This Article Abstract Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Glover, D. K. Articles by Beller, G. A. Search for Related Content PubMed PubMed Citation Articles by Glover, D. K. Articles by Beller, G. A.

(Circulation. 1996;94:1726-1732.)
© 1996 American Heart Association, Inc.

Articles

Pharmacological Stress Thallium Scintigraphy With 2-Cyclohexylmethylidenehydrazinoadenosine (WRC-0470)

A Novel, Short-Acting Adenosine A_2A Receptor Agonist

David K. Glover, ME; Mirta Ruiz, MD; Joo Y. Yang, MD; Bruce A. Koplan, MD; Terry R. Allen, BS; William H. Smith, MS; Denny D. Watson, PhD; Richard J. Barrett, PhD; George A. Beller, MD

the Cardiovascular Division, Department of Medicine, University of Virginia Health Sciences Center, Charlottesville, and Discovery Therapeutics, Inc (R.J.B.), Richmond, Va.

Correspondence to David K. Glover, ME, University of Virginia Health Sciences Center, Box 500, Health Sciences Center, Charlottesville, VA 22908. E-mail dkg2w@virginia.edu.

	Abstract

Top Abstract Introduction Methods Results Discussion References

 
Background Pharmacological stress imaging with adenosine or dipyridamole is associated with a high incidence of side effects, including hypotension, chest pain, AV conduction abnormalities, and bronchospasm. Although the desired coronary vasodilatory response is mediated primarily by the adenosine A_2A receptors, these side effects result from stimulation of the A₁, A_2B, or A₃ adenosine receptors. We hypothesized that a selective adenosine A_2A receptor agonist would induce coronary vasodilatation appropriate for pharmacological stress imaging, without evoking adenosine receptor–mediated side effects.

Methods and Results Infusions of a potent and selective A_2A adenosine receptor agonist, WRC-0470 (0.1 to 3 µg · kg^-1 ·min^-1 for 10 minutes), to five open-chest dogs produced dose-related left anterior descending (LAD) and left circumflex (LCx) coronary artery vasodilatation without altering mean arterial pressure, heart rate, left atrial pressure, or left ventricular dP/dt. In the same dogs, adenosine (300 µg · kg^-1 · min^-1 for 4 minutes) produced coronary vasodilatation that was limited by significant hypotension. To determine the utility of WRC-0470 for pharmacological stress imaging, the hemodynamic responses to WRC-0470 (0.6 µg · kg^-1 · min^-1 for 10 minutes) and adenosine (250 µg · kg^-1 · min^-1 for 4 minutes) were compared in dogs with critical LAD stenoses. ²⁰¹Tl was injected at the peak WRC-0470 stress response. WRC-0470 increased LCx flow nearly fivefold but did not significantly lower mean arterial pressure. Anteroseptal defects were readily apparent in slice images from all dogs. The mean defect ratio (LAD/LCx) was 0.59±0.06.

Conclusions The potent A_2A-selective adenosine receptor agonist WRC-0470 is a short-acting coronary vasodilator with potential utility for pharmacological stress perfusion imaging.

Key Words: adenosine • receptors • radioisotopes • imaging

	Introduction

Top Abstract Introduction Methods Results Discussion References

 
Pharmacological stress is frequently induced with adenosine or dipyridamole in patients with suspected CAD before imaging with ²⁰¹Tl scintigraphy or echocardiography. Since dipyridamole prevents uptake and inactivation of endogenous adenosine by erythrocytes and vascular endothelial cells, both drugs effect dilation of the coronary resistance vessels by the same mechanism: activation of specific cell-surface A₂ receptors. Although pharmacological stress was originally introduced as a means of provoking coronary dilatation in patients unable to exercise, several studies have shown that the prognostic value of ²⁰¹Tl or echocardiographic imaging in patients subjected to pharmacological stress with adenosine or dipyridamole was equivalent to those subjected to traditional exercise stress.^{1 2 3 4} Although the prognostic value is clear, there is an unusually high incidence of drug-related adverse side effects during pharmacological stress imaging with these drugs. A prospective study of 9256 patients subjected to coronary vasodilatation with adenosine during radionuclide imaging reported that 82% experienced adverse side effects, the most common of which were flushing (37%), chest pain (35%), shortness of breath or dyspnea (35%), headache (14%), ECG changes (9%), and AV conduction block (8%).⁵ When associated with adenosine, these events are usually transient, but all produce patient discomfort and are sometimes severe enough to require administration of aminophylline or premature termination of the study.⁵

