Design, synthesis, and antiproliferative activity of 3,4-diarylpyrazole-1-carboxamide derivatives against melanoma cell line

Synthesis of a new series of 3,4-diarylpyrazole-1-carboxamide derivatives is described. Their antiproliferative activity against A375P human melanoma cell line was tested and the effect of substituents on the diarylpyrazole scaffold was investigated. The biological results indicated that five synthesized compounds (Ig, Ii, IIc, IIg, and IIh) exhibited similar activity to Sorafenib. In addition, three compounds (IIa, IIb, and IIi) were more potent than Sorafenib. Among all of these derivatives, compound IIa which has dimethylamino and phenolic moieties showed the most potent antiproliferative activity against A375P human melanoma cell line. Virtual screening was carried out through docking of the most potent compound IIa into the domain of V600E-b-Raf and the binding mode was studied.     

薯蓣皂苷元衍生物的抗肿瘤活性研究

目的研究薯蓣皂苷元衍生物在体外的抗肿瘤活性。方法采用MTT法对人恶性黑色素瘤细胞A375、人肺腺癌细胞A549、人肝癌细胞HepG-2及人慢性髓原白血病细胞株K562进行体外抗肿瘤活性试验。结果薯蓣皂苷元衍生物对4个肿瘤细胞株A375、A549、K562、HepG-2具有不同程度的抗肿瘤活性。结论绝大部分薯蓣皂苷元衍生物对4个肿瘤细胞株有较好的抗肿瘤活性,IC50值都低于30μmol.L-1。化合物22对细胞株A375的IC50=4.48μmol.L-1,化合物9、10对细胞株K562的IC50分别为2.51、2.38μmol.L-1;显示其抗肿瘤活性与对照化合物1-(3β-薯蓣皂苷元)-3-苄基咪唑溴盐相当。     

A new cyclotetrapeptide from marine fungus Trichoderma reesei

A new cyclotetrapeptide, trichoderide A, was isolated from the marine fungus Trichoderma reesei. The structure was identified by spectral methods, and the stereochemical assignments were made by chiral HPLC of the hydrolyzed compound. Trichoderide A showed moderate cytotoxity against human A375-S2 melanoma cell line.     

Synthesis and biological evaluation of 1-cyano-2-amino-benzimidazole derivatives as a novel class of antitumor agents

A series of 1-cyano-2-amino-benzimidazole derivatives were synthesized and evaluated for their cytotoxic activities in vitro against three human cancer cell lines (human lung carcinoma cell line: A549, human leukemia cell line: K562, and human prostate cancer cell line: PC-3). Most of these compounds showed potent activities against these tumor cell lines, especially against A549 and K562 cell lines. The preliminary structure–activity relationships of 1-cyano-2-amino-benzimidazole derivatives were also discussed. The cell cycle analysis was carried out in K562 cells and the results showed that compound 4d caused a marked increase of cells in G ₂/M phase.     

A new piperazine-2,5-dione from the marine fungus Gliocladium sp

A new piperazine-2,5-dione, named gliocladride, was isolated from marine fungus Gliocladium sp. The structure was established on the basis of spectral data, and the stereochemical assignments were made by 1H NMR spectrum and chiral HPLC of the hydrolyzed compound. Gliocladride showed a cytotoxic effect with an IC50 value of 3.86 microg/ml against human A375-S2 melanoma cell line.     

CD147siRNA对恶性黑色素瘤细胞VEGF及血管内皮细胞体外迁移活性的影响

目的研究CD147siRNA对恶性黑色素瘤细胞株A375中VEGF表达水平和血管内皮细胞体外迁移活性的影响。方法采用半定量RT-PCR法检测转染后A375细胞中VEGF mRNA表达的变化。采用ELISA法检测A375细胞与成纤维细胞共同培养后VEGF表达水平的变化。采用细胞迁移实验检测转染CD147siRNA前后的A375细胞对脐静脉内皮细胞株ECV-304在体外迁移活性的影响。结果转染CD147siRNA后,A375细胞中VEGF的mRNA和蛋白表达水平显著降低,其下调率分别为10.77%（P〈0.05）和38.5%（P〈0.01）。转染CD147siRNA前后的A375细胞与成纤维细胞共同培养后VEGF的表达水平均较单独培养时显著升高,其升高率分别为98.5%（P〈0.01）和159.9%（P〈0.01）。转染CD147siRNA后的A375细胞使ECV-304细胞的体外迁移活性下降44.77%（P〈0.01）。结论 CD147siRNA能有效抑制恶性黑色素瘤细胞株A375中VEGF的表达水平和脐静脉内皮细胞株ECV-304在体外的迁移活性。     

