Lillibridge J H, Liang B H, Kerr B M, Webber S, Quart B, Shetty B V, Lee C A
Agouron Pharmaceuticals Inc., San Diego, CA 92121, USA.
Drug Metab Dispos. 1998 Jul;26(7):609-16.
In vitro studies with human liver microsomes and P450 probe substrates were performed to characterize selectivity and mechanism of cytochrome P450 inhibition by nelfinavir mesylate. At therapeutic concentrations (steady-state plasma concentrations approximately 4 microM), nelfinavir was found to be a competitive inhibitor of only testosterone 6beta-hydroxylase (CYP3A4) with a Ki concentration of 4. 8 microM. At supratherapeutic concentrations, nelfinavir competitively inhibited dextromethorphan O-demethylase (CYP2D6), S-mephenytoin 4-hydroxylase (CYP2C19), and phenacetin O-deethylase (CYP1A2) with Ki concentrations of 68, 126, and 190 microM, respectively. Nelfinavir did not appreciably inhibit tolbutamide 4-hydroxylase (CYP2C9), paclitaxel 6alpha-hydroxylase (CYP2C8), or chlorzoxaxone 6beta-hydroxylase (CYP2E1) activities. The inhibitory potency of nelfinavir toward CYP3A4 suggested the possibility of in vivo inhibition of this isoform, whereas in vivo inhibition of other P450s was considered unlikely. In a one-sequence crossover study in 12 healthy volunteers, nelfinavir inhibited the elimination of the CYP3A substrate terfenadine and the carboxylate metabolite of terfenadine. The 24-hr urinary recoveries of 6beta-hydroxycortisol were reduced by an average of 27% during nelfinavir treatment, consistent with CYP3A inhibition by nelfinavir. Inhibition of CYP3A4 by nelfinavir in vitro was NADPH-dependent requiring the catalytic formation of a metabolite or a metabolic intermediate. The catechol metabolite of nelfinavir (M3) was considered unlikely to be responsible for inhibition as the addition of catechol O-methyl transferase, S-adenosyl methionine, and ascorbic acid to the preincubation mixture did not protect against the loss of testosterone 6beta-hydroxylase activity. Also, the addition of M3 to human liver microsomes did not inhibit CYP3A4. Although incubations with nelfinavir showed a time- and concentration-dependent loss of CYP3A4 activity, the partial or complete recovery of enzyme activity upon dialysis indicated that inhibition was reversible. Microsomal incubations with nelfinavir and NADPH did not result in a loss of spectral P450 content compared with the NADPH control. Glutathione, N-acetylcysteine, and catalase did not attenuate CYP3A4 inhibition by nelfinavir. Collectively, these results suggest that the probable mechanism for CYP3A4 inhibition by nelfinavir is a transient metabolic intermediate or stable metabolite that coordinates tightly but reversibly to the heme moiety of the P450.
利用人肝微粒体和P450探针底物进行了体外研究，以表征甲磺酸奈非那韦对细胞色素P450的抑制选择性和机制。在治疗浓度（稳态血浆浓度约为4 microM）下，发现奈非那韦是仅对睾酮6β-羟化酶（CYP3A4）有竞争性抑制作用的抑制剂，其Ki浓度为4.8 microM。在超治疗浓度下，奈非那韦对右美沙芬O-脱甲基酶（CYP2D6）、S-美芬妥因4-羟化酶（CYP2C19）和非那西丁O-脱乙基酶（CYP1A2）有竞争性抑制作用，其Ki浓度分别为68、126和190 microM。奈非那韦对甲苯磺丁脲4-羟化酶（CYP2C9）、紫杉醇6α-羟化酶（CYP2C8）或氯唑沙宗6β-羟化酶（CYP2E1）的活性没有明显抑制作用。奈非那韦对CYP3A4的抑制效力表明其在体内抑制该同工酶的可能性，而对其他P450的体内抑制作用则被认为不太可能。在一项针对12名健康志愿者的单序列交叉研究中，奈非那韦抑制了CYP3A底物特非那定及其羧酸盐代谢物的消除。在奈非那韦治疗期间，6β-羟基皮质醇的24小时尿回收率平均降低了27%，这与奈非那韦对CYP3A的抑制作用一致。奈非那韦在体外对CYP3A4的抑制作用是NADPH依赖性的，需要催化形成一种代谢物或代谢中间体。奈非那韦的儿茶酚代谢物（M3）不太可能是抑制作用的原因，因为在预孵育混合物中加入儿茶酚O-甲基转移酶、S-腺苷甲硫氨酸和抗坏血酸并不能防止睾酮6β-羟化酶活性的丧失。此外，将M3加入人肝微粒体中也不会抑制CYP3A4。尽管用奈非那韦孵育显示CYP3A4活性呈时间和浓度依赖性丧失，但透析后酶活性的部分或完全恢复表明抑制作用是可逆的。与NADPH对照相比，用奈非那韦和NADPH进行微粒体孵育不会导致光谱P450含量的丧失。谷胱甘肽、N-乙酰半胱氨酸和过氧化氢酶不会减弱奈非那韦对CYP3A4的抑制作用。总体而言，这些结果表明，奈非那韦对CYP3A4的抑制作用可能机制是一种短暂的代谢中间体或稳定的代谢物，它与P450的血红素部分紧密但可逆地结合。
