Allosteric activation of midazolam CYP3A5 hydroxylase activity by icotinib - Enhancement by ketoconazole.
Icotinib (ICO), a novel small molecule and a tyrosine kinase inhibitor, was developed and approved recently in China for non-small cell lung cancer. During screening for CYP inhibition potential in human liver microsomes (HLM), heterotropic activation toward CYP3A5 was revealed. Activation by icotinib was observed with CYP3A-mediated midazolam hydroxylase activity in HLM (∼40% over the baseline) or recombinant human CYP3A5 (rhCYP3A5) (∼70% over the baseline), but not in the other major CYPs including rhCYP3A4. When co-incubated with selective CYP3A4 inhibitor CYP3cide or monoclonal human CYP3A4 inhibitory antibody in HLM, the activation was extended to ∼60%, suggesting CYP3A5 might be the isozyme involved. Further, the relative activation was enhanced to ∼270% in rhCYP3A5 in the presence of ketoconazole. The activation was substrate and pathway dependent and observed only in the formation of 1'-OH-midazolam, and not 4-OH-midazolam, 6β-OH-testosterone, or oxidized nifedipine. The activation requires the presence of cytochrome b5 and it is only observed in the liver microsomes of dogs, monkeys, and humans, but not in rats and mice. Kinetic analyses of 1'-OH-midazolam formation showed that ICO increased the Vmaxvalues in HLM and rhCYP3A5 with no significant changes in Kmvalues. By adding CYP3cide with ICO to the incubation, the Vmaxvalues increased 2-fold over the CYP3cide control. Addition of ketoconazole with ICO alone or ICO plus CYP3cide resulted in an increase in Vmaxvalues and decrease in Kmvalues compared to their controls. This phenomenon may be attributed to a new mechanism of CYP3A5 heterotropic activation, which warrants further investigation.
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China. Electronic address: xiaomeizhuang@163.com.
XiaoMei
X
Zhuang
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
TianHong
T
Zhang
International School, Beijing University of Posts and Telecommunications, Beijing, China.
SiJia
S
Yue
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
Juan
J
Wang
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
Huan
H
Luo
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
YunXia
Y
Zhang
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
Zheng
Z
Li
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
JinJing
J
Che
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
HaiYing
H
Yang
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, China.
Hua
H
Li
Bristol Myers Squibb, Wallingford, CT, USA.
MingShe
M
Zhu
Department of DMPK, Biogen, MA, USA. Electronic address: chuang.lu@biogen.com.
Chuang
C
Lu
D000494:Allosteric Regulation; D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology*; D043844:Crown Ethers / Q000378:metabolism / Q000494:pharmacology*; D051544:Cytochrome P-450 CYP3A / Q000378:metabolism*; D004285:Dogs; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D000882:Haplorhini; D006801:Humans; D066298:In Vitro Techniques; D007654:Ketoconazole / Q000378:metabolism / Q000494:pharmacology*; D051379:Mice; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D008874:Midazolam / Q000378:metabolism*; D011799:Quinazolines / Q000378:metabolism / Q000494:pharmacology*; D051381:Rats; D013045:Species Specificity; D013379:Substrate Specificity
D000970:Antineoplastic Agents; D043844:Crown Ethers; D011799:Quinazolines; C531470:icotinib; C510164:CYP3A5 protein, human; D051544:Cytochrome P-450 CYP3A; D008874:Midazolam; D007654:Ketoconazole
1′-OH-MDZ, 1′-hydroxy-midazolam (PubChem CID: 107917); 4-OH-MDZ, 4-hydroxy-midazolam (PubChem CID: 124449); Allosteric activation; Amodiaquine (PubChem CID: 2165); Bupropion (PubChem CID: 444); CYP3A5; CYP3cide (PubChem CID: 71315139); Dextromethorphan (PubChem CID: 5360696); Diclofenac (PubChem CID: 3033); Drug-drug interaction; ICO, icotinib (PubChem CID: 22024915); Icotinib; KTZ, ketoconazole (PubChem CID: 456201); Ketoconazole; MDZ, midazolam (PubChem CID: 4192); NIF, nifedipine (PubChem CID: 4458); Phenacetin (PubChem CID: 4754); Propranolol (PubChem CID: 4946); S-mephenytoin (PubChem CID: 107921); Species differences; TST, testosterone (PubChem CID: 6013)