Preliminary Study On Preclinical Pharmacokinetics Of Recombinant Ganoderma Lucidum Immunoregulatory Protein(rLZ-8)

Ling Zhi-8（LZ-8） is an immunomodulatory protein derived from Ganoderma lucidum. Publications both in China and abroad have conclusively proven that r LZ-8which is produced by gene recombination technology as well as the native LZ-8 has much biological activity such as immune suppression and promoting lymphocyte proliferation, and could have a very high-potential as a drug candidate. However, no research on the preclinical pharmacokinetic of LZ-8 has yet been published.The preclinical pharmacokinetic of r LZ-8 was preliminarily studied in this thesis.The r LZ-8 was labeled with I¹²⁵ by Iodogen method, TCA precipitation was combined to study the dynamic process of r LZ-8 over time and clarify the pharmacokinetics properties and tissue distribution and excretion of rats. This preclinical pharmacokinetics study of r LZ-8 benefits advanced clinical research and could also provide necessary basis for new drug application.After SHPLC determined the average radiochemical purity of I¹²⁵-r LZ-8 was97.44%. The results of biological activity identification experiment showed that Iodogen method nearly has no influence to the biological activity of r LZ-8 and this demonstrates that I¹²⁵-r LZ-8 meets the requirements of pharmacokinetic experiments.This study used two methods of administration, intravenous and subcutaneous for a single dose. I¹²⁵-r LZ-8 concentrations in serum and blood clots were measured for each method. 38 SD rats were divided into eight groups, four rats in each group, 2male and 2 female; Group 1 and group 2 were used for the pharmacokinetics test, the dose was 500 μg/kg for each subject in each group; Group 3 to Group 6 were used for tissue distribution test at 6 h, 12 h, 24 h and 48 h; Group 7 and group 8, respectively was used in fecaluria excretion and biliary excretion test. Each dose was 500 μg/kg and the last 6 rats were used in blank bases.The experiment results show:（1） After I¹²⁵-r LZ-8 was administered intravenously in rats, primary pharmacokinetic parameters were calculated by measuring the drug concentration in the serum. Cmax,AUC0-t and T1/2 were respectively 5487.58 ng/m L, 12884.55 h*ng/m L and 34.62 h;By measuring the drug concentration of blood clots, the Cmax, AUC0-t and T1/2 were respectively 3200.60 ng/m L, 7647.83 h*ng/m L and 21.67 h; After intravenous administration, serum and blood clot drug exposure ratio is 1: 0.59.（2）After I¹²⁵-r LZ-8 was administered subcutaneously in rats, primary pharmacokinetic parameters was calculated by measuring the drug concentration in the serum. Cmax and AUC0-t were 128.20 ng/m L and 5638.81 h*ng/m L; By measuring the drug concentration of blood clots, the Cmax, AUC0-t were respectively 53.46 ng/m L and3040.20 h*ng/m L. After subcutaneous administration, serum and blood clot drug exposure ratio is 1: 0.54. After I¹²⁵-r LZ-8 was administered subcutaneously in rats, the bioavailability（serum plus blood clots） was 42.27%.（3） After administered subcutaneously, r LZ-8 was mainly distributed in the lung,spleen, lymph nodes.whereas in the fat, brain, spinal cord and muscles I¹²⁵-r LZ-8 was less distributed.（4） After intravenous injections in the rats the I¹²⁵-r LZ-8 radioactive metabolites were excreted primarily in the urine, and then in fecal excretion. About 74.27% ± 5.73% of injected radioactive substances were excreted in urine with 2.52%± 0.42% in feces excreted within 12 days. After intravenous injection, biliary excretion of radioactive metabolites within 8 h accounted for 2.00% ± 0.64% of the injected radioactive substances which demonstrate that the radioactive substances were excreted mainly in urine and kidney is the main excretion organ.