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Cell, Tumor and Stem Cell Biology |
1 Department of Medicine, Division of Pulmonary, Allergy and Critical Care Medicine; 2 Department of Hematology and Oncology, Winship Cancer Institute, Emory University School of Medicine; and 3 Atlanta Veterans Affairs Medical Center, Atlanta, Georgia
Requests for reprints: ShouWei Han, Division of Pulmonary, Allergy and Critical Care Medicine, Emory University School of Medicine, Whitehead Bioresearch Building, 615 Michael Street, Suite 205-M, Atlanta, GA 30322. Phone: 4047122661; Fax: 4047122151; E-mail: shan2{at}emory.edu.
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Abstract |
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), both known to down-regulate mTOR. Rapamycin, an inhibitor of mTOR, blocked the fibronectin-induced phosphorylation of p70S6K and 4E-BP1. Akt small interfering RNA (siRNA) and an antibody against the fibronectin-binding integrin 5ß1 also blocked the p70S6K phosphorylation in response to fibronectin. In contrast, an inhibitor of extracellular signal-regulated kinase 1/2 (PD98095) had no effect on fibronectin-induced phosphorylation of p70S6K. Moreover, the combination of rapamycin and siRNA for Akt blocked fibronectin-induced cell proliferation. Taken together, these observations suggest that fibronectin-induced stimulation of NSCLC cell proliferation requires activation of the Akt/mTOR/p70S6K pathway and is associated with inhibition of LKB1/AMPK signaling. (Cancer Res 2006; 66(1): 315-23) |
Introduction |
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5ß1, induction of cyclooxygenase-2 (COX-2), and inhibition of cyclin-dependent kinase inhibitor p21Waf1/Cip1 gene expression (7). These effects were associated with activation of several kinase signaling pathways, including extracellular signal-regulated kinase 1/2 (ERK1/2) and Rho kinase (7, 8). Importantly, these effects are not shared by other matrix components, such as type 1 collagen. More recently, fibronectin was shown to stimulate the phosphatidylinositol 3-kinase (PI3K), which is capable of controlling the expression of cyclin D and related genes involved in cell cycle control (9, 10). The above information suggests that fibronectin promotes lung tumor growth/survival and resistance to therapy and could represent a novel target for the development of new anticancer drugs. This prompted our interest in elucidating the mechanisms by which fibronectin stimulates human lung carcinoma cell growth and led us to investigate the role of the Akt/mammalian target of rapamycin (mTOR)/p70S6K signaling pathway in this process. Members of the mTOR subfamily belong to the PI3K-related kinase family and are inhibited by rapamycin, a feature that has facilitated efforts to study their functions in eukaryotic cells (11, 12). The mTOR serine/threonine kinases play an important role in the translational control of gene expression during cell growth and the overall rate of protein synthesis in response to nutrients and other environmental factors (12, 13). mTOR is also a key intermediary in multiple mitogenic signaling pathways and plays a central role in modulating cellular proliferation and angiogenesis in normal tissues and neoplastic processes (13, 14). The primary pathway by which growth factors and cytokines activate mTOR and its downstream targets seems to be the PI3K/Akt pathway. Accordingly, targeting the Akt/PI3K pathway with mTOR antagonists may increase the therapeutic efficacy of cancer therapy (14).
The signals that up-regulate mTOR activity are frequently activated in human cancers. In mammals, the two best-characterized targets of mTOR are the ribosomal S6 kinases (S6K1 and S6K2) and the eukaryotic initiation factor 4E (eIF4E)–binding protein 1 (4E-BP1). mTOR activity leads to S6K1/2 phosphorylation and activation and to 4E-BP1 phosphorylation and release from the cap-dependent translation initiation factor eIF4E. These two events, likely combined with other mTOR targets, lead to an increase in ribosomal biogenesis and the selective translation of specific mRNA populations (11–15). This ability to increase the protein synthesis capacity of the cell is responsible, at least in part, for the ability of the mTOR proteins to drive cell growth and proliferation (15) and their implication in lung cancer progression (16).
Herein, we report that fibronectin stimulates the phosphorylation of Akt and inhibits LKB1 and AMP-activated protein kinase (AMPK) in NSCLC, thereby promoting activation of mTOR and its downstream targets, such as p70S6K. This results in stimulation of NSCLC cell proliferation in vitro.