Data from animal studies suggest that specific adenosine A_2A subtype receptors on coronary resistance vessels mediate the coronary dilatory responses to adenosine, whereas subtype A_2B receptor stimulation relaxes conductance vessels.⁶ Evidence from human studies suggests that adenosine-induced AV conduction abnormalities and chest pain are due to activation of adenosine A₁ receptors.^{7 8 9 10} Thus, it is encouraging that pretreatment with a selective adenosine A₁ receptor antagonist such as N-0861^{7 8} may prevent at least some of the adverse events associated with adenosine or dipyridamole without hindering the coronary vasodilatation necessary for effective stress imaging.¹¹ Although this strategy may improve patient comfort and safety, a simpler strategy might be to avoid any stimulation of the A₁ receptors by designing a compound that had little or no affinity for adenosine A₁ receptors but that selectively stimulated only the adenosine A₂ receptors responsible for coronary vasodilatation. We postulated that a short-acting, selective adenosine A_2A receptor agonist might produce controlled, transient dilatation of the coronary vasculature without producing systemic hypotension or provoking adenosine A₁ receptor–mediated adverse events. A number of highly potent and selective adenosine A₂ receptor agonists have been synthesized,^{12 13 14} several of which produce long-lasting systemic hypotension and reflex tachycardia in rats.¹⁵ We are not aware of any studies that examined the coronary dilatory responses to these agonists in dogs in vivo.

The goals of the present experimental study were (1) to characterize the systemic hemodynamic and coronary blood flow responses to the intravenous administration of a novel adenosine analogue, WRC-0470 (Fig 1), in dogs and to compare these responses to those after adenosine administration in the same animals and (2) to determine the potential utility of WRC-0470 as a coronary vasodilator for pharmacological stress imaging.

View larger version (15K): [in this window] [in a new window]   Figure 1. Comparison between the molecular structures of adenosine and WRC-0470 (molecular weight of WRC-0470=398.6).

	Methods

Top Abstract Introduction Methods Results Discussion References

 
All experiments described in this article were carried out in accordance with the Guide for the Care and Use of Laboratory Animals as adopted by the National Institutes of Health, in compliance with the position of the American Heart Association, and with the approval of the Animal Care and Use Committee of the University of Virginia.

Twelve fasted adult mongrel dogs of either sex (weight, 21.5±0.97 kg, mean±SEM) were anesthetized with Innovar (0.08 mL/kg IV) and sodium pentobarbital (6.6 mg/kg IV), intubated, and ventilated on a respirator (Harvard Apparatus) with 4 cm H₂O positive end-expiratory pressure. Arterial blood gases were monitored throughout each experiment (model 170, Ciba-Corning), and pH, PCO₂, and HCO₃ levels were maintained at physiological levels. Both femoral veins were cannulated with 8F polyethylene catheters for the administration of adenosine, WRC-0470, and ²⁰¹Tl. Additional Innovar anesthetic was also administered intravenously as needed. The right and left femoral arteries were then isolated and cannulated with similar 8F catheters to serve as sites for the collection of arterial blood for blood gas determinations and microsphere reference samples and to monitor systemic arterial pressure. Next, the left main carotid artery was isolated and cannulated with a Millar high-fidelity pressure recording catheter. While the pressure waveform was observed, the Millar catheter was advanced into the left ventricle.

The heart was then exposed through a left lateral thoracotomy and suspended in a pericardial cradle. A flare-tipped polyethylene catheter was inserted into the left atrial appendage for continuous monitoring of left atrial pressure and for the injection of radiolabeled microspheres. A 1-cm section of the LAD was isolated and encircled with an ultrasonic flow probe (Transonic Systems) and snare ligature. A similar flow probe was placed on the LCx.

