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Precedence and Promise of Covalent Inhibitors of EGFR and KRAS for Patients with Non-Small-Cell Lung Cancer Hengmiao Cheng* and Simon Planken
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Oncology Medicinal Chemistry, La Jolla Laboratories, Pfizer Worldwide Research and Development, 10770 Science Center Drive, San Diego, California 92121, United States ABSTRACT: Epidermal growth factor receptor (EGFR) and Kirsten rat sarcoma viral oncogene homolog (KRAS) oncogenic mutations are leading causes for lung cancer. Extensive drug discovery efforts targeting EGFR have led to the discovery and FDA approval of both reversible and covalent inhibitors. Second and third generation covalent inhibitors for EGFR have also been described, with the latter targeting specific emerging mutations. After decades of extensive effort, KRAS is widely regarded as an intractable therapeutic target; however, recent publications suggest covalent inhibition is a promising strategy to deliver inhibitors of the KRASG12C mutation.
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FDA approval of both second generation and third generation covalent drugs (Figure 1). Afatinib and dacomitinib, representatives of the second generation covalent EGFR inhibitors, demonstrate increased potency against EGFR oncogenic variants compared to the first generation drugs. These compounds also have activity against all three kinase-active members of the ErbB family (EGFR/ HER1, HER2, and HER4). In July 2013, afatinib was approved by theFDA for patients with late stage (metastatic) NSCLC; in January 2018, it was approved for the first line treatment of patients with metastatic NSCLC whose tumors have nonresistant EGFR mutations such as L861Q, G719X, and S768I.4 In April 2018, dacomitinib was granted priority review for the first-line treatment of patients with locally advanced or metastatic NSCLC with EGFR activating mutations.5 Osimertinib is the first approved third-generation covalent EGFR inhibitor. Developed to target the resistant gatekeeper T790M mutation, it also demonstrates potent activities against EGFR oncogenic mutants (L858R, Del) and double mutants (L858R/T790M, Del/T790M), with minimal activity against wild-type EGFR. In 2015, osimertinib received accelerated approval for the treatment of patients with metastatic EGFR T790M mutation positive NSCLC. Full approval for the same indication was granted in March 2017. In April 2018, osimertinib received approval as a first line treatment for EGFR-mutated NSCLC.6 The knowledge and understanding of covalent drug discovery and development acquired from the industry-wide effort to develop third generation EGFR inhibitors can be applied to future programs where a covalent strategy can be employed.
ung cancer is the leading cause of cancer death for both men and women, and non-small-cell lung cancer (NSCLC) accounts for 85% of all lung cancers. Out of the identified genetic causes for NSCLC, mutations of epidermal growth factor receptor (EGFR) and Kirsten rat sarcoma viral oncogene homolog (KRAS) are two of the most prevalent oncogenic drivers.1 EGFR L858R and EGFR Del (exon 19 deletions between amino acids 746 and 750) are two frequent and mutually exclusive primary oncogenic mutations, present in approximately 85% of all mutant EGFR NSCLC cases.2 Patients with these genotypes are treated with first generation EGFR tyrosine kinase inhibitors (gefitinib or erlotinib, Figure 1) which elicit excellent response rates and disease control for 11−14 months. Invariably, responsive patients develop resistance to these therapies, approximately 60% of which are driven by a second-site EGFR kinase domain mutation (gatekeeper residue, T790M), which restores constitutive EGFR-dependent signaling. The resistance mechanism derived from T790M mutation comes from increased ATP binding affinity for T790M mutants, steric clash between the Met790 gatekeeper side chain and the aniline moiety that is utilized by the first generation EGFR drugs, and an altered catalytic domain conformation. To increase the duration of response of EGFR TKIs and to address the resistance arising from the gatekeeper mutation, new treatment options were needed. Covalent inhibitors can achieve complete and sustained target engagement in the presence of high intracellular concentrations of the competitive ligand (ATP) and, more importantly, overcome the increased ATP binding affinity of the emerging mutants. Cys797 in the ATP binding site of EGFR presented an opportunity to pursue a covalent inhibition strategy.3 Extensive efforts were devoted to developing covalent EGFR inhibitors, cumulating in the © XXXX American Chemical Society
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DOI: 10.1021/acsmedchemlett.8b00311 ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX
ACS Medicinal Chemistry Letters
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of GTP.8 Oncogenic KRAS mutations interfere with KRAS GTPase activity, shifting the equilibrium in favor of GTPbound KRAS, which results in activation of downstream signaling pathways. The KRASG12C mutation is found in 15% lung adenocarcinoma. Pioneering work by Shokat and co-workers using tethering approach identified irreversible tool inhibitors that covalently modify Cys12 and bind in the pocket adjacent to SWII region in GDP bound form of KRASG12C. Covalent inhibition traps the inactive KRASG12C−GDP complex, blocking nucleotide exchange, and thereby inhibiting downstream signaling.9,10 ARS-1620 represents a recently described series of KRASG12C inhibitors that illustrate the potential of covalent inhibition targeting the Cys12 residue.11 Illustrated in Figure 2
Figure 2. Cocrystal structure of ARS-1620 in human KRASG12C.
