菜籽饼提取物抑制黑色素生成作用

为了研究菜籽饼提取物对黑色素合成的抑制作用,为菜籽饼提取物作为美白化妆品的天然添加剂提供实验依据。采用ABTS法、DPPH法检测菜籽饼提取物清除自由基的能力,测定A-375黑色素细胞中酪氨酸酶活性和黑色素含量,并利用各自的试剂盒检测A-375细胞中抗氧化相关因子(MDA, SOD, GSH)。研究结果表明:菜籽饼提取物具有清除DPPH和ABTS的能力,对DPPH和ABTS的半数抑制率分别为304μg/mL和305.8μg/mL。菜籽饼提取物具有显著的抑制A-375黑色素的作用,药物作用48 h后,其IC50为113.3μg/m L,同时能够显著的降低黑色素的含量和酪氨酸酶的活性。与空白对照组(17.66 mol/ng)相比,菜籽饼提取物能显著降低A-375细胞内MDA含量,其抑制率为9.21 mol/ng,且能够显著提高SOD的含量,当浓度为125μg/m L时,SOD的含量较空白对照组提高了36.54 U/mL,与此同时,菜籽饼提取物亦能增加GSH的含量,当浓度为62.5μg/mL时,细胞中GSH的含量为6.43 mol/ng,其含量显著高于空白对照组。因此,菜籽饼提取物在降低黑色素含量、抑制酪氨酸酶活性的同时,也能够通过显著的抗氧化能力和清除自由基的能力,抑制A-375细胞中黑色素的生成。     

桦褐孔菌提取物对胃癌MGC-803细胞株的抗增殖与诱导凋亡作用

探讨了桦褐孔菌提取物对胃癌MGC - 80 3细胞株的抗增殖、诱导凋亡作用及对凋亡相关基因表达的影响。MTT比色法结果显示 ,桦褐孔菌提取物在 0 .5～ 16 0 μg/mL范围内 (其IC50 为 4 μg/mL)对胃癌MGC- 80 3细胞株均有抑制作用 ,并表现出浓度依赖性关系 ;凋亡形态学观察结果 ,药物浓度 2 μg/mL ,作用时间 12～ 2 4h后 ,细胞核染色质固缩并凝结成块 ,聚集在核膜周边 ,凋亡小体形成 ;TUNEL法检测结果 ,不同浓度药物均诱导胃癌MGC - 80 3细胞株凋亡 ,细胞凋亡率随药物浓度增加而上升 ,显示明显的量效关系。Ki- 6 7抗原检测结果 ,不同浓度药物均抑制胃癌MGC - 80 3细胞株增殖 ,表现出浓度、时间依赖性关系 ;2μg/mL桦褐孔菌提取物作用胃癌MGC - 80 3细胞株 4 8h之后 ,明显下调Bcl - 2基因蛋白表达。因此 ,通过此项研究可得出 ,桦褐孔菌提取物对胃癌MGC - 80 3细胞株有抗增殖作用和诱导凋亡作用 ,其凋亡的分子生物学机制可能与下调凋亡抑制基因Bcl 2表达有一定关系。     

The metabolic bioactivation of caffeic acid phenethyl ester (CAPE) mediated by tyrosinase selectively inhibits glutathione S-transferase

