■ INTRODUCTION In the relatively young but expanding field of irreversible kinase inhibitor drug discovery, there are two main developments that are of central importance. First, the patients have watched the first wave of low molecular weight protein kinase inhibitors becoming available to them over the past decade. These kinase inhibitors have been approved by the U.S. Food and Drug Administration (FDA) for therapeutic use in oncology indications and constitute an important addition to the arsenal of drugs to combat cancer (Table 1). Without exception, these marketed protein kinase inhibitors have been identified and developed using conventional approaches, i.e., reversible inhibitors that (partly) occlude the ATP pocket in the catalytic domain of the kinase. Although protein kinases are regarded as an attractive drug target family, it took (and still takes) huge efforts to master the human kinome, which comprises more than 500 protein kinases. The search for clinically applicable kinase inhibitors that target the highly conserved ATP pocket has been thwarted by a couple of well-known hurdles. Selectivity, cellular potency, and an increasingly crowded intellectual property arena are major points of attention. The second important development is the renewed interest in covalent binding drugs (reviewed by Potashman and Duggan and by Singh et al). This recent revival results from a better understanding of the benefits of the covalent binding principle and the approval of effective and safe covalent drugs. Historically, drug discoverers have been taught to stay away from small molecular entities that harbor reactive electrophilic groups because these used to be equivalent to promiscuity. Promiscuous hits that relied on reactive groups were traditionally hard to optimize toward leads, since these were more than often interfering with the biochemical assay rather than truly modifying the activity of the target of interest. Even if the target modulation was real, indiscriminant reactivity was believed to trigger insurmountable toxic events that may surface in late stage clinical trials when larger patient populations are involved. As a consequence, “suicide inhibitors”, “warheads”, and covalent irreversible inhibitors developed a negative flavor over time and became almost synonymous with toxicity in some organizations. The skepticism toward irreversible drugs may evaporate as more examples of irreversible drugs progress clinically that demonstrate good efficacy and safety margins. In a nutshell, the therapeutic applicability or the success of irreversible binding kinase inhibitors is dependent on whether or not the covalent bond can be confined solely to the protein kinase of interest. So this approach is in essence a story about two T’s: treatment and toxicity. When relying on the covalent binding principle, it is important to discern adduct-based toxicity and adduct-based treatment, since the adducting molecular entities in question obey overlapping fundamental rules in terms of reactivity. The only key difference is the nature and the function of the proteins that are covalently modified. Covalent kinase inhibitors with well-balanced recognition and reactivity should provide efficacy, selectivity, and ultimately the safety margins that are required for regulatory approval. If we strike the right balance, a third “T” will give enough comfort: therapeutic window. This Perspective aims to give a comprehensive account from a medicinal chemist's point of view on the progress of irreversible kinase inhibitor drug discovery. The “state of the art” is reviewed by means of reported irreversible kinase inhibitors profiles and their chemical structures. The potential upsides and pitfalls that are associated with this concept are highlighted to provide a general understanding of the differences with respect to conventional drug discovery, as well as the future potential of this approach.

Irreversible Protein Kinase Inhibitors: Balancing the Benefits and Risks

The invention features compounds of the general formula (I) in which the variable groups are as defined herein, and to their preparation and use.

Phosphorous derivatives as kinase inhibitors

We present a fast and effective covalent docking approach suitable for large-scale virtual screening (VS) We applied this method to four targets (HCV NS3 protease, Cathepsin K, EGFR, and XPO1) with known crystal structures and known covalent inhibitors We implemented a customized “VS mode” of the Schrodinger Covalent Docking algorithm (CovDock), which we refer to as CovDock-VS Known actives and target-specific sets of decoys were docked to selected X-ray structures, and poses were filtered based on noncovalent protein–ligand interactions known to be important for activity We were able to retrieve 71%, 72%, and 77% of the known actives for Cathepsin K, HCV NS3 protease, and EGFR within 5% of the decoy library, respectively With the more challenging XPO1 target, where no specific interactions with the protein could be used for postprocessing of the docking results, we were able to retrieve 95% of the actives within 30% of the decoy library and achieved an early enrichment factor (EF1%) of 33 The poses

Structure-Based Virtual Screening Approach for Discovery of Covalently Bound Ligands

Covalent linkage formation is a very important mechanism for many covalent drugs to work. However, partly due to the limitations of proper computational tools for covalent docking, most covalent drugs are not discovered systematically. In this article, we present a new covalent docking package, the CovalentDock, built on the top of the source code of Autodock. We developed an empirical model of free energy change estimation for covalent linkage formation, which is compatible with existing scoring functions used in docking, while handling the molecular geometry constrains of the covalent linkage with special atom types and directional grid maps. Integrated preparation scripts are also written for the automation of the whole covalent docking workflow. The result tested on existing crystal structures with covalent linkage shows that CovalentDock can reproduce the native covalent complexes with significant improved accuracy when compared with the default covalent docking method in Autodock. Experiments also suggest that CovalentDock is capable of covalent virtual screening with satisfactory enrichment performance. In addition, the investigation on the results also shows that the chirality and target selectivity along with the molecular geometry constrains are well preserved by CovalentDock, showing great capability of this method in the application for covalent drug discovery.

