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

Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 25 Shattuck Street, Boston, MA 02115, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 03/2008; 105(6):2070-5. DOI: 10.1073/pnas.0709662105
Source: PubMed


Lung cancers caused by activating mutations in the epidermal growth factor receptor (EGFR) are initially responsive to small molecule tyrosine kinase inhibitors (TKIs), but the efficacy of these agents is often limited because of the emergence of drug resistance conferred by a second mutation, T790M. Threonine 790 is the "gatekeeper" residue, an important determinant of inhibitor specificity in the ATP binding pocket. The T790M mutation has been thought to cause resistance by sterically blocking binding of TKIs such as gefitinib and erlotinib, but this explanation is difficult to reconcile with the fact that it remains sensitive to structurally similar irreversible inhibitors. Here, we show by using a direct binding assay that T790M mutants retain low-nanomolar affinity for gefitinib. Furthermore, we show that the T790M mutation activates WT EGFR and that introduction of the T790M mutation increases the ATP affinity of the oncogenic L858R mutant by more than an order of magnitude. The increased ATP affinity is the primary mechanism by which the T790M mutation confers drug resistance. Crystallographic analysis of the T790M mutant shows how it can adapt to accommodate tight binding of diverse inhibitors, including the irreversible inhibitor HKI-272, and also suggests a structural mechanism for catalytic activation. We conclude that the T790M mutation is a "generic" resistance mutation that will reduce the potency of any ATP-competitive kinase inhibitor and that irreversible inhibitors overcome this resistance simply through covalent binding, not as a result of an alternative binding mode.

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Available from: Angela Toms, Oct 02, 2015
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    • "EGFR inhibitors, such as gefitinib and erotinib have been clinically used for cancer treatment [6] [7] [8]. However, the clinical efficacy of EGFR inhibitors in EGFR-mutant non-small-cell lung cancer (NSCLC) is limited by the development of drug-resistance mutations [9] [10] [11]. Therefore, to find novel mutant-selective EGFR inhibitors for developing new drugs for treatment of cancer is urgently needed [12] [13]. "
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    ABSTRACT: A method for screening of inhibitors to epidermal growth factor receptor (EGFR) in natural product extracts with capillary electrophoresis (CE) in conjunction with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) is reported. The method was established by employing 5-carboxyfluorescein labeled substrate peptide, two commercially available EGFR inhibitors OSI-744 and ZD1839, and a small chemical library consisted of 39 natural product extracts derived from the Traditional Chinese Medicines. Biochemical assay of crude natural product extracts was carried out by using CE equipped with a laser induced fluorescence detector. The CE separation allowed an accurately quantitative measurement of the phosphorylated product, hence the measurement of the enzymatic activity as well as the inhibition kinetics. The hits are identified if the peak area of the phosphorylated product is reduced in comparison with the negative control. The active constituents in the natural product extract were then identified by an assay-guided isolation with HPLC-MS/MS system. With the method, the flavonoids component of the Lycopus lucidus extract, namely quercetin and rutin were identified to be the active ingredients. Their IC50 values were determined as 0.88μM and 10.1μM, respectively. This result demonstrated a significant merit of our method in the identification of the bioactive compounds in natural products. Copyright © 2015. Published by Elsevier B.V.
    Journal of Chromatography A 05/2015; 1400. DOI:10.1016/j.chroma.2015.04.055 · 4.17 Impact Factor
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    • "Currently, almost all patients with activating mutations (deletion in exon 19 or point mutation in exon 21) eventually develop acquired resistance while receiving EGFR-TKI therapy,20 and about 50% of these patients have a secondary mutation in exon 20 (p.T790M).86–90 The p.T790M mutation abolishes the influence of EGFR-TKIs by increasing the affinity for adenosine triphosphate (ATP).87 There is a major clinical problem in that NSCLC patients with EGFR mutations achieve acquired resistance after treatment with EGFR-TKIs. "
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    ABSTRACT: Non-small-cell lung cancer (NSCLC) has entered the age of individual treatment, and increasing point mutations of specific oncogenes and rearrangement of some chromosomes are biomarkers used to predict the therapeutic effect of targeted therapy. At present, there is a consensus among clinicians that epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have shown favorable efficacy in NSCLC patients with EGFR mutation, and some relevant research has suggested that the presence of EGFR mutations is a favorable prognostic marker. However, the association of EGFR mutation status with the responsiveness to conventional chemotherapy agents and survival in NSCLC patients is still unclear. This review provides an overview of and assesses the role of EGFR as a prognostic marker for postoperative patients and as a predictive marker for response to cytotoxic chemotherapy. In addition, we review the comparison of response to chemotherapy between EGFR mutations in exon 19 and in exon 21 and the predictive role of p.T790M mutation.
    Drug Design, Development and Therapy 09/2014; 8:1595-1611. DOI:10.2147/DDDT.S69690 · 3.03 Impact Factor
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    • "T790M, which represents threonine-to-methionine substitution at position 790 in the gatekeeper residue of EGFR kinase domain because of a point mutation within exon 20, presents in about 50% of EGFR mutant NSCLCs that become resistant to TKIs after the treatment.109,112,118,119 EGFR of lung cancer cells with both TKI-sensitive mutations and T790M mutation regain affinity to ATP at the hydrophobic pocket, hence abrogating competitive inhibition of the receptor by gefitinib and erlotinib.120 Other secondary mutations in the EGFR kinase domain that are associated with acquired resistance include L747S,121 D761Y,122 and T854A.123 "
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    ABSTRACT: Lung cancer, mostly nonsmall cell lung cancer, continues to be the leading cause of cancer-related death worldwide. With the development of tyrosine kinase inhibitors that selectively target lung cancer-related epidermal growth factor receptor mutations, management of advanced nonsmall cell lung cancer has been greatly transformed. Improvements in progression-free survival and life quality of the patients were observed in numerous clinical studies. However, overall survival is not prolonged because of later-acquired drug resistance. Recent studies reveal a heterogeneous subclonal architecture of lung cancer, so it is speculated that the tumor may rapidly adapt to environmental changes via a Darwinian selection mechanism. In this review, we aim to provide an overview of both spatial and temporal tumor heterogeneity as potential mechanisms underlying epidermal growth factor receptor tyrosine kinase inhibitor resistance in nonsmall cell lung cancer and summarize the possible origins of tumor heterogeneity covering theories of cancer stem cells and clonal evolution, as well as genomic instability and epigenetic aberrations in lung cancer. Moreover, investigational measures that overcome heterogeneity-associated drug resistance and new assays to improve tumor assessment are also discussed.
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