Glutathione S-transferase (GST) and multidrug resistance-associated proteins (MRPs) play major roles in drug resistance in melanoma. In this study, we investigated caffeic acid phenethyl ester (CAPE) as a selective GST inhibitor in the presence of tyrosinase, which is abundant in melanoma cells. Tyrosinase bioactivates CAPE to an o-quinone, which reacts with glutathione to form CAPE-SG conjugate. Our findings indicate that 90% CAPE was metabolized by tyrosinase after a 60-min incubation. LC–MS/MS analyses identified a CAPE-SG conjugate as a major metabolite. In the presence of tyrosinase, CAPE (10–25 μM) showed 70–84% GST inhibition; whereas in the absence of tyrosinase, CAPE did not inhibit GST. CAPE-SG conjugate and CAPE-quinone (25 μM) demonstrated ?85% GST inhibition via reversible and irreversible mechanisms, respectively. Comparing with CDNB and GSH, the non-substrate CAPE acted as a weak, reversible GST inhibitor at concentrations >50 μM. Furthermore, MK-571, a selective MRP inhibitor, and probenecid, a non-selective MRP inhibitor, decrease the IC₅₀ of CAPE (15 μM) by 13% and 21%, apoptotic cell death by 3% and 13%, and mitochondrial membrane potential in human SK-MEL-28 melanoma cells by 10% and 56%, respectively. Moreover, computational docking analyses suggest that CAPE binds to the GST catalytic active site. Caffeic acid, a hydrolyzed product of CAPE, showed a similar GST inhibition in the presence of tyrosinase. Although, as controls, 4-hydroxyanisole and l-tyrosine were metabolized by tyrosinase to form quinones and glutathione conjugates, they exhibited no GST inhibition in the absence and presence of tyrosinase. In conclusion, both CAPE and caffeic acid selectively inhibited GST in the presence of tyrosinase. Our results suggest that intracellularly formed quinones and glutathione conjugates of caffeic acid and CAPE may play major roles in the selective inhibition of GST in SK-MEL-28 melanoma cells. Moreover, the inhibition of MRP enhances CAPE-induced toxicity in the SK-MEL-28 melanoma cells.     

Synthesis and anticancer activity of 5'-chloromethylphosphonates of 3'-azido-3'-deoxythymidine (AZT)

A series of novel N-alkyl 5'-chloromethylphosphonates of 3'-azido-3'-deoxythymidine (6-15) was synthesized by means of phosphonylation of 3'-azido-3'-deoxythymidine (4) with P-chloromethylphosphonic ditriazolide (3) followed by a reaction with the appropriate amine. The synthesized phosphonamidates 6-15 were evaluated for their cytotoxic activity in two human cancer cell lines: oral (KB) and breast (MCF-7) using the sulforhodamine B (SRB) assay. The highest activity in KB human cancer cells was displayed by phosphonamidate 8 (IC(50)=5.8 μg/mL), however, this compound was less potent than the parent AZT (IC(50)=3.1 μg/mL). Phosphonamidate 10 showed only moderate activity (IC(50)=12.1 μg/mL) whereas the other phosphonamidates proved inactive. Similarly, the highest activity in MCF-7 human cancer cells was displayed by phosphonamidate 8 (IC(50)=3.7 μg/mL) but it proved somewhat less active than AZT (IC(50)=2.6 μg/mL). Some activity was also displayed by phosphonamidate 10 (IC(50)=12.8 μg/mL) but the other phosphonamidates were found inactive. Hydrolysis studies indicate that the synthesized phosphonamidates are likely to act as prodrugs of the parent nucleoside (AZT). Transport measurements showed that the most active phosphonamidates (8 and 10) were able to permeate across the intestinal epithelium in vitro. The apparent permeability coefficients determined in Caco-2 cell monolayers indicated that these compounds could be moderately absorbed in humans.     

MicroRNA-330 inhibits growth and migration of melanoma A375 cells: In vitro study

Melanoma skin cancer is one of the main causes of male cancer-related deaths worldwide. It has been suggested that miR-330-5p can act as a tumor suppressor in various types of cancers. So, in this study, we replaced miR-330 in melanoma cancer cells by vector-based miR-330 to evaluate the effects of this microRNA on the growth and migration inhibition of melanoma cancer cells, and to determine the molecular mechanisms underlying its action. By using the MTT assay, the IC₅₀ of Geneticin antibiotic was obtained as 460 µg/mL. The results of the qRT-PCR showed the increased expression level of miR-330 and decreased expression levels of MMP-9, CXCR4, Vimentin, melanoma cell adhesion molecule, AKT1, and E2F1 messenger RNA in A375 transfected cells. The cytotoxicity assay results demonstrated the inhibition of cancer cells proliferation. Furthermore, the wound healing test results showed a migration reduction of transfected cells with miR-330 compared with nontransfected ones. In addition, 4′,6-diamidino-2-phenylindoleLB: Luria-Bertani (DAPI) staining revealed the significant nucleus fragmentation in miR-330 replaced cells, which correspond to apoptosis induction in replaced cells. The results showed that increase in miR-330 expression level could significantly inhibit the tumor cell growth and the migration of melanoma cancer cells.     

