tumor size. Analysis of cell death pathways unexpectedly showed minimal impact of cysmethynil treatment on apoptosis; rather, drug treatment significantly enhanced autophagy and autophagic cell death. Cysmethynil-treated cells displayed reduced mammalian target of rapamycin (mTOR) signaling, providing a potential mechanism for the excessive autophagy as well as G1 cell cycle arrest observed. These results identify a novel mechanism for the antitumor activity of Icmt inhibition. Further, the dual effects of cell death and cell cycle arrest by cysmethynil treatment strengthen the rationale for targeting Icmt in cancer chemotherapy.

Genotype of human carbonyl reductase CBR3 correlates with doxorubicin disposition and toxicity.

Objectives Doxorubicin is a cytotoxic drug with potential for severe myelosuppression that is highly variable and poorly predictable. Methods We correlated CBR1 and CBR3 genotypes with the pharmacokinetics and pharmacodynamics of doxorubicin in 101 Southeast Asian breast cancer patients receiving first-line doxorubicin. Results A common CBR3 11G>A variant was associated with lower doxorubicinol area under the concentration-time curve (AUC)/doxorubicin AUC metabolite ratio (P=0.009, GG vs. AA; trend test, P=0.004), lower CBR3 expression in breast tumor tissue (P=0.001, GG vs. AA), greater tumor reduction (P=0.015, GG vs. AA), and greater percentage reduction of leukocyte and platelet counts at nadir (trend test, P≤0.03). Chinese and Malays had higher frequency of the CBR3 11G>A variant than Indians (P≤0.002). Another variant CBR3 730G>A was associated with higher doxorubicinol AUC (P=0.009, GG vs. AA) and CBR3 expression in breast tumor tissue (P=0.001, GG vs AA). Conclusion Polymorphisms in CBR3 may explain interindividual and interethnic variability of doxorubicin pharmacokinetics and pharmacodynamics.

The use of submicron/nanoscale PLGA implants to deliver paclitaxel with enhanced pharmacokinetics and therapeutic efficacy in intracranial glioblastoma in mice

Pharmacokinetics and therapeutic efficacy of submicron/nanoscale, intracranial implants were evaluated for treating malignant glioblastoma in mice. 9.1% (w/w) paclitaxel-loaded polylactide-co-glycolide (PLGA) nanofiber discs (F3) were fabricated and characterized for morphology and size distribution. Along with F3, three other formulations, 9.1% (w/w) paclitaxel-loaded PLGA submicron-fiber discs (F2), 16.7% (w/w) paclitaxel-loaded PLGA microspheres entrapped in hydrogel matrices (H80 and M80) were intracranially implanted in BALB/c mice and the coronal brain sections were analyzed for bio-distribution of paclitaxel on 14, 28 and 42 days post-implantation. BALB/c nude mice with intracranial human glioblastoma (U87 MG-luc2) were used in the therapeutic efficacy study. Animals were randomized to intracranial implantation of F3 and H80 with paclitaxel dose of 10mg/kg, placebo F3, placebo H80, weekly intratumoral injection of Taxol (10mg/kg) or no treatment and the treatment response was analyzed by bioluminescence imaging and histological (H&E, Ki-67) examinations. Enhanced, therapeutic paclitaxel penetration (approximately 1 microm) in the mouse brain up to 5mm from the implant site even after 42 days post-implantation from F3 and H80 was confirmed and deduced to be diffusion/elimination controlled. F3 and H80 demonstrated significant (approximately 30 fold) tumor inhibition and significantly low tumor proliferation index after 41 days of treatment in comparison to sham and placebo controls. The submicron/nanoscale implants are able to demonstrate optimal paclitaxel pharmacokinetics in the brain/tumor with significant tumor inhibition in a glioblastoma xenograft model in mice and hence could be potentially useful to treat highly recurrent GBM.

