In vitro metabolism, reaction phenotyping, enzyme kinetics, CYP inhibition and induction potential of ataluren

Abstract

Ataluren promotes ribosomal readthrough of premature termination codons in mRNA which result from nonsense mutations. In vitro studies were performed to characterize the metabolism and enzyme kinetics of ataluren and its interaction potential with CYP enzymes. Incubation of [14C]‐ataluren with human liver microsomes indicated that the major metabolic pathway for ataluren is via direct glucuronidation and that the drug is not metabolized via cytochrome P450 (CYP). Glucuronidation was also observed in the incubation in human intestinal and kidney microsomes, but not in human pulmonary microsomes. UGT1A9 was found to be the major uridine diphosphate glucuronosyltransferase (UGT) responsible for ataluren glucuronidation in the liver and kidney microsomes. Enzyme kinetic analysis of the formation of ataluren acyl glucuronide, performed in human liver, kidney, and intestinal microsomes and recombinant human UGT1A9, found that increasing bovine serum albumin (BSA) levels enhanced the glucuronidation Michaelis‐Menten constant (Km) and ataluren protein binding but had a minimal effect on maximum velocity (Vmax) of glucuronidation. Due to the decreased unbound Michaelis‐Menten constant (Km,u), the ataluren unbound intrinsic clearance (CLint,u) increased for all experimental systems and BSA concentrations. Human kidney microsomes were about 3.7‐fold more active than human liver microsomes, in terms of CLint,u/mg protein, indicating that the kidney is also a key organ for the metabolism and disposition of ataluren in humans. Ataluren showed no or little potential to inhibit or induce most of the CYP enzymes.

Drug interaction of ningetinib and gefitinib involving CYP1A1 and efflux transporters in non‐small cell lung cancer patients

“…For comparison of CL int in different tissues, CL int was expressed per gram of tissue by correcting the values for microsomal protein yield, resulting in a scaled CL int. 20 The scaled CL int values of ningetinib in HLMs and HIMs were 36.7 and 4.66 μL·min −1 ·g −1 tissue, respectively, which showed that the liver had a higher metabolic contribution in M1 formation than the intestine.…”

“…V max = 4.30 ± 0.26 pmolÁminÁmg −1 protein, intrinsic clearance CL int = 0.918 μLÁminÁmg −1 protein) than in HIMs (K m = 6.30 ± 1.52 μM, V max = 1.39 ± 0.12 pmolÁmin −1 Ámg −1 protein, CL int = 0.221 μLÁminÁmg −1 protein). For comparison of CL int in different tissues, CL int was expressed per gram of tissue by correcting the values for microsomal protein yield, resulting in a scaled CL int 20. The scaled CL int values of ningetinib in HLMs and HIMs were 36.7 and 4.66 μLÁmin −1 Ág −1 tissue,…”

Ataluren Prevented Bone Loss Induced by Ovariectomy and Aging in Mice Through the BMP-SMAD Signaling Pathway

“…This defective gene product causes many diseases, including muscular dystrophy, cystic brosis, and some cancers (38-40). Ataluren is a small-molecule nonsense repressor that stimulates stop codon reading and thereby acts as a treatment for these diseases (41). The addition of Stop-POST5 (containing the UGA stop codon at the A-site) is shifted to extension.…”

Ataluren—Promising Therapeutic Premature Termination Codon Readthrough Frontrunner

“…However, the concentrations of the free ataluren in human plasma after administration of the most typical dosage regimen (10, 10, 20 mg/kg) is much lower than inhibition constants toward these enzymes due to the high protein binding of the drug. What is more, clinically irrelevant induction of CYP2B6 and CYP2C9 activity in cultured primary human hepatocytes was also observed [ 42 ].…”