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OBJECTIVE To evaluate the safety and efficacy of intratumoral injection of polylactic-co-glycolic acid (PLGA) microspheres containing cobra venom cytotoxin in nude mice with transplanted human hepatoma. METHODS Cytotoxic activity of cytotoxin from cobra venom was determined by using methyl thiazolyl tetrazolium method in vitro. Microspheres containing cobra venom cytotoxin were prepared with a double emulsion-solvent evaporation method. Forty BALB/c nude mice were inoculated subcutaneously in right flank with hepatoma BEL-7404 cells. Thirty-two mice whose tumor size reached about 1.0 cm in diameter, were randomly assigned into normal saline group, blank microsphers group, cytotoxin group and cytotoxin-PLGA group. Nude mice were intratumorally injected with normal saline, blank microspheres, cytotoxin or cytotoxin-PLGA microspheres respectively. Internal echo characteristics and blood flow of tumors were observed by high-frequency ultrasound every week after treatment. Twenty-six days after treatment, the tumors were removed to calculate the inhibition rate of tumor growth. The tumor, heart, liver and kidney tissues were obtained for histopathological examination. RESULTS The cytotoxin separated and purified from crude cobra venom caused intense cytotoxic effects to the BEL-7404 cells in vitro. The diameter of PLGA microspheres containing cobra venom cytotoxin was about (34.45+/-9.85)microm. Encapsulation rate was up to (78.13+/-8.92)%, and cumulative amount of cobra venom cytotoxin released from the PLGA microspheres in vitro during 30 days was up to 84.3%. After intratumoral injection, tumor volumes and weights in the cytotoxin-PLGA group were lower than those in the normal saline group, with a tumor growth inhibition rate of 52.36%. Observed under a light microscope, most tumor tissues were necrotic. No obvious morphological change could be seen on the liver, kidney and heart tissues. CONCLUSION The above findings indicate that intratumoral injection of cytotoxin-PLGA microspheres has strong antitumor effect and can obviously lessen systemic toxicity, which may provide an effective and feasible method for hepatocellular carcinoma treatment.

OBJECTIVE: To explore the efficacy and feasibility of interstitial chemotherapy using thermosensitive gel-coated ricin in hepatoma H22-bearing mice. METHODS: Ricin was purified by chromatography method. The purified ricin was identified by Western blot assay and the purity was determined by high-performance liquid chromatography. BALB/c mice were inoculated subcutaneously in right flank with hepatoma H22 cells. When the tumor size reached about 1.0 cm in diameter, 40 mice were randomly divided into untreated group, thermosensitive gel group, ricin group and thermosensitive gel-coated ricin group. Mice in each group were administered different agents by percutaneous intratumoral injection, including normal saline, thermosensitive hydrogel, ricin and thermosensitive gel-coated ricin. Fifteen days after treatment, the tumors were removed to calculate inhibition rate of tumor growth. The tumor tissues were made into pathological sections to perform histopathological examination. The ultrastructure of tumor tissue was examined by electron microscope examination as well. Blood was collected to detect the hepatic and renal functions. The caspase-3 activity of tumor tissue was determined by using zymologic method with a spectrophotometer. RESULTS: After intratumoral therapy, tumor weight in the thermosensitive gel-coated ricin group was lower than that in the untreated group, with a tumor growth inhibition rate of 71.31%. No obvious hepatic or renal toxicities were detected after thermosensitive gel-coated ricin treatment. Histopathologic observation of the tumor tissue showed massive necrosis and typical apoptosis phenomena, including chromatin margination and apoptotic body. Meanwhile, thermosensitive gel-coated ricin resulted in a significant increase in the caspase-3 activity as compared with the untreated group and the ricin group (P

AIM To investigate the anti-tumor effects and mechanisms of interstitial chemotherapy using intra-tumor injection of thermosensitive gel-coated ricin in nude mice bearing a human hepatoma. METHODS In a subcutaneous mouse model of hepatoma, saline, blank gel, ricin, or thermosensitive gel-coated ricin (TGR) was injected directly into tumors. Fourteen days later, eight mice in each group were sacrificed. The tumors were removed and weighed for calculating tumor growth inhibition rate. Serum alpha-fetoprotein levels, as well as hepatic and renal functions, were measured. Tumor tissue was analyzed under an optical microscope. Terminal deoxynucleotidyl transferase mediated dUTP nick end labeling was used to detect the apoptotic index. Moreover, caspase-3 activity and protein expression in tumor tissue were examined. The survival time of the tumor bearing mice was determined. RESULTS Following interstitial chemotherapy by intra-tumor injection of TGR in nude mice, serum alpha-fetoprotein levels were significantly reduced with no significant impact on hepatic or renal functions. The rate of tumor growth inhibition was 58.5% following a single, local injection. Histological analysis revealed abundant necrosis. The apoptotic index was 45.96 +/- 7.41%. Caspase-3 activity was increased, and caspase-3 protein was significantly activated in tumor cells. Compared to the saline group, the survival time of mice in the TGR group was significantly extended. At the observation terminal time, day 120, two mice were still alive and fully recovered. CONCLUSION Interstitial chemotherapy by intra-tumor injection of TGR was highly efficient and safe for the treatment of nude mice bearing a human hepatoma. Interstitial chemotherapy exhibits inhibitory effects by inducing apoptosis and directly killing tumor cells.