Analysis of the potassium channel openers celikalim, pinacidil and cromakalim in platelet models of thrombosis.

The antihypertensive agents pinacidil, cromakalim and celikalim lower blood pressure by opening potassium channels in vascular smooth muscle. The role of these compounds in inhibiting human platelet aggregation and preventing white thrombus formation in a rabbit arteriovenous shunt model was examined. None of these agents (100 microM), substantially inhibited platelet aggregation induced by epinephrine or arachidonic acid. Only celikalim (100 microM) inhibited collagen (45%), ADP (56%), or serotonin (61%) induced platelet aggregation and ADP- (41%) or epinephrine-potentiated (61%) serotonin-induced platelet aggregation. Celikalim inhibited white thrombus formation at i.v. doses of 0.25 mg/kg (46% inhibition) but not 0.1 mg/kg; (14.6%); equihypotensive doses of pinacidil (0.5 mg/kg; 21.7%) and cromakalim (0.2 mg/kg, 7.5%; 0.4 mg/kg, 33%) were less effective. Glyburide (i.v. dose of 0.5 mg/kg) inhibited the antithrombotic activity of celikalim and to a lesser extent cromakalim. The greater antithrombotic activity of celikalim in vivo may be related to beneficial effects on blood rheology and reduced red blood cell deformability.

Comparative antithrombotic activities of the phosphodiesterase inhibitors pelrinone (AY-26,768), AY-31,390 and milrinone.

The phosphodiesterase (PDE) inhibitors AY-31,390, milrinone and pelrinone (AY-28,768) were analyzed in human platelet aggregatory systems and in a rabbit arteriovenous shunt model to delineate their activity. AY-31,390 showed a remarkably potent capacity to inhibit human antithrombotic platelet aggregation. AY-31,390 inhibited arachidonic acid, U46619, collagen, epinephrine (second phase) and adenosine diphosphate (second phase) induced platelet aggregation (PA) with IC50 values of 0.18, 0.21, 0.54, 0.43 and 0.20 microM, respectively. Milrinone, although less potent than AY-31,390, inhibited PA with IC50 values of 2.1, 2.0, 5.4, 3.7 and 4.1 microM and pelrinone's IC50 values were 2.8, 6.6, 13.3, 18.6 and 11.8 microM, respectively. Platelets which were incubated with AY-31,390, milrinone or pelrinone, washed with Hanks' balanced salt solution and then resuspended in platelet poor plasma, lost their inhibitory activity in collagen and arachidonic acid PA systems. These results suggested that AY-31,390, milrinone and pelrinone did not bind tightly to cAMP PDE. If human platelet-rich plasma was pretreated with adenosine deaminase, an enzyme that degrades adenosine, the inhibitory effect of milrinone and to a lesser extent pelrinone was reversed. AY-31,390 did not produce a loss of activity with adenosine deaminase in the arachidonic acid system and only a small loss in the collagen system. Adenosine did not appear to be a meaningful factor in AY-31,390's inhibitory activity. Pelrinone, milrinone to a greater extent, and AY-31,390 to the greatest extent were effective inhibitors of white thrombus formation in the in vivo rabbit arteriovenous shunt model. These PDE III inhibitors were potent deterrants of platelet aggregation and white thrombus formation; these agents would be expected to be efficacious therapeutic antithrombotics.

Immunomodulating activity of 3-(p-chlorophenyl)thiazolo-[3,2-a] benzimidazole-2-acetic acid (Wy-18,251, NSC 310633) in mice implanted with Lewis lung tumor. Effect of Wy-18,251 on lymphocyte proliferation and phagocytosis.

Proliferation of B lymphocytes is depressed in Lewis lung tumor bearing mice. Treatment of these mice with Wy-18,251 (5 mg/kg) significantly increased the responsiveness of their splenic T cells to concanavalin A in cell culture. Levamisole (5 mg/kg) acted more weakly than Wy-18,251. Neither Wy-18,251 nor levamisole elevated the depressed B cell mitogenesis. Wy-18,251 significantly increased macrophage phagocytosis against 51chromium labeled opsonized chicken red blood cells. Levamisole behaved differently: it either had no effect on phagocytosis, or depressed it.

Immunomodulating and antimetastatic activity of 3-(p-chlorophenyl) thiazolo[3,2-a]benzimidazole-2-acetic acid (Wy-18,251, NSC 310633).

The antimetastatic activity of a thiazolobenzimidazole, Wy-18,251 was investigated using various dosage regimens in mice with implanted Lewis lung tumors. The low doses, 1 and 5 mg/kg (i.p.) given 24 hours after implantation with two subsequent doses at 1 week intervals were most effective in reducing lung metastases. Delayed hypersensitivity reactions in guinea pigs were significantly increased by oral doses of 50 and 150 mg/kg. In normal rats, peripheral blood lymphocytes determined by rosette assay, were significantly increased by oral doses of 50 to 150 mg/kg of Wy-18,251 and in a more sensitive assay using anti-theta serum the lymphocyte levels were increased by doses of 7.5 and 10 mg/kg. When cultured T lymphocytes from CBA/J mouse spleens were incubated with a suboptimal concentration of Concanavalin A, [3H]thymidine uptake was significantly increased in the presence of 0.05 to 1.0 microgram per culture. These results suggest that Wy-18,251 may have potential therapeutic value as an antimetastatic agent through its stimulation of the cellular immune system.

Anti-tumour and anti-metastatic activity of 3-(P-Chlorophenyl)-2,3-Dihydro-3-Hydroxythiazolo (3,2-A)-Benzimidazole-2-acetic acid (WY-13,876).

Extensive investigation of 3-(p-chlorophenyl)-2,3-dihydro-3-hydroxythiazolo(3,2-alpha)-benzimidazole-2-acetic acid (Wy-13,876) in BDF1 mice implanted with Lewis lung tumour has shown that it is an effective anti-tumour and anti-metastatic agent. In vitro examination using HEp-2 human epidermal tumour cells has indicated that Wy-13,876 is not cytotoxic. When mice implanted with Lewis lung tumour and treated with Wy-13,876 are also injected with anti-thymocyte serum, an increase in lung metastases is observed suggesting that thymocyte activity is involved in the drug's mechanism of action. An increase in peripheral T lymphocytes observed in rats 18 h after a single oral dose of Wy-13,876 further supports this possibility. When Wy-13,876 is given to tumour -bearing mice in combination with low, ineffective doses of 5-fluorouracil or cyclophosphamide, further reduction of primary tumour growth is observed.