The ECG, heart rate, arterial and left atrial pressures, LAD and LCx coronary blood flows, and LV pressure and dP/dt were continuously monitored on an eight-channel strip-chart recorder (model 7758D, Hewlett Packard Co).

Protocol 1: Effect of WRC-0470 Dose on Systemic Hemodynamic Parameters (n=5 Dogs)
After baseline hemodynamic measurements of heart rate, arterial and left atrial pressures, LAD and LCx coronary flows, and dP/dt had been collected, adenosine was infused at a dose of 300 µg · kg^-1 · min^-1 to produce a maximal coronary vasodilatory response. Previous experimental work in our laboratory has shown that this dose of adenosine produces such an effect in dogs within 4 minutes.¹⁶ After the peak response, the adenosine infusion was terminated, and all hemodynamic parameters were allowed to return to their baseline values. In our past experience, we have found no evidence of adenosine tachyphylaxis at these doses in dogs. Next, WRC-0470 was infused over a period of 10 minutes at doses of 0.1, 0.3, 0.6, 1.0, and 3.0 µg · kg^-1 · min^-1, and hemodynamic responses were continuously recorded. Sufficient time was allowed between doses for all parameters to return to their baseline levels. No attempt was made to measure adenosine A₁ receptor–mediated ECG changes (eg, PR interval) because it was previously shown that A₁ receptor–mediated responses are not easily evoked in the canine heart.¹⁷

Protocol 2: Administration of WRC-0470 in Dogs With an LAD Stenosis (n=6 Dogs)
The protocol is summarized in Fig 2. After baseline hemodynamic measurements of heart rate, arterial and left atrial pressures, LAD and LCx coronary flows, and LV dP/dt had been collected, the snare ligature was tightened to produce a critical LAD stenosis. A critical stenosis was defined as no change in resting coronary flow, but a complete abolition of the reactive hyperemic response to a transient 10-second total occlusion. After a brief stabilization period, a left atrial injection of radioactive microspheres was given to assess regional myocardial blood flow. Microspheres have been used extensively in our laboratory, and their use has been described previously.¹⁸ Five minutes later, an infusion of adenosine (250 µg · kg^-1 · min^-1 IV) was begun and continued until ultrasonically measured LCx flow was maximal, at which time a second microsphere injection was given and the adenosine infusion was terminated. Thirty minutes later, after all monitored hemodynamic parameters had returned to baseline, a third microsphere injection was given. On the basis of the results of protocol 1, WRC-0470 (0.6 µg · kg^-1 · min^-1) was then infused for 10 minutes. Immediately before the infusion was stopped, ²⁰¹Tl (0.5 to 1.0 mCi) and a fourth microsphere injection were administered simultaneously. The dogs were killed within 5 minutes with an overdose of sodium pentobarbital. Their hearts were removed and sliced into four rings of equal thickness. The slices were trimmed of excess fat and adventitia, placed on a thin piece of cardboard, and covered with cellophane wrap. The slices were imaged directly on the collimator of a conventional gamma camera (model 420, Ohio-Nuclear) with a 25% window centered on the ²⁰¹Tl photopeak for 28 minutes (maximal count time). To quantify the magnitude of the defects on the uncorrected myocardial slice images, a region of interest was drawn on the defect area visible in the anteroapical regions of the LV wall. A second region of interest was drawn on the normal posterior wall. The defect count ratio was defined as the ratio of the average counts in the LAD defect area region of interest to the average counts in the normal region.

View larger version (14K): [in this window] [in a new window]   Figure 2. Experimental protocol 2. Aden indicates adenosine; mic, radioactive microsphere.

