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Combination Osimertinib and Gefitinib in C797S and T790M EGFR-Mutated Non–Small Cell Lung Cancer

  • Surein Arulananda
    Affiliations
    Cancer Immunobiology Laboratory, Olivia Newton-John Cancer Research Institute, Heidelberg, Victoria, Australia

    School of Cancer Medicine, La Trobe University, Bundoora, Victoria, Australia
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  • Hongdo Do
    Affiliations
    School of Cancer Medicine, La Trobe University, Bundoora, Victoria, Australia

    Translational Genomics and Epigenomics Laboratory, Olivia Newton-John Cancer Research Institute, Heidelberg, Victoria, Australia

    Department of Pathology, University of Melbourne, Parkville, Victoria, Australia
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  • Ashan Musafer
    Affiliations
    Translational Genomics and Epigenomics Laboratory, Olivia Newton-John Cancer Research Institute, Heidelberg, Victoria, Australia
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  • Paul Mitchell
    Affiliations
    Cancer Immunobiology Laboratory, Olivia Newton-John Cancer Research Institute, Heidelberg, Victoria, Australia

    Department of Medical Oncology, Olivia Newton-John Cancer and Wellness Centre, Austin Health, Heidelberg, Victoria, Australia
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  • Alexander Dobrovic
    Affiliations
    School of Cancer Medicine, La Trobe University, Bundoora, Victoria, Australia

    Translational Genomics and Epigenomics Laboratory, Olivia Newton-John Cancer Research Institute, Heidelberg, Victoria, Australia

    Department of Pathology, University of Melbourne, Parkville, Victoria, Australia
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  • Thomas John
    Correspondence
    Corresponding author. Address for correspondence: Thomas John, M.B.B.S., PhD, Cancer Immunobiology Laboratory, Olivia Newton-John Cancer Research Institute, 145 Studley Road, Heidelberg 3084. Victoria, Australia.
    Affiliations
    Cancer Immunobiology Laboratory, Olivia Newton-John Cancer Research Institute, Heidelberg, Victoria, Australia

    School of Cancer Medicine, La Trobe University, Bundoora, Victoria, Australia

    Department of Medical Oncology, Olivia Newton-John Cancer and Wellness Centre, Austin Health, Heidelberg, Victoria, Australia
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Open AccessPublished:August 23, 2017DOI:https://doi.org/10.1016/j.jtho.2017.08.006

      Abstract

      Introduction

      Osimertinib, a third-generation EGFR tyrosine kinase inhibitor has demonstrated efficacy in tumors harboring the EGFR T790M resistance mutation. Inevitably, resistance to third-generation inhibitors results in disease progression, with the EGFR C797S mutation being one of several resistance pathways identified to date. On the basis of preclinical data, we report what is the first known case of a patient harboring the T790M and C797S mutations in trans treated with combination gefitinib and osimertinib.

      Methods

      On development of progressive disease after multiple therapies, the patient’s plasma was sequenced using the Oncomine Lung cfDNA Assay (Thermo Fisher Scientific, Waltham, MA). Subsequent monitoring of circulating tumor DNA in plasma was performed by droplet digital polymerase chain reaction.

      Results

      Sequencing showed that the T790M and C797S mutations were in trans. Within 2 weeks of commencement of combination therapy, rapid clinical improvement occurred. Accompanying this, a rapid decline in the C797S mutation subclone in plasma was detected. However, the levels of the EGFR exon 19 deletion driver mutation and the T790M resistance mutation in the circulating tumor DNA continued to rise and the patient died from progressive disease 6 weeks after commencement of combination therapy. There were no adverse events seen with the combination therapy.

      Conclusion

      This is, to the best of our knowledge, the first reported case of combination EGFR tyrosine kinase inhibitor therapy tailored to the allelic conformation of T790M and C797S mutation that resulted in brief clinical improvement without toxicity.

