Major Clinical Response to Afatinib Monotherapy in Lung Adenocarcinoma Harboring EGFR Exon 20 Insertion Mutation

Clinical Practice Points•Exon 20 insertions consist of up to 10% of epidermal growth factor receptor (EGFR) mutations detected in cases of non–small-cell lung cancer. They are considered as a hard-to-treat group of EGFR mutations, where tyrosine kinase inhibitor (TKI) resistance is common. Yet several lines of evidence suggest that exon 20 insertions are a biochemically heterogenous group, and their response to different types of TKIs depends on the individual mutations.•We report the case of a patient with lung adenocarcinoma with a relatively common insertion at EGFR-H773 with a lack of response to the first-generation TKI, gefitinib, which is in striking contrast to the major benefit experienced by afatinib treatment, in agreement with preclinical data.•Considering the underlying biochemistry, treatment of cases with non–small-cell lung cancer with EGFR exon 20 insertions should be considered individually, based on preceding evidence.•Targeted therapy using second-generation EGFR inhibitors is a promising approach for personalized treatment of patients with cancer driven by insertions near H773. •Exon 20 insertions consist of up to 10% of epidermal growth factor receptor (EGFR) mutations detected in cases of non–small-cell lung cancer. They are considered as a hard-to-treat group of EGFR mutations, where tyrosine kinase inhibitor (TKI) resistance is common. Yet several lines of evidence suggest that exon 20 insertions are a biochemically heterogenous group, and their response to different types of TKIs depends on the individual mutations.•We report the case of a patient with lung adenocarcinoma with a relatively common insertion at EGFR-H773 with a lack of response to the first-generation TKI, gefitinib, which is in striking contrast to the major benefit experienced by afatinib treatment, in agreement with preclinical data.•Considering the underlying biochemistry, treatment of cases with non–small-cell lung cancer with EGFR exon 20 insertions should be considered individually, based on preceding evidence.•Targeted therapy using second-generation EGFR inhibitors is a promising approach for personalized treatment of patients with cancer driven by insertions near H773. The essential aim of continuous drug discovery in precision oncology is to develop new generations of targeted therapies, which overcome resistance owing to drug-resistant driver mutations. Therefore, new generation targeted therapies are typically statistically superior in comparative clinical studies and become the new standard of care. However, here we show an example that the choice of the right targeted therapy should be made at the individual mutation level where previous generation inhibitors based on biochemical and preclinical evidence can be the right clinical decision. Most activating mutations of epidermal growth factor receptor (EGFR) in non–small-cell lung cancers (NSCLCs) are either exon 19 deletions or L858R point mutation, which are sensitive to available EGFR tyrosine kinase inhibitors (TKIs).1Hirsch F.R. Scagliotti G.V. Mulshine J.L. et al.Lung cancer: current therapies and new targeted treatments.Lancet. 2017; 389: 299-311Abstract Full Text Full Text PDF PubMed Scopus (1306) Google Scholar A subset, approximately 4% to 10% of EGFR-mutant NSCLC tumors, harbor an in-frame insertion in exon 20 of EGFR.2Vyse S. Huang P.H. Targeting EGFR exon 20 insertion mutations in non-small cell lung cancer.Signal Transduct Target Ther. 2019; 4: 8Crossref PubMed Scopus (125) Google Scholar Activating exon 20 insertions mostly occur either in the C-terminal part of the C-helix of the protein kinase domain or in the directly following loop region. Structural distortions caused by either deletions in exon 19 (upstream of C-helix) or insertions in exon 20 force the C-helix inwards the