Declination of long noncoding RNA paternally expressed gene 10 inhibits A375 cells proliferation,migration, and invasion via mediating microRNA-33a

The importance of long noncoding RNAs (lncRNAs) has been certified in malignant melanoma. Nonetheless, the functions of lncRNA paternally expressed gene 10 (PEG10) in malignant melanoma remain uninvestigated. This research discloses the influence of PEG10 in the biological actions of malignant melanoma cells. The sh-PEG10 plasmid was transfected into A375 cells; meanwhile, the effects of declined PEG10 on cell viability, apoptosis, migration, invasion, and the correlative protein levels were probed. The miR-33a expression in sh-PEG10-transfected cells was examined, and the above biological processes were studied again in miR-33a inhibitor-transfected A375 cells. Phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and mechanistic target of rapamycin (mTOR) pathways were delved via Western blot. We found that the enhancement of PEG10 was discovered in melanoma tissues compared to related nonmelanoma tissues. Declination of PEG10 frustrated cell viability, repressed cyclinD1 and CDK4 expression, and triggered apoptosis, as well as suppressed migration and invasion in A375 cells. A negative correction between PEG10 and miR-33a was confirmed, and repressed miR-33a inverted the functions of PEG10 repression in A375 cells. In addition, PEG10 repression discouraged the activation of PI3K/AKT and mTOR pathways via elevation of miR-33a. These results indicated that declination of PEG10 restrained A375 cell growth, migration, and invasion via adjusting miR-33a and PI3K/AKT and mTOR pathways.     

补骨脂素抑制膀胱癌T24细胞增殖和迁移的作用及机制研究

目的探讨补骨脂素对人膀胱癌T24细胞存活率、细胞周期、细胞凋亡和迁移的影响及其分子机制。 方法分别用细胞培养液、3‰二甲基亚砜（DMSO）和不同浓度（10、30、50、100 μg/mL）补骨脂素处理膀胱癌细胞分成对照组、DMSO组和补骨脂素组,CCK-8检测细胞存活率。流式细胞术检测细胞周期和细胞凋亡。划痕实验检测划痕愈合率。RT-qPCR法检测磷脂酰肌醇3激酶（PI3K）和蛋白激酶B （AKT） mRNA表达水平、Western blot法检测PI3K和AKT蛋白的表达及磷酸化情况。多组间比较采用单因素方差分析,组间两两比较采用LSD-t检验。 结果与DMSO组比较,除10 μg/mL补骨脂素作用24 h外,其余浓度补骨脂素作用不同时间的细胞存活率随着补骨脂素浓度增高、作用时间延长而逐渐降低（P < 0.05）。与DMSO组比较,30、100 μg/mL补骨脂素干预24 h后,G1期细胞比例增多,G2/M期比例减少,细胞凋亡率[（9.16±0.97）%、（15.45±1.57）%比（1.02±0.36）%]升高,划痕愈合率[24 h：（45.00±3.44）%、（27.60±2.21）%比（66.10±2.61）%,48 h：（70.00 ± 3.40）%、（45.17±2.44）%比（85.17±3.85）%]降低,PI3K、AKT mRNA表达以及PI3K、AKT蛋白表达水平和磷酸化水平均降低（P均< 0.05）。 结论补骨脂素降低膀胱癌细胞存活率、阻滞细胞周期、诱导细胞凋亡和抑制细胞迁移,其机制可能与下调PI3K、AKT mRNA、蛋白表达及磷酸化水平有关。     

吴茱萸碱对黑色素瘤细胞的耐药逆转作用

黑色素瘤是目前恶性程度最高的肿瘤之一。临床上,常采用维罗非尼（PLX4032）作为晚期患者的治疗药物,但患者很快就出现了耐药。因此,如何克服耐药、提高患者生存率成为急需解决的问题。本文选用中药吴茱萸碱（EVO）与黑色素瘤A375细胞、对PLX4032耐药的A375细胞（A375/R）进行相关研究。采用CCK-8法检测发现,用EVO的细胞非毒性浓度0.5 μmol/L处理A375/R,PLX4032对A375/R细胞的半数抑制浓度（IC50）降低,逆转倍数为3.85。显示EVO能够增强黑色素瘤A375/R细胞对PLX4032的敏感性。后续实验分为对照组、EVO组、PLX组、PLX + EVO组。流式细胞仪检测结果显示,EVO组细胞凋亡率为5.88%,PLX组细胞凋亡率为17.88%,PLX + EVO组细胞凋亡率为30.28%。细胞集落结果证明,PLX4032联合EVO能够抑制A375/R细胞的克隆形成。免疫印迹法结果证明,联合用药能够上调促凋亡蛋白质Bax、胱天蛋白酶3的表达量,下调p-Akt、p-NF-κB-p65及抗凋亡蛋白Bcl-2的蛋白质水平。以上结果均表明,EVO能够有效逆转黑色素瘤A375/R细胞耐药,并且诱导细胞凋亡,抑制细胞增殖。     

