nitroarginine and Pancreatic-Neoplasms

Both nitric oxide (NO) and vascular endothelial growth factor (VEGF) mediate tumor vascular function. Because these molecules regulate one another's expression, we hypothesized that NO synthase (NOS) inhibition produces effects comparable to those of anti-VEGF therapy on human pancreatic cancer xenografts.. L3.6pl human pancreatic cancer cells were s.c. implanted in nude mice. On day 6, mice were randomized to receive (a) PBS (control), (b) DC101 [VEGF receptor 2 (VEGFR-2) antibody] by i.p. injection, (c) N-nitro-l-arginine (NNLA; NOS inhibitor) in the drinking water, or (d) both DC101 and NNLA. Mice were killed on day 20.. DC101 and NNLA as single agents inhibited tumor growth by approximately 50% to 60% (P < 0.008 for both). Furthermore, combined therapy inhibited mean tumor growth by 89% (P < 0.008). Combined inhibition of VEGFR-2 and NOS also decreased mean vessel counts by 65% (P < 0.03) and vessel area by 80% versus controls (P < 0.001). In contrast to DC101 where vessel diameter was similar to control, NNLA decreased mean vessel diameter by 42% (P < 0.001). NNLA also led to a 54% (P < 0.03) decrease in tumor uptake of the perfusion marker Hoechst 33342 versus controls whereas DC101 decreased Hoechst 33342 staining by 43% (P < 0.03). The combination of inhibitors decreased perfusion by 73% (P < 0.03).. Although VEGFR-2 can mediate NOS activity, the combination of VEGFR-2 and NOS inhibition significantly increased the antivascular effect over single agent therapy. The addition of NOS inhibition led to an even further alteration of tumor vessel morphology and vascular perfusion compared with VEGFR-2 blockade, suggesting that NO and VEGFR-2 have distinct but complementary effects on the tumor vasculature.

Topics: Animals; Antibodies, Monoclonal; Apoptosis; Blood Vessels; Cell Line, Tumor; Cell Proliferation; Enzyme Inhibitors; Humans; Immunohistochemistry; Male; Mice; Mice, Nude; Neovascularization, Pathologic; Nitric Oxide Synthase; Nitroarginine; Pancreatic Neoplasms; Platelet Endothelial Cell Adhesion Molecule-1; Random Allocation; Vascular Endothelial Growth Factor Receptor-2; Xenograft Model Antitumor Assays

The effects of L-arginine and its analogues NG-nitro-L-arginine, NG-methyl-L-arginine, L-homoarginine and D-arginine on cytosolic calcium concentration were investigated to characterise the mechanisms of arginine-induced stimulation and to determine if nitric oxide production played a role in this stimulation. NIT-1 cells, a transgenic beta-cell line, were used for this purpose since they release insulin in response to typical beta-cell stimuli. Our data demonstrate that the arginine-induced increase in cytosolic calcium concentration was completely dependent on the influx of extracellular Ca2+ via verapamil-sensitive voltage-activated Ca2+ channels and that arginine stimulation requires the presence of a nutrient in order to cause an increase in cytosolic calcium concentration. The nutrient likely acted by closing the K+ ATP channels, since its effect could be inhibited by activation of these channels with diazoxide. L-arginine, as well as nitro-arginine and methyl-arginine which are not substrates for the production of nitric oxide, caused similar increases in cytosolic calcium concentration. Non-metabolisable arginine analogues homoarginine and D-arginine also caused increases in the cytosolic calcium concentration although not to the same extent. Insulin secretion was enhanced to the same extent by all analogues of arginine. It can be concluded that the arginine-induced increase in cytosolic calcium concentration in NIT-1 cells is attributable to an electrogenic effect following the transport of arginine leading to depolarisation of the plasma membrane potential, although metabolism of the amino acid itself may also partially contribute to the response.

Topics: Animals; Arginine; Calcium; Cell Line; Cytosol; Diazoxide; Homoarginine; Insulin; Insulin Secretion; Insulinoma; Kinetics; Mice; Mice, Transgenic; Molsidomine; Nitric Oxide; Nitroarginine; omega-N-Methylarginine; Pancreatic Neoplasms; Potassium Channels; Potassium Chloride; Tumor Cells, Cultured; Verapamil