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Materials and Methods |
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, phosphorylated AMPK (p-AMPK, Thr172), p70S6K, p-p70S6K (Thr421/Ser424), Akt, p-Akt (Ser473), phosphatase and tensin homologue deleted on chromosome 10 (PTEN), 4E-BP1, and p-4E-BP1(Thr37/46) were purchased from Cell Signaling (Beverly, MA). Fibronectin derived from human fibroblasts was purchased from Sigma-Aldrich (St. Louis, MO). The mouse monoclonal anti-human 5ß1 integrin (JBS5) and anti-human 2ß1 integrin (AB1944) antibodies were purchased from Chemicon International, Inc. (Temecula, CA). The LightCycler/FastStart DNA Master SYBR Green I kit and the 5' DNA Terminus Labeling System were purchased from Roche Molecular Biochemicals (Indianapolis, IN). All reverse transcription-PCR (RT-PCR) kit components were obtained from Perkin-Elmer Co. (Foster City, CA). All other chemicals were purchased from Sigma (St. Louis, MO), unless otherwise indicated.
Western blot analysis. The procedure was done as previously described (17). Protein concentrations were determined by the Bio-Rad (Richmond, VA) protein assay. Equal amounts of protein from whole-cell lysates were solubilized in 2x SDS/sample buffer and separated on SDS/10% polyacrylamide gels. Blots were incubated with rabbit antibodies raised against mouse LKB1, AMPK, p70S6K, 4E-BP1, Akt, and their phosphorylated forms (1:1,000). The blots were washed and followed by incubation with a secondary goat antibody raised against rabbit IgG conjugated to horseradish peroxidase (1:2,000, Cell Signaling). The blots were washed, transferred to freshly made enhanced chemiluminescence solution (Amersham, Arlington Heights, IL) for 1 minute, and exposed to X-ray film. In controls, the primary antibodies were omitted or replaced with a control rabbit IgG.
RT-PCR. Total RNA was prepared from cells using TRI Reagent (Sigma) according to the manufacturer's instructions. The LKB1 (280 bp) and the glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 200 bp) cDNA fragments were amplified using primers synthesized by Sigma-Genosys (The Woodlands, TX): LKB1 forward, 5'-ACATCTGGTCGGCTG-3'; LKB1 reverse, 5'-GGTCTGTGGTTCCTG-3'; GAPDH forward, 5'-CCATGGAGAAGGCTGGGG-3'; and GAPDH reverse, 5'-CAAAGTTGTCATGGATGACC-3' according to published data (12). RT-PCR was carried out as previously described (7). Briefly, samples were denatured at 95°C for 30 seconds followed by 32 PCR cycles with temperatures as follows: 95°C for 30 seconds, 60°C for 30 seconds, and 72°C for 30 seconds. PCR products were subject to a final extension incubation of 7 minutes at 72°C. PCR products were analyzed on a 1% agarose gel containing 0.2 µg/µL ethidium bromide and visualized under a UV light.
Real-time RT-PCR. This procedure was described previously (7). The primers for LKB1 and GAPDH were the same as those described above for RT-PCR. Final results were expressed as the percentage of LKB1 mRNA in treated cells compared with that in untreated control cells. LKB1 mRNA expression was normalized to endogenous GAPDH mRNA. All PCR reactions were done using the LightCycler/FastStart DNA Master SYBR Green I kit in the Cepheid Smart-Cycler real-time PCR cycler (Sunnyvale, CA; ref. 7). Experiments were done in triplicate for each data point.
Treatment with Akt small interfering RNA. The Akt small interfering RNA (siRNA) duplex target sequences were synthesized by Sigma-Genosys based on published data (18). The Akt target sequence was 5'-CAGCAUGAGGUUCUCCAGCTT-3'. Nonspecific control siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). For the transfection procedure, cells were grown to 50% confluence, and Akt or control siRNAs were transfected using the LipofectAMINE 2000 reagent (Invitrogen) according to the manufacturer's instructions. Briefly, oligofectamine reagent was incubated with serum-free medium for 10 minutes. Subsequently, a mixture of siRNA was added. After incubation for 15 minutes at room temperature, the mixture was diluted with medium and added to each well. The final concentration of siRNA in each well was 100 nmol/L. After culturing for 30 hours, cells were washed, resuspended in new culture media in the presence or absence of fibronectin, and/or incubated with 1 µCi/mL [methyl-3H]thymidine (Amersham; specific activity, 250 Ci/mmol) for an additional 24 hours for Western blot or [methyl-3H]thymidine incorporation assays.