吴茱萸碱对黑色素瘤细胞的耐药逆转作用

黑色素瘤是目前恶性程度最高的肿瘤之一。临床上,常采用维罗非尼（PLX4032）作为晚期患者的治疗药物,但患者很快就出现了耐药。因此,如何克服耐药、提高患者生存率成为急需解决的问题。本文选用中药吴茱萸碱（EVO）与黑色素瘤A375细胞、对PLX4032耐药的A375细胞（A375/R）进行相关研究。采用CCK-8法检测发现,用EVO的细胞非毒性浓度0.5 μmol/L处理A375/R,PLX4032对A375/R细胞的半数抑制浓度（IC50）降低,逆转倍数为3.85。显示EVO能够增强黑色素瘤A375/R细胞对PLX4032的敏感性。后续实验分为对照组、EVO组、PLX组、PLX + EVO组。流式细胞仪检测结果显示,EVO组细胞凋亡率为5.88%,PLX组细胞凋亡率为17.88%,PLX + EVO组细胞凋亡率为30.28%。细胞集落结果证明,PLX4032联合EVO能够抑制A375/R细胞的克隆形成。免疫印迹法结果证明,联合用药能够上调促凋亡蛋白质Bax、胱天蛋白酶3的表达量,下调p-Akt、p-NF-κB-p65及抗凋亡蛋白Bcl-2的蛋白质水平。以上结果均表明,EVO能够有效逆转黑色素瘤A375/R细胞耐药,并且诱导细胞凋亡,抑制细胞增殖。     

早孕因子单克隆抗体对黑色素瘤细胞A-375增殖、凋亡的影响

目的:研究早孕因子单克隆抗体(EPF-McAb)对黑色素瘤细胞A-375增殖、凋亡的影响。方法:体外培养黑色素瘤细胞A-375,通过CCK-8实验检测EPF-McAb对A-375细胞增殖的影响;通过流式细胞术检测EPF-McAb对A-375细胞凋亡的影响;Western Blot检测EPF-McAb对A-375细胞EPF蛋白表达的影响。结果:CCK-8结果显示EPF-McAb可以抑制黑色素瘤细胞A-375的增殖,且随着作用时间延长和药物浓度的增加,对A-375细胞增殖的抑制作用也增强;流式细胞术实验结果显示EPF-McAb可以促进黑色素瘤细胞A-375的凋亡,且调亡率随着药物浓度的增加而升高,0.2、0.4、0.8 mg/m L的EPF单抗作用于A-375细胞24 h后,细胞调亡率分别为:14.68%(P0.01)、19.81%(P0.01)、23.97%(P0.01);Western Blot实验结果显示EPF-McAb可以降低黑色素瘤细胞A-375 EPF蛋白的表达。结论:早孕因子单克隆抗体可以抑制黑色素瘤细胞A-375的增殖,并促进其凋亡。     

Extraction,isolation and in vitro evaluation of affinisine from Tabernaemontana catharinensis in human melanoma cells

