Initial experience with factor-Xa inhibition in percutaneous coronary intervention: the XaNADU-PCI Pilot.

BACKGROUND: Direct factor (F)Xa inhibition is an attractive method to limit thrombotic complications during percutaneous coronary intervention (PCI). OBJECTIVES: To investigate drug levels achieved, effect on coagulation markers, and preliminary efficacy and safety of several doses of DX-9065a, an intravenous, small molecule, direct, reversible FXa inhibitor during PCI. PATIENTS AND METHODS: Patients undergoing elective, native-vessel PCI (n = 175) were randomized 4 : 1 to open-label DX-9065a or heparin in one of four sequential stages. DX-9065a regimens in stages I-III were designed to achieve concentrations of > 100 ng mL-1, > 75 ng mL-1, and > 150 ng mL-1. Stage IV used the stage III regimen but included patients recently given heparin. RESULTS: At 15 min median (minimum) DX-9065a plasma levels were 192 (176), 122 (117), 334 (221), and 429 (231) ng mL-1 in stages I-IV, respectively. Median whole-blood international normalized ratios (INRs) were 2.6 (interquartile range 2.5, 2.7), 1.9 (1.8, 2.0), 3.2 (3.0, 4.1), and 3.8 (3.4, 4.6), and anti-FXa levels were 0.36 (0.32, 0.38), 0.33 (0.26, 0.39), 0.45 (0.41, 0.51), and 0.62 (0.52, 0.65) U mL-1, respectively. Stage II enrollment was stopped (n = 7) after one serious thrombotic event. Ischemic and bleeding events were rare and, in this small population, showed no clear relation to DX-9065a dose. CONCLUSIONS: Elective PCI is feasible using a direct FXa inhibitor for anticoagulation. Predictable plasma drug levels can be rapidly obtained with double-bolus and infusion DX-9065a dosing. Monitoring of DX-9065a may be possible using whole-blood INR. Direct FXa inhibition is a novel and potentially promising approach to anticoagulation during PCI that deserves further study.

Synthesis of phenoxyacetic acid derivatives as highly potent antagonists of gastrin/cholecystokinin-B receptors. III.

In order to improve the biological characteristics of DA-3934 (5), a novel gastrin/cholecystokinin (CCK)-B receptor antagonist, phenoxyacetic acid derivatives replacing the N-methyl-N-phenylcarbamoylmethyl moiety of 5 with various alkyl chains have been synthesized and their biological activity evaluated. The relationship between the structure of these compounds and their human gastrin receptor binding affinity showed that there should be the optimal size among the various N-alkyl chains. Also a significant increase in the receptor binding affinity was achieved by several compounds. Among those compounds, 2-[3-[3- [N-cyclohexylmethyl-N-[2-(N-methyl- N-phenylcarbamoylmethoxy)phenyl]carbamoylmethyl]ureido]pheny l]acetic acid (22c) and (+/-)-2-[3-[3-[N-[2-(N-methyl-N- phenylcarbamoylmethoxy)phenyl]-N-(3-methylpentyl)carbamoy lmethyl]ureido] phenyl]acetic acid (22h) exhibited high affinity for human gastrin receptors and were also more potent inhibitors in a pentagastrin-induced gastric acid secretion model than the parent compound, 5. The ED50 values of these compounds when administered intraduodenally to rats were 0.12 and 0.63 mg/kg, respectively.

Synthesis of phenoxyacetic acid derivatives as highly potent antagonists of gastrin/cholecystokinin-B receptors. II.

A series of phenoxyacetanilide derivatives was synthesized and their antagonist activities for human gastrin/cholecystokinin (CCK)-B and CCK-A receptors were evaluated. Among the compounds synthesized, 2-[3-[3-[N-[2-(N-methyl-N-phenylcarbamoylmethoxy)phenyl]-N-(N-meth yl-N- phenylcarbamoylmethyl)carbamoylmethyl]-ureido]phenyl]acetic acid (20i, DA-3934) exhibited high affinity for gastrin/CCK-B receptors and high selectivity over CCK-A receptors. DA-3934 and its methyl ester derivative inhibited pentagastrin-induced gastric acid secretion in rats in a dose-dependent manner.

Synthesis and pharmacological activities of novel bicyclic thiazoline derivatives as hepatoprotective agents II. (7-Alkoxycarbonyl-2,3,5,6-tetrahydropyrrolo[2,1-b]thiazol-3-ylidend) acetamid derivatives.

