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C. Codony-Servat
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Optimizing targeted therapy in lung cancer (ID 56)
- Event: ELCC 2018
- Type: Poster Discussion session
- Track:
- Presentations: 1
- Moderators:F. Cappuzzo, N. Peled
- Coordinates: 4/12/2018, 16:45 - 17:45, Room W
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3PD - Co-targeting PIM1 or Src to overcome the limits of single MET inhibition (ID 333)
16:45 - 17:45 | Author(s): C. Codony-Servat
- Abstract
Background:
Single MET inhibition has controversial effects in MET addicted tumors. Proviral integration site for Moloney murine leukemia virus-1 (PIM1) is activated upon MET inhibition in MET addicted cells. PIM1 drives the expression of receptor tyrosine kinases (RTKs). SHP2, a non-receptor protein tyrosine phosphatase, is central in RTKs signaling and in Src activation. We have shown that the overexpression of RTKs like AXL and the transmembrane protein CUB domain-containing protein-1 (CDCP1) as well as Src activation, are mechanisms of intrinsic resistance to EGFR inhibition in EGFR mutant lung cancer. We are now testing whether RTKs, SHP2 or CDCP1 expression and activation are driven by PIM1 or Src and cause resistance to MET inhibitors in MET addicted tumors.
Methods:
We studied the inhibitory effect of the class I MET inhibitors tepotinib and savolitinib, the pan-PIM inhibitor AZD1208, and the Src inhibitor dasatinib in five MET addicted cell lines: 2 MET amplified lung cancer cells (EBC1 and H1993), 2 MET exon 14 mutant cells (Hs746T and H596) and a glioma cell line that carries the still not well recognized MET exon 7–8 splicing variant (E98). We assessed the effect of combined MET and PIM or MET and Src inhibition in cell viability and protein immunoblotting.
Results:
All the cell lines were sensitive to single MET inhibition (except H596) and resistant to single AZD1208 or dasatinib. The combination of savolitinib or tepotinib with AZD1208 was synergistic in the EBC1 cell line and slightly synergistic or additive in the H1993, Hs746T and E98 cell lines. The combination of savolitinib or tepotinib with dasatinib was highly synergistic in all four cell lines. The treatment of EBC1 cells with tepotinib monotherapy was not able to inhibit AXL activation while it induced the activation of SHP2 and CDCP1. AXL, CDCP1 and SHP2 expression or activation were downregulated when tepotinib was combined with dasatinib.
Conclusions:
Co-targeting PIM or Src with MET can be more effective than MET inhibition alone in MET addicted cell lines. Overexpression and activation of RTKs, CDCP1 and SHP2 can be mechanisms of resistance to single MET inhibition. The investigation of combinatorial strategies in MET addicted tumors, merits further investigation.
Clinical trial identification:
Legal entity responsible for the study:
IGTP – Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
Funding:
Fundació Obra Social “La Caixa”
Disclosure:
All authors have declared no conflicts of interest.
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Poster Display session (Friday) (ID 65)
- Event: ELCC 2018
- Type: Poster Display session
- Track:
- Presentations: 3
- Moderators:
- Coordinates: 4/13/2018, 12:30 - 13:00, Hall 1
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24P - Proviral integration site for Moloney murine leukemia virus-1 (PIM-1) inhibition with AZD1208 to prevent resistance to osimertinib in EGFR mutant NSCLC (ID 399)
12:30 - 13:00 | Author(s): C. Codony-Servat
- Abstract
Background:
Although treatment with EGFR tyrosine kinase inhibitors (TKIs) is initially effective in a subgroup of lung cancer patients, therapy resistance ultimately occurs. Resistance mechanisms consist of overexpression and co-expression of receptor tyrosine kinases (RTKs) and overexpression of STAT3. PIM1 was shown to be an important regulator of both RTKs and STAT3 expression, and inhibition of PIM1 with the pan-PIM inhibitor, AZD1208, might prevent STAT3 and RTK up-regulation after TKI treatment. This research focuses on combining TKI treatment with AZD1208 to prevent therapy resistance in different EGFR mutant NSCLC cell lines.
Methods:
Using TaqMan quantitative-PCR we studied basal mRNA expression levels of PIM1 and PIM3 in 5 EGFR-mutant NSCLC cell lines. Cells were then exposed to single osimertinib or AZD1208 treatment, or the combination, to determine the combination index (CI) and changes in cell viability. Moreover, changes in protein expression of different RTKs, STAT3 and downstream effectors of STAT3 were studied in 2 cell lines using western blotting.
Results:
The PC9 and H1975 cell lines were shown to have high PIM1 and STAT3 expression compared to the other EGFR mutant NSCLC cell lines. Combined osimertinib and AZD1208 treatment showed moderate synergism in all cell lines, with CIs ranging from 0.75 to 0.86 in triplicate experiments. Western blot experiments indicate that osimertinib treatment leads to upregulation of pCDCP1, pYAP1, pPaxillin and pSTAT3 in the PC9 and H1975 cell lines, suggesting initiation of resistance to single osimertinib treatment. In contrast, single AZD1208 treatment does not induce, or even lower expression of these proteins compared to baseline levels. When combining both treatments, the osimertinib-induced pSTAT3 up-regulation can be prevented with AZD1208. The effect of AZD1208 on RTK expression will be further explored.
Conclusions:
Single TKI treatment in EGFR mutant NSCLC induces expression of RTKs and STAT3, ultimately leading to therapy resistance. Inhibition of PIM1 with AZD1208 can abolish the osimertinib-induced phosphorylation of STAT3, and thereby prevents activation of resistance pathways.
Clinical trial identification:
Legal entity responsible for the study:
IGTP, Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
Funding:
Fundació Obra Social “La Caixa”
Disclosure:
All authors have declared no conflicts of interest.