N-lobe of the kinase domain to stabilize the active state of EGFR even in the absence of ligand. Nevertheless, there is a marked difference in sensitivity to available TKIs between exon 19 and 20 mutants. Yet, although exon 20 insertions are generally considered to be resistant to TKIs, several lines of evidence indicate differential responses of the various insertion mutations to different inhibitors. For example, whereas first-generation EGFR inhibitors are ineffective in the vast majority of patients with exon 20 insertion mutant NSCLC,3Naidoo J. Sima C.S. Rodriguez K. et al.Epidermal growth factor receptor exon 20 insertions in advanced lung adenocarcinomas: clinical outcomes and response to erlotinib.Cancer. 2015; 121: 3212-3220Crossref PubMed Scopus (121) Google Scholar,4Beau-Faller M. Prim N. Ruppert A.M. et al.Rare EGFR exon 18 and exon 20 mutations in non-small-cell lung cancer on 10 117 patients: a multicentre observational study by the French ERMETIC-IFCT network.Ann Oncol. 2014; 25: 126-131Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar they were effective in patients carrying the A763_Y764insFQEA mutation (within the C-helix).5Arcila M.E. Nafa K. Chaft J.E. et al.EGFR exon 20 insertion mutations in lung adenocarcinomas: prevalence, molecular heterogeneity, and clinicopathologic characteristics.Mol Cancer Ther. 2013; 12: 220-229Crossref PubMed Scopus (263) Google Scholar,6Voon P.J. Tsui D.W.Y. Rosenfeld N. Chin T.M. EGFR exon 20 insertion A763-Y764insFQEA and response to erlotinib—letter.Mol Cancer Ther. 2013; 12: 2614-2615Crossref PubMed Google Scholar Similarly, preclinical models identified differential responses to second-generation inhibitors across distinct exon 20 insertions.7Kosaka T. Tanizaki J. Paranal R.M. et al.Response heterogeneity of EGFR and HER2 exon 20 insertions to covalent EGFR and HER2 inhibitors.Cancer Res. 2017; 77: 2712-2721Crossref PubMed Scopus (85) Google Scholar,8Robichaux J.P. Elamin Y.Y. Tan Z. et al.Mechanisms and clinical activity of an EGFR and HER2 exon 20-selective kinase inhibitor in non-small cell lung cancer.Nat Med. 2018; 24: 638-646Crossref PubMed Scopus (214) Google Scholar A patient-derived xenograft model, harboring EGFR-H773_V774insNPH mutation (following the C-helix), showed poor responses to the third-generation EGFR inhibitors, osimertinib and rociletinib.9Yang M. Xu X. Cai J. Ning J. Wery J.P. Li Q.X. NSCLC harboring EGFR exon-20 insertions after the regulatory C-helix of kinase domain responds poorly to known EGFR inhibitors.Int J Cancer. 2016; 139: 171-176Crossref PubMed Scopus (36) Google Scholar In contrast, preclinical models harboring this mutation indicate favorable responses to the second-generation TKI, afatinib.8Robichaux J.P. Elamin Y.Y. Tan Z. et al.Mechanisms and clinical activity of an EGFR and HER2 exon 20-selective kinase inhibitor in non-small cell lung cancer.Nat Med. 2018; 24: 638-646Crossref PubMed Scopus (214) Google Scholar,9Yang M. Xu X. Cai J. Ning J. Wery J.P. Li Q.X. NSCLC harboring EGFR exon-20 insertions after the regulatory C-helix of kinase domain responds poorly to known EGFR inhibitors.Int J Cancer. 2016; 139: 171-176Crossref PubMed Scopus (36) Google Scholar Therefore, charting the clinical utility of different TKIs in the hard-to-treat group of exon 20 mutants is of primary importance for facilitating precision oncology in NSCLC. Here, we present the case of a patient with exon 20 insertion carrier NSCLC with strikingly differential responses to first- and second-generation TKI treatments (Figure 1A). In June 2016, a 61-year-old male never-smoker had an abnormality discovered on a routine x-ray examination of the chest. Computed tomography (CT) scan set up the preliminary diagnosis of pulmonary tumor in the superior lobe of the left lung with metastases in the right upper lobe and in the left lung (Stage IVa, T2N0M1a) (Figure 1B). In June, tumor mass reductive surgery