Ryuta Yamazaki

Selective cyclooxygenase‐2 inhibitors show a differential ability to inhibit proliferation and induce apoptosis of colon adenocarcinoma cells

Although the influence of selective cyclooxygenase (COX)‐2 inhibitors on the proliferation of colon adenocarcinoma cells have been the subject of much investigation, relatively little research has compared the effects of different COX‐2 inhibitors. Celecoxib strongly suppressed the proliferation of COX‐2 expressing HT‐29 cells at 10–40 μM. NS‐398 and nimesulide also inhibited cell proliferation, whereas rofecoxib, meloxicam, and etodolac did not. Only celecoxib induced apoptosis of HT‐29 cells, as detected on the basis of DNA fragmentation, TUNEL positivity, and caspase‐3/7 activation. DNA fragmentation was also increasd in COX‐2 non‐expressing cell lines (SW‐480 and HCT‐116) by exposure to celecoxib for 6–24 h. All six COX‐2 inhibitors suppressed the production of prostaglandin E2 by HT‐29 cells, suggesting that the pro‐apoptotic effect of celecoxib was unrelated to inhibition of COX‐2. Inactivation of Akt might explain the differential pro‐apoptotic effect of these selective COX‐2 inhibitors on colon adenocarcinoma cells.

Novel acrylonitrile derivatives, YHO-13177 and YHO-13351, reverse BCRP/ABCG2-mediated drug resistance in vitro and in vivo

Breast cancer resistance protein (BCRP/ABCG2) confers resistance to anticancer drugs such as 7-ethyl-10-hydroxycamptothecin (SN-38, an active metabolite of irinotecan), mitoxantrone, and topotecan. In this study, we examined the reversing effects of YHO-13177, a novel acrylonitrile derivative, and its water-soluble diethylaminoacetate prodrug YHO-13351 on the BCRP-mediated drug resistance. YHO-13177 potentiated the cytotoxicity of SN-38, mitoxantrone, and topotecan in both BCRP-transduced human colon cancer HCT116 (HCT116/BCRP) cells and SN-38–resistant human lung cancer A549 (A549/SN4) cells that express BCRP, but had little effect in the parental cells. In addition, YHO-13177 potentiated the cytotoxicity of SN-38 in human lung cancer NCI-H460 and NCI-H23, myeloma RPMI-8226, and pancreatic cancer AsPC-1 cells that intrinsically expressed BCRP. In contrast, it had no effect on P-glycoprotein–mediated paclitaxel resistance in MDR1-transduced human leukemia K562 cells and multidrug resistance-related protein 1–mediated doxorubicin resistance in MRP1-transfected human epidermoid cancer KB-3-1 cells. YHO-13177 increased the intracellular accumulation of Hoechst 33342, a substrate of BCRP, at 30 minutes and partially suppressed the expression of BCRP protein at more than 24 hours after its treatment in both HCT116/BCRP and A549/SN4 cells. In mice, YHO-13351 was rapidly converted into YHO-13177 after its oral or intravenous administration. Coadministration of irinotecan with YHO-13351 significantly increased the survival time of mice inoculated with BCRP-transduced murine leukemia P388 cells and suppressed the tumor growth in an HCT116/BCRP xenograft model, whereas irinotecan alone had little effect in these tumor models. These findings suggest that YHO-13351, a prodrug of YHO-13177, could be clinically useful for reversing BCRP-mediated drug resistance in cancer chemotherapy. Mol Cancer Ther; 10(7); 1252–63. ©2011 AACR.

Diarylheptanoids suppress expression of leukocyte adhesion molecules on human vascular endothelial cells

Diarylheptanoids possess potent anti-inflammatory properties. However, the mechanism of their action is not fully understood. In this study, we found that three diarylheptanoids, 1-(3, 5-dimethoxy-4-hydroxyphenyl)-7-phenylhept-1-en-3-one (YPE-01), yakuchinone B and demethyl-yakuchinone B, reduced the adhesion of both human monocytic cell line U937 and human eosinophilic cell line EoL-1 cells to tumor necrosis factor-alpha (TNF-alpha)-treated human umbilical vein endothelial cells. In addition, they suppressed interleukin-1beta- or TNF-alpha-induced expression of E-selectin, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) on the surface of the endothelial cells. Since YPE-01 reduced both VCAM-1 and ICAM-1 mRNA induction in TNF-alpha-stimulated endothelial cells, diarylheptanoids appeared to suppress adhesion molecule expression at the transcriptional level. Furthermore, YPE-01 suppressed both VCAM-1 and ICAM-1 mRNA induction as well as edema in 12-O-tetradecanoylphorbol 13-acetate (TPA)-inflamed mice ears in vivo. These results suggest that the anti-inflammatory action of diarylheptanoids is, at least in part, due to their suppressive effect on the surface expression of inducible adhesion molecules in endothelial cells, and subsequent leukocyte adhesion.