A derivative of aminopeptidase inhibitor (BE15) has a dual inhibitory effect of invasion and motility on tumor and endothelial cells

Bestatin is an inhibitor of aminopeptidase N (APN)/CD13 and aminopeptidase B. In our previous report, bestatin inhibited the tumor cell invasion and the angiogenesis induced by the inoculation of B16-BL6 melanoma cells into mice and capillary formation on human umbilical vein endothelial cells (HUVECs) in vitro. The results show that the enzymatic activity of APN is deeply involved in tumor invasion and angiogenesis. We investigated the effect of three bestatin derivatives on A375 human melanoma cells and in vitro. All the derivatives inhibited the activity of APN, but BE15 was most effective and controlled the migration of A375 cells and HUVECs and capillary formation of HUVECs. Furthermore, the bestatin derivatives had an inhibitory effect not only on aminopeptidase activity but also on cell motility. Compared with bestatin and the other derivatives, BE15 had a marked inhibitory effect on the formation of capillary structure by HUVECs in vitro. These results suggest that new anti-metastatic and anti-angiogenic agents, which have a dual inhibitory effect on the degradation of the extra cellular matrix and cell motility, may be developed from bestatin.     

The experimental chemotherapeutic N-furfuryladenosine (kinetin-riboside) induces rapid ATP depletion, genotoxic stress, and CDKN1A (p21) upregulation in human cancer cell lines

Cytokinins and cytokinin nucleosides are purine derivatives with potential anticancer activity. N⁶-furfuryladenosine (FAdo, kinetin-riboside) displays anti-proliferative and apoptogenic activity against various human cancer cell lines, and FAdo has recently been shown to suppress tumor growth in murine xenograft models of human leukemia and melanoma. In this study, FAdo-induced genotoxicity, stress response gene expression, and cellular ATP depletion were examined as early molecular consequences of FAdo exposure in MiaPaCa-2 pancreas carcinoma, A375 melanoma, and other human cancer cell lines. FAdo, but not adenosine or N⁶-furfuryladenine (FA), displayed potent anti-proliferative activity that was also observed in human primary fibroblasts and keratinocytes. Remarkably, massive ATP depletion and induction of genotoxic stress as assessed by the alkaline comet assay occurred within 60-180 min of exposure to low micromolar concentrations of FAdo. This was followed by rapid upregulation of CDKN1A and other DNA damage/stress response genes (HMOX1, DDIT3, and GADD45A) as revealed by expression array and Western analysis. Pharmacological and siRNA-based genetic inhibition of adenosine kinase (ADK) suppressed FAdo cytotoxicity and also prevented ATP depletion and p21 upregulation suggesting the importance of bioconversion of FAdo into the nucleotide form required for drug action. Taken together our data suggest that early induction of genotoxicity and energy crisis are important causative factors involved in FAdo cytotoxicity.     

Antimelanoma activity of the redox dye DCPIP (2,6-dichlorophenolindophenol) is antagonized by NQO1

