Comparative metabolic study between two selective estrogen receptor modulators, toremifene and tamoxifen, in human liver microsomes.

Toremifene (TOR) and Tamoxifen (TAM) are widely used as endocrine therapy for estrogen receptor positive breast cancer. Poor metabolizers of TAM are likely to have worse clinical outcomes than patients who exhibit normal TAM metabolism due to lower plasma level of its active metabolite, 4-hydroxy-N-desmethyl (4OH-NDM) tamoxifen (endoxifen). In this study, we examined the role of individual cytochrome P450 (CYP) isoforms in the metabolism of TOR to N-desmethyl (NDM), 4-hydroxy (4OH) and 4OH-NDM metabolites in comparison with TAM using human liver microsomes (HLMs) with selective chemical inhibitors for each CYP isoform and recombinant CYP proteins. Similar levels of NDM metabolites were formed for both TOR and TAM, and N-demethylation of both compounds was primarily carried out by CYP3A4. We found that the formation of 4OH-NDM-TOR was catalyzed both by CYP2C9 and CYP2D6, whereas the formation of 4OH-TAM and endoxifen was specifically catalyzed by CYP2D6 in HLMs. Our results suggest that the potential contribution of CYP2D6 in the bioactivation pathway of TOR may be lower compared to TAM, and may have a different impact on clinical outcome than CYP2D6 polymorphisms.

A novel compound, NK150460, exhibits selective antitumor activity against breast cancer cell lines through activation of aryl hydrocarbon receptor.

Antiestrogen agents are commonly used to treat patients with estrogen receptor (ER)-positive breast cancer. Tamoxifen has been the mainstay of endocrine treatment for patients with early and advanced breast cancer for many years. Following tamoxifen treatment failure, however, there are still limited options for subsequent hormonal therapy. We discovered a novel compound, NK150460, that inhibits 17ß-estradiol (E2)-dependent transcription without affecting binding of E2 to ER. Against our expectations, NK150460 inhibited growth of not only most ER-positive, but also some ER-negative breast cancer cell lines, while never inhibiting growth of non-breast cancer cell lines. Cell-based screening using a random shRNA library, identified aryl hydrocarbon receptor nuclear translocator (ARNT) as a key gene involved in NK150460's antitumor mechanism. siRNAs against not only ARNT but also its counterpart aryl hydrocarbon receptor (AhR) and their target protein, CYP1A1, dramatically abrogated NK150460's growth-inhibitory activity. This suggests that the molecular cascade of AhR/ARNT plays an essential role in NK150460's antitumor mechanism. Expression of ERα was decreased by NK150460 treatment, and this was inhibited by an AhR antagonist. Unlike two other AhR agonists now undergoing clinical developmental stage, NK150460 did not induce histone H2AX phosphorylation or p53 expression, suggesting that it did not induce a DNA damage response in treated cells. Cell lines expressing epithelial markers were more sensitive to NK150460 than mesenchymal marker-expressing cells. These data indicate that NK150460 is a novel AhR agonist with selective antitumor activity against breast cancer cell lines, and its features differ from those of the other two AhR agonists.

2-Aryl-1,4-naphthoquinone-1-oxime methyl ethers: their cytotoxic activity.

Preliminary examination for the structure-activity relationship of quinone monooxime derivatives on cytotoxicity against HeLa S3 cell and further trials using eight different cell lines suggested that 2-aryl-6,7-methylenedioxy-1,4-naphthoquinone-1-oxime methyl ethers, carrying 2-methoxy-4,5-methylenedioxyphenyl, 7-methoxy-2-methylbenzofuran-4-yl, and 2-methoxycarbonyl-3,4-dimethoxyphenyl as the 2-aryl substituent, were potential candidates for anti-cancer drugs.

Growth inhibition of human colon cancer cell line HCT116 by bis[2-(acylamino)phenyl] disulfide and its action mechanism.

