inositol-1-4-5-trisphosphate and Lung-Neoplasms

Platelets are increasingly recognized for their contributions to tumor metastasis. Here, we show that the phosphoinositide signaling modulated by phosphatidylinositol transfer protein type α (PITPα), a protein which shuttles phosphatidylinositol between organelles, is essential for platelet-mediated tumor metastasis. PITPα-deficient platelets have reduced intracellular pools of phosphoinositides and an 80% reduction in IP

Topics: Animals; Anticoagulants; Blood Platelets; Fibrin; Gene Deletion; Hemostasis; Hyperplasia; Immunity, Mucosal; Inositol 1,4,5-Trisphosphate; Integrases; Lung Neoplasms; Lymphoid Tissue; Male; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Phospholipid Transfer Proteins; Platelet Aggregation; Signal Transduction; Thrombin; Thrombosis

As ectopic expression of the neuronal inositol-1,4,5-trisphosphate-3-kinase A (InsP3Kinase) in tumor cells increases the metastatic potential, InsP3Kinase is an interesting target for tumor therapy. Recently, we have identified a membrane-permeable InsP3Kinase inhibitor (BAMB-4) exhibiting an IC50-value of 20 μM. Here we characterized a new InsP3Kinase inhibitor which shows a 130-fold lower IC50 value (157 ± 57 nM) as compared to BAMB-4. We demonstrate that this nitrophenolic compound, BIP-4, is non-competitive to ATP but competitive to InsP3, thus exhibits a high selectivity for inhibition of InsP3Kinase activity. Docking analysis suggested a putative binding mode of this molecule into the InsP3Kinase active site. Determination of cellular uptake in lung cancer cells (H1299) revealed that 6% of extracellular BIP-4 is internalized by non-endosomal uptake, showing that BIP-4 is not trapped inside endo/lysosomes but is available to inhibit cellular InsP3Kinase activity. Interestingly, we found that BIP-4 mediated inhibition of InsP3Kinase activity in the two lung cancer cell lines H1299 and LN4323 inhibited proliferation and adhesion at IC50 values of 3 μM or 2 μM, respectively. InsP3Kinase inhibition did not alter ATP-induced calcium signals but significantly reduced the level of Ins(1,3,4,5,6)P5. From these data we conclude that the inhibitory effect of BIP-4 on proliferation and adhesion of lung cancer cells does not result from alterations of calcium but from alterations of inositol phosphate signals. In summary, we reveal that inhibition of cellular InsP3Kinase by BIP-4 impairs proliferation and adhesion and therefore BIP-4 might be a promising compound to reduce the metastatic potential of lung carcinoma cells.

Topics: Antineoplastic Agents; Apoptosis; Benzamides; Benzoxazoles; Calcium Signaling; Cell Adhesion; Cell Line, Tumor; Cell Proliferation; Drug Screening Assays, Antitumor; High-Throughput Screening Assays; Humans; Inositol 1,4,5-Trisphosphate; Lung Neoplasms; Molecular Docking Simulation; Naphthalimides; Phosphotransferases (Alcohol Group Acceptor); Pyrazoles

Recent studies have identified novel actions for 2-aminoethoxydiphenyl borate (2-APB) in triggering calcium release and enhancing calcium influx induced by the depletion of intracellular calcium stores. In this study, we have examined the effects of 2-APB on the human lung adenocarcinoma A549 cell line, which we have previously shown displays a unique calcium influx response, when ER calcium stores are depleted by thapsigargin (TG) treatment. Here, we show that low concentrations of 2-APB failed to induce the rapid augmentation of TG-activated calcium influx previously reported for other cell types. We observed that store-operated calcium (SOC) channels in the A549 cell line exhibited short-term sensitivity to low doses of 2-APB, perhaps reflecting a delayed augmentation of SOC channel activity or the recruitment of 2-APB-insensitive SOC channels. In both intact and permeabilized cells, 2-APB effectively discharged a subset of A549 calcium pools corresponding to the hormone-sensitive intracellular calcium stores. The 2-APB-induced calcium release produced a long-lasting perturbation of the adenosine triphosphate (ATP)-releasable calcium pools, effectively uncoupling ATP-activated calcium release even, when stores are replenished with calcium. In contrast to previous reports, we found that disruption of either the actin or microtubule-based cytoskeleton failed to block the 2-APB-induced effects on calcium signaling in A549 cells. Our study describes novel cytoskeletal-independent effects of 2-APB on Ca2+-signaling pathways, revealing differentially sensitive Ca2+-influx pathways and long-term perturbation of hormone-sensitive Ca2+ stores.