Plant compounds have been identified as new drug prototypes. In this line, this work aimed to isolate the indole alkaloid affinisine from Tabernaemontana catharinensis and test its antitumor activity. The alkaloid was isolated by silica gel open column chromatography from the ethanolic extract of the stem of T. catharinensis. Afterwards, this molecule was characterized by high-resolution mass spectrometry and nuclear magnetic resonance. In the next step, the cytotoxicity of the compound was tested against human melanoma cell lines (A375, WM1366 and SK-MEL-28) and a normal skin cell line (CCD-1059Sk) using a MTT (3-4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Cells treated with affinisine were evaluated by flow cytometry to analyze apoptosis and the induction of cell cycle arrest, to evaluate the dead mechanism. The metabolite was isolated in a 0.2% yield relative to the extract. Cytotoxic activity of the molecule was observed at 48 h, resulting in considerable growth inhibition rates in melanoma cells, especially in WM1366, which had the lowest IC₅₀ (32.86 ± 2.54 µg/mL). The apoptosis rate was lower in A375 (56.66 and 86.71% with 57 and 65 µg/mL, respectively). Moreover, affinisine was able to significantly induce cell cycle arrest in different phases in the A375 and WM1366 cell lines. However, in SK-MEL-28 cells, cycle arrest was not observed. In summary, this compound significantly decreased the viability of tumor cells in a dose- and time-dependent manner for all evaluated lineages, reduced cell viability by the apoptosis mechanism and presented prominent activities of cell cycle arrest. In this way, the use of antineoplastic agents is among the most widely used therapeutic measures for the control and treatment of cancer. Affinisine is a promising prototype in the search for new drugs to treat cancer.     

新狼毒素A诱导人黑色素瘤A375细胞凋亡的机制研究

为探讨新狼毒素A抑制人黑色素瘤A375细胞增殖及诱导凋亡的作用。本实验采用噻唑蓝(MTT)法检测新狼毒素A对人黑色素瘤A375细胞增殖的影响,Hoechst33258法观察细胞形态,流式细胞仪检测细胞凋亡率和线粒体膜电位,Westernblot检测凋亡相关蛋白表达。结果显示新狼毒素A以时间剂量依赖性的方式显著抑制A375细胞的增殖;不同浓度新狼毒素A(0、15、30、45μmol/L)作用于A375细胞48h后细胞出现显著凋亡特征,新狼毒素A增加A375细胞的凋亡率且降低了线粒体膜电位,上调Bax、caspase-3和细胞色素C(CytochromeC)蛋白的表达,下调Bcl-2蛋白的表达。以上结果说明新狼毒素A抑制A375细胞增殖,诱导细胞凋亡,其机制可能与诱导线粒体途径的凋亡有关。     

Synthesis, biological evaluation and molecular docking studies of resveratrol derivatives possessing curcumin moiety as potent antitubulin agents

A series of resveratrol derivatives possessing curcumin moiety were synthesized and evaluated for their antiproliferative activity against three cancer cell lines including murine melanoma B16-F10, human hepatoma HepG2 and human lung carcinoma A549. Among them, compound C5 displayed the most potent in vitro antiproliferative activity against B16-F10 with IC(50) value of 0.71 μg/mL. Compound C5 also exhibited good tubulin polymerization inhibitory activity with IC(50) value of 1.45 μg/mL. Furthermore, docking simulation was carried out to position C5 into the tubulin-colchicine binding site to determine the probable binding mode.     

CB-RAF600E-1 exerts efficacy in vemurafenib-resistant and non-resistant-melanoma cells via dual inhibition of RAS/RAF/MEK/ERK and PI3K/Akt signaling pathways

Background and AimPredicting novel dual inhibitors to combat adverse effects such as the development of resistance to vemurafenib in melanoma treatment due to the reactivation of MAPK and PI3K/AKT signaling pathways is studied to help in reversal of cancer symptoms.Reversal of cancer symptoms in melanoma associated with vemurafenib resistance is driven by reactivation of MAPK and PI3K/Akt signaling pathways. Novel dual inhibitors targeting these proteins would be beneficial to combat resistance.MethodsHigh-throughput virtual screening of the ChemBridge library against B-RAFV600E and Akt was performed using an automated protocol with the AutoDock VINA program. Luminescence and time-resolved fluorescence kits were used to measure enzyme activities. The MTT assay was used to determine proliferation in normal and vemurafenib-resistant A375 cells. Flow cytometry was used to examine apoptosis, cell cycle, and phosphorylation of ERK/Akt signaling pathway.ResultsHigh-throughput screening from the ChemBridge library identified 15 compounds with high binding energy towards B-RAFV600E; among these, CB-RAF600E-1 had the highest ΔG_binding score −11.9 kcal/mol. The compound also had a high affinity towards Akt, with a ΔG_binding score of −11.5 kcal/mol. CB-RAF600E-1 dose-dependently inhibited both B-RAFV600E and Akt with IC₅₀ values of 635 nM and 154.3 nM, respectively. The compound effectively controlled the proliferations of normal and vemurafenib-resistant A375 cells, with GI₅₀ values of 222.3 nM and 230.5 nM, respectively. A dose-dependent increase in the sub G₀/G₁ phase of the cell cycle and total apoptosis was observed following compound treatment in both normal and vemurafenib-resistant melanoma cells. Treatment with CB-RAF600E-1 decreased the pERK/pAkt dual-positive populations in normal and vemurafenib-resistant A375 cells.ConclusionCB-RAF600E-1, identified as a novel dual inhibitor effective against normal and vemurafenib-resistant melanoma cells, requires further attention for development as an effective chemotherapeutic agent for melanoma management.     