A major metabolite of aceclofenac, 4'-hydroxy aceclofenac, suppresses the production of interstitial pro-collagenase/proMMP-1 and pro-stromelysin-1/proMMP-3 by human rheumatoid synovial cells.

Abstract.Objective and Design: We examined the effects of aceclofenac and its metabolites on the production of pro-collagenase-1/pro-matrix metalloproteinase-1 (proMMP-1), pro-gelatinase A/proMMP-2, pro-stromelysin-1/proMMP-3 and tissue inhibitor of metalloproteinases-1 (TIMP-1) by rheumatoid synovial cells.¶Materials: Synovial cells were obtained from patients with rheumatoid arthritis.¶Treatment: Cultures of confluent cells were treated with interleukin-1β (IL-1β)(1 ng/ml) and/or test drugs (0.3-30 μM) for 48 h.¶Methods: Production of proMMPs and TIMP-1 was monitored by Western blotting or gelatin zymography. Prostaglandin E2 (PGE2) was measured by an enzyme immunoassay.¶Results: 4′-Hydroxy aceclofenac, a major metabolite of aceclofenac, down-regulated both basal and IL-1β-induced production of proMMP-1 and proMMP-3 at a concentration sufficient to suppress PGE2 production without modulating proMMP-2 or TIMP-1, whereas aceclofenac itself had no marked effect on the production of proMMPs.¶Conclusions: Down-regulation of proMMP-1 and proMMP-3 production by 4′-hydroxy aceclofenac may contribute to the therapeutic effect of aceclofenac on rheumatoid arthritis and osteoarthritis.

Synthesis of phenanthroindolizidine alkaloids and evaluation of their antitumor activities and toxicities.

We previously reported that phenanthroindolizidine alkaloid 3 and its derivatives had markedly potent in vitro cytotoxicity. However, they had low in vivo antitumor activities and high in vivo toxicities, which was a serious problem. To address this problem, new phenanthroindolizidine derivatives were synthesized and their antitumor activities and toxicities were evaluated. This study describes the relationship between the chemical structures, antitumor activities, and toxicities of these phenanthroindolizidine derivatives. Based on its properties, compound 8 was found to be the most suitable potential antitumor agent.

Aspirin and sodium salicylate inhibit proliferation and induce apoptosis in rheumatoid synovial cells

Aspirin has been reported to induce apoptosis in a variety of cell lines. In this study, we examined whether aspirin and sodium salicylate inhibit cell growth and induce apoptosis in rheumatoid synovial cells. Synovial cells were obtained from patients with rheumatoid arthritis, and the cells were treated with aspirin or sodium salicylate (0.1–10 mm) for 24 h. Cell proliferation and viability were assessed by 5‐bromo‐2′‐deoxyuridine incorporation and by 4‐[3‐(4‐iodophenyl)‐2‐(4‐nitrophenyl)‐2H‐5‐tetrazolio]‐1,3‐benzene disulfonate (WST‐1) assay, respectively. The apoptosis of synovial cells was identified by DNA fragmentation assay and terminal deoxynucleotidyl transferase‐mediated dUTP nick end labelling (TUNEL) assay. Aspirin and sodium salicylate suppressed the proliferation (IC50 (concentration causing 50% inhibition of cell proliferation): 2.1 and 1.2 mm, respectively) and reduced the viability (IC50: 2.0 and 1.4 mm respectively) of synovial cells in a concentration‐dependent manner at 0.3–10 mm. Furthermore, they induced DNA fragmentation and increased the number of TUNEL‐ positive synovial cells. These results suggest that aspirin and sodium salicylate can inhibit the proliferation of rheumatoid synovial cells through induction of apoptosis.