A series of exomethylenic bicyclic thiazoline derivatives (3a--i) was synthesized and evaluated for hepatoprotective activity against galactosamine-induced and monoclonal antibody-induced acute liver injuries in rats. The structure-activity relationships were investigated. Among the compounds synthesized, N-methyl-(7-isopropoxy-carbonyl-6,6-dimethyl-2,3,5,6- tetrahydropyrrolo[2,1-b]thiazol-3-ylidene)acetamide (3i) exhibited the most potent hepatoprotective activity. This compound suppressed galactosamine-induced hepatic injury at 100 mg/kg by oral administration and further prevented monoclonal antibody-induced hepatic injury at 30 mg/kg by intraperitoneal injection, as judged from the changes in serum transaminase activities.

Structural determinants of the melanocortin peptides required for activation of melanocortin-3 and melanocortin-4 receptors.

The melanocortins are peptide products of proopiomelanocortin post-translational processing that, among other functions, are thought to influence cognition. Recently, we isolated genes encoding two human melanocortin receptors, the melanocortin-3 receptor (hMC3R) and the melanocortin-4 receptor (hMC4R), which are expressed primarily in brain. We undertook the present studies to examine the structural features of melanocortins that determine activation of these two receptors. For our studies we expressed the coding regions of the hMC3R and hMC4R genes in Hepa cells using the eukaryotic expression vector CMVneo and examined the generation of intracellular cyclic 3',5'-adenosine monophosphate in response to stimulation with various melanocortins. Our findings indicate that the core heptapeptide sequence common to most of the melanocortins (amino acids 4-10 of adrenocorticotropic hormone [ACTH]) is the primary determinant for activation of hMC3R but, in addition, tyrosine2 is necessary for maximal response. Activity of hMC4R is heavily dependent on proline12, but full activity also requires a contribution by tyrosine2. These findings may provide insight into the development of targeted ligands for the brain melanocortin receptors.

Synthesis and pharmacological activities of novel bicyclic thiazoline derivatives as hepatoprotective agents. I. 8-Ethoxycarbonyl-5,6-dihydrothiazolo[2,3-c][1,4]thiazine derivatives.

A series of bicyclic thiazoline derivatives (4a-s) was synthesized and evaluated for hepatoprotective activity against galactosamine-induced and monoclonal antibody-induced acute liver injuries in rats. The structure-activity relationships were investigated. Among the compounds synthesized, ethyl 3-(N-methylcarbamoyl)-5,6-dihydrothiazolo[2,3-c][1,4]thiazin e-8- carboxylate (4p) exhibited remarkable hepatoprotective activity and lower toxicity. This compound suppressed galactosamine-induced hepatic injury at 100 mg/kg by gavage and further prevented monoclonal antibody-induced hepatic injury at 30 mg/kg by intraperitoneal injection, as evaluated by measuring changes in serum transaminase activities.

Interaction of dual intracellular signaling pathways activated by the melanocortin-3 receptor.

We undertook these studies to explore the intracellular signaling mechanisms activated by a newly described human brain melanocortin receptor (hMC3R). Hepa cells transfected with the hMC3R gene responded to stimulation with alpha-melanocyte stimulation hormone (alpha-MSH) and adrenocorticotropic hormone (ACTH) with dose-dependent increases in cellular content of cyclic 3',5'-adenosine monophosphate (cAMP) reaching a maximum of over 1500% of control cells at the 10(-8) M dose (EC50 = 10(-11) M). In contrast, the production of [3H]inositol phosphates in cells prelabeled with myo-[2-3H]inositol exhibited a biphasic dose-response curve with increases as high as 155% of basal at 10(-11) M alpha-MSH or ACTH, but beyond that a dose-dependent decrease was observed. The inhibitory component of the dose-response curve could be abolished by pretreatment of transfected cells with the cAMP antagonist (Rp)-adenosine 3',5'-monophosphorothioate (Rp-cAMP) or the protein kinase A inhibitor H-89. Increases in intracellular calcium induced in transfected cells by alpha-MSH in doses ranging from 10(-11) to 10(-7) M could not be observed unless the cells were pretreated with H-89. By replacing the third intracytoplasmic loop of the canine H2-histamine receptor with that of hMC3R the biphasic characteristic of agonist-induced production of [3H]inositol phosphates was conferred to the chimeric receptor. These data indicate that the hMC3R is coupled to both cAMP and inositol phospholipid/Ca(2+)-mediated post-receptor signaling systems and that the latter response is regulated by protein kinase A activity.