Altered redox homeostasis involved in the control of cancer cell survival and proliferative signaling represents a chemical vulnerability that can be targeted by prooxidant redox intervention. Here, we demonstrate that the redox dye 2,6-dichlorophenolindophenol (DCPIP) may serve as a prooxidant chemotherapeutic targeting human melanoma cells in vitro and in vivo. DCPIP-apoptogenicity observed in the human melanoma cell lines A375 and G361 was inversely correlated with NAD(P)H:quinone oxidoreductase (NQO1) expression levels. In A375 cells displaying low NQO1 activity, DCPIP induced apoptosis with procaspase-3 and PARP cleavage, whereas G361 cells expressing high levels of enzymatically active NQO1 were resistant to DCPIP-cytotoxicity. Genetic (siRNA) or pharmacological (dicoumarol) antagonism of NQO1 strongly sensitized G361 cells to DCPIP apoptogenic activity. DCPIP-cytotoxicity was associated with the induction of oxidative stress and rapid depletion of glutathione in A375 and NQO1-modulated G361 cells. Expression array analysis revealed a DCPIP-induced stress response in A375 cells with massive upregulation of genes encoding Hsp70B’ (HSPA6), Hsp70 (HSPA1A), heme oxygenase-1 (HMOX1), and early growth response protein 1 (EGR1) further confirmed by immunodetection. Systemic administration of DCPIP displayed significant antimelanoma activity in the A375 murine xenograft model. These findings suggest feasibility of targeting tumors that display low NQO1 enzymatic activity using DCPIP.     

Inhibitors of 5-lipoxygenase inhibit expression of intercellular adhesion molecule-1 in human melanoma cells

INTRODUCTIONAlthough malignant melanoma accounts for lessthan 5 % of all cancers, it is among the most lethalbecause of its extremely high metastatic propensity[1-3].Metastasis develops after melanoma cancer cells com-plete a complex sequence of events which include dis-sociation from the primary cancer, invasion and sur-vival in the circulation and extravasation and prolifera-tion at a second site[4]. At each step during the cascade,melanoma cancer cells interact with various host cells.…     

N‐(Indazolyl)benzamido Derivatives as CDK1 Inhibitors: Design,Synthesis, Biological Activity,and Molecular Docking Studies

A series of N‐1H‐indazole‐1‐carboxamides has been synthesized and their effects on both CDK1 / cyclin B and the K‐562 (human chronic myelogenus leukemia) cell line were evaluated. Using a computational model, we have observed that all the most active compounds 9e , f , i – n exhibited the same binding mode of purvanalol A in the ATP‐binding cleft. Although they were able to moderately inhibit the leukemic cell line K‐562 and to show inhibitory activity against the Cdc2‐Cyclin B kinase in the low micromolar range, they turned out to be non‐cytotoxic against HuDe (IZSL) primary cell cultures from human derm. These preliminary results are quite encouraging in view of the low toxicity demonstrated by the above‐mentioned compounds.     

Oridonin induces human melanoma A375-S2 cell death partially through inhibiting insulin-like growth factor 1 receptor signaling

Our previous studies indicated that oridonin, a diterpenoid isolated from Rabdosia rubescens, induced human melanoma A375-S2 cell apoptosis. In this study, we investigated whether the proapoptotic effect of oridonin on A375-S2 cells would depend on an interference with function of the insulin-like growth factor 1 (IGF-1) receptor, a plasma membrane receptor critical for the survival or antiapoptotic ability in melanoma cells. We found that IGF-1 receptor (IGF-1R) signaling was a potential survival pathway against a low concentration of 20 μmol/L oridonin-induced apoptosis in A375-S2 cells. The activation of Ras or its downstream effector p38 mitogen-activated protein kinase (p38 MAPK) was shown to be necessary for IGF-1-mediated protection, but the activation of phosphatidylinositol-3-OH kinase (PI3 kinase) or extracellular signal-regulated kinase (ERK) did not correlate with the regulation of survival. However, in the presence of 40 μmol/L (IC₅₀ at 24 h) oridonin, A375-S2 cells could not be protected by IGF-1 from apoptosis, accompanied by a severe impairment of IGF-1R expression. Therefore, we concluded that the proapoptotic activity of oridonin was partially attributed to its repression of IGF-1R signaling. In addition, p53 was supposed to be a pivotal transducer of proapoptotic and survival signaling pathway in this system.     