Bevacizumab and rapamycin induce growth suppression in mouse models of hepatocellular carcinoma

BACKGROUND/AIMS Hepatocellular carcinoma is a leading cause of global cancer mortality, with standard chemotherapy being minimally effective in prolonging survival. We investigated if combined targeting of vascular endothelial growth factor protein and expression might affect hepatocellular carcinoma growth and angiogenesis. METHODS We treated patient-derived hepatocellular carcinoma xenografts with (i) bevacizumab; (ii) rapamycin; and (iii) bevacizumab plus rapamycin. Western blotting was employed to determine changes in the proteins. Apoptosis, vascular endothelial growth factor expression, microvessel density, and cell proliferation were analyzed by immunohistochemistry. RESULTS Hepatocellular carcinoma growth was inhibited by bevacizumab plus rapamycin treatment to a significantly greater degree than bevacizumab or rapamycin monotherapy. Reductions in tumor growth by bevacizumab plus rapamycin were associated with inhibition of downstream targets of the mammalian target-of-rapamycin pathway, reductions in vascular endothelial growth factor expression, and tumor microvessel density. Potentially additive effects of bevacizumab plus rapamycin included reductions in vascular endothelial growth factor expression, cyclin D1, and cyclin B1. In an intra-peritoneal model of hepatocellular carcinoma, bevacizumab plus rapamycin potently inhibited both intra-liver and intra-abdominal tumor growth, reduced ascites levels, and significantly prolonged mouse survival. CONCLUSIONS Bevacizumab and rapamycin, which are both clinically approved drugs, may represent a novel molecularly-targeted combination treatment for hepatocellular carcinoma.

Dose- and schedule-dependent protective efficacy of celgosivir in a lethal mouse model for dengue virus infection informs dosing regimen for a proof of concept clinical trial.

Celgosivir (6-O-butanoyl castanospermine), a pro-drug of the naturally occurring castanospermine, is an inhibitor of α-glucosidase I and II that is found to be a potent inhibitor of several enveloped viruses including all four serotypes of dengue virus. We showed previously that the compound fully protected AG129 mice from lethal infection with a mouse adapted dengue virus at a dose of 50mg/kg twice daily (BID) for 5days and was effective even after 48h delayed treatment. Here we show that the protection by celgosivir is dose- and schedule-dependent and that a twice-a-day regimen of 50, 25 or 10mg/kg is more protective than a single daily dose of 100mg/kg. Treatment with 50mg/kg BID castanospermine had comparable efficacy as 25mg/kg BID celgosivir, suggesting that celgosivir is approximately twice as potent as castanospermine with respect to in vivo antiviral efficacy. Pharmacokinetics (PK) studies of celgosivir in mice showed that it rapidly metabolized to castanospermine. Simulation of the PK data with the survival data for the various doses of celgosivir tested suggests that the steady-state minimum concentration is a critical parameter to note in choosing dose and schedule. These results influenced the selection of the dose regimen for a proof-of-concept clinical trial of celgosivir as a treatment against dengue fever.

A Pharmacodynamic Model for the Time Course of Tumor Shrinkage by Gemcitabine + Carboplatin in Non–Small Cell Lung Cancer Patients

Purpose: This tumor response pharmacodynamic model aims to describe primary lesion shrinkage in non–small cell lung cancer over time and determine if concentration-based exposure metrics for gemcitabine or that of its metabolites, 2′,2′-difluorodeoxyuridine or gemcitabine triphosphate, are better than gemcitabine dose for prediction of individual response. Experimental Design: Gemcitabine was given thrice weekly on days 1 and 8 in combination with carboplatin, which was given only on day 1 of every cycle. Gemcitabine amount in the body and area under the concentration-time curves of plasma gemcitabine, 2′,2′-difluorodeoxyuridine, and intracellular gemcitabine triphosphate in white cells were compared to determine which best describes tumor shrinkage over time. Tumor growth kinetics were described using a Gompertz-like model. Results: The apparent half-life for the effect of gemcitabine was 7.67 weeks. The tumor turnover time constant was 21.8 week·cm. Baseline tumor size and gemcitabine amount in the body to attain 50% of tumor shrinkage were estimated to be 6.66 cm and 10,600 mg. There was no evidence of relapse during treatment. Conclusions: Concentration-based exposure metrics for gemcitabine and its metabolites were no better than gemcitabine amount in predicting tumor shrinkage in primary lung cancer lesions. Gemcitabine dose-based models did marginally better than treatment-based models that ignored doses of drug administered to patients. Modeling tumor shrinkage in primary lesions can be used to quantify individual sensitivity and response to antitumor effects of anticancer drugs.