Molecular cloning, expression, and characterization of a fifth melanocortin receptor.

We report the isolation of a gene encoding a novel member of the family of melanocortin receptors. The mouse melanocortin-5 receptor (mMC5R) responds to melanocortins with an increase in intracellular cyclic 3',5'-adenosine monophosphate (cAMP) concentrations. Stimulation of the mMC5R by the melanocortins revealed a hierarchy of potency in which alpha-melanocyte stimulating hormone (alpha-MSH) > beta-melanocyte stimulating hormone (beta-MSH) > adrenocorticotropic hormone (ACTH) > gamma- melanocyte stimulating hormone (gamma-MSH). Further structure-activity studies indicated that amino- and carboxyl-terminal portions of alpha-MSH appear to be key determinants in the activation of mMC5R whereas the melanocortin core heptapeptide sequence is devoid of pharmacological activity. Northern blot analysis demonstrated the expression of mMC5R mRNA in mouse skeletal muscle, lung, spleen, and brain.

Molecular cloning, expression, and gene localization of a fourth melanocortin receptor.

The recent cloning of three melanocortin receptors suggests an unexpected diversity in this family of seven transmembrane G-protein linked receptors. Herein, we report the cloning, expression, and gene localization of a fourth human melanocortin receptor, the melanocortin-4 receptor. By Northern blot analysis and in situ hybridization, this receptor is expressed primarily in the brain, but its expression is notably absent in the adrenal cortex, melanocytes, and placenta. Agonist stimulation of COS-1 cells transiently transfected and L-cells permanently transfected with the coding region of the cloned melanocortin-4 receptor leads to increases in intracellular cyclic 3',5'-adenosine monophosphate. The profile of the responses of the melanocortin-4 receptor to different melanocortins distinguishes it from melanocortin receptors previously described. Using the technique of fluorescent in situ hybridization, the gene encoding the melanocortin-4 receptor was localized to chromosome 18 (q21.3).

Molecular cloning of a novel melanocortin receptor.

Using the technique of the polymerase chain reaction primed with oligonucleotides based on the homologous transmembrane regions of seven transmembrane G protein-linked receptors, we isolated three full-length human genes that encode a novel subgroup of this receptor family. Recently, two of these receptors were identified as specific for alpha-melanocyte-stimulating hormone (alpha-MSH) and adrenocorticotropic hormone. We report the molecular cloning and pharmacologic characterization of a third member of this subgroup. The gene for this receptor encodes a protein of 361 amino acids in length. Its pharmacology characterizes it as an MSH receptor specific to the heptapeptide core common to adrenocorticotropic hormone and alpha-, beta-, and gamma-MSH. By Northern blot hybridization and polymerase chain reaction, it is expressed in brain, placental, and gut tissues but not in melanoma cells or in the adrenal gland. These findings may yield insight into the physiology of peptides derived from pro-opiomelanocortin post-translational processing.

Alleviation of acetylsalicylic acid-induced fetal toxicity by calcium.

Alleviation of aspirin-induced fetotoxicity by calcium was investigated in rats. ASA (500 mg/kg s.c.) decreased the plasma Ca level in pregnant rats and that in the feto-placenta units. 0.05 M CaCl2 given to dams as tap water on days 8 through 20 of gestation inhibited ASA-induced hypocalcemic effect in maternal plasma. Tap water of 0.002-0.05 M CaCl2, Ca(OH)2 and Ca-lactate on days 8 through 20 of gestation alleviated malformations elicited by the administration of ASA (500 mg/kg s.c.) on the 11th day of gestation resulting in a decrease in the fetotoxicity in rats.

Species differences in hypocalcemia induced by acetyl-salicylic acid.

The study was performed to explore species differences in hypocalcemia induced by acetylsalicylic acid (ASA). ASA decreased plasma and serum calcium (Ca) level at the dose of more than 200 mg/kg s.c. and p.o. in mouse, rat and guinea pig, and a correlation between serum Ca and salicylic (SA) levels was recognized in these species. However, hypocalcemia was not observed in rabbit and dog by the oral administration of ASA 400 mg/kg though high serum levels were found. Species differences were recognized in ASA-induced hypocalcemia. It was suggested that the differences were probably due to the different activity of calcitonin decreased serum Ca level in rabbit.