P38 MAPK activation, JNK inhibition, neoplastic growth inhibition, and increased gap junction communication in human lung carcinoma and Ras-transformed cells by 4-Phenyl-3-butenoic acid
Department of Pharmaceutical Sciences, College of Pharmacy and Health Sciences, Mercer University, Atlanta, Georgia 30341, USA.Journal of Cellular Biochemistry (Impact Factor: 3.26). 01/2012; 113(1):269-81. DOI: 10.1002/jcb.23353
Human lung neoplasms frequently express mutations that down-regulate expression of various tumor suppressor molecules, including mitogen-activated protein kinases such as p38 MAPK. Conversely, activation of p38 MAPK in tumor cells results in cancer cell cycle inhibition or apoptosis initiated by chemotherapeutic agents such as retinoids or cisplatin, and is therefore an attractive approach for experimental anti-tumor therapies. We now report that 4-phenyl-3-butenoic acid (PBA), an experimental compound that reverses the transformed phenotype at non-cytotoxic concentrations, activates p38 MAPK in tumorigenic cells at concentrations and treatment times that correlate with decreased cell growth and increased cell-cell communication. H2009 human lung carcinoma cells and ras-transformed rat liver epithelial cells treated with PBA showed increased activation of p38 MAPK and its downstream effectors which occurred after 4 h and lasted beyond 48 h. Untransformed plasmid control cells showed low activation of p38 MAPK compared to ras-transformed and H2009 carcinoma cells, which correlates with the reduced effect of PBA on untransformed cell growth. The p38 MAPK inhibitor, SB203580, negated PBA's activation of p38 MAPK downstream effectors. PBA also increased cell-cell communication and connexin 43 phosphorylation in ras-transformed cells, which were prevented by SB203580. In addition, PBA decreased activation of JNK, which is upregulated in many cancers. Taken together, these results suggest that PBA exerts its growth regulatory effect in tumorigenic cells by concomitant up-regulation of p38 MAPK activity, altered connexin 43 expression, and down-regulation of JNK activity. PBA may therefore be an effective therapeutic agent in human cancers that exhibit down-regulated p38 MAPK activity and/or activated JNK and altered cell-cell communication.
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- "Thus, it appears that in vitro-aged oocytes have selectively deactivated the MAPK14 stress pathway while increasing the MAPK8/9/10 pathway. Intriguingly, a similar phenomenon is seen in some proliferative human cancers that down-regulate MAPK14 and up-regulate MAPK8/9/10 (Matesic et al., 2012). It is clear that the JNK pathway could play several different roles in response to in vitro aging, the details of which have yet to be worked out. "
ABSTRACT: Post-ovulatory aging of oocytes results in the progressive loss of fertilization and developmental competence. This degradation of oocyte quality has been the object of numerous investigations, primarily focused on individual signaling pathways which provide limited insight into the status of global signaling events. The purpose of the present investigation was to comprehensively assess broad patterns of signaling pathway activity during in vitro aging as an initial step in defining control points that can be targeted to prevent the reduction in oocyte quality during prolonged culture. An antibody microarray-based phospho-proteome analysis performed on oocytes before and after eight hours of culture revealed significant changes in the abundance or activation state of 43 proteins that function in a wide variety of protein kinase-mediated signaling pathways. Several of the most significantly affected kinases were studied by Western blot and confocal immunofluorescence to corroborate the array results. Prolonged culture resulted in global changes in the abundance and activity of protein kinases that regulate the response to calcium, stress, and cell-cycle control. Examination of intracellular structures revealed a previously unrecognized increase in the abundance of large autophogagic lysosomes, which correlates with changes in protein kinase pathways. These results provide insight into the stresses experienced by oocytes during culture and the diversity of responses that results from them. The observed increase in autophagy-related activity, together with the disruptions in calcium signaling, cell-cycle, and stress-response pathways, have the potential to negatively impact oocyte quality by interfering with the normal sequence of biochemical changes that constitute egg activation following fertilization. Mol. Reprod. Dev. 2014. © 2014 Wiley Periodicals, Inc.