[Methyl-3H]thymidine incorporation assay. Human NSCLC cells were incubated with 1 µCi/mL [methyl-3H]thymidine (Amersham; specific activity, 250 Ci/mmol) in the presence or absence of indicated reagents for up to 24 hours. The medium was removed, and the attached cells were washed with 1x PBS. Afterwards, the attached cells were treated with ice-cold 6% trichloroacetic acid (TCA) at 4°C for 20 minutes and washed once with 6% TCA. The cells were then solubilized with 0.1 N NaOH and counted in a liquid scintillation counter in 4 mL of scintillation fluid.
Statistical analysis. All experiments were repeated a minimum of three times. Data collected from Western blot, RT-PCR, and [methyl-3H]thymidine incorporation assay were expressed as means ± SD. The data presented in some figures are from a representative experiment, which was qualitatively similar in the replicate experiments. Statistical significance was determined with Student's t test (two tailed) comparison between two groups of data sets. Asterisks indicate significant differences of experimental groups compared with the corresponding control condition (P < 0.05, see figure legends).
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Results |
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protein in a dose-dependent and time-dependent manner, whereas it had no effect on the total content of AMPK protein as determined by Western blot analysis (Fig. 3A and B). We also examined whether inhibition of either PI3K/Akt or mitogen-activated protein/ERK kinase-1 (MEK-1)/ERK signaling would block the effect of fibronectin on AMPK protein expression. As shown in Fig. 3C, the effect of fibronectin on reduction of AMPK protein expression was unaffected by inhibitors of PI3K (wortmannin) or MEK-1/ERK (PD98095), suggesting that PI3K and ERK signaling pathways are not involved in this process. Similar effects were observed in H1838 cells (data not shown)
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5ß1. mTOR activation stimulates the phosphorylation of downstream kinases, such as p70S6K and 4E-BP1. Therefore, we examined the effects of fibronectin on p70S6Kand 4E-BP1 expression. As shown in Fig. 4A and B, H1792 cells exposed to the indicated concentrations of fibronectin showed increased phosphorylation of p70S6K protein. The stimulatory effect of fibronectin was dose and time dependent with optimal increases noted when 20 µg/mL of fibronectin were used for 24 hours; no effect was detected in the abundance of p70S6K total protein. Fibronectin also increased the phosphorylation 4E-BP1 (Thr37/46), another downstream target of mTOR, in a time-dependent and dose-dependent manner with maximal induction at 24 hours, whereas it had little effect on total 4E-BP1 protein (Fig. 4C and D). Similar results were obtained in an additional NSCLC cell line, H1838 (data not shown).
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5ß1 (7). Therefore, we tested the effects of blocking 5ß1 in our system. We found that antibodies against the integrin 5ß1 blocked fibronectin-induced p70S6K phosphorylation (Fig. 4E). In contrast, neither control IgG (data not shown) nor antibodies against integrin 2ß1 (a collagen-binding integrin) mimicked this effect (Fig. 4E). In separate experiments, we found that a specific inhibitor of ERK1/2 (PD98095) did not affect fibronectin-induced p70S6K phosphorylation (Fig. 4F). These results suggest that the effect of fibronectin on p70S6k protein activity is mediated, at least in part, through PI3K/Akt activation but not through ERK1/2 signaling. Similar effects were observed in H1838 cells (data not shown). Akt siRNA and rapamycin inhibit fibronectin-induced phosphorylation of p70S6K and 4E-BP1 and lung carcinoma cell proliferation. We next tested whether knockdown of Akt expression could prevent the stimulatory effect of fibronectin on p70S6K phosphorylation. To this end, we depleted Akt expression in cells using the small RNA interference method. As shown in Fig. 5A, Akt siRNA greatly diminished endogenous Akt protein production; no changes were noted in cells transfected with control siRNA. The Akt siRNA also blocked the stimulatory effects of fibronectin on p70S6K phosphorylation; the Akt siRNA alone or a control siRNA had no effect (Fig. 5B).
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Next, we investigated whether Akt played a role in lung carcinoma cell proliferation induced by fibronectin. H1792 cells were transfected with Akt siRNA duplexes. Afterwards, the cells were exposed to fibronectin for an additional 24 hours followed by evaluation of cell proliferation. As shown in Fig. 5E, Akt siRNA significantly reduced fibronectin-induced cell proliferation; the control siRNA had no effect. We next examined if blocking mTOR affects the stimulatory effect of fibronectin on NSCLC cell growth. The mTOR inhibitor, rapamycin, inhibited, at least in part, fibronectin-induced NSCLC cell growth; rapamycin alone showed a small inhibitory effect on cell growth when used for 48 hours. A higher dose of rapamycin had no further effect (data not shown). Furthermore, rapamycin enhanced the inhibitory effect of Akt siRNA on cell proliferation (Fig. 5E). Similar effects were observed in H1838 cells (data not shown).
