n-(n-(3-5-difluorophenacetyl)alanyl)phenylglycine-tert-butyl-ester and Lung-Neoplasms

Although pterostilbene (PTE) has been shown to have potent antitumor activities against various cancer types, the molecular mechanisms of these activities remain unclear. In this study, we investigated the antitumor activity of PTE against human lung adenocarcinoma in vitro and in vivo and explored the role of the Notch1 signaling pathway in this process. PTE treatment resulted in a dose- and time-dependent decrease in the viability of A549 cells. Additionally, PTE exhibited strong antitumor activity, as evidenced not only by a reduced mitochondrial membrane potential (MMP) and a decreased intracellular glutathione content but also by increases in the apoptotic index and the level of reactive oxygen species (ROS). Furthermore, PTE treatment induced the activation of the Notch1 Intracellular Domain (NICD) protein and activated Hes1. DAPT (a gamma secretase inhibitor) and Notch1 siRNA prevented the induction of NICD and Hes1 activation by PTE treatment and sensitized the cells to PTE treatment. The down-regulation of Notch signaling also prevented the activation of pro-survival pathways (most notably the PI3K/Akt pathway) after PTE treatment. In summary, lung adenocarcinoma cells may enhance Notch1 activation as a protective mechanism in response to PTE treatment. Combining a gamma secretase inhibitor with PTE treatment may represent a novel approach for treating lung adenocarcinoma by inhibiting the survival pathways of cancer cells.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Shape; Cell Survival; Dipeptides; Drug Resistance, Neoplasm; Drug Synergism; Glutathione; Humans; Lung Neoplasms; Male; Membrane Potential, Mitochondrial; Mice; Mice, Nude; Proto-Oncogene Proteins c-akt; Reactive Oxygen Species; Receptor, Notch1; Signal Transduction; Stilbenes; Tumor Burden; Xenograft Model Antitumor Assays

Acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) occurs in non-small cell lung cancer (NSCLC) patients who initially respond to TKI treatment but whose cancer then progresses. Recent studies have shown that Notch signal is associated with drug resistance. However, the exact mechanism of Notch during acquisition of resistance to EGFR-TKI in human lung cancer remains unclear. In the present study, we showed that the expression of Notch-1 was highly upregulated in EGFR-TKI acquired resistant lung cancer cells. More importantly, Notch-1 contributed to the acquisition of the epithelial-mesenchymal transition (EMT) phenotype, which was critically associated with acquired resistance to EGFR-TKI. Silencing of Notch-1 using siRNA resulted in mesenchymal-epithelial transition (MET), which was associated with impaired invasion and anchorage-independent growth of lung cancer and resensitisation to gefitinib in acquired resistant NSCLC cells. Finally, gefitinib treatment of Balb/c nu/nu with acquired resistant lung cancer xenografts in combination with Notch inhibitor N-[N-(3,5-difluorophenacetyl)-L-alanyl]-(S)-phenylglycine t-butyl ester (DAPT) resulted in effective tumour growth retardation, with decreased proliferative activity and increased apoptotic activity. Collectively, these data suggest that Notch-1 might play a novel role in acquired resistance to gefitinib, which could be reversed by inhibiting Notch-1.

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Proliferation; Dipeptides; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; ErbB Receptors; Female; Gefitinib; Humans; Lung Neoplasms; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Invasiveness; Protein Kinase Inhibitors; Quinazolines; Receptor, Notch1; RNA Interference; Signal Transduction; Time Factors; Transfection; Xenograft Model Antitumor Assays

Notch receptor signaling pathway (NRSP) is increasingly linked to carcinogenesis. Non-small cell lung cancer (NSCLC) appears to actively utilize this conserved developmental pathway. The aims of this study are to determine whether or not Notch 1-4 are overexpressed in NSCLC tissues compared with normal lung tissues and whether inhibiting NRSP could induce caspase-dependent or caspase-independent apoptosis. Immunohistochemistry was used to evaluate the expression of Notch 1-4 in 101 NSCLC tissue samples and 30 normal lung tissue samples. DAPT was used to repress NRSP in SK-MES-1 cells. Apoptosis was determined by Annexin V and PI staining. Cleaved poly ADP-ribose polymerase (PARP) was measured by Western blot; X-linked inhibitor of apoptosis protein (XIAP) and Survivin were assessed by qRT-PCR and Western blot; the release of second mitochondria-derived activator of caspase (Smac) from mitochondria to cytoplasm was evaluated by Western blot; the subcellular locations of endonuclease G (Endo G) and apoptosis inducing factor (AIF) were observed by Western blot and indirect immunofluorescence analysis. (Mech Dev, 98, 2000, 95) Notch 1-4 are up-regulated in NSCLC tissues and Notch 1, 2 are positively correlated with lymph node metastasis, (Proc Natl Acad Sci U S A, 106, 2009, 22293) DAPT treatment could inhibit NRSP and induce apoptosis, with a marked increase in cleaved PARP, decreases in XIAP and Survivin proteins and concomitant release of Smac, EndoG, and AIF from mitochondria, indicating that inhibiting NRSP by DAPT triggers caspase-dependent and caspase-independent apoptosis.

Topics: Aged; Amyloid Precursor Protein Secretases; Apoptosis; Apoptosis Regulatory Proteins; Carcinoma, Squamous Cell; Caspases; Cell Line, Tumor; Cytoplasm; Dipeptides; Down-Regulation; Endodeoxyribonucleases; Female; Humans; Inhibitor of Apoptosis Proteins; Intracellular Signaling Peptides and Proteins; Lung Neoplasms; Lymph Nodes; Lymphatic Metastasis; Male; Mitochondria; Mitochondrial Proteins; Poly(ADP-ribose) Polymerases; Receptors, Notch; Signal Transduction; Survivin; Up-Regulation; X-Linked Inhibitor of Apoptosis Protein

A disintegrin and metalloproteinase 10 (ADAM10) was identified as a key protease in the ectodomain shedding of various substrates, such as Notch1 protein, ErbB2 and E-cadherin, which are important in the development of non-small cell lung cancer (NSCLC). The aim of this study was to investi-gate the role of ADAM10 in NSCLC metastasis.We characterized the expression of ADAM10 and Notch1 in human NSCLC tissues in vivo. Immunohistochemical analysis indicated that ADAM10 expression was significantly increased in the NSCLC tissues, particularly in the metastatic tissues. Futhermore, ADAM10 overexpression positively correlated with Notch1 expression in the NSCLC tissues. The in vitro downregulation of ADAM10 expression using ADAM10 short hairpin RNA (shRNA) reduced the migration and invasion of NSCLC cells. We present further evidence that ADAM10 promotes NSCLC cell migration and invasion via the activation of the Notch1 signaling pathway. Taken together, our results suggest that ADAM10 may serve as a potential target for the therapeutic intervention of NSCLC metastasis. The data provided in this study may aid in the further understanding of the function of ADAM10 in the progression of NSCLC and open new perspectives for the diagnosis and treatment of NSCLC.

Topics: ADAM Proteins; ADAM10 Protein; Amyloid Precursor Protein Secretases; Carcinoma, Non-Small-Cell Lung; Cell Movement; Dipeptides; Female; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Male; Membrane Proteins; Middle Aged; Neoplasm Invasiveness; Neoplasm Metastasis; Receptor, Notch1; RNA Interference; RNA, Small Interfering; Signal Transduction