Topics: Actins; Adenocarcinoma; Adenosine Triphosphate; Antineoplastic Agents, Phytogenic; Boron Compounds; Calcium; Calcium Channels; Calcium Signaling; Cell Line, Tumor; Cytochalasin D; Cytoskeleton; Digitonin; Dose-Response Relationship, Drug; Enzyme Inhibitors; Green Fluorescent Proteins; Humans; Indicators and Reagents; Inositol 1,4,5-Trisphosphate; Ionomycin; Lung Neoplasms; Microscopy, Confocal; Nucleic Acid Synthesis Inhibitors; Paclitaxel; Thapsigargin; Time Factors

Gelsolin expression is frequently downregulated in lung cancer and several types of different human cancers. To examine the effects of gelsolin restoration on tumorigenicity, we here stably expressed various levels of gelsolin via gene transfer in lung cancer cells (squamous cell carcinoma line, PC10). We observed the alterations in tumorigenicity in vivo when implanted in nude mice, and the changes in growth properties in vitro. As compared to parental cells and control clones, gelsolin transfectants highly reduced tumorigenicity and repressed cell proliferation. Moreover, we investigated bradykinin-induced responses in gelsolin-overexpressing clones, because agonist-stimulated activation of the phospholipases C (PLC)/protein kinase C (PKC) signal transduction pathway is critical for cell growth and tumorigenicity. Bradykinin promotes phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis by PLC and translocation of various PKC isoforms from the cytosolic fraction to the particulate fraction. Bradykinin treatment did not increase inositoltriphosphate (IP3) production and induce the membrane fractions of PKC alpha and PKC gamma in gelsolin tranfectants, while it induced PIP2 hydrolysis and increased the fractions in parental and control clones. These results suggest that gelsolin suppressed the activation of PKCs involved in phospholipid signalling pathways, inhibiting cell proliferation and tumorigenicity.

Topics: Animals; Apoptosis; Bradykinin; Cell Division; Enzyme Activation; Gelsolin; Humans; Inositol 1,4,5-Trisphosphate; Isoenzymes; Lung Neoplasms; Mice; Mice, Nude; Models, Biological; Neoplasm Transplantation; Protein Kinase C; Protein Transport; Time Factors; Transfection; Tumor Cells, Cultured

The intracellular pathway of polymeric immunoglobulin receptor (pIgR) is governed by multiple signals that lead to constitutive transcytosis. In addition, in transfected polarized MDCK cells, polymeric immunoglobulin A (pIgA) binding stimulates rabbit pIgR-transcytosis, owing to phospholipase-C gamma 1 activation and increase of intracellular calcium. Transcytosis of rat pIgR across hepatocytes is similarly accelerated by pIgA injection. In contrast we show here that human Madrin-Darby Canine Kidney (pIgR)-transcytosis, in human Calu-3 and human pIgR-transfected MDCK cells, is not promoted by pIgA, as monitored by a continuous apical release of its secreted ectodomain. However, the incubation of cells expressing human or rabbit pIgR with pIgA induces a comparable IP3 production, and pIgR-transcytosis of either species is accelerated by the protein kinase C (PKC)-activator phorbol myristate acetate. Without pIgA, mimicking phospholipase-C activation by combining low concentrations of phorbol myristate acetate with ionomycin, or high concentrations of ionomycin alone, stimulates the rabbit, but not the human, pIgR transcytosis. These data suggest that the species difference in pIgA-induced pIgR-transcytosis does not stem from the defective production of second messengers, but from a different sensitivity of pIgR to intracellular calcium. Our results outline the danger of extrapolating to humans the abundant data obtained from mucosal vaccination of laboratory animals.

Topics: Adenocarcinoma; Animals; Calcium Signaling; Cell Line; DNA, Complementary; Dogs; Enzyme Activation; Humans; Immunoglobulin A; Inositol 1,4,5-Trisphosphate; Ionomycin; Ionophores; Kidney Tubules, Proximal; Lung Neoplasms; Protein Kinase C; Protein Transport; Rabbits; Rats; Receptors, Polymeric Immunoglobulin; Recombinant Fusion Proteins; Second Messenger Systems; Signal Transduction; Species Specificity; Tetradecanoylphorbol Acetate; Transfection; Tumor Cells, Cultured; Vaccination

The signal transduction involved in growth regulation of cells originally from a human giant cell lung carcinoma(PG) was investigated. The purinergic receptor agonists, ATP and its analogues, as well as bombesin all played a significant growth regulatory role on PG cells. ATP showed a growth inhibitory effect while bombesin showed a growth stimulatory effect on PG cells in vitro. Further investigation showed that ATP and bombesin activated phosphatidylinositol turnover/calcium mobilization signal transduction pathways in PG cells. and increased production of inositol-(1,4,5) trisphosphate and mobilization of intracellular free calcium, in which ATP and bombesin effects were dose-dependent. The purinergic receptor on PG cells was characterized by P2y subtype according to the order of PG cell responses to a series of ATP analogues. Pretreatment with cholera toxin showed different effects on the functions of ATP and bombesin, suggesting that the major difference between ATP and bombesin on signal transduction pathways may be at the G protein level.

Topics: Adenosine Triphosphate; Bombesin; Calcium; Carcinoma; Cell Division; Cholera Toxin; Humans; Inositol 1,4,5-Trisphosphate; Lung Neoplasms; Phosphatidylinositols; Signal Transduction; Tumor Cells, Cultured