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ABSTRACT: Summary BACKGROUND AND PURPOSE: The Maillard Reaction Products (MRPs) are known to be effective in chemoprevention. Here we focused on the anti-cancer mechanism of (E)-2, 4-bis (p-hydroxyphenyl)-2-butenal (MRP), in a concentrations (10-40 μg/ml) and time (30 min-72 h) dependent manner on the human non-small-cell lung cancer (NSCLC)cells (NCI-H460 and A549). EXPERIMENTAL APPROACH: Here we analyzed the activity by, western blot for major apoptotic protein, MAPK, NF-κB and death receptor expression, RT-PCR for death receptor mRNA expression, EMSA for NF-κB DNA binding activity and effect of inhibitors by colony formation assay. KEY RESULTS: Our studies show that this compound has a concentration- and time-dependent inhibitory effect on the growth of NSCLC cells due to induction of apoptosis. Concomitantly, there is also a significant increase in apoptotic proteins like cleaved caspase-3, cleaved caspase-9, Bax and p53 but down regulation of anti-apoptotic proteins like Bcl-2, cIAP1 and cIAP2. This activity is due to up regulation of mitogen-activated protein kinases (MAPK) and death receptor (DR) proteins like DR3, DR5 and DR6, but inactivation of nuclear transcriptional factor-κB (NF-κB). Among the all DR activated, only DR5 knock down with siRNA reversed the effect. Even though all MAPKinases are activated, only the pre-treatment of p38 MAPKinase inhibitor reversed (E)-2, 4-bis (p-hydroxyphenyl)-2-butenal-induced cell growth inhibition, cleaved caspase-3, -9 and DR5 expression, NF-κB inactivation. CONCLUSIONS AND IMPLICATIONS: Thus, these data show that p38 MAPK mediated NF-κB and DR5 signals are critical in the anti-cancer activity of (E)-2, 4-bis (p-hydroxyphenyl)-2-butenal.
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ABSTRACT: Cell-cell communication through gap junctions is aberrant or absent in a majority of human cancer cells, compared to cells in corresponding normal tissues. This and other evidence has led to the hypothesis that gap junction channels, comprised of connexin proteins, are important in growth control and cancer progression. The major goal of this ongoing study was to identify bioactive compounds that specifically upregulate gap junction channel-mediated cell-cell communication as potential anti-tumor therapies. Control of cell-cell communication is linked to growth regulatory intracellular signaling pathways; we therefore further aimed to identify signaling pathways modulated by these compounds in order to assess their potential as targeted anti-tumor therapies. Compounds were screened for their ability to upregulate gap junction-mediated cell-cell communication by using a fluorescent dye transfer assay to measure cell-cell communication between tumor promoter-treated astroglial cells or ras-transformed epithelial cells. Western blotting using connexinspecific and phosphorylation site-specific antibodies was used to monitor phosphorylation changes in signaling pathway proteins. Our results identified three compounds that upregulate gap junction-mediated cell-cell communication in our screening assays, chaetoglobosin K(ChK), 4-phenyl-3-butenoic acid (PBA) and the methyl ester of PBA (PBA-Me). Further analyses demonstrated that in tumorigenic cells, ChK downregulates phosphorylation of Akt kinase, an enzyme in the PI3-kinase signaling pathway that is found to be upregulated in a number of human cancers, on a key activation site. However, ChK did not inhibit PI-3 kinase in vitro as did the classic PI-3 kinase inhibitor, Wortmannin. PBA and PBA-Me were found to upregulate phosphorylation of p38 MAPK on a key activation site in tumorigenic cells, which is downregulated in several human cancer cell types. ChK and PBA also decreased activation of SAPK/JNK, another kinase found to be upregulated in a number of human cancers. These studies highlight the potential of monitoring gap junction intercellular communication for identifying experimental anti-tumor compounds.
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