New ionic derivatives of betulinic acid as highly potent anti-cancer agents

Betulinic acid is a natural compound with high in vitro cytotoxicity toward many cancer cells. However, the poor water solubility of this compound hampers an effective in vivo cancer study. We prepared new ionic derivatives of betulinic acid with higher water solubilities, without losing the structural integrity and functionality of this compound. As a result, these new ionic derivatives have shown much higher inhibitory effects against different cancer cell lines such as melanoma A375, neuroblastoma SH-SY5Y and breast adenocarcinoma MCF7. For A375 cell lines, the derivative 5 exhibited a low IC(50) value of 36 μM (vs 154 μM for betulinic acid); for MCF7 cell lines, the derivative 5 also exhibited a low IC(50) value of 25 μM (vs 112 μM for betulinic acid). The high cytotoxicity of these new derivatives can be linked to their greatly improved water solubility. Our assay method used little DMSO in aiding the dissolution of these derivatives to demonstrate the advantage of improved water solubility and to mimic the in vivo study conditions. The cell viability studies based on both MTT and LDH assay methods have confirmed the high inhibitory effect of our ionic derivatives of betulinic acid (particularly 4 and 5) against different cancer cells.     

Down-regulation of cellular FLICE-inhibitory protein (Long Form) contributes to apoptosis induced by Hsp90 inhibition in human lung cancer cells

Background

Breast cancer is the most common cancer in the Arab world and it ranked first among Saudi females. Doxorubicin (DOX), an anthracycline antibiotic is one of the most effective anticancer agents used to treat breast cancer. chronic cardiotoxicity is a major limiting factor of the use of doxorubicin. Therefore, our study was designed to assess the role of a natural product resveratrol (RSVL) on sensitization of human breast cancer cells (MCF-7) to the action of DOX in an attempt to minimize doxorubicin effective dose and thereby its side effects.

Methods

Human breast cancer cell line MCF-7, was used in this study. Cytotoxic activity of DOX was determined using (sulforhodamine) SRB method. Apoptotic cells were quantified after treatment by annexin V-FITC- propidium iodide (PI) double staining using flow-cytometer. Cell cycle disturbance and doxorubicin uptake were determined after RSVL or DOX treatment.

Results

Treatment of MCF-7 cells with 15 μg/ml RSVL either simultaneously or 24 h before DOX increased the cytotoxicity of DOX, with IC50 were 0.056 and 0.035 μg/ml, respectively compared to DOX alone IC50 (0.417 μg/ml). Moreover, flow cytometric analysis of the MCF-7 cells treated simultaneously with DOX (0.5 μg/ml) and RSVL showed enhanced arrest of the cells in G₀ (80%). On the other hand, when RSVL is given 24 h before DOX although there was more increased in the cytotoxic effect of DOX against the growth of the cells, however, there was decreased in percentage arrest of cells in G₀, less inhibition of DOX-induced apoptosis and reduced DOX cellular uptake into the cells.

Conclusion

RSVL treatment increased the cytotoxic activity of DOX against the growth of human breast cancer cells when given either simultaneously or 24 h before DOX.     