Our laboratory has been investigating the use of compounds which disrupt beta-catenin/T cell factor (TCF) binding to treat human colon cancer. There are several cysteine residues on the surface of beta-catenin where it binds to TCF. Some bis[2-(acylamino)phenyl] disulfides might have the ability to form a disulfide bond with the cysteine residues of beta-catenin, leading to inhibition of the growth of human colon cells. Bis[2-(acylamino)phenyl] disulfides were screened to inhibit the growth of cancer cells. Among them, bis[2-(2,2-dimethylpropanoylamino)phenyl] disulfide (1) had promising inhibitory effects (HCT116, IC50: 9.7 microM; DLD-1, IC50: 6.9 microM) on cell proliferation, and did not show any cytotoxicity among normal human fibroblast CCD-1059SK cells even at 200 microM. This derivative reduced the beta-catenin/TCF4 association in the HCT116 cells to ca. 50% at 150 microM. Furthermore, it activated markedly the phosphorylation of c-Jun N-terminal kinase (JNK) connected to stress-activated apoptosis at a lower concentration (30 microM). In view of cell cycle analyses, Hoechst staining, and terminal deoxynucleotidyl transferase-mediated dUTP-biotin Nick end-labeling (TUNEL) assays along with the above results, it is likely that 1 inhibited the growth of HCT116 cells through pathways including the JNK-mediated apoptosis.

OSW-1 analogues: modification of the carbohydrate moiety.

4 OSW-1 analogues featuring modified carbohydrate moieties were prepared. The purpose of these modifications was to assess the importance of certain chemical functions with respect to biological activity. The synthesis and biological activity of the target molecules are shown.

Cytotoxicity of lissoclibadins and lissoclinotoxins, isolated from a tropical ascidian Lissoclinum cf. badium, against human solid-tumor-derived cell lines.

The in vitro growth inhibitory activity of lissoclibadins and lissoclinotoxins isolated from the tropical ascidian Lissoclinum cf. badium against nine human cancer cell lines was examined to evaluate their potential anticancer efficacy. Lissoclibadins 1 (1) and 2 (2), and lissoclinotoxin F (4) showed the strongest activity of the six compounds tested, which were more potent than the anticancer drug cisplatin. Compound 1 has a trimeric structure, and compounds 2 and 4 are structural isomers possessing dimeric structures connected by disulfide and sulfide bonds of trans- and cis-orientations, respectively. Lissoclibadin 3 (3), a dimeric compound connected by two sulfide bonds, and two monomeric compounds (5, 6) were less active than 1, 2, and 4. Lissoclibadin 2 (2) was the most interesting compound possessing potent inhibitory activity against colon (DLD-1 and HCT116), breast (MDA-MB-231), renal (ACHN), and non-small-cell lung (NCI-H460) cancer cell lines and showing no toxicity following a 50 mg/kg single treatment to mice, and preferable stability in rat plasma.

Identification of a potent and stable antiproliferative agent by the prodrug formation of a thiolate histone deacetylase inhibitor.

To identify prodrugs of a thiolate histone deacetylase inhibitor NCH-31 that show potent antiproliferative activity and are stable in human plasma, we synthesized several candidate prodrugs of NCH-31. Among these compounds, S-2-methyl-3-phenylpropanoyl compound 2 showed more potent antiproliferative activity and higher stability in human plasma than S-isobutyryl compound NCH-51.

Lysocellin, a metabolite of the novel drug 'alopestatin', induces G1 arrest and prevents cytotoxicity induced by etoposide.

We report here that lysocellin, a polyether antibiotic from a streptomycete, induces G1 phase arrest in human osteosarcoma MG63 cells. Lysocellin up-regulates p21WAF1/Cip1 and down-regulates cyclin D1 at the mRNA level. In addition, cyclin D1 is down-regulated by the proteasome-dependent signal pathway in MG63 cells. In drug combination studies, we found that lysocellin treatment weakened the cytotoxic activity of etoposide in MG63 cells using a colony-formation assay. To study the in vivo efficacy of lysocellin, we isolated a novel compound related to lysocellin from the same streptomycete, and found that the novel drug is converted to lysocellin in vivo and decreases etoposide-induced alopecia in a neonatal rat model. We raise the possibility that this novel drug, named 'alopestatin', may be a promising agent against alopecia.

Novel inhibitors of human histone deacetylases: design, synthesis, enzyme inhibition, and cancer cell growth inhibition of SAHA-based non-hydroxamates.

To find novel non-hydroxamate histone deacetylase (HDAC) inhibitors, a series of compounds modeled after suberoylanilide hydroxamic acid (SAHA) was designed and synthesized. In this series, compound 7, in which the hydroxamic acid of SAHA is replaced by a thiol, was found to be as potent as SAHA, and optimization of this series led to the identification of HDAC inhibitors more potent than SAHA. In cancer cell growth inhibition assay, S-isobutyryl derivative 51 showed strong activity, and its potency was comparable to that of SAHA. The cancer cell growth inhibitory activity was verified to be the result of histone hyperacetylation and subsequent induction of p21(WAF1/CIP1) by Western blot analysis. Kinetical enzyme assay and molecular modeling suggest the thiol formed by enzymatic hydrolysis within the cell interacts with the zinc ion in the active site of HDACs.

