Characterization Of A Novel CYP1A2 Knockout Rat Model Constructed By CRISPR/Cas9 And Related Drug Metabolism Study

Cytochrome P450 1A2(CYP1A2),as one of the most important CYP isoforms,is involved in the biotransformation of many important endogenous and exogenous substances.Since CYP1A2 can catalyze the biotransformation of many precancerous substances and poisons,it plays an important role in the occurrence and development of many diseases.Although the generation of Cyp1a2 knockout(KO)mouse model has been reported and used in pharmacology and toxicology research,there are still no relevant rat models for the study of CYP1A2-mediated pharmacokinetics and diseases.In this report,homozygous CYP1A2 KO rat model was established successfully by using CRISPR/Cas9 technology without any detectable off-target effect for the first time.Compared with wild-type rats,this model showed a loss of CYP1A2 protein expression.The results of liver microsomes incubation experiments in vitro and pharmacokinetics experiments in vivo of two CYP1A2 specific substrates,namely caffeine and tizanidine,further confirmed the lack of function of CYP1A2 in KO rats.To determine the potential compensatory effects of other relevant genes,several important CYP isoforms and nuclear receptor-related genes were compared in the m RNA levels between KO and WT rats.Our results showed that CYP1A1 was significantly up-regulated,and CYP2E1,CYP3A2 and LXR? were down-regulated in KO rats.In addition,CYP1A2 KO rats exhibited a significant increase in serum cholesterol and free testosterone,accompanied by mild liver damage and lipid deposition,suggesting CYP1A2 deficiency affects lipid metabolism and liver function in rats to some extent.In summary,we successfully constructed the CYP1A2 KO rat model,which provides a very useful tool for studying the metabolic function and physiological function of CYP1A2.