| HCPT(Hydroxycamptothecin), a compound isolated from extracts of the Asian tree Camptotheca Acuminata in the1960s(6), is a cytotoxic quinoline alkaloid and a promising class of anti-tumor agent, which was proved usefully in the treatment of a broad spectrum of human tumors. Pharmacodynamics studies with HCPT showed that when inhaled the same dosage of drug by mice, a more significant inhibition was observed than that given by intravenous injection (i.v.) or intraperitoneal injection (i.p.) administration. But the research accordance of influence of aerosol administration to the pulmonary metastasis cancer, preclinical pharmacokinetics, excretion and metabolism are lacked in present. As aerosol delivery system and HCPT developed more and more deeply, it is necessary to investigate aerosol delivery and lung cancer therapy.On the basis of preview research, we investigate the pharmacokinetics, tissue distribution and excretion of HCPT in mice with lung cancer after aerosol administration. Then the correlation between CYP450and HCPT is studied. Finally we explore the antitumor mechanism of HCPT to improve the effect.We established B16F10melanoma pulmonary metastasis mice model and to investigated the drug concentrations of lactone and carboxylate in the lung, plasma and other tissues of the mice with pulmonary cancer from i.v. and aerosol treatment. After aerosol administration, the lactone content was only23.2%and18.6%of the carboxylate and total HCPT in the plasma, respectively. Comparatively, the lactone and carboxylate level in the plasma was lower after i.v. administration. For aerosol administration, the lactone concentration-time profile of lung declined in parallel with the profile of lactone in plasma with lactone concentration in lung remaining higher than lactone concentration in plasma. The pharmacokinetics parameters in the lungs showed the superiority of this treatment.The AUC value of total HCPT in liver, heart and kidney after i.v. administration were higher than that of aerosol treatment. The AUC value and MRT of total HCPT in the liver were highest after aerosol treatment. The AUC value of total HCPT in the liver was highest after i.v. administration, but the MRT of the total HCPT in the heart was longest.After aerosol administration of HCPT, the ratios of cumulative excretion in feces and urine were8.27%and1.21%, respectively. The major of exeretion in urine and feces turned into metabolites. Then four metabolites were speculated according to the molecular ion peak and the structural fragments by using LC-MS.In the study about the correlation between CYP450activity and HCPT metabolism, the metabolic kinetics parameters showed that the enzyme from CYP450had a great affinity for L-HCPT and the reaction rate of L-HCPT was fast, so L-HCPT was chosen as a substrate. L-HCPT from50～200μM inhibited the metabolism of testosterone and acetaminophen and had no effect on the metabolism of metoprolol tartrate, phenacetin, diclofenac sodium and omeprazole. So we concluded that HCPT was affected by CYP3A4and CYP2E1in the metabolic processes and competitively inhibited the metabolism of testosterone and acetaminophen in liver microsomes. In order to verify this result we added specific CYP3A4inhibitor and CYP2E1inhibitor in the incubation system, the metabolism of HCPT was inhibited.Finally, in order to enhance the antitumor effect and clinical application of HCPT, real-time fluorescent PCR technology was used to study the expression of BCL-2and P53at different concentrations of HCPT to B16F10melanoma cells in different time. P53protein was not expressed and after HCPT treatment the expression level was improved in a concentration-and time-dependent manner. After HCPT treatment the BCL-2protein expression decreased in a concentration-and time-dependent manner. HCPT may be inhibit the proliferation of B16F10melanoma cells through increasing expression of P53and reducing the expression of BCL-2, thus inducing cell apoptosis. |
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