was performed, although complete excision was not possible. His diagnosis was well-differentiated lung adenocarcinoma. Positron emission tomography (PET) scan showed a residual suspicious lesion in the right upper lobe (80 × 34 mm); fine needle aspiration cytology results confirmed metastasis of the excised adenocarcinoma. The presence of an exon 20 insertion, EGFR-H773_V774insNPH (also referred to as: EGFR-N771_H773dup; genotype c.2319_2320insAACCCCCAC) was detected in the histologic sample by Sanger sequencing. No further alterations were detected either in hotspot regions of 50 genes by next-generation sequencing or in the ALK and MET genes by fluorescence in situ hybridization. Therefore, first-line gefitinib therapy was started. PET-CT in September 2016 showed progressive disease (Figure 1B), but the presence of the resistance mutation EGFR-T790M was not detected by circulating tumor DNA analysis. A reduction in performance status could be observed (Eastern Cooperative Oncology Group [ECOG] performance status 1). The patient was taken off of gefitinib, and paclitaxel + carboplatin + bevacizumab therapy was started, because an intrinsic resistance to EGFR inhibition was suspected. A week later, he developed pulmonary complications and was hospitalized with a life-threatening condition requiring intensive care. Symptoms included high temperature (39.5°C), wheezing, and dyspnea. Radiographic imaging showed massive infiltration in both lobes, indicating inflammatory reaction (Figure 1C). Antibiotic and steroid therapy was applied, and the patient gradually recovered. After his health status was stabilized, afatinib therapy was started. Subsequently, the patient regained fitness (ECOG performance status 0) and demonstrated partial response (50 × 20 mm; −37.5%) lasting for over 3 years now (Figure 1B). He is still on afatinib therapy, having no side effects, with strict monitoring by PET-CT and circulating tumor DNA tracking. TKIs have proven dramatic clinical benefit in patients with NSCLC harboring ‘classical’ activating mutations in EGFR. Exon 20 insertion mutations, on the other hand, are generally considered as poor responders. Nevertheless, emerging evidence highlights marked differences in the efficacy of different types of TKIs against different specific exon 20 insertion mutations, implying biochemical heterogeneity of this mutation type that may require individual evaluation in precision oncology. ‘First-generation’ EGFR inhibitors, such as gefitinib and erlotinib, are reversible, ATP-competitive compounds, whereas second-generation inhibitors, such as afatinib and dacomitinib, form irreversible, covalent complexes with EGFR at C797. Third-generation EGFR inhibitors, such as osimertinib and rociletinib, also bind covalently to C797 and in addition maintain activity for the secondary mutation T790M. ‘Classical’ mutations induce conformational changes that cause an equilibrium shift towards the active state.10Eck M.J. Yun C.H. Structural and mechanistic underpinnings of the differential drug sensitivity of EGFR mutations in non-small cell lung cancer.Biochim Biophys Acta. 2010; 1804: 559-566Crossref PubMed Scopus (138) Google Scholar, 11Yun C.H. Boggon T.J. Li Y. et al.Structures of lung cancer-derived EGFR mutants and inhibitor complexes: mechanism of activation and insights into differential inhibitor sensitivity.Cancer Cell. 2007; 11: 217-227Abstract Full Text Full Text PDF PubMed Scopus (764) Google Scholar, 12Landau M. Ben-Tal N. Dynamic equilibrium between multiple active and inactive conformations explains regulation and oncogenic mutations in ErbB receptors.Biochim Biophys Acta. 2008; 1785: 12-31PubMed Google Scholar Moreover, these mutations reduce the ATP-binding affinity of EGFR, thereby alleviating competitive pressure and enhancing relative affinity to reversible ATP-competitive (first-generation) inhibitors.13Carey K.D. Garton A.J. Romero