Virtual Library

 x 
My Library Virtual Library FAQ

Start Your Search

Kristine Raaby Jakobsen



Author of

  • +

    P2.01 - Advanced NSCLC (ID 618)

    • Event: WCLC 2017
    • Type: Poster Session with Presenters Present
    • Track: Advanced NSCLC
    • Presentations: 1
    • +

      P2.01-075c - FGFR1 as an Intrinsic Resistance Mechanism in Erlotinib Treated EGFR Mutated NSCLC (ID 9344)

      09:00 - 09:00  |  Presenting Author(s): Kristine Raaby Jakobsen

      • Abstract

      Background:
      A major obstacle of NSCLC targeted treatment is the occurrence of resistance. Drug inhibiting the Epidermal Growth Factor Receptor (EGFR) has been a treatment for more than a decade and is given as first line treatment in EGFR mutated patients. However, up to 30 % of the patients with EGFR mutations experience no objective response to EGFR-TKIs and hence appear as intrinsic resistant. We and others have discovered, that increased FGFR1 expression occurs in the lung adenocarcinoma cell lines with acquired resistance to EGFR inhibitors, but no studies have investigated high FGFR1 expression as an intrinsic resistance mechanism to erlotinib.

      Method:
      CRISPR-Cas9 SAM was used for genetically upregulation of FGFR1 in the adenocarcinoma cell lines HCC827 and PC9. Proliferation and response to erlotinib was investigated with CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay (Promega). Protein expression and was investigated using western blotting. FGF2 levels was investigated in serum samples from 36 EGFR-mutated patients using the Quantikine HS ELISA kit for Human FGF basic Immunoassay (R&D) and FGFR1 expression will be investigated with IHC in a cohort of 30 EGFR mutated patients.

      Result:
      FGFR1 upregulation in PC9 and HCC827 (FGFR1+) did not by itself decrease the sensitivity to erlotinib. However, when the cells were treated with the FGFR1 ligand FGF2 the cells became significantly (p<0.05) more resistant compared to control cells. FGF2 in itself also made the control cells less sensitive to erlotinib, hence we hypothesized that serum levels of FGF2 may influence the response to erlotinib in EGFR mutated patients. FGF2 ELISA performed on 36 EGFR mutated patients, however, revealed that serum-FGF2 did not correlate with PFS. At this moment, we are conducting WB analysis of the FGFR1 pathway for FGFR1+ cells and control cells treated with FGF2 and erlotinib to reveal the functional bypass signalling. We are also preparing IHC for FGFR1 on tissue from EGFR mutated patients obtained prior to erlotinib treatment to confirm our in vitro findings.

      Conclusion:
      FGFR1 overexpression is a putative intrinsic resistance mechanism to erlotinib treatment. FGF2 levels may influence the activity and resistance mediation by FGFR1, but serum levels of FGF2 are not determining for erlotinib response in EGFR mutated patients.

  • +

    P3.02 - Biology/Pathology (ID 620)

    • Event: WCLC 2017
    • Type: Poster Session with Presenters Present
    • Track: Biology/Pathology
    • Presentations: 1
    • +

      P3.02-050 - Mechanisms of Acquired Resistance to the ALK Inhibitor Lorlatinib in ALK-Rearranged NSCLC Cell Lines (ID 9787)

      09:30 - 09:30  |  Author(s): Kristine Raaby Jakobsen

      • Abstract

      Background:
      Lorlatinib is a potent third-generation ALK tyrosine kinase inhibitor, which has demonstrated promising efficacy in ALK-rearranged NSCLC patients and excellent preclinical activity against ALK resistance mutations. However, as with all targeted treatments, resistance inevitably emerges. The molecular mechanisms underlying lorlatinib resistance remain largely undescribed, except for the L1198F mutation in ALK that was presented in a case study involving a single patient. Thus, the present study is performed to discover novel mechanisms of resistance to lorlatinib treatment.

      Method:
      H3122 (EML4-ALK v1) and H2228 (EML4-ALK v3) NSCLC cell lines were treated with increasing doses of lorlatinib or the first-generation ALK inhibitor crizotinib (10 nM - 1 µM). The resulting cell lines were investigated using MTS viability assays with ALK inhibitors to confirm resistance. Targeted next generation sequencing (NGS) using the Oncomine Focus Panel (Thermo Fisher) will be used to examine potential ALK-dependent resistance mutations and/or bypass mechanisms. This panel detects variants in 52 different genes – 35 genes are investigated for hotspot mutations, 19 genes for focal CNV gains, and 23 genes for fusions. The ALK gene is covered by both the hotspot, CNV, and fusion analyses, thus ensuring that all types of ALK-dependent resistance mechanisms will be discovered.

      Result:
      The two cell lines were treated with either lorlatinib or crizotinib in triplicates resulting in 12 resistant cell lines. Resistance occurred following approximately 4-6 months of drug treatment and was confirmed using MTS assays. The lorlatinib-resistant H3122 cell lines were insensitive to both crizotinib and lorlatinib treatment. This indicates that the mechanism of resistance is of a novel character, as the known L1198F ALK mutation is sensitive to crizotinib treatment. Hence, we are currently performing NGS analyses of all 12 resistant cell lines in order to discover potential resistance mechanisms.

      Conclusion:
      Using the comprehensive NGS Oncomine Focus Panel, we will be able to discover novel resistance mechanisms to the ALK-targeted drug lorlatinib, which will provide new knowledge of the nature of acquired resistance. This will help determining which mechanisms of resistance to look for in lorlatinib-resistant patients in future investigations, ultimately improving the treatment of this subset of patients.