is the cocrystal structure of ARS-1620 bound to KRASG12C. The structure reveals the inhibitor covalently modifying the target Cys12 residue, and the Switch II loop closed around the inhibitor, forming the inducible pocket. The nitrogen at the 2 position of the quinazoline core forms a hydrogen bond with His95 side chain. The fluorophenol binds in the hydrophobic SWII binding pocket that is formed by the side chains of Val9, Met72, I100, and Val103, and the phenol is oriented toward the SWII loop. This forms a hydrogen bond network with three residues from the SWII loop (Asp69 side chain carboxylate, Ser65 backbone carbonyl, and Glu63 backbone carbonyl) via recruitment of two water molecules and effectively “stitches” the inducible pocket together. Substitution of the core with halogens at the 6 and 8 positions restricts the conformation of the fluorophenol, resulting in the existence of two stable atropisomers. ARS-1620 is the S-atropisomer, being nearly 1000-fold more potent than the R-conformational atropisomer. A piperazine linker connects the quinazoline core and an acrylamide warhead. Critically, the carbonyl of the warhead forms an activating hydrogen bond with the side chain of Lys16 and positions the acrylamide perfectly to engage the thiol group of the target cysteine. In contrast to EGFR T790M inhibitors, the activity of ARS1620 is primarily driven by KRAS-mediated catalysis of the chemical reaction with Cys12, with kinact of >0.066 ± 0.019 s−1 and kinact/Ki of 1100 ± 300 M−1 s−1.12 The intrinsic binding affinity is weak with an estimated Ki > 64 μM.12 The covalent complex of ARS-1620 and KRASG12C cannot undergo nucleotide exchange with GTP, and thus, the protein remains trapped in its inactive state. ARS-1620 inhibits KRASG12C signaling in tumor cells with nanomolar to low-micromolar potency, and in vivo studies show tumor regressions in KRASG12C tumor cell lines and patient-derived mouse xenograft models.
Figure 1. Structures of EGFR inhibitors.
Activating mutations in KRAS (a small GTPase) are among the most common mutations found in human cancer. With unmet medical need for drugs to treat KRAS mutant driven NSCLC, development of inhibitors has been actively pursued in both academia and industry for decades. However, mutant KRAS has remained an intractable therapeutic target due to (a) the extremely high affinity (pM) of the natural ligand (GTP) and (b) the lack of clearly defined binding sites outside of the nucleotide-binding site for developing high affinity small molecule inhibitors for both GTP and GDP bound states.7 KRAS controls a complex cellular signaling cascade and is a driver of cellular proliferation and survival. Upon binding of the GTP nucleotide, two disordered protein regions (termed “switch I” and “switch II”) undergo conformational changes enabling Ras to interact with effector enzymes (via a Rasbinding domain (RBD)), initiating signaling cascades and altering gene expression. The KRAS GTP/GDP cycle is governed by GTPase activating proteins (GAPs) that accelerate the intrinsic conversion of GTP to GDP (rendering the protein in an inactive state, leading to the signal termination), and guanine nucleotide exchange factors (GEFs), which remove GDP from Ras and allow the entry B
DOI: 10.1021/acsmedchemlett.8b00311 ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX
ACS Medicinal Chemistry Letters
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ARS-1620 represents a first-in-class tool compound, which was used for preclinical validation of the covalent inhibition strategy to target KRASG12C and to provide early solid evidence of the druggability of this mutant form. It will be exciting to see progression of the first wave of KRASG12C inhibitors into clinical trials bringing cancer medicine to the specific subset of NSCLC patients that currently have limited options.
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oncogenic KRAS output with small molecules targeting the inactive state. Cancer Discovery 2016, 6, 316−329. (11) Janes, M. R.; Zhang, J.; Li, L.; Hansen, R.; Peters, U.; Guo, X.; Chen, Y.; Babbar, A.; Firdaus, S. J.; Darjania, L.; Feng, J.; Chen, J. H.; Li, S.; Li, S.; Long, Y. O.; Thach, C.; Liu, Y.; Zarieh, A.; Ely, T.; Kucharski, J. M.; Kessler, L. V.; Wu, T.; Yu, K.; Wang, Y.; Yao, Y.; Deng, X.; Zarrinkar, P.; Brehmer, D.; Dhanak, D.; Lorenzi, M. V.; HuLowe, D.; Patricelli, M. P.; Ren, P.; Liu, Y. Targeting KRAS mutant cancers with a covalent G12C-specific inhibitor. Cell 2018, 172, 578− 589. (12) Hansen, R.; Peters, U.; Babbar, A.; Chen, Y.; Feng, J.; Janes, M. R.; Li, L.; Ren, P.; Liu, Y.; Zarrinkar, P. P. The reactivity-driven biochemical mechanism of covalent KRASG12C inhibitors. Nat. Struct. Mol. Biol. 2018, 25, 454−462.
AUTHOR INFORMATION
Corresponding Author
*E-mail: henry.cheng@pfizer.com. ORCID
Hengmiao Cheng: 0000-0003-1761-4503 Simon Planken: 0000-0001-8901-8992 Notes
The authors declare the following competing financial interest(s): Both authors for this manuscript are employees and stockholders of Pfizer.
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ACKNOWLEDGMENTS We are grateful to Dr. Alexei Brooun for reading the manuscript and providing helpful comments.
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REFERENCES
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DOI: 10.1021/acsmedchemlett.8b00311 ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX
