Abstract
Background
Activating mutations in the epidermal growth factor receptor (EGFR), particularly exon 19 deletions and L858R mutation, are frequently observed in non-small cell lung cancer (NSCLC) and confer sensitivity to EGFR-tyrosine kinase inhibitors (EGFR-TKIs). Among these EGFR-TKIs, osimertinib is currently the standard of care for patients with NSCLC harboring the activating mutations. However, resistant mutations often arise, leading to resistance to osimertinib. The resistant mutation that most frequently occurs in EGFR during osimertinib treatment is the C797S mutation. Another major resistant mutation arising in EGFR during treatment with other EGFR-TKIs, such as gefitinib and afatinib, is the T790M mutation. Currently, no approved EGFR-TKIs are effective in patients who simultaneously develop the T790M and C797S mutations. Additionally, brain metastasis often causes disease progression due to reduced drug penetration into the brain.
Methods
We conducted preclinical evaluations of TAS3351, a fourth-generation EGFR-TKI, including biochemical, structural, and in vitro/in vivo pharmacological assays. We also evaluated the efflux transporter susceptibility and brain penetrability of TAS3351 in male mice.
Results
Here, we demonstrate that TAS3351 overcomes resistance due to T790M and C797S mutations while sparing wild-type EGFR activity. Furthermore, TAS3351 is not a substrate of P-glycoprotein (P-gp) and the breast cancer-resistant protein (BCRP) and exhibits significant brain penetrability, resulting in anti-tumor efficacy in mice with intracranial allografts.
Conclusions
These findings indicate that TAS3351 is a promising therapeutic candidate for patients with NSCLC whose tumors have relapsed or are refractory to treatment due to the C797S and T790M mutations, and the brain metastases.
Plain language summary
Changes in a protein called the epidermal growth factor receptor (EGFR) are known to play a crucial role in the development of non-small cell lung cancer (NSCLC). Many drugs target EGFR, and these are called EGFR-tyrosine kinase inhibitors. Whilst they are often effective as a treatment for NSCLC initially, often the cancers develop resistance, which means that they stop responding to the treatment. Also, cancer often occurs in the brain, because many cancer drugs cannot reach the brain due to a barrier in blood transfer to the brain. Here we describe a EGFR-tyrosine kinase inhibitor that works against some of the cancers resistant to other EGFR-tyrosine kinase inhibitors and also can reach the brain effectively. These data suggest our drug, called TAS3351, has potential as an improved treatment option for patients with NSCLC.
Similar content being viewed by others
Data availability
The crystallographic data generated in this study have been deposited in Research Collaboratory for Structural Bioinformatics Protein Data Bank (RCSB PDB) with the accession codes (9KL4 and 9KLW). The crystallographic data and refinement statistics are available in the Supplementary Information file as Supplementary Tables 1 and 2. The coordinate Crystallographic Information Files for 9KL4 and 9KLW are found in Supplementary Data 1 and 3, respectively. The structure-factor Crystallographic Information Files for 9KL4 and 9KLW are found in Supplementary Data 2 and 4, respectively. All numerical results underlying the graphs and table presented in this study are available in Supplementary Data 5–8. The uncropped immunoblotting data with molecular markers for Figs. 6a, b are available in the Supplementary Information file as Supplementary Fig. 6 and 7, respectively. All additional relevant data and information in this study are available from corresponding author upon request.
References
Lynch, T. J. et al. Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib. N. Engl. J. Med. 350, 2129–2139 (2004).
Paez, J. G. et al. EGFR mutations in lung cancer: correlation with clinical response to gefitinib therapy. Science 304, 1497–1500 (2004).
Pao, W. et al. EGF receptor gene mutations are common in lung cancers from ‘never smokers’ and are associated with sensitivity of tumors to gefitinib and erlotinib. Proc. Natl. Acad. Sci. USA 101, 13306–13311 (2004).
Shigematsu, H. et al. Clinical and biological features associated with epidermal growth factor receptor gene mutations in lung cancers. J. Natl. Cancer Inst. 97, 339–346 (2005).
Dearden, S., Stevens, J., Wu, Y. L. & Blowers, D. Mutation incidence and coincidence in non small-cell lung cancer: meta-analyses by ethnicity and histology (mutMap). Ann. Oncol. 24, 2371–2376 (2013).
Shi, Y. et al. A prospective, molecular epidemiology study of EGFR mutations in Asian patients with advanced non-small-cell lung cancer of adenocarcinoma histology (PIONEER). J. Thorac. Oncol. 9, 154–162 (2014).
Midha, A., Dearden, S. & McCormack, R. EGFR mutation incidence in non-small-cell lung cancer of adenocarcinoma histology: a systematic review and global map by ethnicity (MutMapII). Am. J. Cancer Res. 5, 2892–2911 (2015).
Maemondo, M. et al. Gefitinib or chemotherapy for non-small-cell lung cancer with mutated EGFR. N. Engl. J. Med. 362, 2380–2388 (2010).
Rosell, R. et al. Erlotinib versus standard chemotherapy as first-line treatment for European patients with advanced EGFR mutation-positive non-small-cell lung cancer (EURTAC): a multicentre, open-label, randomised phase 3 trial. Lancet Oncol. 13, 239–246 (2012).
Park, K. et al. Afatinib versus gefitinib as first-line treatment of patients with EGFR mutation-positive non-small-cell lung cancer (LUX-Lung 7): a phase 2B, open-label, randomised controlled trial. Lancet Oncol. 17, 577–589 (2016).
Wu, Y.-L. et al. Dacomitinib versus gefitinib as first-line treatment for patients with EGFR-mutation-positive non-small-cell lung cancer (ARCHER 1050): a randomised, open-label, phase 3 trial. Lancet Oncol. 18, 1454–1466 (2017).
Ramalingam, S. S. et al. Overall survival with osimertinib in untreated, EGFR-mutated advanced NSCLC. N. Engl. J. Med. 382, 41–50 (2020).
Sequist, L. V. et al. Genotypic and histological evolution of lung cancers acquiring resistance to EGFR inhibitors. Sci. Transl. Med. 3, 75ra26 (2011).
Kobayashi, S. et al. EGFR mutation and resistance of non–small-cell lung cancer to gefitinib. N. Engl. J. Med. 352, 786–792 (2005).
Yun, C.-H. et al. The T790M mutation in EGFR kinase causes drug resistance by increasing the affinity for ATP. Proc. Natl. Acad. Sci. USA 105, 2070–2075 (2008).
Cross, D. A. E. et al. AZD9291, an irreversible EGFR TKI, overcomes T790M-mediated resistance to EGFR inhibitors in lung cancer. Cancer Discov. 4, 1046–1061 (2014).
Jänne, P. A. et al. AZD9291 in EGFR inhibitor-resistant non-small-cell lung cancer. N. Engl. J. Med. 372, 1689–1699 (2015).
Soria, J.-C. et al. Osimertinib in untreated EGFR-mutated advanced non-small-cell lung cancer. N. Engl. J. Med. 378, 113–125 (2018).
Thress, K. S. et al. Acquired EGFR C797S mutation mediates resistance to AZD9291 in non–small cell lung cancer harboring EGFR T790M. Nat. Med. 21, 560–562 (2015).
Oxnard, G. R. et al. Assessment of resistance mechanisms and clinical implications in patients with EGFR T790M–positive lung cancer and acquired resistance to osimertinib. JAMA Oncol. 4, 1527 (2018).
Schmid, S., Li, J. J. N. & Leighl, N. B. Mechanisms of osimertinib resistance and emerging treatment options. Lung Cancer 147, 123–129 (2020).
Uchibori, K. et al. Brigatinib combined with anti-EGFR antibody overcomes osimertinib resistance in EGFR-mutated non-small-cell lung cancer. Nat. Commun. 8, 14768 (2017).
Yang, Z. et al. Investigating novel resistance mechanisms to third-generation EGFR tyrosine kinase inhibitor osimertinib in non-small cell lung cancer patients. Clin. Cancer Res. 24, 3097–3107 (2018).
Jia, Y. et al. Overcoming EGFR(T790M) and EGFR(C797S) resistance with mutant-selective allosteric inhibitors. Nature 534, 129–132 (2016).
To, C. et al. Single and dual targeting of mutant EGFR with an allosteric inhibitor. Cancer Discov. 9, 926–943 (2019).
To, C. et al. An allosteric inhibitor against the therapy-resistant mutant forms of EGFR in non-small cell lung cancer. Nat. Cancer 3, 402–417 (2022).
Izumi, H. et al. 997P Phase I study of brigatinib plus panitumumab in patients with advanced EGFR-mutated non-small cell lung cancer resistant to osimertinib (BEBOP): early termination due to severe early onset pneumonitis by brigatinib. Ann. Oncol. 33, S1009–S1010 (2022).
Eno, M. S. et al. Discovery of BLU-945, a reversible, potent, and wild-type-sparing next-generation EGFR mutant inhibitor for treatment-resistant non-small-cell lung cancer. J. Med. Chem. 65, 9662–9677 (2022).
Elamin, Y. Y. et al. BLU-945 monotherapy and in combination with osimertinib (OSI) in previously treated patients with advanced EGFR -mutant (EGFRm) NSCLC in the phase 1/2 SYMPHONY study. J. Clin. Oncol. 41, 9011–9011 (2023).
Nayak, L., Lee, E. Q. & Wen, P. Y. Epidemiology of brain metastases. Curr. Oncol. Rep. 14, 48–54 (2012).
Planchard, D. et al. Osimertinib with or without chemotherapy in EGFR-mutated advanced NSCLC. N. Engl. J. Med. 389, 1935–1948 (2023).
Ballard, P. et al. Preclinical comparison of osimertinib with other EGFR-TKIs in EGFR-mutant NSCLC brain metastases models, and early evidence of clinical brain metastases activity. Clin. Cancer Res. 22, 5130–5140 (2016).
Ito, K. et al. TAS-121, a selective mutant EGFR inhibitor, shows activity against tumors expressing various EGFR mutations, including T790M and uncommon mutations G719X. Mol. Cancer Ther. 18, 920–928 (2019).
Hasako, S. et al. TAS6417, a novel EGFR inhibitor targeting exon 20 insertion mutations. Mol. Cancer Ther. 17, 1648–1658 (2018).
Yung-Chi, C. & Prusoff, W. H. Relationship between the inhibition constant (KI) and the concentration of inhibitor which causes 50 per cent inhibition (I50) of an enzymatic reaction. Biochem. Pharmacol. 22, 3099–3108 (1973).
Zhai, X., Ward, R. A., Doig, P. & Argyrou, A. Insight into the therapeutic selectivity of the irreversible EGFR tyrosine kinase inhibitor osimertinib through enzyme kinetic studies. Biochemistry 59, 1428–1441 (2020).
Schwartz, P. A. et al. Covalent EGFR inhibitor analysis reveals importance of reversible interactions to potency and mechanisms of drug resistance. Proc. Natl. Acad. Sci. USA 111, 173–178 (2014).
Bell, D. W. et al. Inherited susceptibility to lung cancer may be associated with the T790M drug resistance mutation in EGFR. Nat. Genet. 37, 1315–1316 (2005).
Yu, H. A. et al. Germline EGFR T790M mutation found in multiple members of a familial cohort. J. Thorac. Oncol. 9, 554–558 (2014).
Morrison, J. F. Kinetics of the reversible inhibition of enzyme-catalysed reactions by tight-binding inhibitors. Biochim. Biophys. Acta. 185, 269–286 (1969).
Kuzmic, P., Hill, C., Kirtley, M. P. & Janc, J. W. Kinetic determination of tight-binding impurities in enzyme inhibitors. Anal. Biochem. 319, 272–279 (2003).
Murphy, D. J. Determination of accurate KI values for tight-binding enzyme inhibitors: an in silico study of experimental error and assay design. Anal. Biochem. 327, 61–67 (2004).
Moyer, J. D. et al. Induction of apoptosis and cell cycle arrest by CP-358,774, an inhibitor of epidermal growth factor receptor tyrosine kinase. Cancer Res. 57, 4838–4848 (1997).
Aguilar, H. N., Zielnik, B., Tracey, C. N. & Mitchell, B. F. Quantification of rapid myosin regulatory light chain phosphorylation using high-throughput in-cell western assays: comparison to western immunoblots. PLoS ONE 5, e9965 (2010).
Li, D. et al. BIBW2992, an irreversible EGFR/HER2 inhibitor highly effective in preclinical lung cancer models. Oncogene 27, 4702–4711 (2008).
Engelman, J. A. et al. PF00299804, an irreversible pan-ERBB inhibitor, is effective in lung cancer models with EGFR and ERBB2 mutations that are resistant to gefitinib. Cancer Res. 67, 11924–11932 (2007).
Ellis, P. M. et al. Dacomitinib compared with placebo in pretreated patients with advanced or metastatic non-small-cell lung cancer (NCIC CTG BR.26): a double-blind, randomised, phase 3 trial. Lancet Oncol. 15, 1379–1388 (2014).
Miller, V. A. et al. Afatinib versus placebo for patients with advanced, metastatic non-small-cell lung cancer after failure of erlotinib, gefitinib, or both, and one or two lines of chemotherapy (LUX-Lung 1): a phase 2b/3 randomised trial. Lancet Oncol. 13, 528–538 (2012).
Jiang, J. et al. Epidermal growth factor–independent transformation of Ba/F3 cells with cancer-derived epidermal growth factor receptor mutants induces gefitinib-sensitive cell cycle progression. Cancer Res. 65, 8968–8974 (2005).
Godin-Heymann, N. et al. Oncogenic activity of epidermal growth factor receptor kinase mutant alleles is enhanced by the T790M drug resistance mutation. Cancer Res. 67, 7319–7326 (2007).
Merlino, G. T. et al. Amplification and enhanced expression of the epidermal growth factor receptor gene in A431 human carcinoma cells. Science 224, 417–419 (1984).
Pardridge, W. M. Drug transport across the blood-brain barrier. J. Cereb. Blood Flow Metab. 32, 1959–1972 (2012).
Brouwer, K. L. R. et al. In vitro methods to support transporter evaluation in drug discovery and development. Clin. Pharmacol. Ther. 94, 95–112 (2013).
International Transporter Consortium Giacomini, K. M. et al. Membrane transporters in drug development. Nat. Rev. Drug Discov 9, 215–236 (2010).
Loryan, I. et al. Unbound brain-to-plasma partition coefficient, Kp,uu,brain-a game changing parameter for CNS drug discovery and development. Pharm. Res. 39, 1321–1341 (2022).
Zou, L. et al. Considerations in Kp,uu,brain-based strategy for selecting CNS-targeted drug candidates with sufficient target coverage and substantial pharmacodynamic effect. AAPS J. 27, 52 (2025).
Wu, Y.-L. et al. Osimertinib in resected EGFR-mutated non-small-cell lung cancer. N. Engl. J. Med. 383, 1711–1723 (2020).
Chmielecki, J. et al. Candidate mechanisms of acquired resistance to first-line osimertinib in EGFR-mutated advanced non-small cell lung cancer. Nat. Commun. 14, 1070 (2023).
Osoegawa, A. et al. High incidence of C797S mutation in patients with long treatment history of EGFR tyrosine kinase inhibitors including osimertinib. JTO Clin. Res. Rep. 2, 100191 (2021).
Starrett, J. H. et al. Drug sensitivity and allele specificity of first-line osimertinib resistance EGFR mutations. Cancer Res. 80, 2017–2030 (2020).
