Chul-Hoon Kwon

An N-hydroxylated derivative of cyanamide that inhibits yeast aldehyde dehydrogenase

A stable, N,O-dibenzoyl derivative (DBHC) of N-hydroxycyanamide, the latter the postulated bioactivation product of the alcohol deterrent agent, cyanamide, has been synthesized. DBHC was an effective inhibitor of yeast aldehyde dehydrogenase (AIDH) in vitro and inhibited this enzyme in a concentration-dependent manner with an IC50 of 25 microM. Hydrolysis of the benzoate moiety of DBHC with dilute NaOH gave rise to the formation of nitroxyl (HN = O), detected by gas chromatography as nitrous oxide (N2O), the end-product of nitroxyl dimerization and disproportionation. It is postulated that the nitroxyl liberated by esterase action on DBHC by yeast AIDH may be the reactive species that inhibits AIDH.

Caspase-dependent cell death mediates potent cytotoxicity of sulfide derivatives of 9-anilinoacridine.

9-Anilinoacridine contains a tricyclic and planar aromatic structure that enables DNA intercalation and inhibition of topoisomerase II. Two recently developed sulfide derivatives of 9-anilinoacridines, 2-({4-[4-(acridin-9-ylamino)phenylthio]phenyl}(2-hydroxyethyl)amino)ethan-1-ol (CK0402) and 3-({4-[4-(acridin-9-ylamino)phenylthio]phenyl}(3-hydroxypropyl)amino)propan-1-ol (CK0403), displayed potent cytotoxic activity in multiple cancer cell lines. In-vitro enzymatic assay demonstrated that CK0402 and CK0403 directly inhibit decatenation reaction of topoisomerase II&agr;. Cells exposed to CK0403 showed DNA fragmentation, and activation of caspase-3 and caspase-2, indicating that it triggers caspase-dependent apoptosis. This was further supported by the fact that cytotoxicity of these drugs is attenuated by pharmacological inhibition of caspases with z-VAD-FMK. Studies with wild-type and p53−/− primary mouse embryonic fibroblasts demonstrated that p53 does not play a significant role in cell death process initiated by this kind of drug. In addition, pharmacological inhibition of poly(ADP-ribose) polymerase-1activity moderately enhanced cytotoxic activity of sulfide 9-anilinoacridine, suggesting that poly(ADP-ribose) polymerase-1 may have a protective function against 9-anilinoacridine-induced cell death process.

Facile Synthesis of Substituted 2-Iminohydantoins

Abstract A practical method for the preparation of substituted 2-iminohydantoins by cyclization of N-alkoxycarbonyl-α-amino acyl cyanamides is described.

CK0403, a 9‑aminoacridine, is a potent anti‑cancer agent in human breast cancer cells

3‑({4‑[4‑(Acridin‑9‑ylamino)phenylthio]phenyl}(3‑hydroxypropyl)amino)propan‑1‑ol (CK0403) is a sulfur‑containing 9‑anilinoacridine analogue of amsacrine and was found to be more potent than its analogue 2-({4-[4-(acridin-9-ylamino)phenylthio]phenyl}(2‑hydroxyethyl)amino)ethan‑1‑ol (CK0402) and amsacrine in the inhibition of the topoisomerase II‑catalyzed decatenation reaction. A previous study by our group reported that CK0402 was effective against numerous breast cancer cell lines, and the combination of CK0402 with herceptin enhanced its activity in HER2(+) SKBR‑3 cells. In order to identify novel chemotherapeutic agents with enhanced potency, the present study explored the potential of CK0403 in the treatment of breast cancer. First, the growth inhibitory activity of CK0403 in the breast cancer cell lines MCF‑7, MDA‑MB‑231, BT474 and SKBR‑3, as well as in the non‑cancerous MCF‑10A cell line, was examined using a sulforhodamine B assay. The results showed that CK0403 exerted more potent growth inhibitory activity than CK0402 in all of the breast cancer cell lines except MCF‑7. SKBR‑3 and MDA‑MB‑231 were the most sensitive cell lines tested, and the combination of CK0403 with herceptin in HER2(+) SKBR‑3 cells enhanced the growth inhibitory activity of CK0403. Analysis of cell cycle alterations induced by CK0403 in SKBR‑3 cells revealed that, similarly to CK0402, CK0403 induced G2/M‑phase arrest with a decreased S- and G0/G1-phase ratio. In addition, it was shown that CK0403 induced apoptosis more effectively than CK0402 in SKBR‑3 cells. Further analysis of autophagy protein 5 (Atg5) indicated that CK0403 induced more cleaved Atg5 than CK0402 and other chemotherapeutic agents tested. Of note, although still relatively potent, CK0403 exhibited reduced growth inhibitory activity under hypoxic conditions, which can induce autophagy. Collectively, the present results supported that CK0403 is highly potent and more effective than CK0402 against estrogen receptor-negative and HER2‑overexpressing breast cancer cell lines, suggesting its future application for chemotherapy in breast cancer.

N3-methyl-mafosfamide as a chemically stable, alternative prodrug of mafosfamide.