CovalentDock: Automated covalent docking with parameterized covalent linkage energy estimation and molecular geometry constraints

The epidermal growth factor receptor (EGFR) has an essential role in multiple signaling pathways, including cell proliferation and migration, through extracellular ligand binding and subsequent activation of its intracellular tyrosine kinase (TK) domain. The non-small cell lung cancer (NSCLC)-associated EGFR mutants, L858R and G719S, are constitutively active and oncogenic. They display sensitivity to TK inhibitors, including gefitinib and erlotinib. In contrast, the secondary mutation of the gatekeeper residue, T790M, reportedly confers inhibitor resistance on the oncogenic EGFR mutants. In this study, our biochemical analyses revealed that the introduction of the T790M mutation confers gefitinib resistance on the G719S mutant. The G719S/T790M double mutant has enhanced activity and retains high gefitinib-binding affinity. The T790M mutation increases the ATP affinity of the G719S mutant, explaining the acquired drug resistance of the double mutant. Structural analyses of the G719S/T790M double mutant, as well as the wild type and the G719S and L858R mutants, revealed that the T790M mutation stabilizes the hydrophobic spine of the active EGFR-TK conformation. The Met790 side chain of the G719S/T790M double mutant, in the apo form and gefitinib- and AMPPNP-bound forms, adopts different conformations that explain the accommodation of these ligands. In the L858R mutant structure, the active-site cleft is expanded by the repositioning of Phe723 within the P-loop. Notably, the introduction of the F723A mutation greatly enhanced the gefitinib sensitivity of the wild-type EGFR in vivo, supporting our hypothesis that the expansion of the active-site cleft results in enhanced gefitinib sensitivity. Taken together, our results provide a structural basis for the altered drug sensitivities caused by distinct NSCLC-associated EGFR mutations.

Structural basis for the altered drug sensitivities of non-small cell lung cancer-associated mutants of human epidermal growth factor receptor.

HTS hit 7 was modified through hybrid design strategy to introduce a chiral side chain followed by introduction of Michael acceptor group to obtain potent EGFR kinase inhibitors 11 and 19. Both 11 and 19 showed over 3 orders of magnitude enhanced HCC827 antiproliferative activity compared to HTS hit 7 and also inhibited gefitinib-resistant double mutant (DM, T790M/L858R) EGFR kinase at nanomolar concentration. Moreover, treatment with 19 shrinked tumor in nude mice xenograft model.

https://ir.nctu.edu.tw:443/bitstream/11536/32051/1/000283106300005.pdf

Design and synthesis of tetrahydropyridothieno[2,3-d]pyrimidine scaffold based epidermal growth factor receptor (EGFR) kinase inhibitors: the role of side chain chirality and Michael acceptor group for maximal potency.

Simple Summary The risk of lung cancer occurrence among patients with chronic kidney disease (CKD) is significantly higher than patients with normal renal function. Moreover, the long-term outcomes of CKD patients with lung cancer are poor. We conducted the present nationwide population based study to assess the association between the risk of lung cancer and influenza vaccination. The results showed that among CKD patients, potentially risk reduction of lung cancer was observed after influenza vaccination. In addition, the potentially protective effect of influenza vaccination on lung cancer risk reduction appeared to be dose dependent. The findings of present study provided the additional effect of influenza vaccination among patients with CKD. Abstract Chronic kidney disease (CKD) is significantly associated with lung cancer incidence. The aim of this study was to elucidate whether influenza vaccination reduces the incidence of lung cancer in patients with CKD. This cohort study enrolled patients with a record of CKD diagnosis from 2000 to 2012 in Taiwan’s National Health Insurance Research Database. Included patients were divided into vaccinated and unvaccinated groups. In total 12,985 patients with CKD were enrolled. Among these patients, 5495 were vaccinated and 7490 were unvaccinated. The risk of lung cancer was significantly lower in the influenza vaccination group after adjusting for age, sex, dialysis status, lung diseases, comorbidities, level of urbanization, and monthly income (adjusted hazard ratio (HR): 0.50, 95% confidence interval (CI; 0.38–0.65), p < 0.05). Lower risk of lung cancer was observed in both sexes, all age groups, dialysis status and co-existed lung diseases. The association between the risk of lung cancer and vaccination appeared to be dose-dependent (adjusted HRs: 0.91 (0.66–1.25), 0.49 (0.34–0.71), and 0.25 (0.17–0.38) for patients who received 1, 2 or 3, and ≥4 vaccinations during the follow-up period, respectively). In conclusion, Influenza vaccination decreased the risk of lung cancer in patients diagnosed with CKD. This potentially protective effect against lung cancer appeared to be dose dependent.

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Chia Hsien Wu

Papers

Design and synthesis of tetrahydropyridothieno[2,3-d]pyrimidine scaffold based epidermal growth factor receptor (EGFR) kinase inhibitors: the role of side chain chirality and Michael acceptor group for maximal potency.

Influenza Vaccination and Risk of Lung Cancer in Patients with Chronic Kidney Disease: A Nationwide, Population-Based Cohort Study