After imaging, the heart slices were divided into 96 endocardial, midwall, and epicardial segments and counted in a gamma well counter (model 5550, Packard Instruments) as previously described.¹⁶ The tissue samples were counted within 24 hours for 5 minutes each with ²⁰¹Tl window settings (50 to 100 keV). Three weeks later, when the ²⁰¹Tl had decayed, the tissue samples were recounted for microsphere blood flow. The window settings used were ¹¹³Sn, 340 to 440; ¹⁰³Ru, 450 to 550; ⁹⁵Nb, 680 to 840; and ⁴⁶Sc, 842 to 1300 keV. The tissue counts were corrected for background, decay, and isotope spillover, and regional myocardial blood flow was calculated with specialized computer software (PCGERDA, Packard Instruments). Transmural activity and flow values were calculated as the weighted average of the corresponding epicardial, midwall, and endocardial samples.

Statistics
All statistical computations were made with SYSTAT software (Systat Inc), and the results are expressed as mean±SEM. Differences between hemodynamic responses to adenosine with their corresponding baseline values were assessed by use of a paired t test. WRC-0470 comparisons over time were assessed with a repeated measures ANOVA with values of P<.05 considered significant.

Drug Preparation
WRC-0470 was originally synthesized by Dr Ray Olsson, University of South Florida¹⁴ ; additional material was synthesized by Dr Ronald Wysocki, Chemistry Department, Discovery Therapeutics Inc, Richmond, Va. Stock solutions were made in distilled water and diluted in 0.9% NaCl. Adenosine was obtained from Sigma Chemical Co.

	Results

Top Abstract Introduction Methods Results Discussion References

 
Protocol 1: WRC-0470 Dose Response in Normal Dogs
Results from these experiments are summarized in Table 1 and illustrated in Fig 3. Each of the five dogs in this protocol was treated with adenosine before WRC-0470. Within 1 minute of the start of the infusion (0.96±0.4 minutes), adenosine (300 µg · kg^-1 · min^-1) markedly increased flow through the LAD and LCx and decreased mean systemic blood pressure significantly, from 106 to 77 mm Hg. In addition, in these anesthetized dogs, adenosine produced a slight reflex rise in heart rate, although this change did not reach statistical significance (P=.06). There were no significant effects on either left atrial pressure or LV dP/dt. All hemodynamic parameters returned to their baseline levels within 5 minutes of the end of the adenosine infusion. These results are consistent with those produced previously in our laboratory.^{11 16} Subsequent sequential 10-minute infusions of WRC-0470 (0.1, 0.3, 0.6, 1, and 3 µg · kg^-1 · min^-1) produced dose-related increments in LAD and LCx flow; the maximal coronary vasodilatory effect of WRC-0470, which was produced by 1 µg · kg^-1 · min^-1, was considerably greater than that produced by adenosine (Fig 3). The time to the peak flow response at the 0.6-µg · kg^-1 · min^-1 dose rate was 2.4±0.5 minutes. Despite the marked coronary vasodilatory response, WRC-0470 did not produce significant systemic hypotension. The highest dose of WRC-0470 tested (3 µg · kg^-1 · min^-1) reduced mean systemic blood pressure from 104±6 to 88±13 mm Hg (after 10 minutes), but this fall did not achieve statistical significance. As was seen with adenosine, there was a slight reflex rise in heart rate with WRC-0470 that did not reach statistical significance, and there was a trend toward a dose-related increase in LV dP/dt; however, this increase was also not statistically significant. All parameters began to return to baseline levels immediately after the infusion was stopped. After cessation of the 0.1, 0.3, 0.6, 1, and 3 µg · kg^-1 · min^-1 infusions, LAD and LCx flows returned to within 10% of baseline by 3, 5, 15, 30, and 35 minutes, respectively.

View this table: [in this window] [in a new window]   Table 1. Hemodynamic Parameters: Protocol 1

View larger version (23K): [in this window] [in a new window]   Figure 3. Comparison of the changes in mean LCx coronary flow (Qcx) and mean arterial pressure (MAP) in response to either adenosine (A; 300 µg · kg-1 · min-1) or multiple doses of WRC-0470 in anesthetized dogs (protocol 1, n=5). Points are shown at 1.0-minute intervals during the adenosine infusion and at 1.0, 2.5, and 5.0 minutes after termination. For WRC-0470, points are shown every 2.5 minutes during infusion and afterward until the responses had returned to near baseline. The 0.6-µg ·kg-1 · min-1 dose of WRC-0470 was selected for use in protocol 2 because it produced at least as great an increase in coronary flow as did adenosine but without significant systemic hypotension.