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Discussion |
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, upstream down-regulators of mTOR. These events induce the activation of mTOR as highlighted by the phosphorylation of a downstream target of mTOR, p70S6K, and result in increased NSCLC proliferation.
The demonstration that fibronectin stimulates Akt phosphorylation supports a role for fibronectin in activating the PI3K signaling pathway. Activation of PI3K seems to play a key role in both malignant transformation and resistance to chemotherapy by perpetuating processes that positively influence cell survival, growth, migration, and angiogenesis. Aberrations of PI3K/Akt, which increase signaling through mTOR, are found in several types of malignancies, most notably prostate and breast cancers (12). Moreover, gene amplifications of these kinase signals have been identified in cervical, ovarian, gastric, pancreatic, head and neck, and NSCLCs (12, 23). Of note, it seems that the primary pathway by which most growth factors and cytokines activate mTOR and its downstream targets is the PI3K/Akt pathway. Consistent with this statement, PI3K inhibitors, wortmannin or LY294002, have been shown to inhibit mTOR in several cell systems (24, 25). For example, wortmannin reduced the epidermal growth factor (EGF)–stimulated phosphorylation of both Akt and p70S6K, whereas it had no effect on EGF-induced ERK phosphorylation in human trophoblast cells (26). LY294002 was shown to inhibit mTOR and block the human inducible nitric oxide synthase gene transcription in lung epithelial adenocarcinoma (A549) cells (25). As we found in NSCLCs (this report), fibronectin, through its integrin 5ß1, has been shown to activate the PI3K/Akt pathway in several other cell systems (27, 28). Recently, others have shown that wortmannin and LY294002 blocked fibronectin-induced activation of mTOR/S6K signaling in human arterial smooth muscle cells (29). We also found that fibronectin reduced the protein levels of PTEN. PTEN suppresses tumor cell growth by antagonizing protein tyrosine kinases, such as PI3K/Akt, and regulates tumor cell invasion and metastasis through interactions at focal adhesions (30). Loss of PTEN function results in Akt phosphorylation with subsequent stimulation of cell proliferation (30). PTEN heterozygous mice spontaneously develop neoplasia, associated with loss of the normal PTEN allele and increased activation of AKT/mTOR/P70S6K signaling (31). Therefore, up-regulation of mTOR can be related to loss of the tumor suppressor gene PTEN and activation of Akt, and both seem to be affected by fibronectin.
To test the role of Akt in fibronectin-induced NSCLC proliferation, we blocked Akt expression using Akt siRNA. The targeting of Akt by Akt siRNA inhibited basal NSCLC cell growth and significantly blocked fibronectin-induced cell proliferation suggesting a role for Akt activation in this process. This is particularly interesting in view of related work showing that knocking out Akt using specific siRNAs increases the sensitivity of cancer cells towards cisplatin and enhances programmed cell death (27).
We also examined the effects of fibronectin on the tumor suppressor gene LKB1, a serine/threonine kinase that is inactivated by mutation in the autosomal-dominant Peutz-Jeghers syndrome (20) as well as in some sporadic lung adenocarcinomas (24). As a major upstream kinase of AMPK, LKB1 directly phosphorylates Thr172 in the activation loop of AMPK, a modification that is absolutely required for AMPK catalytic activity (20). Activation of AMPK inhibits mTOR signaling and is associated with inhibition of cancer cell growth (22, 32). In cell culture–based systems, LKB1 modulation of mTOR is mediated through phosphorylation of AMPK in response to energy stress. AMPK has been shown to diminish protein synthesis by activating TSc2, leading to inhibition of mTOR and subsequent inhibition of p70S6K protein activity (20, 33). We found that fibronectin inhibited LKB1 protein expression as well as the phosphorylation of AMPK in NSCLC cells. Because this observation was associated with activation of p70S6k phosphorylation, we postulate that fibronectin stimulates mTOR through activation of PI3K and Akt phosphorylation, while concomitantly inhibiting LKB1/AMPK signals. In another work, we explored the connection between the mTOR downstream signal p70S6K and AMPK. We found that AICAR, a specific AMPK activator (34), abrogated not only the effect of fibronectin on reduction of AMPK but also the fibronectin-induced phosphorylation of p70S6K, suggesting a cross-talk between the AMPK and p70S6K in mediating the effect of fibronectin in stimulation of cell growth.4 In contrast with other studies, which showed cross-talk among Akt, MEK-1/ERK, and AMPK (32, 35, 36), we found that the effect of fibronectin on reduction of AMPK protein expression was unaffected by inhibitors of PI3K (wortmannin) or MEK-1/ERK (PD98095), suggesting that PI3K and ERK signaling pathways are not involved in this process.