      Keywords

      Introduction

      EGFR-mutated NSCLC is exquisitely responsive to targeted tyrosine kinase inhibitor (TKI) therapy with erlotinib, gefinitib, or afatinib. However, resistance to the EGFR TKIs invariably occurs, most frequently because of the T790M mutation, which increases the receptor affinity for adenosine triphosphate (ATP) binding and hence reduction in primary activity of ATP-competitive TKIs.
      • Minari R.
      • Bordi P.
      • Tiseo M.
      Third-generation epidermal growth factor receptor-tyrosine kinase inhibitors in T790M-positive non-small cell lung cancer: review on emerged mechanisms of resistance.
      Osimertinib, a third-generation TKI, is a potent inhibitor of activating EGFR mutations and the T790M resistance mutation. The phase III AURA 3 study confirmed a significant progression-free survival (PFS) benefit (hazard ratio = 0.32, 95% confidence interval: 0.21–0.39) of osimertinib compared with platinum-pemetrexed chemotherapy in T790M-positive patients after progression while taking a first-generation TKI.
      • Mok T.S.
      • Wu Y.L.
      • Ahn M.J.
      • et al.
      Osimertinib or platinum-pemetrexed in EGFR T790M-positive lung cancer.
      In the original AURA dose expansion study, the median PFS noted with osimertinib was 9.6 months.
      • Jänne P.A.
      • Yang J.C.
      • Kim D.W.
      • et al.
      AZD9291 in EGFR inhibitor-resistant non-small-cell lung cancer.
      On the AURA 3 study, the median PFS was 10.1 months.
      • Mok T.S.
      • Wu Y.L.
      • Ahn M.J.
      • et al.
      Osimertinib or platinum-pemetrexed in EGFR T790M-positive lung cancer.
      The C797S mutation occurs in exon 20. The cysteine at position 797 is used by third-generation TKIs such as osimertinib to covalently bind with the receptor. Hence the EGFR C797S mutation leads to ineffective binding and subsequent resistance to osimertinib.
      • Zhou W.
      • Ercan D.
      • Chen L.
      • et al.
      Novel mutant-selective EGFR kinase inhibitors against EGFR T790M.
      Unlike T790M, which is the most common mechanism of resistance to first-generation TKIs, the C797S mutation was initially reported in only six of 19 patients progressing during osimertinib therapy (31%).
      • Thress K.S.
      • Paweletz C.P.
      • Felip E.
      • et al.
      Acquired EGFR C797S mutation mediates resistance to AZD9291 in non-small cell lung cancer harboring EGFR T790M.
      In our own series of patients, the C797S mutation was detected in four of 10 patients with the T790M mutation, but it occurred in trans in only one.
      Intriguing preclinical data in a T790M-positive cultivated first-generation– and third-generation–resistant cell line demonstrated that the configuration of the T790M and C797S mutations affected the cellular response to therapy.
      • Niederst M.J.
      • Hu H.
      • Mulvery H.E.
      • et al.
      The allelic context of the C797S mutation acquired upon treatment with third-generation EGFR inhibitors impacts sensitivity to subsequent treatment strategies.
      If the two EGFR mutations were in cis (on the same DNA strand), the cells were refractory to combination first- and third-generation TKIs. However, when the two mutations were in trans (on different DNA strands), the combination of first- and third-generation EGFR inhibitors led to inhibition in EGFR signalling and cell death.
      With this background, we report a patient with metastatic EGFR-mutant NSCLC treated with the combination of gefitinib and osimertinib after development of the T790M and C797S mutations in the trans allelic context.