It is apparent from the data shown in Fig 3 that the 0.6-µg · kg^-1 · min^-1 dose of WRC-0470 produced coronary vasodilatation that was nearly equivalent to that produced by adenosine (300 µg · kg^-1 · min^-1). Therefore, this dose was used to determine the suitability of WRC-0470 to produce a pharmacological stress response conducive to imaging with ²⁰¹Tl.

Protocol 2: Administration of WRC-0470 in Dogs With an LAD Stenosis
Hemodynamic Data
The measured hemodynamic parameters are summarized in Table 2 for six of the seven dogs instrumented in this protocol. One dog was eliminated from the final analysis because of the lack of a coronary stenosis. As shown in Table 2, setting a critical stenosis on the LAD had no effect on any of the hemodynamic measurements. Each dog was given adenosine (250 µg · kg^-1 · min^-1 IV) 30 minutes before WRC-0470 (0.6 µg · kg^-1 · min^-1). All measured hemodynamic parameters had returned to their baseline values before WRC-0470 infusion.

View this table: [in this window] [in a new window]   Table 2. Hemodynamic Parameters: Protocol 2

The effects of adenosine and WRC-0470 infusions on mean coronary blood flow and systemic arterial blood pressure are compared in Figs 4 and 5, respectively. Adenosine infusion caused LCx flow to increase approximately threefold, from 34±5 to 96±11 mL/min (P<.01), whereas flow in the critically stenotic LAD coronary artery did not change (23±3 versus 23±3 mL/min) (Fig 4, left). At the same time, adenosine caused mean systemic blood pressure to fall from 100±5 to 76±4 mm Hg (P<.01) (Fig 5, left) but had no effect on left atrial pressure, heart rate, or dP/dt. In contrast, WRC-0470 infusion caused LCx flow to increase fivefold, from 26±1 to 124±18 mL/min (P<.01) (Fig 4, right). This marked increase in flow was not accompanied by hypotension (Fig 5) or by changes in any other of the measured hemodynamic parameters (Table 2). Responses to both adenosine and WRC-0470 were nearly identical to those evoked in the first protocol (Fig 3), except that LAD dilatation was prevented by imposition of the stenosis.

View larger version (14K): [in this window] [in a new window]   Figure 4. Ultrasonically measured flow in the critically stenotic LAD and normal LCx of protocol 2 dogs. Left, LAD and LCx flows at rest (solid bars) and during pharmacological stress (hatched bars) with adenosine (250 µg · kg-1 · min-1); right, coronary flow responses to WRC-0470 (0.6 µg · kg-1 · min-1). LCx flow increased threefold with adenosine and fivefold with WRC-0470. Flow in the critically stenotic LAD did not change with either stress.

View larger version (36K): [in this window] [in a new window]   Figure 5. Comparison between the decrease in mean arterial pressure produced by adenosine or WRC-0470 in protocol 2 dogs. Solid bars represent mean arterial pressure at rest; hatched bars show pressure during pharmacological stress. Adenosine infusion resulted in a significant hypotensive response (100 to 76 mm Hg), whereas WRC-0470 produced no hypotension. *P<.0001 vs rest; +P=.02 adenosine vs WRC-0470.

Regional Myocardial Blood Flow
Table 3 summarizes regional myocardial blood flow in the endocardium, midwall, epicardium, and transmurally in the LAD and LCx zones of the heart, as determined by radioactive microspheres. Baseline flow was similar before and after adenosine in both the LAD and LCx zones. In the LAD zone, there was slightly lower endocardial flow during WRC-0470 infusion compared with adenosine (0.69±0.08 versus 0.88±0.11 mL · min^-1 · g^-1), suggesting a greater degree of coronary steal, although this difference did not achieve statistical significance. Also, with both adenosine and WRC-0470 infusions, there was a significant increase in epicardial flow in the critically stenotic LAD zone; however, endocardial, midwall, and transmural average flow did not increase significantly. In the LCx zone, both adenosine and WRC-0470 caused significant increases in absolute coronary flow across the LV wall. Although there was a trend toward a greater percent increase in transmural flow during WRC-0470 (274.7±74%) versus adenosine (172.4±48%) infusion, this difference did not reach statistical significance (P=.08).