Regulation of Heart Function by Endogenous Gaseous Mediators—Crosstalk Between Nitric Oxide and Hydrogen Sulfide

Both nitric oxide (NO) and hydrogen sulfide (H(2)S) are two important gaseous mediators regulating heart function. The present study examined the interaction between these two biological gases and its role in the heart. We found that l-arginine, a substrate of NO synthase, decreased the amplitudes of myocyte contraction and electrically induced calcium transients. Sodium hydrogen sulfide (an H(2)S donor), which alone had minor effect, reversed the negative inotropic effects of l-arginine. The effect of l-arginine + sodium hydrogen sulfide was abolished by three thiols (l-cysteine, N-acetyl-cysteine, and glutathione), suggesting that the effect of H(2)S + NO is thiol sensitive. The stimulatory effect on heart contractility was also induced by GYY4137, a slow-releasing H(2)S donor, when used together with sodium nitroprusside, an NO-releasing donor. More importantly, enzymatic generation of H(2)S from recombinant cystathionine-γ-lyase protein also interacted with endogenous NO generated from l-arginine to stimulate heart contraction. In summary, our data suggest that endogenous NO may interact with H(2)S to produce a new biological mediator that produces positive inotropic effect. The crosstalk between H(2)S and NO also suggests an intriguing potential for the endogenous formation of a thiol-sensitive molecule, which may be of physiological significance in the heart.

Pharmacokinetics of intravenous vancomycin in patients with end-stage renal failure

The pharmacokinetics of a 500-mg dose of i.v. vancomycin were studied in six Chinese patients with end-stage renal failure. Serum vancomycin concentrations were determined by high-performance liquid chromatography. Observed peak and trough (at 168 h postinfusion) concentrations were in the range of 14.2–35.0 μg/ml and 2.8–5.5 μg/ml, respectively. The data were analyzed using the PCNONLIN. In all six patients, the data could be fitted well by both the biexponential and triexponential models, but in three patients the latter model provided a better fit. Two-compartment pharmacokinetic parameters obtained from the six patients were t1/2α 1.13 ± 0.25 h (mean ± SEM), t1/2β 121.3 ± 8.2 h, Vc0.45 ± 0.09 L/kg, Vss 1.00 ± 0.12 L/kg, ClT5.90 ± 0.69 ml/kg/h, and the calculated Cmax 25.0 ± 6.1 μg/ml. The mean vancomycin serum protein binding was 18.5 ± 12.0% as compared with a mean of 46.0% in pooled serum from normal controls. Hemodialysis had no significant effect on vancomycin protein binding or clearance. On the basis of our kinetic study, 500 mg of vancomycin given every seven days is probably adequate treatment for methicillin resistant Staphylococcus aureus infection in end-stage renal failure patients, but further clinical studies are necessary to confirm this.

Clinical evaluation and plasma clozapine concentrations in Chinese patients with schizophrenia

The relationships between clozapine dosages, plasma concentrations, and clinical responses in Chinese schizophrenics were studied. Fourteen treatment-refractory schizophrenic patients were treated with clozapine for 12 weeks. Patients were assessed before and after 6 and 12 weeks of treatment using the Brief Psychiatric Rating Scale (BPRS), the Clinical Global Impression (CGI), and the Simpson-Angus Scale for Extrapyramidal Side Effect. Plasma clozapine concentrations were determined by high-performance liquid chromatography. Ten patients (71.4%) responded after 12 weeks of treatment. Although the mean daily dosage at week 12 (373 +/- 90 mg/day) was lower than that reported in American trials (444 mg/day), the mean plasma clozapine concentration attained (1,078 +/- 385 ng/ml) was higher. This higher concentration may be due to the lower body wight and the preponderance of women among our patients, absence of smoking and alcohol use, and/or ethnic difference between Chinese and non-Chinese. There was wide interindividual variation in the plasma clozapine concentrations. Compared with other studies, the plasma clozapine concentrations and the response rate were higher. Although the sample size was small, the findings are suggestive of pharmacokinetic and pharmacodynamic ethnic differences in Chinese with clozapine therapy.

Hoarding and clozapine-risperidone combination.

There is a school of thought that the growing, if not insatiable, public appetite for euthanasia may be considerably blunted if health care providers simply did a better job of providing compassionate care to terminally ill patients (2). The Rights of the Terminally Ill Bill, passed recently by the Northern Territory parliament in Australia, allows mentally competent, terminally ill patients to request voluntary assistance from a physician to terminate the patient’s life humanely (3). Critics opine that the push toward physician-assisted dying may be based largely on ignorance of the availability of good palliative care services in Australia (4). Others have suggested that inadequate pain control in cancer patients may be a factor associated with the high rate of euthanasia in the Netherlands (5).