PLGA/O-CMC载药纳米微粒的体外降解及释药行为研究

本研究以聚乳酸-乙醇酸共聚物（PLGA）和自行制备的O-羧甲基壳聚糖（O-CMC）为原料，以5-氟尿嘧啶（5-FU）为抗癌药物模型，采用自身设计的改良复乳法制备了载药纳米微粒。微粒平均粒径为98.5nm，粒径分布指数为0.192，粒子表面∈电位为61．48eV，载药率高达18.9％。然后用SEM动态监测载药纳米粒子降解过程中表面形貌的变化，并连续追踪粒子降解过程中的质量损失和降解介质的pH变化。载药纳米粒子在PBS中的释药行为研究表明，（1）前12h的释药动力学符合Huguchi方程，具有一级释放特性；（2）在20d内的释药动力学符合零级释放特性。细胞凋亡实验结果表明载药纳米粒子对TJ905脑胶质瘤细胞增殖有明显的抑制作用。     

地高辛-PLGA抗肿瘤纳米粒子的制备及其抗肿瘤活性的初步研究

目的 制备载地高辛的聚乳酸-羟基乙酸共聚物(PLGA)纳米粒子,提高地高辛的生物利用度,降低其毒副作用.方法 建立测定地高辛-PLGA纳米粒子载药量和包封率的高效液相色谱法;采用乳化溶剂挥发法制备地高辛-PLGA纳米粒子,并通过单因素实验优化制备条件;采用噻唑蓝法评价地高辛和地高辛-PLGA纳米粒子的抗肿瘤能力.结果 以粒径为筛选条件的单因素实验结果表明,制备地高辛-PLGA纳米粒子的最佳条件为PLGA 30 mg,地高辛2 mg,二氯甲烷3 ml,聚乙烯醇质量分数0.5％,超声功率200 W.此制备条件下得到的地高辛-PLGA纳米粒子的粒径约231 nm,包封率为74.61％,载药量为5.37％,且其抗肿瘤活性优于地高辛,差异具有统计学意义(P＜0.05).结论 以PLGA为载体材料制备地高辛-PLGA纳米粒子可增强地高辛的抗肿瘤作用.     

Targeting efficiency of RGD-modified nanocarriers with different ligand intervals in response to integrin αvβ3 clustering

Receptor change induced by ligand binding is a new issue to face in the field of targeted delivery. Receptor clustering, the main pattern of receptor changes, decreases the affinity between ligand and receptor due to the redistribution of receptor position. In an attempt to respond to such challenge, we designed and constructed three RGD-modified nanocarriers with different ligand intervals: stealth liposomes modified with the monomeric RGD (moRGD-LP), dimeric RGD (diRGD-LP) and a special dimeric RGD with a linker between two cyclic RGD motifs (P-diRGD-LP). The αvβ3-positive and -negative tumor cells (Melanoma B16 and MCF-7) were used as the cell models. As a result, P-diRGD-LP demonstrated strongest interaction with B16 cells in surface plasmon resonance study and highest cellular uptake in B16 cells in real-time confocal analysis. The enhanced endocytosis of P-diRGD-LP was found to be αvβ3-mediated and P-diRGD-LP increased the involvement of the clathrin-dependent pathway. Importantly, P-diRGD-LP demonstrated the best targeting effect in B16-tumor bearing mice in both in vivo and ex vivo near-infrared fluorescent images, about 2.4-fold that of moRGD-LP and 2.8-fold that of diRGD-LP at 3 h. Further, we validated integrin αvβ3 clustering on B16 cells via a single-molecule imaging by a total internal reflection fluorescence microscopy. Finally, the 3D models of αvβ3 clustering suggested a receptor interval within 41.916–65.779 Å, while the molecular computation revealed an RGD ligand interval of 20.944 Å, 42.753 Å and 78.196 Å for diRGD-LP, P-diRGD-LP and moRGD-LP, respectively, confirming the best matching between clustered αvβ3 and P-diRGD-LP. In conclusion, P-diRGD-LP could achieve higher targeting to αvβ3-positive tumor via the enhanced interaction based on the better ligand-receptor compatibility. The design of targeted nanocarriers against receptor clustering might provide new insight into the nanotechnology-based anticancer therapy.     