View this table: [in this window] [in a new window]   Table 3. Regional Myocardial Blood Flow

²⁰¹Tl Uptake During WRC-0470 Vasodilator Stress
Fig 6 depicts the transmural microsphere flow ratios (LAD/LCx) at baseline and during WRC-0470 stress and the ²⁰¹Tl activity ratio (LAD/LCx) as determined by gamma well counting. The ²⁰¹Tl was injected during peak WRC-0470 stress. The LAD/LCx flow ratio fell from 0.93±0.04 to 0.35±0.06 (P<.01) with WRC-0470 infusion. The ²⁰¹Tl LAD/LCx transmural activity ratio (0.50±0.05) underestimated the flow disparity as measured by microspheres (0.35±0.06) (P<.01) because of the plateau in tracer extraction at high flow. On ex vivo images of heart slices, anteroseptal defects were readily apparent in all dogs. The mean defect count ratio (LAD/LCx) was 0.59±0.06.

View larger version (25K): [in this window] [in a new window]   Figure 6. Transmural microsphere flow ratios (LAD/LCx) at rest and during WRC-0470 stress (left pair of bars) vs the transmural 201Tl activity ratio (LAD/LCx) (right bar) as determined by gamma well counting. 201Tl was injected at peak WRC-0470 stress. WRC-0470 stress produced a large flow heterogeneity between the stenotic LAD and normal LCx perfusion territories. Although the magnitude of the flow disparity was underestimated by 201Tl uptake, anteroseptal defects (mean defect magnitude=41%) were observed in all dogs. *P<.01 vs rest; +P=.01 vs stress flow.

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
The data from these experiments demonstrate that the highly potent A_2A-selective adenosine agonist WRC-0470 produced marked coronary vasodilatation without hypotension that was conducive to ²⁰¹Tl scintigraphic imaging of a myocardial perfusion defect.

Adenosine Receptor Selectivity
Previous studies in isolated guinea pig hearts established that WRC-0470 is 16 000-fold more selective for the activation of the A_2A than A₁ receptor.¹⁴ WRC-0470 is 11-fold more potent than NECA, a nonspecific adenosine analogue, as a coronary dilator. WRC-0470 is 150-fold less potent than NECA at inducing A_2B adenosine receptor–mediated relaxations of the guinea pig aorta. The nonselective adenosine receptor antagonists 8-phenyltheophylline and 8-p-sulfophenyltheophylline antagonized both the coronary (A_2A) and the aortic (A_2B) vasodilatory responses to WRC-0470, indicating that these actions were mediated by adenosine receptors and were not a result of a nonspecific effect.¹⁴ Activation of myocardial adenosine A₁ receptors evokes negative inotropic, chronotropic, and dromotropic responses in the heart. WRC-0470 activates adenosine A₁ receptors only at concentrations exceeding 30 µmol/L (Discovery Therapeutics, unpublished data). Because the maximum inhibitory response produced by WRC-0470 in the electrically paced left atrium was less than that produced by NECA, it was concluded that WRC-0470 was a partial agonist of A₁ receptors. Furthermore, WRC-0470 antagonized the responses to NECA with a dissociation constant of 21 µmol/L, indicating that WRC-0470 is acting at the same A₁ receptor as NECA to induce negative inotropy but that WRC-0470 has a very low affinity for binding to the A₁ receptors and has a low intrinsic efficacy for activation of these receptors. When the tissue-dependent variables are eliminated, the comparative molar potency ratios for WRC-0470 and NECA in the isolated atrial and aortic adenosine receptor assay systems used in this study and in the previously published Langendorff heart assay¹⁴ may be used to calculate relative selectivities for one receptor subtype versus another. WRC-0470 was thus 5500-fold more selective for activation of the coronary A_2A adenosine receptors than the cardiac A₁ receptors, 1600-fold more selective for activation of the A_2A receptors than the aortic A_2B receptors, and 3.5-fold more selective for the A_2B than the A₁ receptors.