      Case Report

      A 41-year-old man of Asian ethnicity with lymphangitic pulmonary disease and metastases to the mediastinum and bone presented in August 2014 with symptomatic cough and dyspnoea on exertion. He began receiving erlotinib, 150 mg, (Fig. 1A) after the detection of an exon 19 deletion within the EGFR gene, which resulted in a radiological response and rapid clinical benefit lasting 6 months (Fig. 1B). Subsequent tissue biopsy at the time of relapse confirmed the original exon 19 deletion in addition to an acquired T790M in exon 20.
      Figure thumbnail gr1
      Figure 1Sequence of the patient’s anticancer treatment and radiological imaging before and after therapies. (A) Time line of anticancer therapies received by the patient. (B) Computed tomography (CT) scan (left) shows a left lower lobe lung primary 3 months into erlotinib therapy in November 2014. CT scan (right) shows progressive disease during erlotinib therapy in March 2015. (C) CT scan (left) shows response in the lung 3 months into osimertinib therapy in June 2015. CT scan (right) shows progressive disease with lymphangitis carcinomatosis in March 2016. (D). Chest radiograph (left) obtained before commencement of combination gefitinib and osimertinib therapy. Chest radiograph (right) obtained 2 weeks after combination gefitinib and osimertinib therapy.
      The patient then received osimertinib, 80 mg, as part of the AURA3 trial; he achieved a rapid clinical and radiological response lasting 8 months (Fig. 1C). Because of ongoing clinical benefit, the patient continued receiving osimertinib for a further 4 months but was switched to carboplatin and paclitaxel after worsening lymphangitis. Again, chemotherapy resulted in a partial response after two cycles but disease progression after the fourth cycle.
      The patient went on to receive nivolumab, 3 mg/kg; however, after four cycles, his condition worsened and he required hospitalization for dyspnea on minimal exertion and a debilitating cough. High-resolution chest computed tomography (CT) demonstrated worsening of the lymphangitis carcinomatosis. Restaging brain, abdominal, and pelvic CT revealed new metastases in the brain, liver, and pleura. The patient received one cycle of pemetrexed chemotherapy, but his condition continued to deteriorate.
      To study the cause of disease progression after osimertinib, a blood sample was collected and double-spun to separate plasma. Plasma DNA samples isolated using the QIAamp Circulating Nucleic Acid Kit (Qiagen, Hilden, Germany) were tested for the EGFR exon 19 deletion, T790M, and C797S mutations using droplet digital polymerase chain reaction assays. An EGFR C797S (c.2390G>C) mutation was detected. To determine the cis or trans allelic relationship of the C797S with T790M (Fig. 2A), the patient’s plasma DNA was deep sequenced by using the Oncomine Lung cfDNA Assay (Thermo Fisher Scientific, Waltham, MA). A C797S (c.2390G>C) mutation in addition to the original exon 19 deletion and T790M mutations was detected in the EGFR gene. However, the C797S and T790M mutations were detected on different DNA strands, indicating the trans allelic relationship (Fig. 2B).
      Figure thumbnail gr2
      Figure 2Allelic relationship of EGFR T790M and C797S mutations. (A) Diagram showing the cis (on the same DNA strand) and trans (on different DNA strands) allelic relationships of the EGFR T790M and C797S mutations. (B) Deep sequencing data on the patient’s plasma DNA showing the EGFR T790M and C797S mutations detected in a trans allelic relationship.
      As the C797S mutation was detected in trans with the T790M mutation, the patient was treated with a combination of gefitinib and osimertinib. Within 3 days, his clinical situation improved and he was discharged. His condition was reviewed 2 weeks later, and his dyspnea and cough were well controlled. A chest radiograph showed some improvement of the lymphangitis carcinomatosis (Fig. 1D). However, a month after commencement of combination TKI therapy, the patient re-presented to the hospital with relapse of his cough and dyspnea. A chest CT scan showed significant progression of the lymphangitis carcinomatosis; the patient's condition continued to deteriorate, and he died 2 weeks later (6 weeks after commencement of the combination TKI and 2 years and 2 months after the diagnosis of metastatic lung cancer). Two blood samples were collected during the combination therapy to monitor the levels of the EGFR exon 19 deletion, T790M, and C797S mutations. The level of the C797S mutation was substantially reduced during the combination therapy, whereas the EGFR exon 19 deletion and T790M mutations steadily increased (Fig. 3).
      Figure thumbnail gr3
      Figure 3Monitoring of the EGFR mutations in blood. Three blood samples were collected before and during the combination therapy consisting of gefitinib and osimertinib (days 3 and 16). The levels of EGFR exon 19 deletion (ex19del), T790M, and C797S mutations were measured by using droplet digital polymerase chain reactions assays. Levels of the EGFR ex19del and T790M steadily increased, whereas the level of the C797S mutation rapidly decreased.

