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J.J. Riehm
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P1.04 - Poster Session/ Biology, Pathology, and Molecular Testing (ID 233)
- Event: WCLC 2015
- Type: Poster
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 1
- Moderators:
- Coordinates: 9/07/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P1.04-051 - C. Elegans, an in Vivo Model for Lung Cancer: Effect of Chronic Exposure of Nicotine on Specific Mutants Relevant to Lung Cancer (ID 1552)
09:30 - 17:00 | Author(s): J.J. Riehm
- Abstract
Background:
There are a number of genetic abnormalities that can occur in lung cancer. We, and others, have shown that receptor tyrosine kinases (RTKs), such as EGFR, c-MET, RON, and Eph, frequently harbor gain-of-function mutations in addition to being overexpressed in lung cancer. We also have shown that the soil nematode C. elegans overexpressing a c-Met mutant revealed an abnormal vulval phenotype with hyperplasia. Interestingly, exposure to nicotine significantly aggravated the phenotype suggesting that C. elegans can be used as an in vivo model for rapid screening of RTK mutants as well as carcinogens.
Methods:
C. elegans strains vab-1 (Eph receptor), RB2088 (MET receptor), SD551 (temperature sensitive strain expressing constitutively active form of KRAS), and three sli-1 mutants PS2728, PS1258, and MT13032 (inactive, c-CBL) were compared to wild-type N2 worms for survival, fertility, egg-laying capacity, locomotion, and phenotypic changes in the absence or presence of nicotine. Gene expression analysis was also performed on each of the strains in the absence or presence of nicotine.
Results:
Nicotine treatment reduced the lifespan of worms for all strains (p=.0034). Nicotine treatment adversely affected egg-laying capacity of all strains, including N2 control, reducing egg production by 13% at 50μM nicotine and 31% at 500 μM nicotine. Furthermore, the fertility (the number of eggs laid/worm) was significantly reduced in SD551 mutant worms compared to N2 worms (p=.003). Overall locomotion velocity did not change with increasing concentration of nicotine except in MT13032, a c-Cbl mutant. Heat map analysis of gene expression profiling data clearly revealed that the various kinases and phosphatases in C. elegans that are marginally expressed in N2 worms were significantly enhanced upon chronic nicotine exposure. The expression of these genes was already elevated in SD551 and that was further increased in response to nicotine.
Conclusion:
Taken together, chronic nicotine exposure adversely affects various biological functions of C. elegans and these effects are exaggerated in the mutants. Interestingly, nicotine treatment also upregulates the expression of various kinases and phosphatases thereby strengthening our contention that the initial screening studies for the oncogenic mutants detected in humans can be rapidly carried out in C. elegans.
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P1.08 - Poster Session/ Thymoma, Mesothelioma and Other Thoracic Malignancies (ID 224)
- Event: WCLC 2015
- Type: Poster
- Track: Thymoma, Mesothelioma and Other Thoracic Malignancies
- Presentations: 1
- Moderators:
- Coordinates: 9/07/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P1.08-005 - Met and PI3K/mTOR as a Potential Combinatorial Therapeutic Target in Malignant Pleural Mesothelioma (ID 1700)
09:30 - 17:00 | Author(s): J.J. Riehm
- Abstract
Background:
There are a number of genetic alterations such as BAP1 and NF2 that can occur in malignant pleural mesothelioma (MPM). Various studies have shown that both MET and its downstream key intracellular signaling partners PI3K and mTOR are known to be overexpressed and frequently mutated in MPM. Here we have examined the therapeutic efficacy of a new generation small molecule inhibitor of MET receptor tyrosine kinase ARQ 197 and phosphatidylinositol 3-kinase and mTOR (PI3K/mTOR) inhibitors BEZ-235 and GDC-0980 in MPM.
Methods:
The mesothelioma cells were treated with ARQ 197, NVP-BEZ235, or GDC-0980 alone or in combination for 72 hours and cell proliferation was measured by using Alamar Blue assay. Synergistic efficacy was determined by isobologram and combination-index methods of Chou and Talalay. Signaling was assessed by immunoblotting. The mechanism of inhibition was further studied by using apoptosis assays and cell cycle analysis. Cell motility was studied by using scratch assays. We also examined efficacy of the combination of ARQ 197 and GDC-0980 on in vivo tumor growth by using mouse xenograft models.
Results:
MPM cell lines over-express MET and its active form p-MET, PI3K, and p-AKT and total AKT. ARQ 197, NVP-BEZ235, and GDC-0980, when used alone, significantly inhibited the cell proliferation of mesothelioma cells in a dose dependent manner. The combination of MET and PI3K/mTOR inhibitors was synergistic in suppressing MPM cell growth as compared to any single drug alone. Treatment of ARQ 197, NVP-BEZ235, and GDC-0980 alone or in combination inhibited the phosphorylation of AKT and S6 kinase in mesothelioma cells. MET and PI3K/mTOR inhibitors affect cell growth of mesothelioma cells by cell cycle inhibition (cyclin D1) and induction of apoptosis (presence of cleaved PARP, by IF/ confocal microscopy). MET inhibitor ARQ 197 alone inhibits the cell motility of mesothelioma cells in scratch assay. The combination of ARQ 197/ GDC-0980 was much more effective than each single agent alone in inhibiting the tumor growth of mesothelioma xenografts in nude mice. Compared to the control mice (2946±403 mm[3]), the tumors of mice treated with ARQ 197(2262±317 mm[3]) and GDC-0980 (1631±229.57mm[3]) alone had a significant decrease in the tumor volume. The tumor volume of mice treated with the combination of ARQ 197 and GDC-0980 further decreased it to six fold (475±97.43 mm[3]) compared to the control mice.
Conclusion:
Our results suggest that the combined use of ARQ 197/NVP-BEZ235 and ARQ 197/GDC-0980 is far more effective than single drug use in suppressing MPM cell motility and growth in vitro and tumor growth in vivo and therefore merits further translational studies.
