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Zhenhao Fang



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    MA27 - Novel Drugs and PDX Models (ID 931)

    • Event: WCLC 2018
    • Type: Mini Oral Abstract Session
    • Track: Targeted Therapy
    • Presentations: 1
    • Moderators:
    • Coordinates: 9/26/2018, 13:30 - 15:00, Room 206 BD
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      MA27.07 - Lung Adenocarcinoma Harboring BRAF G469V Mutation is Uniquely Sensitive to EGFR Tyrosine Kinase Inhibitors (ID 12771)

      14:10 - 14:15  |  Author(s): Zhenhao Fang

      • Abstract
      • Presentation
      • Slides

      Background

      BRAF mutations occur in 2-5% of non-small cell lung cancers with ~50% being non-V600E. Previous studies reported that two BRAF G469 mutations, G469V and G469A increase kinase activity and MAPK activation, thus are likely oncogenic. Patients with non-V600E mutations are mostly not sensitive to approved BRAF inhibitors vemurafenib or dabrafenib. We established a lung adenocarcinoma (LUAD) patient derived xenograft (PDX) that is epidermal growth factor receptor (EGFR) wild type and non-amplified, but harbors BRAF G469V mutation, yet is sensitive to gefitinib. We performed functional studies to characterize the oncogenicity and sensitivity of BRAF G469 mutations to EGFR tyrosine kinase inhibitors (TKIs).

      a9ded1e5ce5d75814730bb4caaf49419 Method

      PDX12 was established in NOD-SCID mice from a resected stage IIIA LUAD. The XDC12 cell line was established from PDX12. NCI-H1395 and -H1755 LUAD cell lines with BRAF G469A mutation were obtained from ATCC. BRAF mutant driver activity was characterized by shRNA knockdown of BRAF in LUAD cell lines and the ability of the mutants to promote IL3-independent growth when expressed in Ba/F3 cells. PDX12 responsiveness to TKIs was evaluated by tumor volume shrinkage while cell line sensitivity was quantified using the MTS assay. Drug effects on signaling were assessed by phospho-immunoblotting. Computational modeling was used to predict how the mutations promote BRAF activation and sensitivity to EGFR-TKIs, while purified BRAF proteins were used to validate predictions.

      4c3880bb027f159e801041b1021e88e8 Result

      Knockdown of BRAF by shRNA inhibited growth of all BRAF mutant cell lines, while ectopic BRAF G469V and G469A expression in Ba/F3 cells promoted IL3-independent MAPK activation and growth, supporting both mutations being oncogenic drivers. The XDC12 cell line was sensitive to EGFR-TKIs (gefitinib, erlotinib, afatinib, and osimertinib), but resistant to the BRAF inhibitor dabrafenib, which correlated with inhibition of MAPK phosphorylation. By contrast, H1395 and H1755 cell lines with BRAF G469A mutations were resistant to both the EGFR-TKIs and the BRAF inhibitor. Similarly, only Ba/F3 cells expressing BRAF G469V, but not G469A, were sensitive to EGFR-TKIs. Consistent with the in vitro data and our initial PDX findings with gefitinib, multiple EGFR-TKIs induced tumor shrinkage in PDX12 in vivo.

      8eea62084ca7e541d918e823422bd82e Conclusion

      BRAF G469V/A mutations are oncogenic drivers but are insensitive to BRAF inhibitors. However, only BRAF G469V, but not G469A mutation, is sensitive to EGFR-TKIs. Thus, two different driver alterations affecting the same BRAF codon can lead to distinct drug sensitivities.

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