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S. Chatterjee
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P2.04 - Poster Session/ Biology, Pathology, and Molecular Testing (ID 234)
- Event: WCLC 2015
- Type: Poster
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 1
- Moderators:
- Coordinates: 9/08/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P2.04-101 - Ganetespib Resistance in KRAS Mutant NSCLC Is Mediated through Reactivation of the RAF/MEK/ERK and PI3K/MTOR Pathways (ID 1626)
09:30 - 09:30 | Author(s): S. Chatterjee
- Abstract
Background:
One third of all malignancies and approximately 25% of non-small cell lung cancer (NSCLC) patients have KRAS mutations which leads to the activation of several growth regulatory signaling pathways including RAF/MEK/ERK and PI3K/AKT/MTOR. Unfortunately, there are no current therapies targeting this critical oncogene. Heat shock protein 90 (HSP90) is a molecular chaperone required for the stability of ‘client’ oncoproteins, many of which are effectors of KRAS. Unfortunately, limited efficacy was observed in early clinical studies of single agent HSP90 inhibitors (HSP90i) in KRAS mutant NSCLC. Here, we examined the mechanism(s) of acquired resistance to ganetespib, a Phase 3 HSP90i, in KRAS mutant NSCLC to develop rationale combinations with ganetespib.
Methods:
Growth inhibition was determined through the colony formation and MTS assays. Ganetespib resistant (GR)-KRAS mutant NSCLC cell lines were derived to identify resistance mechanism(s). Flow-cytometry was performed to generate cell-cycle profiles. Genetic (shRNA) and pharmacologic inhibition of candidate mediators of resistance was performed.
Results:
Ganetespib was cytotoxic in a panel of KRAS mutant NSCLC cell lines and decreased expression and activity of both RAF/MEK/ERK and PI3K/AKT/MTOR pathways. In order to identify the mechanisms of ganetespib resistance in KRAS mutant NSCLC, we derived three KRAS mutant NSCLC ganetespib resistant (GR) cell lines. GR cells were cross-resistant to a first generation HSP90i, 17-AAG, suggesting that altered metabolism of ganetespib is unlikely to explain this resistance. Moreover, the ganetespib-induced G~2~/M checkpoint arrest observed in A549 parental cells was significantly diminished in A549-GR cells. These results suggest that bypass of this checkpoint may contribute to the observed ganetespib resistance. Furthermore, we demonstrated that GR cells were cross-resistant to docetaxel, an anti-microtubule agent. In addition, expression and activity of the PI3K/AKT/MTOR pathway members as well as the RAF/MEK/ERK pathway members were significantly increased suggesting that reactivation of these pathways may be responsible for the observed resistance. To test this hypothesis, we treated parental and GR cells with inhibitors of the PI3K/AKT/MTOR pathway (dual PI3K/mTOR inhibitor, BEZ235 and PI3K inhibitor PX866) or the RAF/MEK/ERF pathway (ERK inhibitor, SCH772984). Remarkably, GR cells were more sensitive to these inhibitors compared to the parental ones suggesting that the acquired ganetespib resistance lead to increased dependence on the both RAF/MEK/ERK and PI3K/MTOR pathways. Interestingly, the expression/activity of the key ERK and PDK1 substrate and activator of the PI3K/MTOR pathway, p90 ribosomal S6 kinase (RSK) was strikingly increased in the GR cells. Since RSK has been implicated as a key mediator of crosstalk between these two pathways, as well as in promoting G2/M progression, we examine the effect of genetic (shRNA) or pharmacologic (BI-D1870 and SL0101) inhibition of RSK in two GR cell lines. Remarkably, the GR cells showed increased dependency on RSK activity compared to the parental cell lines.
Conclusion:
These data suggests that the combination of inhibitors for HSP90 and PI3K/mTOR or a RSK inhibitor may prevent ganetespib resistance and/or help overcome the resistance after single agent treatment, providing the preclinical rationale for our planned Phase I/II trial of the combination of ganetespib and a dual PI3K/MTOR inhibitor in KRAS mutant NSCLC.
