Due to a lack of physiologic cytochrome P450 (P450) isoform content, P450 activity is typically only determined at the microsomal level (per milligram of microsomal protein) and not at the isoform level (per picomole of P450 isoform), which could result in the misunderstanding of variations in P450 activity between individuals and further hinder development of personalized medicine. We found that there were large variations in protein content, mRNA levels, and intrinsic activities of the 10 P450s in 100 human liver samples, in which CYP2E1 and CYP2C9 showed the highest expression levels. P450 gene polymorphisms had different effects on activity at two levels: CYP3A5*3 and CYP2A6*9 alleles conferred increased activity at the isoform level but decreased activity at the microsomal level; CYP2C9*3 had no effect at the isoform level but decreased activity at the microsomal level. The different effects at each level stem from the different effects of each polymorphism on the resulting P450 protein. Individuals with CYP2A6*1/*4, CYP2A6*1/*9, CYP2C9*1/*3, CYP2D6 100C>T TT, CYP2E1 7632T>A AA, CYP3A5*1*3, and CYP3A5*3*3 genotypes had significantly lower protein content, whereas CYP2D6 1661G>C mutants had a higher protein content. In conclusion, we first offered the physiologic data of 10 P450 isoform contents and found that some single nucleotide polymorphisms had obvious effects on P450 expression in human normal livers. The effects of gene polymorphisms on intrinsic P450 activity at the isoform level were quite different from those at the microsomal level, which might be due to changes in P450 protein content.

The lack of information concerning individual variation in content and activity of human liver microsomal protein is one of the most important obstacles for designing personalized medicines. We demonstrated that the mean value of microsomal protein per gram of liver (MPPGL) was 39.46 mg/g in 128 human livers and up to 19-fold individual variations existed. Meanwhile, the metabolic activities of 10 cytochrome P450 (CYPs) were detected in microsomes and liver tissues, respectively, which showed huge individual variations (200-fold). Compared with microsomes, the activities of liver tissues were much suitable to express the individual variations of CYP activities. Furthermore, individual variations in thein vivoclearance of tolbutamide were successfully predicted with the individual parameter values. In conclusion, we offer the values for MPPGL contents in normal liver tissues and build a new method to assess thein vitroCYP activities. In addition, large individual variations exist in predicted hepatic clearance of tolbutamide. These findings provide important physiological parameters for physiologically-based pharmacokinetics models and thus, establish a solid foundation for future development of personalized medicines.

Significant change of cytochrome P450s activities in patients with hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is one of the most common types of primary liver cancer and the third most frequent cause of cancer death worldwide. Diethylnitrosamine (DEN) is one of the recognized risk factors for hepatocarcinogenesis likely due to cytochrome P450 2E1 (CYP2E1)-mediated metabolic activation. However, CYP2E1-mediated DEN metabolic activity in non-neoplastic liver tissue from HCC patients has not been determined; the role of CYP2E1 activity, in particular CYP2E1 constitutive activity and CYP2E1 inhibited activity, with respect to the hepatocarcinogenesis induced by DEN is yet not clear. Herein, we determined CYP2E1-mediated DEN metabolic activity in non-neoplastic liver tissue from HCC patients and found that CYP2E1-mediated DEN metabolic activity was significantly elevated with a 43.3% positive rate and clinico-pathological parameters did not affect the activity. Then, using a Sprague-Dawley rat liver tumor model induced by DEN, the relationship between CYP2E1 constitutive/inhibited activity and hepatocarcinogenesis was explored. The results showed that the CYP2E1 constitutive activity was strongly correlated with tumor incidence and severity of liver tumorigenesis (nodule numbers and size), while inhibition of CYP2E1 activity decreased hepatocyte proliferation, liver injury, and liver carcinogenesis in the presence of DEN. In conclusion, the higher CYP2E1 activity would lead to an increased incidence of HCC as a result of CYP2E1-mediated DEN activation. Therefore, higher CYP2E1 activity might be a risk factor for HCC induced by DEN.