Thomas B. McClanahan

The antithrombotic effects of CI-1028, an orally bioavailable direct thrombin inhibitor, in a canine model of venous and arterial thrombosis.

Direct thrombin inhibitors represent a new class of drug that may offer a therapeutic alternative for the treatment and prevention of thrombembolic conditions, especially on the venous side of the systemic circulation. CI-1028 (PD 172524/LB30057) is a potent, highly selective inhibitor of thrombin that is orally bioavailable. The efficacy of this compound has been demonstrated in animal models in which intra-venous administration was used. The objective of this study was to evaluate the efficacy of CI-1028 after oral administration in a canine electrolytic injury model of venous and arterial thrombosis. CI-1028 was administered via oral gavage, and animals received either saline or 10, 15, 20, or 30 mg/kg of drug. Fifteen minutes later, the dogs were anesthetized and a femoral artery and vein were exposed and instrumented to induce electrolytic injury and thrombosis while continuously monitoring blood flow in the vessels. Maximum blood CI-1028 concentrations of 0.88±0.27, 1.8±0.3, 2.2±0.5, and 3.2±0.5 μg/mL were generally achieved 15 to 30 minutes after administering the compound in the 10-, 15-, 20-, and 30-mg/kg groups, respectively. Administration of CI-1028 increased the time to occlusion (TTO), the principal efficacy end point, in a dose-dependent manner in both arteries and veins. The TTO in the control group (n=8) averaged 66±11 minutes in the arteries and 69±6 minutes in the veins. In dogs treated with 10 mg/kg (n=8), the TTO was not significantly different from that of the control group. In the 15-mg/kg group (n=9) TTO averaged 140±27 minutes in the arteries (p=not significant) and 125±15 minutes (p<0.05) in the veins. In the 20-mg/kg group (n=8), TTO was significantly longer than controls in both types of vessels, averaging 168±30 minutes in the arteries (p=0.05) and 155±21 minutes (p<0.05) in the veins. Likewise, at 30 mg/kg (n=8) both the arterial (179±17 minutes) and venous (188±15 minutes) TTO was significantly prolonged compared with controls. Surgical blood loss and template bleeding times tended to increase in a dose-dependent manner but a statistically significant elevation was detected for template bleeding time only at the highest dose. Dramatic changes in thrombin time were detected, consistent with the CI-1028 mechanism of action. Virtually no changes were detected in prothrombin time. Maximum activated partial thromboplastin time (aPTT) and activated clotting time changes were detected approximately 30 minutes after dosing, and they were approximately twofold and fivefold baseline values, respectively, at the highest dose. In conclusion, these results demonstrate that CI-1028 provides dose-dependent antithrombotic efficacy after oral administration in a canine model of venous and arterial thrombosis.

Hemodynamic effects of intraatrial administration of deferoxamine or deferoxamine-pentafraction conjugate to conscious dogs.

Summary: Deferoxamine (DFX) is a specific Fe3+ chelator that is used to manage iron overload, and is being evaluated as an agent to reduce ischemic organ damage that involves iron-mediated -OH formation. However, high intravascular doses cause significant hemodynamic changes that may limit or counteract beneficial effects. We used conscious, closed-chest dogs to test the hypothesis that conjugating DFX to pentafraction, a high molecular weight fraction of pentastarch, could reduce such hemodynamic changes. We infused 50 mg/kg of body weight of native DFX, or an equivalent dose as DFX-pentafraction, intraatrially over 15 min. Within 10 min of starting the infusion, DFX increased heart rate from predrug values of 105 ± 11 (mean ± SEM; N = 9) to 158 ± 13 beats/min, and reduced left ventricular (LV) systolic pressure from 131 ± 3 to 99 ± 16 mm Hg, LV enddiastolic pressure from 12 ± 3 to 3 ± 3 mm Hg, and mean arterial pressure (MABP) from 101 ± 5 to 74 ± 13 mm Hg. In two dogs, MABP decreased to ≤35 mm Hg. These parameters returned to predrug values by 60 min after infusion. All of these changes were statistically significant (p<0.05). In contrast, infusing DFX-pentafraction (N=9) caused no significant cardiac or hemodynamic changes other than a transient and slight (approximately 7%) increase in systolic arterial pressures. This conjugate, which prolongs the plasma half-life and does not alter the iron-chelating activity of native DFX, eliminates many undesirable hemodynamic actions. It may be a useful therapeutic alternative to native DFX in some settings

Increase in experimental infarct size with digoxin in a canine model of myocardial ischemia-reperfusion injury

In the present study, dogs were pretreated with intravenous digoxin, 0.0125 mg/kg/day, for 6 to 7 consecutive days to achieve clinically relevant serum concentrations; untreated animals were used as control subjects. After pretreatment, nine digoxin-pretreated dogs and nine control dogs were anesthetized and subjected to a 60-minute occlusion of the left circumflex coronary artery, followed by 6 hours of reperfusion. Anatomic myocardial infarct size, expressed as a percentage of the areas at risk of infarction and as a percentage of the total left ventricle were: 20.2 +/- 3.3% control vs 35.4 +/- 6.2% digoxin-pretreated (p less than 0.05) and 8.6 +/- 1.3% control vs 14.7 +/- 2.5% digoxin-pretreated (p less than 0.05), respectively (2.04 +/- 0.37 ng/ml serum digoxin). Regional myocardial blood flow in the nonischemic and ischemic zones tended to be lower in digoxin-pretreated than in control animals at baseline testing and were significantly reduced in the anterior subendocardial sites of digoxin-pretreated dogs during ischemia and reperfusion. These data suggest that an exacerbation or enhancement of myocardial ischemia-reperfusion injury may occur in the presence of clinically observable serum digoxin concentrations.

