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Limei Hu



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    MA09 - EGFR & MET (ID 128)

    • Event: WCLC 2019
    • Type: Mini Oral Session
    • Track: Targeted Therapy
    • Presentations: 1
    • Now Available
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      MA09.03 - Identification of Mechanisms of Acquired Resistance to Poziotinib in EGFR Exon 20 Mutant Non-Small Cell Lung Cancer (NSCLC) (Now Available) (ID 2904)

      15:15 - 16:45  |  Author(s): Limei Hu

      • Abstract
      • Presentation
      • Slides

      Background

      Insertions/mutations in exon 20 of EGFR occur in ~2% Insertions/mutations in exon 20 of EGFR occur in ~2% of all lung adenocarcinomas. These alterations are characterized by primary resistance to approved tyrosine kinase inhibitors (TKIs) with response rates of <12%. We have shown that exon 20 insertions restrict the size of the drug-binding pocket, limiting binding of large inhibitors. However, poziotinib can circumvent these steric changes and is a potent inhibitor of EGFR exon 20 mutants. In our investigator-initiated phase 2 trial of EGFR exon 20 mutant NSCLC, poziotinib was associated with a best objective response rate of 55% (Heymach et al, 19th WCLC). Herein, we use preclinical models and clinical samples from our phase 2 study to identify mechanisms of acquired poziotinib resistance (NCT03066206).

      Method

      EGFR exon 20 insertion (D770insNPG) genetically engineered mice (GEM) were treated with poziotinib until progression. Upon progression, tumor DNA and protein were analyzed using whole exome sequencing (WES) and reverse phase protein assay (RPPA). Mandatory and optional biopsies were obtained at baseline and progression, respectively, from patients treated in our phase 2 trial of poziotinib in EGFR exon 20 mutant NSCLC. Serial cfDNA was collected at baseline, 8 weeks of therapy, and on progression. Patient samples were analyzed using targeted next generation sequencing or WES.

      Result

      Poziotinib acquired-resistance GEM tumors acquired mutations in ErbB4, KRAS, and other genes which represent potential targetable bypass pathways. Resistant GEM tumors displayed increased activation of MAPK, AKT, ERK and MEK compared to sensitive tumors, suggesting that poziotinib acquired resistance is associated with reactivation of the MAPK/PI3K pathways. We enrolled 50 EGFR exon 20 mutant patients in our phase 2 trial. Analysis of matched pre-poziotinib and on-progression samples from 20 responding patients revealed acquired EGFR tyrosine kinase domain point mutations in 4 patients (T790M (2), V774A (1), D770A, (1)). Ba/F3 cells co-expressing EGFR exon 20 insertion (S768supSVD) and T790M were resistant to poziotinib, suggesting that T790M is a poziotinib resistance driver. Potential acquired EGFR-independent resistance mechanisms identified in patients to date include PIK3CA E545K (1), MAP2K2 S94L (1), MET amplification (1), EGFR amplification (2), and CDK6 amplification (2).

      Conclusion

      Parallel to acquired resistance mechanisms seen in classical EGFR mutation, acquired resistance to poziotinib can be mediated through EGFR-dependent mechanisms, notably T790M and other EGFR tyrosine kinase domain point mutations. EGFR-independent resistance mechanisms include activation of bypass pathways. Preclinical validation of resistance mechanisms and additional analysis of patient samples will be presented at the meeting.

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    P1.14 - Targeted Therapy (ID 182)

    • Event: WCLC 2019
    • Type: Poster Viewing in the Exhibit Hall
    • Track: Targeted Therapy
    • Presentations: 1
    • Moderators:
    • Coordinates: 9/08/2019, 09:45 - 18:00, Exhibit Hall
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      P1.14-08 - Activity of Poziotinib and Other 2nd-Gen Quinazoline EGFR TKIs in Atypical Exon18 and Acquired Osimertinib Resistance Mutants (ID 2694)

      09:45 - 18:00  |  Author(s): Limei Hu

      • Abstract

      Background

      In EGFR, exon 18 encodes for the P-loop (L718-V726), and mutations in this region (G719S/A, L718Q/V, G724S) are known to reduce sensitivity to osimertinib and first-generation EGFR TKIs. Osimertinib resistance is associated with a number of acquired mutations in exons 19 and 20 (S784F, L747S, C797S and L792H). We investigated the frequency and drug sensitivity of these and other osimertinib-resistant EGFR mutations

      Method

      We generated ~50 different Ba/F3 cell line models expressing classical and/or atypical EGFR mutations (exons 18-21) and evaluated the transforming ability and sensitivity to 14 EGFR TKIs including non-covalent (first-generation), afatinib, dacomitinib, and poziotinib (quinazoline and covalent, second-generation), and covalent T790M-specific (third-generation) inhibitors. Impact of atypical mutations was analyzed by in silico modeling.

      Result

      We found 3.6% (N=32/895) of EGFR-mutant patients had atypical, exon 18, P-loop mutations in the MD Anderson GEMINI database. Modeling of classical EGFR mutations revealed osimertinib has distinct interactions between the solvent front of osimertinib and residues within the P-loop of EGFR, whereas second-generation quinazoline TKIs, such as poziotinib, extend into the pocket, near T790, lacking these interactions. Mutations in the P-loop were predicted to shift osimertinib out of alignment with V726 and F723, causing resistance to osimertinib but not quinazoline-based TKIs. Atypical exon 18 mutations (G719S/A, L718Q/V, G724S) had IC50 values of 113.6nM, 1.6nM, and 137.5nM for first-, second-, and third-generation TKIs, respectively. Second-generation TKIs inhibited G719S/A-T790M mutations at concentrations 2-fold lower than third-generation TKIs (IC50 = 23.4nM and 46nM). Osimertinib-resistance mutations (L747S, S784F, C797S, L792H) co-occurring with classical sensitizing mutations (L858R or ex19del) had IC50 values of 56.8nM, 1.4nM, and 996nM to first, second and third-generation inhibitors. Of the second-generation TKIs tested, poziotinib was the most potent for atypical exon 18 P-loop mutations; G719S/A-T790M mutations; and classical mutants with acquired osimertinib-resistance mutations (IC50= 0.4nM, 3.2nM, 0.8nM).

      Conclusion

      Exon 18 atypical P-loop mutations and osimertinib-resistance mutations demonstrated high sensitivity to second-generation quinazoline TKIs, compared to first- and third-generation inhibitors. Mutations in the P-loop of EGFR confer resistance to third-generation TKIs by destabilizing solvent front interactions of the molecule, and osimertinib-resistance mutations interfere with covalent binding at C797. Second-generation TKIs, especially poziotinib, are potent inhibitors of these mutations because they have increased hydrophobic interactions at the back of the drug binding cleft that are retained without covalent binding. Together, these data indicate that poziotinib and other second-generation TKIs may be useful for the treatment of NSCLC patients with atypical P-loop and selected osimertinib-resistant EGFR mutations.