Potent histone deacetylase inhibitors: N-hydroxybenzamides with antitumor activities.

The screening tests of N-hydroxybenzamides for their HDAC-inhibitory activities led to the discovery of the promising compounds with a 2-naphthylcarbonyl group and with a 1,4-biphenylcarbonyl group. These compounds were further modified to optimize their physico-chemical profile. As a result, the inhibitor with a 6-amino-2-naphthylcarbonyl was obtained, which showed not only promising growth inhibitions against a panel of tumor cells, but also an improved water solubility. It exhibited the maximal 185% of survival rate (%T/C) in a in vivo experiment with P388 cell-inoculated mice.

Total synthesis of 12-methoxydihydrochelerythrine and anti-tumour activity of its quaternary base: toward an efficient synthetic route for 12-alkoxybenzo[c]phenanthridine bases via naphthoquinone monooxime from 2-benzofuranyl-1-tetralone derivative.

A concise total synthesis of 12-methoxydihydrochelerythrine (6), isolated from Bocconia integrifolia, is described. The synthesis features an efficient route to a 12-alkoxybenzo[c]phenanthridine skeleton via naphthoquinone monooxime 11 as a key compound. Starting from 7-methoxy-2-methylbenzo[b]furan (9), 3-aryl-1-tetralone 10 was synthesised, followed by aromatisation to 3-aryl-1-naphthol 17. After oxidative cleavage of the furan ring, basic nitrosation of naphthol 22 gave the naphthoquinone 11. The benzo[c]phenanthridine skeleton was formed by reductive cyclisation of 11. Deoxygenation of the lactam moiety in 23 afforded nor-base 32 and methylation of 32 under reductive conditions gave the target dihydro base 6 (23 steps from benzofuran 9 in 10% overall yield). The corresponding quaternary base 7 showed moderate anti-tumour activity against cancer cell lines; on NCI-H460: IC50 4.5 microM and on MDA-MB-231: IC50 1.2 microM. Introduction of a methoxy group into the 12-position of the benzo[c]phenanthridine skeleton could cause enhanced activity against MDA-MB-231 by comparison of 7 with chelerythrine (35) (IC50 5.3 microM).

[Combined effects of toremifene and paclitaxel on human breast cancer cell lines].

Effects of toremifene (TOR) in combination with paclitaxel (TXL) on various human breast cancer cell lines were evaluated. TOR and TXL exhibited additive effects on estrogen receptor (ER)-positive cancer cell lines, MCF-7 and T-47D, and a sub-additive effect on a tamoxifen (TAM)-resistant line, T-47D/TAM. To all three ER-negative cancer cell lines, the combined treatment also showed additive effects on MDA-MB-134VI, MDA-MB-231 and MDA-MB-453. Furthermore, a synergistic effect was observed on a multi-drug resistant (MDR) line, Adr. This synergistic effect was more potent in the combination with TOR than that with TAM. The combined treatment increased intracellular TXL, and the accumulation by TOR was 1.5-fold that by TAM. Consequently, the ratio of G2M arrested cells was higher, with statistical significance, in the TOR combination than in the TAM combination. In addition, these synergistic effects in MDR cells were also observed in the combination of TXL with major clinical active metabolites, N-desmethyl-TOR (TOR-1) and 4-hydroxy-TOR (TOR-2). These results suggest that the combination therapy of TOR and TXL might be an effective clinical treatment for breast cancer patients.

Studies on the chemical constituents of stem bark of Millettia leucantha: isolation of new chalcones with cytotoxic, anti-herpes simplex virus and anti-inflammatory activities.

Four new chalcone derivatives (1, 4, 7, 10) were isolated from the stem bark of Millettia leucantha KURZ (Leguminosae) along with two known ones (2, 6) and five known flavones (3, 5, 8, 9, 11). Structure elucidation and unambiguous assignment of the isolates were achieved with the aid of 1D and 2D NMR extensive studies. Correlation of 10 to 4 was successfully done by reduction with Et(3)SiH/CF(3)CO(2)H. Moderate cytotoxic activity was observed in chalcones (1, 10), whereas dihydrochalcones (4, 6) showed moderate anti-Herpes Simplex Virus (HSV) activity. Interestingly, flavone 8 showed significant anti-inflammatory effects inhibiting both cyclooxygenase (COX)-1 and -2.