| Summary: | 博士 === 國防醫學院 === 生命科學研究所 === 102 === Anti-tuberculosis drug-induced hepatotoxicity (ATDIH) is the most common adverse drug reaction caused by first-line anti-TB drugs, i.e. isoniazid (INH), pyrazinamide (PZA) and rifampin (RIF). Among them, INH is the most potent single agent against TB but is also the most frequent cause of ATDIH. Prior researches indicated that hepatotoxic free radicals and hydrazine, deriving from catalysis of INH by CYP2E1 and amidase, are the major contributions to INH induced liver injury. The hepatotoxic side effects should rationally be avoided through suitable modifications on CYP2E1 and amidase enzyme activity. In addition, it has been reported that certain genetic polymorphisms of the metabolic enzymes are also the risk factors contributed to ATDIH. Therefore, it is important to understand the correlations between the genetic variation of these metabolic enzymes and the clinical outcome in patients.
The primary objectives of this study are: (1) To screen of effective novel amidase inhibitors from common pure compounds derived from food, natural products or pharmaceutical excipients, so that to inhibit amidase activities from human liver and intestines and to improve clinical benefits by avoiding unwanted side effects. (2) To investigate the genetic polymorphisms of metabolic enzymes and correlate them with the susceptibility of anti-TB drugs induced hepatotoxicity in TB patients, so as to select representative high risk genotypes in patients with the hope for applying the genetic information in routine rapid diagnosis.
In the current study, we use INH as the model drug to establish an in vitro screening test for amidase inhibition. Among 94 Chinese herbal medicine, pure compounds and pharmaceutical excipients tested, 7 were found potent on inhibiting amidase activities by at least 50% or above. HUCHE033, a pure compound with 65.9% in vitro amidase inhibition, was chosen for subsequent animal study in vivo. Effect of protection from INH or INH/RIF induced hepatotoxicity has been demonstrated in mice animal model. Furthermore, it also showed in vivo inhibitory effects on the intrinsic amidase and CYP2E1 in pharmacokinetic studies, both in human and mice.
On the other hand, we analyzed the allele frequency of 43 SNP genetic polymorphisms on INH related metabolic enzymes, including NAT2, CYP2E1 and CES1. Risks for developing ATDIH among different genotypes were also compared. Results showed that, 7 NAT2 SNPs and 4 CES1 SNPs were highly correlated to the incidence of ATDIH, while 7 CYP2E1 SNPs were shown correlated with the intensity (or severity) of ATDIH. Among the best predictive genotypes, patients with high risk genotypes had 3.8 to 14.0 folds higher odds to develop ATDIH, as defined according different levels of liver injury, as compared to those without the genotypes.
In conclusion, the study successfully identified amidase inhibitors with competent potential for developing new anti-TB drug combinations with low hepatotoxic side effects. Several SNP genotypes from the drug metabolic enzyme NAT2, CYP2E1 and CES1 were also demonstrated predicting values on susceptibilities of ATDIH in patients. These results should help to provide novel pathways on development of new diagnosis or medications for caregivers to combat with TB.
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