Virtual Library
Start Your Search
F. Lennon
Author of
-
+
MINI 21 - Novel Targets (ID 133)
- Event: WCLC 2015
- Type: Mini Oral
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 1
- Moderators:B.P. Levy, D.S. Tan
- Coordinates: 9/08/2015, 16:45 - 18:15, Mile High Ballroom 2a-3b
-
+
MINI21.06 - Role of the Focal Adhesion Protein Paxillin in Lung Cancer - From Genetic Alterations to Novel Mitochondrial Functionality (ID 2188)
16:45 - 18:15 | Author(s): F. Lennon
- Abstract
- Presentation
Background:
Cytoskeletal and focal adhesion abnormalities are observed in several types of cancer including lung cancer, which is attributed to a greater number of deaths than prostate, breast and colorectal cancers combined. Paxillin is a 68 kDa protein that is an integral part of the focal adhesion and acts as an adaptor molecule. We initially cloned the gene for paxillin, and localized it to chromosome 12q24. We have previously reported that paxillin can be mutated (approximately 8%), amplified (5-7%), and/or overexpressed in almost 80% of lung cancer patient samples. Paxillin protein is upregulated in more advanced stages of lung cancer compared with earlier stages and is a prognostic factor for non-small cell lung cancer (NSCLC). Paxillin gene is amplified in some pre-neoplastic lung lesions as well as neoplastic lesions. We identified 22 different variants of paxillin mutation in our initial investigation especially between the LD and the LIM domains (Jagadeeswaran et al. 2008). There are mutations that have been validated in the TCGA set. We selected six mutants to perform further studies ((P52L, A127T, P233L, T255I, D399N, and P487L as well as wild-type as control). Our investigations focused on an effort to understand the contribution of molecular abnormalities found in paxillin and their relationship to mitochondrial functionality.
Methods:
HEK293 cells as well as a paxillin null NSCLC cell line H522 was used to overexpress the above paxillin mutants and wild-type paxillin. Live cell confocal microscopy was performed to evaluate cell motility, immunoprecipitation to determine interaction with other proteins, and gene expression analysis was performed to evaluate effects on gene expression.
Results:
Among the mutations we investigated, we found that the most common paxillin mutant A127T in lung cancer cells enhanced cell proliferation, focal adhesion formation and co-localized with the anti-apoptotic protein B cell CLL/Lymphoma 2 (BCL-2), which among other sites also localizes to the mitochondria. We further found that when these variant clones of activating mutations were expressed in HEK293 cells, they conferred phenotypic changes resembling neoplastic cells. In gene chip microarrays assay investigating gene expression modulation conferred by these mutations in these same HEK293 cells, we found that P52L, A127T, T255I, P233L and D399N mutations, compared to wild-type paxillin, indeed modulated the expression of a significant number of genes. In particular, there were a number of mitochondrial signature proteins that were altered in the various mutants. Analyzing mitochondrial functions by measuring the interaction of these mutants with mitochondrial proteins MFN2, and DRP1, we identified that they alter mitochondrial dynamics, with significant fission rather than fusion. Paxillin also translocated from the focal adhesion to the mitochondrial membrane. In relationship to cisplatin responsiveness, PXN and mutant overexpression lead to cisplatin resistance.
Conclusion:
These data suggest that wild-type and mutant paxillin variants play a prominent role in neoplastic changes with direct implications in lung cancer progression and hence, its potential as a therapeutic target needs to be explored further.
Only Members that have purchased this event or have registered via an access code will be able to view this content. To view this presentation, please login, select "Add to Cart" and proceed to checkout. If you would like to become a member of IASLC, please click here.
-
+
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)
-
+
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): F. Lennon
- 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.