槲皮素对大鼠脑胶质瘤抑瘤作用的体内实验研究

目的观察槲皮素及顺铂对大鼠脑胶质瘤生长的抑制作用,初步探讨其作用机制。方法将30只颅内接种C6胶质瘤细胞的SD大鼠随机分为3组:分别给予顺铂、槲皮素及对照治疗7 d,观察各组肿瘤生长情况,于接种后17 d处死各组大鼠,收集肿瘤标本进行肿瘤大小、HE染色及电镜观察,并通过免疫组化检测肿瘤组织增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)的表达。结果顺铂组[(66.2±5.9)mm3]、槲皮素组[(59.6±5.4)mm3]肿瘤体积低于对照组[(101.1±9.6)mm3]槲皮素组优于顺铂组(t=10.61,P<0.01),槲皮素组、顺铂组肿瘤抑制率分别为41.3%、34.8%。光镜观察槲皮素组、顺铂组肿瘤细胞密度降低,电镜显示有细胞凋亡。PCNA免疫组化结果显示槲皮素组(42.2%±5.1%)、顺铂组(50.0%±6.7%)肿瘤PCNA表达减少,与对照组(79.6%±8.6%)相比,具有统计学差异(F=416.12,P<0.01),两两比较,槲皮素组治疗效果优于顺铂组(q=55.25,P<0.01)。结论槲皮素和顺铂均可显著抑制大鼠颅内C6胶质瘤细胞生长,其机制可能与诱导肿瘤细胞凋亡和抑制肿瘤细胞增殖有关。     

Transcatheter arterial chemoembolization with anticancer drug in iodized oil for primary hepatic carcinoma

Transcatheter arterial chemoembolization using a suspension of anticancer drug in iodized oil combined with Gelfoam embolization was performed in 85 patients with advanced primary hepatic carcinoma. Long-term retention of iodized oil in the tumor was observed in 90% of the cases, and changes in tumor size were observable by abdominal X-ray film on follow-up studies. There was a decrease of tumor size in all cases and the overall 1-year survival rate was 43%. In tumors whose diameter was less than 20 cm and in patients without portal vein thrombosis the survival rate was nearly 60%. Combined treatment with anticancer drug in iodized oil and Gelfoam embolization of the proper hepatic artery was superior to use of the suspension alone. There was also better detection of small cancer nodules by the suspension as compared to conventional angiography.     

Analysis of metastatic spread and growth of tumor cells in mice with depressed natural killer activity by anti-asialo GMl antibody or anticancer agents

Summary The mechanism of artificial and spontaneous metastases of tumor was analyzed in B16 melanoma cells and C57BL/6 mice by using anti-asialo GM1 antibody and anticancer agents. Single administrations of 500 g anti-asialo GM1 antibody resulted in significantly decreased NK activity in spleen cells of C57BL/6 mice, lasting 10 days from the day following administration. Treatment with anti-asialo GM1 antibody never decreased the function of T lymphocytes measured by blastogenesis with phytohemagglutinin or T cell growth factor. The tumoricidal functions of activated macrophages but not of resident macrophages were decreased by in vivo treatment with anti-asialo GM1 antibody.The anti-asialo GM1 antibody was evaluated in terms of the enhancing effect on pulmonary metastases with regard to the timing of administration. Treatment with anti-asialo GM1 antibody 1 day before or on the day of tumor inoculation resulted in a substantial increase in the number of artificial pulmonary metastases. In the experimental system of spontaneous metastases, anti-asialo GM1 antibody most effectively increased the number of pulmonary metastases when administered 1–2 weeks before the removal of primary tumor, when the tumor cells are thought to be released into blood circulation from the primary site. In addition, accelerated growth of transplanted tumors at the primary site was observed in mice treated with anti-asialo GM1 antibody. These results strongly suggest that anti-asialo GM1 antibody enhances the incidence of in vivo tumor metastases and the growth of transplanted tumor mainly by suppressing the function of NK cells.The maximum effective dose (MED) of mitomycin C or its derivative (M-83) suppressed NK activity significantly, and pretreatment with these anticancer agents enhanced the growth of the artificial pulmonary and liver metastases. In contrast, the MED of cDDP showed no effect on the NK activity or the numbers of pulmonary and liver metastases. These results indicate that the depression of NK activity induced by chemotherapy results in the promotion of metastatic disease.From these studies it can be concluded that NK cells have a key role in the control of metastases of malignant disease, and that support of NK activity is very important for the prevention of metastases.     