The presence of an alkyl substituent at N3 in the oxazaphosphorine ring stabilizes N-substituted 4-(alkylthio)cyclophosphamides from spontaneous decomposition. Based on this finding, N3-methyl-mafosfamide was synthesized and examined as a chemically stable, biooxidative prodrug of mafosfamide. This prodrug was stable in aqueous buffer (pH 7.4, 37 degrees C) and underwent N-demethylation in a time dependent manner when incubated with rat hepatic microsomes. N3-Methyl-mafosfamide was 10-fold more cytotoxic in vitro than cyclophosphamide against mouse embryo Balb/c 3T3 cells (LC50 = 3.6 microM). Preliminary in vivo antitumor evaluation against L1210 leukemia in mice showed that this prodrug was active [Increase of life span (ILS) > 29%].

Abstract 4576: Quantitative proteomic analysis of 2-({4-[4-(acridin-9-ylamino)phenylthio]phenyl}(2-hydroxyethyl)amino)ethan-1-ol (CK0402) modulated protein expression in breast cancer SKBR-3 cells

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC 2-({4-[4-(acridin-9-ylamino)phenylthio]phenyl}(2-hydroxyethyl)amino)ethan-1-ol (CK0402) was selected as potential anticancer agent among the series of sulfur-containing 9-aminoacridine analogs previously synthesized by our group. CK0402 is a topoisomerase II inhibitor and has been shown to exert anti-cancer activities in both in vitro and in vivo assays. Our previous studies demonstrated that CK0402 exhibited highest activity in SKBR-3 cells among a panel of established human breast cancer cell lines; moreover, CK0402-induced responses of G2/M arrest, apoptosis and autophagy were stronger in ER-(−)/HER2-(+) SKBR-3 cells than in ER-(+)/HER2-(−) MCF-7 cells. These findings suggest that CK0402 may be effective against HER-2 (+) breast cancer. The objective of this study is to identify molecular makers in response to CK0402 treatment in HER-2 (+) cells. These treatment markers may be important for clinical application since they can be used to avoid lethal toxic side effects or unnecessary treatment to insensitive patients. To this end, we analyzed the global protein expression modulated by CK0402 treatment in SKBR-3 cells using LC-MS/MS with a quantitative MS-based iTRAQ labeling method. SKBR-3 cells were exposed to various concentrations of CK0402 for 24 and 72 hrs; in which, 2 plates of cells received control vehicle, 3 plates of cells received 1 μmol/L, and another 3 plates of cells received 5 μmol/L of CK0402. The cell lysates were denatured, reduced, alkylated, and then digested with trypsin. The resulting peptides were labeled with 8-plex iTRAQ™ reagents, fractionated by cation exchange, and subjected to a LC-MS/MS analysis. The levels of differentially expressed proteins between control and treated sample were compared using students t-test. Selected proteins were then validated by Western blot analysis. We found that that a total of 219 proteins were differentially expressed in SKBR-3 cells (p < 0.05) after treated with 5 μmol/L of CK0402 for 72 hours. Among those, the levels of S100 proteins such as A7 and A8/A9 were increased 4.7- and 3.2-fold, respectively; in addition, the increases of these proteins were in a dose- and time- dependent manner, which is consistent with the growth inhibition effect of CK0402 in SKBR-3 cells. These results suggest that the S100 A7 and A8/A9 proteins are responders of CK0402 treatment in SKBR-3 cells. Current mechanistic studies are focusing on determining the roles of these identified proteins in the anticancer activities or toxic side effects of CK0402. The S100 proteins together with other proteins identified in this study could be used as effective indicators of the clinical response of patients to CK0402 treatment in future clinical studies. : Support: Susan G. Koman Breast Cancer Foundation Grant BCTR0707876 and Penn State HMC Deans Feasibility Grant. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4576.

An Improved Synthetic Method of 1-Chloro-7-methoxy-9H-thioxanthen-9-one

Abstract 1-Chloro-7-methoxy-9H-thioxanthen-9-one (3) was prepared via cyclization of 2-chloro-6-(4-methoxyphenylthio)-benzonitrile (1) in a one pot, two step reaction with improved yield (60%) over the previous literature report.

Rationale for Alcoholism Treatment Based on Inhibition of Aldehyde Dehydrogenase

It has been long recognized that certain individuals, particularly of Oriental origin, present a pronounced facial flush on consumption of only modest quantities of alcoholic beverages, thereby sustaining a natural aversion to alcohol (ethanol). These individuals lack a functional low Km hepatic mitochondrial aldehyde dehydrogenase (A1DH2) that metabolizes acetaldehyde, the first product of ethanol metabolism (1). This inborn trait observed in 40–50% of Orientals has been shown to be due to a genetic defect manifested by a point mutation on A1DH2, with the amino acid lysine replacing a glutamate residue near the C-terminal end of the enzyme (2). The consumption of ethanol by such individuals gives rise to elevated blood acetaldehyde levels and a flushing reaction reminiscent of the disulfiram-ethanol reaction (3), leading to alcohol avoidance.