We previously reported that fibronectin induced NSCLC proliferation through effects on its integrin receptor 5ß1 (7). This is consistent with data reported here showing that antibodies against 5ß1 blocked the stimulatory effect of fibronectin on p70S6K phosphorylation. In another work, we also reported that fibronectin/5ß1 binding results in ERK activation, and this is the reason we tested the role of ERK in this system (7). However, the MEK-1/ERK inhibitor PD98095 did not block fibronectin-induced p70S6K phosphorylation, suggesting the induction of an ERK-independent pathway. This is consistent with another work showing that the MEK-1/ERK inhibitor U0126 decreased cell migration and ERK phosphorylation but did not influence p70S6K phosphorylation in response to EGF in human trophoblast cells (25).
We assume that fibronectin stimulates mTOR based on the observed phosphorylation of the mTOR downstream targets p70S6K and 4E-BP1. p70S6K is a key regulator of cell size and growth that is modulated via both PI3K and mTOR. Overexpression of p70S6K has been found in many tumor cells and thus results in increased tumor size in vivo, whereas its blockade of p70S6K prevents cell proliferation (37–40). Interestingly, we also found increased phosphorylation of 4E-BP1 protein in cells exposed to fibronectin. elF4F consists of three polypeptides (elF4A, elf4G, and elF4E) and is responsible for recruiting ribosomes to mRNA. elF4E recognizes the mRNA 5'-cap structure (m7GpppN) and plays a pivotal role in control of translation initiation, which is the rate-limiting step in translation. Phosphorylation of 4E-BP1 results in the release of elF4E, and activation of elF4E is considered an essential component of the malignant phenotype in breast carcinoma (41). Furthermore, growth and survival factors positively regulate assembly of the eIF4F complex by promoting sequential phosphorylation of the 4E-BPs on six serine/threonine sites through multiple Ras-dependent protein kinase cascades (42).
We then tested the effects of the mTOR inhibitor rapamycin in our system. Rapamycin and its analogues are highly specific inhibitors of mTOR (12). Treatment with rapamycin inhibits EGF-induced phosphorylation and activation of p70S6K (43). Recently, Sullivan et al. found that fibronectin induced rapid activation of mTOR and p70S6K in human arterial smooth muscle cells, and rapamycin inhibited this effect (34). Consistent with this, we found that rapamycin blocked the fibronectin-induced phosphorylation of p70S6K and 4E-BP1, and these effects were associated with inhibition of fibronectin-induced NSCLC cell growth. Blockade of Akt with Akt siRNA enhanced the effect of rapamycin, suggesting that they participate in these pathways. This work and a recent study showing that dominant-negative Akt expression or inhibitors of PI3K/Akt are sufficient to increase the sensitivity of tumor cells to rapamycin show that disruption of the PI3K/Akt signaling pathway could greatly enhance the effectiveness of mTOR inhibitors (44). Therefore, therapeutic targeting of mTOR might be a more efficient strategy against malignancies driven by activation of Akt and/or PTEN mutations (45). Rapamycin and several of its analogues with more favorable pharmaceutical properties have shown impressive growth inhibitory effects against a broad range of human cancers in both preclinical and early clinical evaluations, including NSCLC (12).
Taken together, these findings show that fibronectin, by binding to its integrin receptor 5ß1, stimulates NSCLC proliferation through activation of the Akt/mTOR/p70S6K pathway, while inhibiting the PTEN and LKB1/AMPK signals. These studies raise new questions about the potential role of the lung extracellular matrix in lung cancer. Furthermore, they suggest that targeting these signaling pathways might aid in the development of novel agents with therapeutic potential for the prevention and treatment of human lung carcinoma. As inhibitors of mTOR and Akt are brought forward into the clinic, agents blocking fibronectin-mediated lung cancer signaling may provide intriguing complementary effects.