体外转染Kiss-1基因对人黑色素瘤细胞A375生长增殖的影响

目的 探讨转染Kiss-1基因对人黑色素瘤细胞A375生长增殖的影响,筛选出有意义的肿瘤治疗的生物学靶标.方法 在人黑色素瘤细胞A375中转染Kiss-1基因,经G418筛选,建立稳定高表达Kiss-1蛋白的细胞系.Western blot方法 检测Kiss-1蛋白以证实转染成功.流式细胞术检测Kiss-1基因对A375生长增殖的影响.结果 稳定转染Kiss-1基因的A375细胞中不仅Kiss-1蛋白稳定表达（1.20±0.21）高于对照组（0.60±0.41）,Kiss-1基因转染A375细胞48 h后对其具有凋亡的作用（凋亡率为28.42%）高于对照组（2.12%）.结论 外源性Kiss-1基因导入A375细胞后,可诱导A375细胞其凋亡,从而抑制肿瘤细胞增殖.     

Multidirectional anti-melanoma effect of galactolipids (MGDG-1 and DGDG-1) from Impatiens parviflora DC. and their synergy with doxorubicin

The anti-melanoma potential of galactolipids: MGDG-1 and DGDG-1, isolated from Impatiens parviflora, and their synergistic effect with anticancer drug – doxorubicin (DOX) was investigated. Both compounds demonstrated time- and dose-dependent cytotoxicity against human melanoma cells of different metastatic potential. MGDG-1 was more effective than DGDG-1, with the highest activity against A375 cell line (IC₅₀ = 15.14 μg/mL). Both compounds acted selectively, were devoid of hepatotoxicity or mutagenicity. Additionally, MGDG-1 proved to be a tyrosinase inhibitor.Co-administration of MGDG-1 and DGDG-1 with DOX revealed a synergistic cytotoxic effect on melanoma cells. The cytotoxicity of all tested MGDG-1/DOX and DGDG-1/DOX cocktails was considerably higher than that of each agent administered alone. MGDG-1/DOX (Mix3) reduced the viability of A375 melanoma cells almost totally and this effect was 2-fold more potent as compared to DOX alone. Our study indicates that the overall effect is enhanced with the increasing concentration of MGDG-1 in the cocktail. These results open up a possibility for lowering therapeutic doses of chemotherapeutics such as doxorubicin when co-administrated with galactolipids. Thus, MGDG-1 can be prospectively considered as multidirectional anti-melanoma agent and can be recommended for further in vitro and in vivo studies, especially in search for effective combined therapy.     

紫草素对人黑素瘤细胞增殖及黑素合成的影响

目的探讨紫草素对人黑素瘤细胞增殖及黑素合成的影响。方法采用MTT法测定紫草素对人A375黑素瘤细胞增殖的影响,NaOH裂解法测定黑素合成。并分成实验组（紫草素组）、对照组（空白组）和阳性对照组[甲氧沙林（8-MOP）组],对其结果进行比较分析。结果紫草素浓度在0.25～1μmol/L对人A375黑素瘤细胞增殖无影响（P〉0.05）,2μmol/L和4μmol/L的紫草素对于A375黑素瘤细胞的增殖有一定的抑制作用（P〈0.05或P〈0.01）。紫草素对于人A375黑素瘤细胞中酪氨酸酶活性呈剂量依赖性激活作用。实验组与对照组比较,0.25、0.5及1μmol/L紫草素对人A375黑素瘤细胞酪氨酸酶活性有激活作用,差异有统计学意义（P〈0.01）;且阳性对照组紫草素作用优于对照组,差异有统计学意义（P〈0.05）;实验组浓度为0.25、0.5及1μmol/L紫草素溶液和阳性对照组与对照组比较,对于人A375黑素瘤细胞中黑素合成有激活作用,差异有统计学意义（P〈0.05或P〈0.01）。结论紫草素对于培养的人黑素瘤细胞酪氨酸酶活性及黑素生成有较强的剂量相关性激活作用。     

Synthesis and evaluation in vitro of 1-[2-(10-dihydroartemisininoxy) ethyl]-3-phenylurea derivatives as potential agents against cancer

In order to develop potent and selective anticancer agents, a series of novel artemisinin derivatives bearing urea moiety 1a–n were facilely synthesized herein and screened for their activities in vitro against ten human tumor cell lines (HeLa, MCF-7, U937, K562, HL60, HCT116, HepG2, A549, A375-S2, and HT1080). The pharmacological results indicated that some compounds showed excellent activity against cancer cell lines and good selectivity, especially the compound 1c which proved to be the most active against the cancer cells as well as distinctive patterns of selectivity.     