      Discussion

      Lung tumors responding to osimertinib inevitably develop resistance through EGFR-dependent and EGFR-independent mechanisms (Fig. 4). The EGFR-dependent resistance mechanisms include acquired tertiary EGFR mutations such as EGFR C797S and loss of the T790M mutation. The acquired EGFR C797S mutation that affects the binding site of osimertinib was predicted in preclinical models
      • Niederst M.J.
      • Hu H.
      • Mulvery H.E.
      • et al.
      The allelic context of the C797S mutation acquired upon treatment with third-generation EGFR inhibitors impacts sensitivity to subsequent treatment strategies.
      • Ercan D.
      • Choi H.G.
      • Yun C.H.
      • et al.
      EGFR mutations and resistance to irreversible pyrimidine-based EGFR inhibitors.
      and is the most common resistance mechanism found.
      • Thress K.S.
      • Paweletz C.P.
      • Felip E.
      • et al.
      Acquired EGFR C797S mutation mediates resistance to AZD9291 in non-small cell lung cancer harboring EGFR T790M.
      • Yu H.A.
      • Tian S.K.
      • Drilon A.E.
      • et al.
      Acquired resistance of EGFR-mutant lung cancer to a T790M-specific EGFR inhibitor: emergence of a third mutation (C797S) in the EGFR tyrosine kinase domain.
      Thress et al. identified EGFR C797S mutations in six of 19 patients with lung cancer who had progressed while taking osimertinib (31%).
      • Thress K.S.
      • Paweletz C.P.
      • Felip E.
      • et al.
      Acquired EGFR C797S mutation mediates resistance to AZD9291 in non-small cell lung cancer harboring EGFR T790M.
      Figure thumbnail gr4
      Figure 4Mechanisms of resistance to osimertinib, a third-generation EGFR tyrosine kinase inhibitor, including EGFR-dependent and EGFR-independent pathways. MET, Mesenchymal to Epithelial Transition gene; HER2, erb-b2 receptor tyrosine kinase 2 gene.
      Although EGFR C797S mutations are the most commonly detected, other EGFR mutations that interfere with the covalent binding of osimertinib can also cause resistance to osimertinib. Bersanelli et al. identified an EGFR L718Q mutation in an osimertinib-resistant lung tumor.
      • Bersanelli M.
      • Minari R.
      • Bordi P.
      • et al.
      L718Q mutation as new mechanism of acquired resistance to AZD9291 in EGFR-mutated NSCLC.
      Loss of the EGFR T790M mutation with retention of the initial activating EGFR mutation was reported in four of 15 patients with osimertinib-resistant T790M-mutant lung cancer.
      • Thress K.S.
      • Paweletz C.P.
      • Felip E.
      • et al.
      Acquired EGFR C797S mutation mediates resistance to AZD9291 in non-small cell lung cancer harboring EGFR T790M.
      Our group previously reported such a case, in which the plasma level of EGFR exon 19 deletion mutation rapidly increased without detectable EGFR T790M mutation in the blood when the patient progressed during osimertinib treatment.
      • Chia P.L.
      • Do H.
      • Morey A.
      • Mitchell P.
      • Dobrovic A.
      • John T.
      Temporal changes of EGFR mutations and T790M levels in tumour and plasma DNA following AZD9291 treatment.
      Several EGFR-independent mechanisms of resistance to osimertinib that activate bypass pathways have also been reported. Mesenchymal to Epithelial Transition gene (MET) amplification is as an important mechanism of resistance to first-generation EGFR TKIs through phosphorylation of erb-b2 receptor tyrosine kinase 3 and downstream activation of the phosphoinositide 3-kinase/AKT pathway. Recent preclinical and clinical studies have also demonstrated that MET amplification causes resistance to osimertinib.
      • Shi P.
      • Oh Y.T.
      • Zhang G.
      • et al.
      Met gene amplification and protein hyperactivation is a mechanism of resistance to both first and third generation EGFR inhibitors in lung cancer treatment.
      • Ou S.H.
      • Agarwal N.
      • Ali S.M.
      High MET amplification level as a resistance mechanism to osimertinib (AZD9291) in a patient that symptomatically responded to crizotinib treatment post-osimertinib progression.
      In addition, BRAF and KRAS mutations, as well as transformation to small cell carcinoma, have also been identified as mechanisms of acquired resistance to osimertinib.
      • Ho C.C.
      • Liao W.Y.
      • Lin C.A.
      • Shih J.Y.
      • Yu C.J.
      • Chih-Hsin Yang J.
      Acquired BRAF V600E mutation as resistant mechanism after treatment with osimertinib.
      • Ortiz-Cuaran S.
      • Scheffler M.
      • Plenker D.
      • et al.
      Heterogeneous mechanisms of primary and acquired resistance to third-generation EGFR inhibitors.
      • Li L.
      • Wang H.
      • Li C.
      • Wang Z.
      • Zhang P.
      • Yan X.
      Transformation to small-cell carcinoma as an acquired resistance mechanism to AZD9291: a case report.
      Although resistance to third-generation EGFR TKIs inevitably occurs, strategies for managing this scenario have largely been limited to chemotherapy. On the basis of preclinical data, we report for (to the best of our knowledge) the first time, the clinical efficacy of combination first- and third-generation EGFR TKIs in a patient with the C797S mutation occurring in trans with the T790M mutation. Although initial clinical benefit occurred, our patient progressed rapidly and died within a short time. The combination TKIs were well tolerated, with no rash, diarrhea, transaminitis, or pneumonitis (which are adverse events known to occur with each TKI individually). Our results of monitoring circulating tumor DNA suggest multiple EGFR-mutant clones detectable within the plasma. The initial decline in C797S paralleled the clinical improvement; however, the steady rise in the plasma T790M and activating exon 19 deletion levels indicate rapid progression of another resistant clone.
      Recently, a novel TKI that inhibited the triple mutation consisting of the activating EGFR, T790M, and C797S mutations was reported.
      • Jia Y.
      • Yun C.H.
      • Park E.
      • et al.
      Overcoming EGFR(T790M) and EGFR(C797S) resistance with mutant-selective allosteric inhibitors.
      In this preclinical study, a genetically engineered L858R/T790M/C797S mouse was treated with the combination of cetuximab, an antibody that blocks EGFR dimerization, and EAI045, an allosteric inhibitor that synergizes with cetuximab and overcomes the enhanced ATP affinity conferred by the T790M mutation and is not affected by the C797S mutation, as the residue is remote from the allosteric binding pocket. After two doses of treatment, significant tumor shrinkage was noted.

