Covalent Tyrosine Kinase Inhibitor Research Articles

Protein structure prediction based on deep learning: HER2 in complex with a covalent inhibitor

HER2 protein overexpression is associated with the malignant degree and poor prognosis of breast cancer. HER2 levels are elevated in 20% of breast tumors. Several covalent tyrosine kinase inhibitors have been found to reduce tumor cell survival and proliferation in vitro and inhibit downstream HER2 signaling. In the field of protein structure prediction, AlphaFold2, which achieved excellent results in CASP14, can periodically predict protein structures with atomic precision in the absence of similar protein structures. In this study, AlphaFold2 was used to predict the monomeric structure of the HER2 protein. This predicted structure was compared to the conformation of HER2 in complex with a covalent inhibitor, allowing for an examination of the conformational changes induced by the inhibitor. By combining the conformational changes of HER2 protein with the docking results of Protein-Ligand Interaction Profiler, other potential binding sites were identified, which could further reveal the mechanism of drug discovery.

Inhibition of Cytochrome P450 2J2-Mediated Metabolism of Rivaroxaban and Arachidonic Acid by Ibrutinib and Osimertinib.

Covalent tyrosine kinase inhibitors (TKIs) ibrutinib and osimertinib are associated with cardiac arrhythmia. The interactions between these TKIs with CYP2J2 that is highly expressed in the human heart are unknown. In vitro metabolism experiments were performed to characterize CYP2J2-mediated metabolism of ibrutinib and osimertinib. Unbound distribution coefficient (Kpuu) for both TKIs was determined in AC16 cardiomyocytes. In vitro reversible and time-dependent CYP2J2 inhibition experiments were conducted with exogenous and endogenous substrates, namely rivaroxaban and arachidonic acid (AA), respectively, where kinetic parameters were estimated via one-site and multisite kinetic modeling. Ibrutinib was efficiently metabolized by CYP2J2 to a hydroxylated metabolite, M35, following substrate inhibition kinetics. Osimertinib is not a substrate of CYP2J2. Both TKIs depicted Kpuu values above 1 and equipotently inhibited CYP2J2-mediated hydroxylation of rivaroxaban in a concentration-dependent manner without time-dependency. The mode of reversible inhibition of CYP2J2-mediated metabolism of rivaroxaban and AA by osimertinib was described by Michaelis-Menten kinetics, whereas a two-site kinetic model recapitulated the atypical inhibitory kinetics of ibrutinib, assuming multiple substrate-binding domains within the CYP2J2 active site. The inhibition of ibrutinib and osimertinib on cardiac AA metabolism could be clinically significant considering the preferable distribution of both TKIs to cardiomyocytes with R cut-off values of 1.160 and 1.026, respectively. The dysregulation of CYP2J2-mediated metabolism of AA to cardioprotective epoxyeicosatrienoic acids by ibrutinib and osimertinib serves as a novel mechanism for TKI-induced cardiac arrhythmia. Mechanistic characterization of CYP2J2-mediated typical and atypical enzyme kinetics further illuminates the unique catalytic properties of CYP2J2. SIGNIFICANCE STATEMENT: We reported for the first time that ibrutinib is efficiently metabolized by CYP2J2. By using rivaroxaban and arachidonic acid (AA) as substrates, we characterized the typical and atypical inhibition kinetics of CYP2J2 by ibrutinib and osimertinib. The inhibition of both drugs on cardiac AA metabolism could be clinically significant considering their preferable distribution to cardiomyocytes. Our findings serve as a novel mechanism for drug-induced cardiac arrhythmia and shed insights into the multisite interactions between CYP2J2 and ligands.

Metabolite Identification in the Preclinical and Clinical Phase of Drug Development.

Metabolite identification plays a critical role in the phases during drug development. Drug metabolites can contribute to efficacy, toxicity, and drug-drug interaction. Thus, the correct identification of metabolites is essential to understand the behavior of drugs in humans. Drug administration authorities (e.g., FDA, EMA, and NMPA) emphasize evaluating the safety of human metabolites with exposure higher than 10% of the total drugrelated components. Many previous reviews have summarized the various methods, tools, and strategies for the appropriate and comprehensive identification of metabolites. In this review, we focus on summarizing the importance of identifying metabolites in the preclinical and clinical phases of drug development. Summarized scenarios include the role of metabolites in pharmacokinetics/pharmacodynamics (PK/PD) analysis, disproportional exposure of metabolites that contribute to drug toxicity, changes in metabolite exposure in renal-impaired patients, covalent tyrosine kinase inhibitors (anticancer drugs), and metabolite identification of drug candidates from natural medicines. This review is aimed to provide meaningful insight into the significant role of metabolite identification in drug development.