Anti-inflammatory effect of YPE-01, a novel diarylheptanoid derivative, on dermal inflammation in mice

Objective and Design: We investigated the anti-inflammatory effect of YPE-01, a novel diarylheptanoid derivative in vitro and in vivo.¶Material: In the in vitro study, rat basophilic leukemia (RBL-1) cells were used. For the in vivo study, ICR and ddY mice (male, 7 weeks old) were used.¶Treatment: In the in vitro study, the supernatant of RBL-1 cells lysate was incubated with 50 μM arachidonic acid (AA) and 0.01-100 μM test drugs for 15 min. RBL-1 cells were preincubated with 0.01–100 μM test drugs for 10 min before incubation with 0.5 μM calcium ionophore A23187 for 10 min. In the in vivo study, YPE-01 (0.1–3 mg/ear) was applied to the ear of mice at the same time as a 12-O-tetradecanoylphorbol 13-acetate (TPA) application or 1 h before an AA application.¶Methods: In the in vitro study, the amounts of 5-hydroxyeicosatetraenoic acid and leukotrienes were measured by high-performance liquid chromatography and an enzyme immunoassay, respectively. In the in vivo study, a circular tissue sample from the ear of the mice was weighed. Statistical analysis was done using Dunnetts test.¶Results: YPE-01 inhibited the 5-lipoxygenase activity (IC50, 0.28 μM) and the leukotriene B4 (IC50, 0.035 μM) and C4 (IC50, 0.046 μM) production by RBL-1 cells without any inhibition of cyclooxygenase activity in vitro. The topical application of YPE-01 significantly suppressed both the AA- and TPA-induced ear edemas in vivo.¶Conclusions: YPE-01 is a selective 5-lipoxygenase inhibitor with a suppressive effect against dermal inflammation.

Pro-apoptotic effect of nonsteroidal anti-inflammatory drugs on synovial fibroblasts

Rheumatoid arthritis (RA) is a systemic inflammatory disease that mainly affects the articular synovial tissues. Although the etiology of RA has not yet been elucidated, physical and biochemical inhibition of synovial hyperplasia, which is the origin of articular destruction, may be an effective treatment for RA. Nonsteroidal anti-inflammatory drugs (NSAIDs) have long been used for the treatment of RA. The mechanism of action of NSAIDs generally involves the inhibition of cyclooxygenase (COX) at sites of inflammation. Thus, NSAIDs were not generally considered to have a so-called anti-rheumatic effect, including inhibition of progressive joint destruction and induction of remission. However, certain conventional NSAIDs and celecoxib, a selective COX-2 inhibitor, have been reported to inhibit synovial hyperplasia by inducing the apoptosis of human synovial fibroblasts. Therefore, it has been suggested that such NSAIDs may not only have an anti-inflammatory effect but also an anti-rheumatic effect. In this review, we summarize findings about the pro-apoptotic effect, in other words, anti-proliferative effect of NSAIDs on synovial fibroblasts from patients with RA.

YPC-21661 and YPC-22026, novel small molecules, inhibit ZNF143 activity in vitro and in vivo

Zinc‐finger protein 143 (ZNF143) is a transcription factor that is involved in anticancer drug resistance and cancer cell survival. In the present study, we identified a novel small molecule N‐(5‐bromo‐2‐methoxyphenyl)‐3‐(pyridine‐3‐yl) propiolamide (YPC‐21661) that inhibited ZNF143 promoter activity and down‐regulated the expression of the ZNF143‐regulated genes, RAD51, PLK1, and Survivin, by inhibiting the binding of ZNF143 to DNA. In addition, YPC‐21661 was cytotoxic and induced apoptosis in the human colon cancer cell line, HCT116 and human prostate cancer cell line, PC‐3. 2‐(pyridine‐3‐ylethynyl)‐5‐(2‐(trifluoromethoxy)phenyl)‐1,3,4‐oxadiazole (YPC‐22026), a metabolically stable derivative of YPC‐21661, induced tumor regression accompanied by the suppression of ZNF143‐regulated genes in a mouse xenograft model. The present study revealed that the inhibition of ZNF143 activity by small molecules induced tumor regression in vitro and in vivo; therefore, ZNF143 is a promising target of cancer therapeutics.