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25P - A non-pathway-specific approach in EGFR and KRAS mutant or squamous cell histology non-small cell lung cancer (NSCLC) (ID 551)
12:30 - 13:00 | Author(s): C. Codony-Servat
- Abstract
Background:
p21-activated kinase 1 (PAK1) stimulates growth and metastasis in several types of tumors, including NSCLC. Protein kinase C iota (PKCi) is an enzyme highly expressed in NSCLC that regulates PAK1 signaling. We have previously shown that cancer pathway-specific intervention, like EGFR inhibitors in EGFR mutant NSCLC, results in parallel compensatory activation of other pathways, including the receptor tyrosine kinases AXL and MET, the transmembrane protein CUB domain-containing protein-1 (CDCP1) or the transcriptional regulators STAT3 and YAP1. We have now explored whether a non-pathway-specific approach can be efficient in three subsets of NSCLC.
Methods:
Three lung cancer cell lines were used: HCC827 and H23 lung adenocarcinoma cells that carry EGFR and KRAS mutations respectively, and H520 PAK1 amplified squamous NSCLC cells. Cell viability assays and western blotting were applied to evaluate the effect of IPA-3 (PAK1 inhibitor) plus auranofin (PKCi inhibitor). We used a Chou-Talalay modified method for drug combination studies that offers quantitative definition for additive effect (combination index [CI] = 1), synergism (CI < 1), and antagonism (CI > 1).
Results:
We found a differential PAK1 expression or activation profile in the three models, with H520 cells being the ones with the highest PAK1 expression and activation. IPA-3 plus auranofin was highly synergistic in HCC827, H23 and H520 cells with CIs of less than 0.4. In the EGFR mutant HCC827 cell line, IPA-3 plus afatinib or osimertinib was additive (CI = 0.9), or slightly synergistic (CI = 0.8). In the same cell line, the combination of IPA-3 plus auranofin abrogated EGFR and downstream signaling (ERK, AKT, STAT3, YAP1) and inhibited the expression and activation of AXL, MET and CDCP1. IPA-3 plus auranofin was, similarly, highly synergistic in H23 (CI = 0.3) and H520 (CI = 0.3) cells.
Conclusions:
For the first time, we are reporting that a non-pathway specific combination can be effective in EGFR and KRAS mutant as well as squamous NSCLC. The combination could control the counter-regulatory pathways that are made apparent and ultimately cause resistance when a pathway specific-intervention is applied.
Clinical trial identification:
Legal entity responsible for the study:
IGTP, Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
Funding:
International Association for the Study of Lung Cancer (IASLC)
Disclosure:
All authors have declared no conflicts of interest.
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58P - Co-treatment of trametinib (MEK inhibitor) with TPX-0005 (Src/FAK/JAK2 inhibitor) synergistic in KRAS mutant NSCLC cell lines and CDCP1 acts as a biomarker in KRAS mutant patients (p) (ID 407)
12:30 - 13:00 | Presenting Author(s): C. Codony-Servat
- Abstract
Background:
KRAS mutant lung adenocarcinoma has a dismal prognosis. In the current study, we identified combination targets for trametinib, a MEK inhibitor, which acts downstream of KRAS to suppress signaling through MAPK cascade. However, we have previously shown that selumetinib (MEK inhibitor) in KRAS mutant NSCLC cells caused a rebound of ERK, AKT and STAT3, as well as YAP phosphorylation (Y357), NOTCH3 and activation of RTKs, AXL and MET. We hypothesize that, in KRAS mutant NSCLC, suppression of MAPK could lead to activation of Src-YAP1-AXL-MET. Since CUB domain-containing protein 1 (CDCP1) activates Src, we consider that CDCP1 could be a biomarker of Src activation.
Methods:
Cell viability assay, colony formation assay and western blotting were performed to assess the treatment of trametinib plus TPX-0005 in a panel of 8 NSCLC cell lines: A549, Calu6, H23, H460, H2009, H2030, H441, H727. Synergy between trametinib and TPX-0005 was assessed via the Chou-Talalay method to estimate the combination index (CI). CI values <0.7 were considered synergistic, with decreasing CI values indicating greater synergy. We examined tumor samples of 32 KRAS mutant NSCLC p for CDCP1 mRNA levels.
Results:
The combination of trametinib with TPX-0005 was synergistic or additive in all cell lines tested. H23 and Calu6 were the most synergistic, followed by H441 and H2030. In the majority of cell lines examined, the colony formation assay resulted in an almost complete abrogation of tumor cell colonies, particularly in the H23 and A549. Western blotting indicated that the combination of trametinib with TPX-0005 abolished the phosphorylation of STAT3 (Y705), paxillin (Y118), a readout of FAK, and Src (Y416). The median PFS of 32 KRAS mutant NSCLC p was 2.5 months and the overall survival was 13.4 months. According to CDCP1 levels, the median PFS was 3.5 months for those with low CDCP1 and 1.4 months for those with high CDCP1 (P = 0.012). The median survival was 16.3 months for p with low CDCP1, and only 3.2 months for those with high CDCP1 (P = 0.023).
Conclusions:
The combination of trametinib plus TPX-0005 shows significant in-vitro activity in the majority of KRAS mutant NSCLC cell lines and the mRNA levels of CDCP1 could be a biomarker in KRAS mutant NSCLC p, indicating the activation of Src-YAP signaling. Clinical trials with the combination of MEK inhibitors with TPX-0005 are warranted.
Clinical trial identification:
Legal entity responsible for the study:
IGTP, Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
Funding:
Fundació Obra Social “La Caixa”
Disclosure:
All authors have declared no conflicts of interest.