M.S. et al.Kinetic analysis of epidermal growth factor receptor somatic mutant proteins shows increased sensitivity to the epidermal growth factor receptor tyrosine kinase inhibitor, erlotinib.Cancer Res. 2006; 66: 8163-8171Crossref PubMed Scopus (324) Google Scholar,14Mulloy R. Ferrand A. Kim Y. et al.Epidermal growth factor receptor mutants from human lung cancers exhibit enhanced catalytic activity and increased sensitivity to gefitinib.Cancer Res. 2007; 67: 2325-2330Crossref PubMed Scopus (147) Google Scholar In contrast, the exon 20 insertion, D770_N771insNPG, activates EGFR without diminishing ATP affinity,15Yasuda H. Park E. Yun C.H. et al.Structural, biochemical, and clinical characterization of epidermal growth factor receptor (EGFR) exon 20 insertion mutations in lung cancer.Sci Transl Med. 2013; 5: 216ra177Crossref PubMed Scopus (324) Google Scholar and it also pushes the C-helix and the phosphate-binding loop (P-loop) into the drug-binding pocket of the kinase8Robichaux J.P. Elamin Y.Y. Tan Z. et al.Mechanisms and clinical activity of an EGFR and HER2 exon 20-selective kinase inhibitor in non-small cell lung cancer.Nat Med. 2018; 24: 638-646Crossref PubMed Scopus (214) Google Scholar, thus abolishing sensitivity. Moreover, 3-dimensional modeling revealed that the large terminal group of osimertinib reduces its ability to reach C797 in EGFR exon 20 insertion mutants, whereas afatinib has a smaller terminal group, which is indirectly linked to the quinazoline core, enabling afatinib to fit into the sterically hindered binding pocket.8Robichaux J.P. Elamin Y.Y. Tan Z. et al.Mechanisms and clinical activity of an EGFR and HER2 exon 20-selective kinase inhibitor in non-small cell lung cancer.Nat Med. 2018; 24: 638-646Crossref PubMed Scopus (214) Google Scholar Accordingly, osimertinib and rociletinib had minimal activity in a patient-derived xenograft model of NSCLC driven by EGFR-H773_V774insNPH.9Yang M. Xu X. Cai J. Ning J. Wery J.P. Li Q.X. NSCLC harboring EGFR exon-20 insertions after the regulatory C-helix of kinase domain responds poorly to known EGFR inhibitors.Int J Cancer. 2016; 139: 171-176Crossref PubMed Scopus (36) Google Scholar To further complicate the picture, first-generation inhibitors were effective against A763_Y764insFQEA, an insertion within the C-helix just one residue from the exon 19-20 border.5Arcila M.E. Nafa K. Chaft J.E. et al.EGFR exon 20 insertion mutations in lung adenocarcinomas: prevalence, molecular heterogeneity, and clinicopathologic characteristics.Mol Cancer Ther. 2013; 12: 220-229Crossref PubMed Scopus (263) Google Scholar,6Voon P.J. Tsui D.W.Y. Rosenfeld N. Chin T.M. EGFR exon 20 insertion A763-Y764insFQEA and response to erlotinib—letter.Mol Cancer Ther. 2013; 12: 2614-2615Crossref PubMed Google Scholar Moreover, preclinical data show that a nearby C-helix insertion on the other side of the border in exon 19, D761_E762insEAFQ, is sensitive to gefitinib and erlotinib.16Kohsaka S. Nagano M. Ueno T. et al.A method of high-throughput functional evaluation of EGFR gene variants of unknown significance in cancer.Sci Transl Med. 2017; 9: eaan6566Crossref PubMed Scopus (108) Google Scholar These results suggest that exon 20 insertions in the loop sequence may cause different structural alterations than those in the C-helix, the latter exerting distortions more similar to exon 19 deletions, further highlighting the importance of critical consideration of the appropriate treatment choice for rare EGFR mutations. An emerging theme in exon 20 insertion mutant treatment is afatinib + cetuximab combination therapy.17van Veggel B. de Langen A.J. Hashemi S.M.S. et al.Afatinib and cetuximab in four patients with EGFR exon 20 insertion–positive advanced NSCLC.J Thorac Oncol. 2018; 13: 1222-1226Abstract Full Text Full Text PDF PubMed Scopus (39) Google Scholar,18Fang W. Huang Y. Gan J. Shao Y.W. Zhang L. Durable response of low-dose afatinib plus cetuximab in an adenocarcinoma patient with a novel EGFR exon 20 insertion mutation.J Thorac Oncol. 