Effects of endothelin-receptor antagonism with PD156707 on myocardial stunning in swine

Endothelin (ET) has been proposed to play a role in pathogenesis of myocardial ischemia/reperfusion injury. The potential role of ET in myocardial stunning has not been examined. Therefore we tested the hypothesis that selective blockade of ETA receptors with PD156707 {sodium 2-benzo[1,3]dioxol-5-yl-4-(4-methoxy-phenyl) -4-oxo-3-(3,4,5-trimethoxy-benzyl)-but-2-enoate} could improve postischemic contractile dysfunction in open-chest pigs. Myocardial stunning was achieved by a sequence of three 10-min left anterior descending (LAD) occlusions interspersed with 15 min of reperfusion. All pigs received either an intravenous saline vehicle (n =6) or PD156707 (n = 6) at a loading dose infusion of 10 mg/kg/h for 1 h before the first occlusion followed by a maintenance dose of 7 mg/kg/h for 4 h. Systolic wall thickening (percentage of baseline) was measured with sonomicrometers. There was no significant difference in systolic thickening between groups at baseline, at the end of the final stunning occlusion, or at any of the time points during reperfusion. PD156707 significantly reduced arterial blood pressure before myocardial ischemia and throughout reperfusion. There was no significant difference in size of the region at risk between groups. In conclusion, selective blockade of ETA receptors with PD156707 did not significantly alter postischemic contractile function in open-chest pigs. These results suggest that activation of ETA receptors by endogenous ET does not play a significant role in the pathogenesis of myocardial stunning.

Effects of ischemia on epicardial segment shortening

To evaluate the effects of nontransmural ischemia on epicardial contractile function, we implanted sonomicrometers in 15 open-chest, anesthetized (halothane) dogs. One cylindrical crystal (radiating ultrasound 360 degrees) was used as a transmitter for three conventional flat plate crystals arrayed to measure epicardial segment shortening along three different axes that were deviated 0 degree (parallel), 45 degrees (oblique), and 90 degrees (perpendicular) from surface fiber orientation in the anteroapical or posterior-basal left ventricle. During baseline conditions, epicardial shortening was maximal parallel with fiber orientation. Shortening decreased in a non-linear manner as deviation from fiber orientation increased, but there were significant differences between the two left ventricular regions suggesting that more substantial lateral strain occurs in the anterior-apical than the posterior-basal area. During coronary inflow restriction, changes in epicardial segment shortening also varied greatly depending on location and alignment. At levels of wall thickening impairment associated with normal subepicardial perfusion, changes in epicardial function were restricted to the segments aligned perpendicular to fiber orientation whereas the parallel and oblique segments displayed moderate dysfunction or none at all. Thus, transmural tethering modifies epicardial segmental motion during coronary inflow restriction, but the severity of the influence depends on the alignment and location of the epicardial measurements.

Changes in contractility fail to alter the size of the functional border zone in anesthetized dogs.

The functional border zone is nonischemic myocardium that exhibits reduced function adjacent to an ischemic area. To determine if the functional border zone can be modified by pharmacologic interventions that alter contractility, we infused isoproterenol (0.04-0.10 micrograms/kg/min) or administered propranolol (2 mg/kg) during circumflex coronary occlusion in nine anesthetized, open-chest dogs. We measured systolic wall thickening on both sides of the perfusion boundary, which was delineated with myocardial blood flow (microsphere) maps constructed from small tissue samples. By fitting sigmoid curves to the composite systolic wall thickening data after coronary occlusion, we modeled the distribution of functional impairment across the perfusion boundary. Defined as the distance from the perfusion boundary to 97.5% of the nonischemic asymptote of the sigmoid fits, the functional border zone was 31 degrees of circumference after coronary occlusion alone. Isoproterenol increased +dP/dt by 58% and augmented nonischemic systolic wall thickening without changing the lateral extent of the functional border zone (32 degrees). Propranolol reduced +dP/dt by 24% and depressed nonischemic systolic wall thickening, but the size of the functional border zone remained limited to 28 degrees. Within the functional border zone, wall thickening was significantly but only moderately reduced (-28%) compared with thickening in nonischemic myocardium more than 10 mm away from the perfusion boundary. The ratio of nonischemic border zone to central nonischemic area wall thickening remained the same with each intervention. We conclude that the dimensions of the functional border zone are fixed early after coronary occlusion and that inotropic interventions do not modify the extent or relative severity of nonischemic regional dysfunction.

Does Adenosine Deaminase Inhibition Protect Ischemic Myocardium

The protective effects of adenosine in myocardial ischemia are well established (1, 2). Exogenous adenosine, however, can produce hypotension, bradycardia, and AV block. Since ischemic myocardium itself is a significant endogenous source of adenosine, the rationale has developed that inhibition of adenosine catabolism to inosine with adenosine deaminase (ADA) inhibition could augment or sustain local adenosine levels during ischemic conditions. The elevated adenosine levels would be largely restricted to the ischemic area, minimizing potentially adverse side effects. The phrase “site and event specific intervention” has evolved to describe this appealing concept and ADA inhibitors such as erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) and 2-deoxycoformycin (pentostatin) have been used to test it in experimental models of myocardial ischemia.