Dose Responses of WRC-0470 in Normal Dogs (Protocol 1)
When administered to anesthetized dogs in the present study, WRC-0470 produced dose-related coronary vasodilatation. In each experiment, we evoked maximal coronary vasodilatation with adenosine before administering WRC-0470, and in each case, WRC-0470 produced significantly greater coronary dilatation than adenosine. It was obvious in these studies that the hyperemic responses to adenosine were limited by concurrent hypotension. The differences in the responses between adenosine and WRC-0470 were striking. As coronary blood flow increased during WRC-0470 infusion, systemic blood pressure changed little. Since the pressure head driving coronary perfusion remained constant, the dilatory responses achievable with WRC-0470 exceeded those inducible with adenosine. Only the highest dose of WRC-0470 tested (3 µg·kg^-1·min^-1) produced an appreciable hypotensive response (mean blood pressure fell from 104±6 to 88±13 mm Hg) and, although this response was not statistically significant, it appears that the effects on systemic blood pressure became more pronounced as the dose was increased. If WRC-0470 is infused clinically to induce pharmacological stress, a reduction in systemic blood pressure might indicate that the maximal coronary vasodilatory dose has been exceeded.

There are a number of possible explanations for the ability of WRC-0470 to induce coronary vasodilatation at concentrations that did not affect systemic blood pressure. One possibility may be that there is a predominance of A_2A receptors in the coronary resistance vessels compared with the systemic circulation. A second possibility is that the A_2A receptors in the coronary arteries are more abundant or highly coupled than those in other organs. It might be that the same concentrations of WRC-0470 that evoke coronary vasodilatation also dilate other vascular beds but that the magnitude of that vasodilatation is insufficient to reduce overall systemic blood pressure. A third possibility is that the hypotensive response to adenosine may be mediated in part by A₃ adenosine receptors, since specifically blocking these receptors in rats eliminated the hypotensive response to an adenosine analogue.¹⁹ In addition, Hannon et al²⁰ recently showed that A₃ receptor–mediated hypotension may involve mast cells.

Comparison of Adenosine and WRC-0470
Adenosine-induced vasodilatation in resistance and conductance vessels appears to be mediated by the A_2A and A_2B receptors, respectively. This is illustrated by the findings that WRC-0470 and CGS-21680 (another highly specific A_2A receptor agonist) are extremely potent agonists in an assay for adenosine A_2A activity (coronary vasodilatation in the guinea pig isolated heart)¹² but are weak agonists in an assay for adenosine A_2B activity (relaxation of guinea pig isolated aortas).²¹ As a highly specific adenosine A_2A receptor agonist, WRC-0470 produces marked dilatation of coronary resistance vessels, resulting in diversion of blood flow away from the myocardial perfusion zone distal to a stenosis that produces regional flow heterogeneity and a perfusion defect on a myocardial scintigram.

The present study indicates that the selective adenosine A_2A receptor agonist WRC-0470 produced a myocardial perfusion defect (mean, 41% reduction) that was easily detected with ²⁰¹Tl scintigraphy and similar in magnitude (38%) to the ²⁰¹Tl perfusion defects observed after adenosine infusion in the same coronary stenosis model.¹⁶ The magnitude of flow heterogeneity induced by adenosine in the critically stenosed canine preparation used in this study mimics human responses quite well, and the accuracy of ²⁰¹Tl scintigraphy in detecting myocardial perfusion defects in dogs is excellent.^{11 16} In the present study, ²⁰¹Tl activity underestimated the size of the true flow disparity as measured by microspheres. This is because of the roll-off in extraction of ²⁰¹Tl at high flow, which is characteristic of all diffusible flow tracers and has been reported previously by us and others.^{11 16 22} Despite the underestimation of the flow disparity, the mean magnitude of the ²⁰¹Tl defect (41%) was significant and well within clinically detectable limits.