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Acknowledgments |
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The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
We thank Dr. Shi-Yong Sun (Department of Hematology/Oncology, Emory University, Atlanta, GA) for providing the H1792 NSCLC cell line.
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Footnotes |
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Received 7/ 6/05. Revised 9/30/05. Accepted 10/19/05.
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References |
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is regulated by mammalian target of rapamycin. J Biol Chem 2003;278:33637–44.5ß1. J Cell Physiol 2005;202:205–14.[CrossRef][Medline]5ß1 promotes survival of growth-arrested breast cancer cells: an in vitro paradigm for breast cancer dormancy in bone marrow. Cancer Res 2004;64:4514–22.This article has been cited by other articles:
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  J. A. Quinn, C. T. Graeber, A. R. Frackelton Jr., M. Kim, J. E. Schwarzbauer, and E. J. Filardo Coordinate Regulation of Estrogen-Mediated Fibronectin Matrix Assembly and Epidermal Growth Factor Receptor Transactivation by the G Protein-Coupled Receptor, GPR30 Mol. Endocrinol., July 1, 2009; 23(7): 1052 - 1064. [Abstract] [Full Text] [PDF]
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 S. P. Narayanan, A. I. Flores, F. Wang, and W. B. Macklin Akt Signals through the Mammalian Target of Rapamycin Pathway to Regulate CNS Myelination J. Neurosci., May 27, 2009; 29(21): 6860 - 6870. [Abstract] [Full Text] [PDF]
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 C.-X. Xu, D. Jere, H. Jin, S.-H. Chang, Y.-S. Chung, J.-Y. Shin, J.-E. Kim, S.-J. Park, Y.-H. Lee, C.-H. Chae, et al. Poly(ester amine)-mediated, Aerosol-delivered Akt1 Small Interfering RNA Suppresses Lung Tumorigenesis Am. J. Respir. Crit. Care Med., July 1, 2008; 178(1): 60 - 73. [Abstract] [Full Text] [PDF]
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 Y. Zheng, J. D. Ritzenthaler, J. Roman, and S. Han Nicotine Stimulates Human Lung Cancer Cell Growth by Inducing Fibronectin Expression Am. J. Respir. Cell Mol. Biol., December 1, 2007; 37(6): 681 - 690. [Abstract] [Full Text] [PDF]
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 N. Khan, F. Afaq, M.-H. Kweon, K. Kim, and H. Mukhtar Oral Consumption of Pomegranate Fruit Extract Inhibits Growth and Progression of Primary Lung Tumors in Mice Cancer Res., April 1, 2007; 67(7): 3475 - 3482. [Abstract] [Full Text] [PDF]
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 S. Han, J. D. Ritzenthaler, B. Wingerd, H. N. Rivera, and J. Roman Extracellular Matrix Fibronectin Increases Prostaglandin E2 Receptor Subtype EP4 in Lung Carcinoma Cells through Multiple Signaling Pathways: THE ROLE OF AP-2 J. Biol. Chem., March 16, 2007; 282(11): 7961 - 7972. [Abstract] [Full Text] [PDF]
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Y.-P. Sher, C.-C. Chou, R.-H. Chou, H.-M. Wu, W.-S. Wayne Chang, C.-H. Chen, P.-C. Yang, C.-W. Wu, C.-L. Yu, and K. Peck Human Kallikrein 8 Protease Confers a Favorable Clinical Outcome in Non-Small Cell Lung Cancer by Suppressing Tumor Cell Invasiveness Cancer Res., December 15, 2006; 66(24): 11763 - 11770. [Abstract] [Full Text] [PDF]
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S. Han, J. D. Ritzenthaler, S. V. Sitaraman, and J. Roman Fibronectin Increases Matrix Metalloproteinase 9 Expression through Activation of c-Fos via Extracellular-regulated Kinase and Phosphatidylinositol 3-Kinase Pathways in Human Lung Carcinoma Cells J. Biol. Chem., October 6, 2006; 281(40): 29614 - 29624. [Abstract] [Full Text] [PDF]
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 K. R. Laderoute, K. Amin, J. M. Calaoagan, M. Knapp, T. Le, J. Orduna, M. Foretz, and B. Viollet 5'-AMP-Activated Protein Kinase (AMPK) Is Induced by Low-Oxygen and Glucose Deprivation Conditions Found in Solid-Tumor Microenvironments. Mol. Cell. Biol., July 1, 2006; 26(14): 5336 - 5347. [Abstract] [Full Text] [PDF]
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