Design, synthesis, and antiproliferative activity of new 1H-pyrrolo[3,2-c]pyridine derivatives against melanoma cell lines. Part 2

A new series of diarylureas and diarylamides possessing 1H-pyrrolo[3,2-c]pyridine scaffold was designed and synthesized. Their in vitro antiproliferative activities against A375P human melanoma cell line and NCI-9 human melanoma cell line panel were tested. All the target compounds, except three amino derivatives 8g, h and 9h, demonstrated superior potencies against A375P to Sorafenib. In addition, compounds 8a and 9b-f demonstrated higher potencies than Vemurafenib against A375P. Compounds 8c and 9b were 7.50 and 454.90 times, respectively, more selective towards A375P melanoma cells over NIH3T3 fibroblasts. Furthermore, compounds 8d, e and 9a-d, f demonstrated very high potencies against the nine tested melanoma cell lines at the NCI. The bisamide derivatives 9a-c, f showed 2-digit nanomolar IC(50) values over different cell lines of the NCI-9 melanoma cell lines.     

布雷菲德菌素A通过线粒体凋亡途径增强顺铂抗肺癌GLC-82细胞的作用

本研究证明了线粒体凋亡途径在布雷菲德菌素A（brefeldin A,BFA）联合顺铂（cis-dichlorodiamine platinum,CDDP）抗非小细胞肺癌（non-small cell lung cancer,NSCLC）中的作用。MTT结果显示,BFA对肺癌GLC-82和NCI-H1299细胞的半数有效抑制浓度（half maximal inhibitory concentration,IC50）分别是100 ng/mL和400 ng/mL,CDDP对GLC-82和NCI-H1299细胞的IC50分别是4 μg/mL和15 μg/mL;而分别采用半量的BFA和CDDP联合处理GLC-82或NCI-H1299细胞后,抑制作用均进一步加强。DAPI染色结果进一步证明了二者的协同作用——与单独用药组相比,细胞核染色质固缩加剧,核裂解碎片增多,乃至形成凋亡小体,表明细胞凋亡的发生。与单药组比较,联合用药导致肺癌GLC-82细胞线粒体膜电位显著下降;q-RT-PCR及Western印迹结果显示,在联合用药早期（24 h）,GLC-82细胞可能通过提高Bcl2表达以促进存活;而在联合用药晚期（48 h）,细胞已发生不可逆转的凋亡,Bcl2表达受抑制,同时二者通过促进Bax表达来诱导细胞色素C释放,使胱天蛋白酶 3发生剪切激活,最终诱导细胞凋亡发生。提示线粒体凋亡途径可能是BFA协同CDDP抗非小细胞肺癌的分子机制之一,为肺癌的临床治疗方案提供了更多的理论依据。     

负载雷公藤甲素的TPGS-b-(PCL-ran-PGA)纳米制剂体外抑制宫颈癌细胞生长的研究

雷公藤甲素（triptolide,TPL）是传统中药雷公藤的主要活性成分,具有抗炎、抗肿瘤活性,但其毒副作用限制了临床上的广泛使用。为了探讨以TPGS-b-(PCL-ran-PGA)为载体制备的TPGS-b-(PCL-ran-PGA)/TPL纳米粒的表征和体外对宫颈癌细胞的抑制作用,采用乳化/溶剂挥发法,优化TPGS-b-(PCL-ran-PGA)与TPL比例,制备TPGS-b-(PCL-ran-PGA)/TPL纳米粒,对纳米粒进行表征,包括粒径大小、ζ电位、包封率、累积释放率,用MTS法体外研究游离型TPL和TPGS-b-(PCL-ran-PGA)/TPL纳米粒对宫颈癌细胞半数抑制浓度（IC50）,用克隆形成实验分析TPGS-b-(PCL-ran-PGA)/TPL纳米粒对宫颈癌细胞HeLa的抑制作用,用流式细胞仪分析纳米粒对HeLa细胞凋亡的影响。结果显示：当TPGS-b-(PCL-ran-PGA)与TPL为50∶1时制备的纳米粒粒径为(95.3±5.2)nm,zeta电位为(-12.2±0.9)mV,其累积释放曲线呈双相分布,TPGS-b-(PCL-ran-PGA)纳米粒对HeLa细胞在24、48和72 h的IC50（2.8、1.8、0.9 μg·L-1）远远低于游离型TPL（P<0.01）,克隆形成实验证明纳米粒能显著抑制肿瘤细胞生长,并能显著诱导HeLa细胞凋亡。研究结果表明,TPGS-b-(PCL-ran-PGA)/TPL纳米粒能抑制宫颈癌细胞HeLa的生长,其作用主要通过TPL和TPGS共同诱导细胞凋亡,可以作为抗宫颈癌等肿瘤的候选药物。