Jänne, P. A. et al. CNS efficacy of osimertinib with or without chemotherapy in epidermal growth factor receptor-mutated advanced non-small-cell lung cancer. J. Clin. Oncol. 42, 808–820 (2024).
Yun, J. et al. Antitumor activity of amivantamab (JNJ-61186372), an EGFR–MET bispecific antibody, in diverse models of EGFR exon 20 insertion–driven NSCLC. Cancer Discov. 10, 1194–1209 (2020).
Cho, B. C. et al. A phase 1/2 study of lazertinib 240 mg in patients with advanced EGFR T790M-positive NSCLC after previous EGFR tyrosine kinase inhibitors. J. Thorac. Oncol. 17, 558–567 (2022).
Ahn, M.-J. et al. Lazertinib in patients with EGFR mutation-positive advanced non-small-cell lung cancer: results from the dose escalation and dose expansion parts of a first-in-human, open-label, multicentre, phase 1-2 study. Lancet Oncol. 20, 1681–1690 (2019).
Cho, B. C. et al. Amivantamab plus lazertinib in previously untreated EGFR-mutated advanced NSCLC. N. Engl. J. Med. 391, 1486–1498 (2024).
Park, S. et al. EGFR C797S as a resistance mechanism of lazertinib in non-small cell lung cancer with EGFR T790M mutation. Cancer Res. Treat. 52, 1288–1290 (2020).
Jänne, P. A. et al. Efficacy and safety of patritumab deruxtecan (HER3-DXd) in EGFR inhibitor–resistant, EGFR -mutated non–small cell lung cancer. Cancer Discov. 12, 74–89 (2022).
Colclough, N. et al. Preclinical comparison of the blood-brain barrier permeability of osimertinib with other EGFR TKIs. Clin. Cancer Res. 27, 189–201 (2021).
Dardenne, E. et al. Abstract 1229: BDTX-1535: a MasterKey EGFR inhibitor targeting classical and non-classical oncogenic driver mutations and the C797S acquired resistance mutation to address the evolving molecular landscape of EGFR mutant NSCLC. Cancer Res. 84, 1229–1229 (2024).
Acknowledgements
The authors thank Drs. Teruhiro Utsugi, Takeshi Sagara, Yoshikazu Iwasawa, Kazuhiko Yonekura, and Kazutaka Miyadera for mentoring this work. We would also like to thank all those who contributed to this work at the Discovery and Preclinical Research Division of Taiho Pharmaceutical Co., Ltd. We also thank the MD Anderson Cancer Center for their invaluable support in conducting this research. PC-9 and Ba/F3 cells were provided by the RIKEN BRC through the National Bio-Resource Project of the MEXT, Japan. This study was funded by Taiho Pharmaceutical Co., Ltd., a wholly owned subsidiary of Otsuka Holdings Co., Ltd.
Author information
Authors and Affiliations
Contributions
H.K., F.Y., and S.M. conceived, designed, and supervised the entire experiment. H.K., F.Y., Y.K., F.Y., S.T., R.M., and T.S. implemented experiments, acquired data, and all authors participated in data interpretation. H.K. wrote the manuscript, and all authors reviewed the manuscript.
Corresponding author
Ethics declarations
Competing interests
All authors are employees of Taiho Pharmaceutical Co., Ltd., a wholly owned subsidiary of Otsuka Holdings Co., Ltd.
Peer review
Peer review information
Communications Medicine thanks Yi-Chen Zhang and Kimio Yonesaka for their contribution to the peer review of this work. A peer review file is available.
Additional information
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary information
Rights and permissions
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.
About this article
Cite this article
Kasuga, H., Kataoka, Y., Yamamoto, F. et al. TAS3351 is a brain penetrable EGFR-TKI that overcomes T790M and C797S resistant mutations. Commun Med (2026). https://doi.org/10.1038/s43856-026-01546-1
Received:
Accepted:
Published:
DOI: https://doi.org/10.1038/s43856-026-01546-1