Characterization of covalent binding of tyrosine kinase inhibitors to plasma proteins

Eight covalent tyrosine kinase inhibitors (TKIs) were investigated to determine the characteristics of their covalent binding to plasma proteins. The data revealed that their covalent binding to plasma proteins is of species difference. In addition to the reports on neratinib and pyrotinib, osimertinib, alflutinib, AST5902, and ibrutinib were confirmed to covalently bind to the Lys-190 of human serum albumin (HSA). Molecular docking was used to simulate the binding mode of TKIs to HSA. The results exhibited the non-covalent interactions between covalent TKIs and HSA, which stabilize the TKIs-HSA complex and explain the selectivity of covalent binding. The t1/2 values of TKIs that are covalently bound to HSA or human plasma proteins were studied in vitro, and the features highly correlated with the t1/2 were determined by quantitative calculations and linear modeling. Reversibility of the covalent binding and the factors affecting the process of reversibility were evaluated. In conclusion, acrylamide moiety of covalent TKIs can covalently bind to lysine residue of HSA, most of which were determined to be Lys-190. The covalent binding is of species difference, especially between animal and human. Except for osimertinib, covalent binding between TKIs and HSA are reversible.

Integrating pharmacokinetic-pharmacodynamic modeling in preclinical assessment of mutant selective EGFR covalent tyrosine kinase inhibitor EGF816

Integrating pharmacokinetic-pharmacodynamic modeling in preclinical assessment of mutant selective EGFR covalent tyrosine kinase inhibitor EGF816

First-in-human evaluation of CO-1686, an irreversible, highly selective tyrosine kinase inhibitor of mutations of EGFR (activating and T790M).

8010^ Background: Efficacy of existing EGFR tyrosine kinase inhibitors (TKIs) in NSCLC is limited by emergence of the T790M mutation in approximately 60% of patients, and significant skin rash and diarrhea, caused by wild-type (WT)-EGFR inhibition. CO-1686 is an oral, covalent TKI that targets common activating EGFR mutations and T790M, while sparing WT-EGFR. Methods: This is a completed dose finding study in patients with EGFR mutated advanced NSCLC. Patients were previously treated with EGFR TKI and had a tumor biopsy in screening for central EGFR genotyping. CO-1686 was administered twice daily. Endpoints included safety, pharmacokinetics (PK), and efficacy. Results: As of 17thJanuary 2014, 88 patients were treated: 57 with CO-1686 free base (up to 900 mg BID); 31 with CO-1686 HBr (500 to1000 mg BID). 10 transitioned from free base to HBr. 63% were T790M+, median age 61 years, 77% female, 76% white, and 72% ECOG 1. Median number of previous therapies was 3 (1- 7); 40% had >1 prior line of EGFR TKI. PK of the CO-1686 HBr formulation was dose proportional with three times greater exposure than the equivalent free base dose. The dose limiting toxicity (DLT) rate at all doses was <33%. Related AEs (all grades) in ≥ 20% patients were: nausea (25%), fatigue (21%), impaired glucose tolerance/hyperglycemia (21%). Hyperglycemia was well managed with oral hypoglycemics and/or dose reduction. A recommended phase 2 dose of 750 mg BID has been selected. Nine T790M+ patients treated with 900 mg BID (free base) were evaluable for response; 6 (67%) achieved PRs, 2 (22%) achieved SD, one of whom subsequently achieved a PR after transition to CO-1686 HBr. Eight of nine progressed on EGFR TKI immediately before CO-1686. PRs have occurred among patients treated with CO-1686 HBr, however the majority of patients have not reached the first restaging. Efficacy data for at least 41 patients on CO-1686 HBr will be presented at the meeting. Conclusions: CO-1686 has demonstrated promising efficacy against T790M+ EGFR mutant NSCLC. CO-1686 HBr delivered higher exposures than free base and was equally well tolerated. Dose-related WT-driven diarrhea and rash has not been seen. The phase 2/3 program will open in 2014. Clinical trial information: NCT01526928.