吲哚脲类化合物的设计合成及其体外抗肿瘤活性研究

目的以索拉非尼为先导物,设计并合成一系列吲哚脲类化合物,并对其体外抗肿瘤活性进行初步评价。方法以5-硝基吲哚-2-甲酸为起始原料,采用BOP法合成酰胺,再将硝基还原成胺基,最后与异氰酸酯缩合,共3步反应制备目标化合物;采用MTT法评价目标化合物对4种肿瘤细胞株(MX-1、A375、HepG2、Ketr3)的生长抑制作用。结果与结论合成了28个吲哚脲类新化合物,其结构经1H-NMR和HR-MS确证。体外活性结果表明,与索拉非尼相比多数化合物选择性地作用于MX-1细胞株,显示出较强的抑制肿瘤细胞增殖的活性。其中含甲基哌啶的化合物26、30和31抑制MX-1和A375细胞生长的作用显著强于索拉非尼。尤其是化合物31抑制A375细胞增殖的作用是索拉非尼的10倍,对HepG2的抑制活性与索拉非尼相当,IC50值均达到微摩尔级水平,值得进一步研究。     

Induction of growth inhibition and G1 arrest in human cancer cell lines by relatively low-toxic diamantane derivatives

We describe the discovery of a novel series of antitumor diamantane derivatives which induces G1 arrest in Colo 205 cells. Eight diamantane derivatives were screened for their activity in vitro against 60 human cancer cell lines in the National Cancer Institute (NCI)'s anticancer drug screen. The relationships between structure and in vitro antitumor activity are discussed. The structure-activity relationship (SAR) study of diamantane derivatives clarified that the conformation of 1,6-bis(4-(4-aminophenoxy)-phenyl)diamantane (1,6-DPDONH2) was essential for significant antitumor activity. Very strong growth inhibition of 1,6-DPDONH2 (NSC-706829) was observed against one colon cancer line (Colo 205), four melanoma lines (MALME-3M, M14, SK-MEL-5 and UACC-257) and two breast cancer lines (MDA-MB-435 and MDA-N) with GI50 <1.0 microM, i.e. below 0.01, 0.23, 0.48, 0.5, 0.32, 0.26 and 0.28 microM, respectively. 1,6-DPDONH2 also exhibited particular selectivity against one colon cancer line (Colo 205), four melanoma lines (MALME-3M, M14, SK-MEL-5 and UACC-257) and two breast cancer lines (MAD-MB-435 and MDA-N) with GI50 < or=0.5 microM. In the same cancer subpanel, the selectivity of 1,6-DPDONH2 between these seven most sensitive lines and the least sensitive line ranged from 40- to 100-fold. With the exception of melanoma lines, 1,6-bis(4-(4-amino-3-hydroxyphenoxy)-phenyl)diamantane (1,6-DPD/OH/NH2) (NSC-706831) possessed stronger activity than 1,6-DPDONH2 against almost all tested cancer lines. Very strong growth inhibition of 1,6-DPD/OH/NH2 was observed against one leukemia line (HL-60(TB)), one NSCLC line (HOP-92), one ovarian cancer line (OVCAR-8) and one breast cancer line (T-47D) with GI50 <1.0 microM, i.e. 0.50, 0.85, 0.62 and 0.75 microM, respectively.     

薯蓣皂苷元抗肿瘤衍生物的设计、合成与抗肿瘤活性

薯蓣皂苷元对A375、K562等细胞株具有抑制生长并诱导其凋亡的作用。其作用靶点之一是线粒体中的Bcl-2亚家族蛋白。本文以Bcl-2为靶点,运用Autodock设计并合成了一系列全新的薯蓣皂苷元衍生物,希望完善相关化合物的构效关系及提高其抗肿瘤活性。MTT法体外抗肿瘤活性研究结果表明,所设计的化合物大多对K562、A375、A549等3种肿瘤细胞株有较好的抑制作用,而对H293、L02等2种正常细胞株无明显作用。其中,化合物1、6～8对K562表现出了较好的活性(IC50值为1.96～4.35μmol.L-1)。