      Conclusion

      The fluidity of the cancer genome enables rapid evolution of resistance mechanisms on exposure to targeted therapies. Development of the C797S mutation to osimertinib is a relatively new finding, with newer therapeutic strategies looking to target this mutation. Tailoring a therapeutic approach according to the allelic conformation of the T790M and C797S mutations demonstrated some efficacy in our patient, although the improvement was brief. As tumors evolve in the host and become exposed to subsequent drug therapies, resistant subclones emerge. This temporal heterogeneity, which differs between patients undergoing the same therapies, can result in single or multiple resistance mechanisms. The durability of the response to therapy likely depends on the relative quantities of each. Although our patient did not obtain substantial benefit, more data are needed to determine whether the efficacy of combination EGFR TKIs can be taken beyond a novel in vitro observation.

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      Linked Article

      • C797S and T790M EGFR Mutations in Non–Small Cell Lung Cancer: In Trans or in Separate Clones?
        Journal of Thoracic OncologyVol. 13Issue 2
        • Preview
          I read with great interest the article “Combination Osimertinib and Gefitinib in C797S and T790M EGFR-Mutated Non–Small Cell Lung Cancer” by Arulananda et al. that was recently published in the Journal of Thoracic Oncology.1 Using up-to-date methodologies, the authors monitored the EGFR mutation status in circulating tumor DNA (ctDNA) in the plasma of a patient with progressive disease after multiple lines of therapy, including tyrosine kinase inhibitors (TKIs). They detected T790M and C797S resistance mutations on different DNA strands and postulated a trans configuration.
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        • PDF
        Open Archive
      • Resistance in trans-ition
        Journal of Thoracic OncologyVol. 12Issue 11
        • Preview
          The tyrosine kinase inhibitors (TKIs) gefitinib, erlotinib, and afatinib have led to superior outcomes in patients with EGFR-mutated NSCLC compared to conventional chemotherapy. However, resistance to these drugs develops in nearly all patients, most commonly through the evolution of an EGFR T790M mutation, which is detected in more than 50% of patients at time of disease progression.1,2 Osimertinib is a potent, irreversible third-generation TKI that is active against the T790M resistance mutation—along with activating EGFR mutations—and is approved for patients with a T790M mutation who have progressed during treatment with a first-generation TKI.
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        Open Access