Yeast on drugs: <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> as a tool for anticancer drug research

Abstract The budding yeast Saccharomyces cerevisiae is being widely used as a model for investigating fundamental processes relevant to all living organisms. Many of these processes are affected by genetic and epigenetic alterations in cancer such as cell cycle progression, DNA replication and segregation, maintenance of genomic integrity and stress responses. Therefore, yeast emerges as an attractive model for anticancer drug research. The genetic tractability of budding yeast, its ease of manipulation and the wealth of functional genomics tools available in this organism makes it ideal for genome-wide analysis of biological functions and chemical screenings. The present review will discuss some of the innovative advantages based on yeast genetics and genomics for antitumour drug target identification and drug discovery. *Supported by an unrestricted educational grant from Roche Farma S.A.     

Sensitivity of Human Cancer Cells to the New Anticancer Ribo-nucleoside TAS–106 Is Correlated with Expression of Uridine-cytidine Kinase 2

TAS–106 [l–(3– C -ethynyl-β- d - ribo -pentofuranosyl)cytosine] is a new anticancer ribo -nucleoside with promising antitumor activity. We have previously presented evidence suggesting that the TAS–106 sensitivity of cells is correlated with intracellular accumulation of the triphosphate of TAS–106, which may be affected both by cellular membrane transport mechanisms and uridine-cytidine kinase (UCK) activity. Since the presence of a UCK family consisting of two members, UCK1 and UCK2, has recently been reported in human cells, we investigated the relation between expression of UCK1 and UCK2 at both the mRNA and protein levels and UCK activity (TAS–106 phosphorylation activity) in a panel of 10 human cancer cell lines. Measurement of UCK activity in these cell lines revealed that it was well correlated with the cells' sensitivity to TAS–106. In addition, the mRNA or protein expression level of UCK2 was closely correlated with UCK activity in these cell lines, but neither the level of expression of UCK1 mRNA nor that of protein was correlated with enzyme activity. We therefore compared the protein expression level of UCK2 in several human tumor tissues and the corresponding normal tissues. Expression of UCK2 protein was barely detectable in 4 of the 5 human tumor tissues, but tended to be high in the pancreatic tumor tissue. It could not be detected at all in any of the normal tissues. Thus, expression of UCK2 appeared to be correlated with cellular sensitivity to TAS–106, and it may contribute to the tumor-selective cytotoxicity of TAS–106.     

Preclinical oral antitumor activity of BMS-185660, a paclitaxel derivative

Purpose: A water soluble paclitaxel derivative, BMS-185660, identified previously as having parenteral activity comparable with that of the parent drug, was evaluated for antitumor activity when given orally. Methods: Staged subcutaneous (s.c.) tumor models of both murine and human origin were used for this purpose. Results: BMS-185660 achieved levels of activity following oral administration which were comparable with those maximum effects obtained using intravenous (i.v.) paclitaxel. Consecutive daily oral administrations of BMS-185660 resulted in maximum gross log cell kill (LCK) values of 1.7–2.0 in two experiments involving the s.c. Madison 109 murine lung tumor model, which were comparable with the best effects of the derivative injected intravenously, and 0.3 to 0.9 LCK greater than the maximum effects obtained with i.v. paclitaxel; paclitaxel given orally was inactive. Against a human ovarian tumor model with developed resistance to cisplatin (A2780/cDDP), oral BMS-185660 achieved a maximum LCK of 1.8 compared with i.v. paclitaxel, which produced a maximum 2.4 LCK. Also, in the human HCT-116 colon carcinoma model, oral BMS-185660 cured a maximum of seven of eight mice compared with six of seven mice cured with i.v. paclitaxel. The loss in potency between comparably effective intravenously and orally administered doses of BMS-185660 was about four- to five-fold, but since no drug-associated lethality was ever observed following the oral administration of the highest doses of BMS-185660, further dose escalation may have been tolerated. The intermediate metabolite between BMS-185660 and paclitaxel is BMS-181681. This compound was also evaluated orally and found not to be active versus s.c. M109, despite demonstrating good activity by the i.v. route. Conclusion: The comparable activities of both intravenously and orally administered BMS-185660 to intravenously administered paclitaxel, combined with the attribute of improved water solubility, provides a good basis for further derivative development. Received: 10 September 1999 / Accepted: 16 March 2000