2019; 14: e220-e221Abstract Full Text Full Text PDF PubMed Scopus (7) Google Scholar Although this approach demonstrated progression-free survival between 2.7 and 17.6 months, the combination is associated with toxicity issues. Here, we present a clinical case of differential response to first- and second-generation EGFR inhibitors in a patient with lung adenocarcinoma harboring EGFR-H773_V774insNPH, an exon 20 insertion mutation located 8 amino acids downstream of the C-helix of EGFR. The lack of response to first-line gefitinib therapy and the rapid decrease in ECOG performance status is in striking contrast with the lasting benefits of afatinib treatment experienced without any side effects by the patient. This case provides a clinical confirmation to previous preclinical findings revealing marked differences in efficacy of first- and third- versus second-generation TKIs against EGFR mutations in this region. In line with our observations, treatment of a patient with advanced lung adenocarcinoma harboring ERBB2-V777_G778insGSP (corresponding to V769_D770 in EGFR, a position also located in the loop following the C-helix) with afatinib resulted in significant tumor shrinkage.7Kosaka T. Tanizaki J. Paranal R.M. et al.Response heterogeneity of EGFR and HER2 exon 20 insertions to covalent EGFR and HER2 inhibitors.Cancer Res. 2017; 77: 2712-2721Crossref PubMed Scopus (85) Google Scholar The response was sustained for 7 months with eventual clinical progression owing to leptomeningeal carcinomatosis. It is also important to note, that although ERBB2 (human epidermal growth factor receptor 2 [HER2]) mutations are much less common than EGFR mutations (about 2% of patients with NSCLC), exon 20 insertions are the most prevalent type of HER2 alterations in NSCLC (>90% of all HER2 mutations).19Arcila M.E. Chaft J.E. Nafa K. et al.Prevalence, clinicopathologic associations, and molecular spectrum of ERBB2 (HER2) tyrosine kinase mutations in lung adenocarcinomas.Clin Cancer Res. 2012; 18: 4910-4918Crossref PubMed Scopus (319) Google Scholar,20Mazières J. Peters S. Lepage B. et al.Lung cancer that harbors an HER2 mutation: epidemiologic characteristics and therapeutic perspectives.J Clin Oncol. 2013; 31: 1997-2003Crossref PubMed Scopus (456) Google Scholar Taken together, emerging preclinical and clinical evidence supports that second-generation TKIs are effective against exon 20 insertions located in the loop region following the C-helix (downstream of A767) of EGFR. The case described is, to our knowledge, the first to demonstrate the clinical benefit of afatinib in a patient with lung adenocarcinoma with an EGFR-H773_V774 insertion mutation, whereas it also demonstrates a lack of activity for gefitinib. The presented case also illustrates the value of mutation type-specific structural, biochemical, and preclinical evidence for ‘hard-to-treat’ uncommon mutations. These findings warrant further investigation into second-generation EGFR inhibitors for NSCLC driven by insertions within this region of EGFR. Clinical trial data are usually not informative with regards to rare mutations, yet about 20% of exon 20 insertions occur at position H773, and more than 10% at the neighboring P772 and V774 positions.2Vyse S. Huang P.H. Targeting EGFR exon 20 insertion mutations in non-small cell lung cancer.Signal Transduct Target Ther. 2019; 4: 8Crossref PubMed Scopus (125) Google Scholar This translates to a substantial number of patients with NSCLC who can potentially benefit from tumor sequencing and appropriate treatment decisions in precision oncology. The authors have stated that they have no conflicts of interest. This work was supported by the Hungarian Innovation Agency (grant numbers NVKP_16-1-2016-0005 , KFI_16-1-2016-0048 , and 2019-1.1.1-PIACI-KFI-2019-00367 ).