Study Limitations
We chose a canine model for these experiments because of our extensive experience with it and to facilitate comparisons with our previous work, which used adenosine or dipyridamole.^{16 23} Because the canine adenosine A₁ receptor is atypical, we could not assess the effects of WRC-0470 on A₁ receptor responses. In addition, receptor binding studies of WRC-0470 to canine A_2A and A_2B receptors were not performed in these studies. Thus, the preferential binding of the agent to A_2A receptors can be inferred only indirectly. Another limitation was that the hemodynamic and coronary flow responses were evaluated in anesthetized rather than conscious dogs. Whether the hemodynamic effects of anesthesia might be different with adenosine compared with WRC-0470 is unknown. Finally, ²⁰¹Tl imaging was not undertaken with adenosine infusion in this study. Since imaging could be performed only once, it was undertaken only with WRC-0470 administration.

Clinical Implications
Vasodilator stress perfusion imaging is increasingly being used as an alternative to exercise scintigraphy for detection of CAD and risk stratification. Sensitivity and specificity for detection of CAD are similar for dipyridamole and adenosine, ranging between 85% and 90%. The major potential clinical advantage of WRC-0470 over dipyridamole or adenosine is the reduction in A₁, A_2B, and A₃ receptor–mediated side effects. In a study by Abreu et al,²⁴ 79% of 607 patients receiving adenosine for stress perfusion imaging had at least one side effect, including facial flushing in 35%, chest or neck pain in 43%, headache in 21%, and dyspnea in 19%. In the same study, systolic blood pressure fell from 138 to 121 mm Hg and diastolic pressure fell from 78 to 67 mm Hg. O'Keefe et al²⁵ reported that 28 of 340 consecutive patients who received intravenous adenosine for ²⁰¹Tl imaging developed either second-degree or third-degree AV block. In the study by Gupta et al,⁴ 82% of patients receiving intravenous adenosine had side effects, with flushing in 41%, chest pain in 24%, and dizziness in 20%. Cerqueira et al⁵ reported the safety profile of adenosine stress perfusion imaging from a registry comprising 9256 consecutive patients. Side effects were reported in 81% of these patients. Although 72 patients experienced transient third-degree AV block, 706 (7%) had either first-, second-, or third-degree AV block, and chest pain occurred in 3207 (35%). In 7% of the patients, the protocol was terminated prematurely. Significant side effects can also occur with dipyridamole infusion. In 3715 patients receiving dipyridamole, the mean systolic blood pressure fell by 14.07 mm Hg, with 11% of patients demonstrating a 20% drop in systolic pressure.²⁶ Thus, side effects, although mostly tolerated, are common with myocardial perfusion imaging with adenosine or dipyridamole. However, it would be desirable to have a pharmacological stress agent capable of producing the required vasodilatation without the adverse side effects. In the present study, we found that WRC-0470 produced greater vasodilatation at its peak pharmacological dose than did adenosine, and without significant systemic hypotension. Clinically, this would be beneficial in preventing a suboptimal stress test and might translate to an enhanced detection rate, especially in CAD patients with mild coronary stenoses. In addition, because of its marked selectivity for the A_2A adenosine receptor, it would be expected that fewer of the other A₁, A_2B, and A₃ receptor–mediated side effects (AV conduction abnormalities, chest pain, and perhaps breathing difficulties) would occur during treatment with WCR-0470. It has been shown that both AV block and chest pain are A₁ receptor–mediated in humans, because they were prevented by the A₁-selective antagonist N-0861.⁸ Since WRC-0470 has a very low affinity for the A₁ adenosine receptors in isolated heart tissues and produces negative inotropic and negative chronotropic responses in vitro and in vivo in rats only at very high concentrations, it is unlikely that the concentrations of WRC-0470 that produce coronary vasodilatation will produce any side effects attributable to adenosine A₁ receptor stimulation, such as AV conduction abnormalities and pain.^{7 8 9 10} Some recent evidence also suggests that the adenosine A₃ receptor may play a role in bronchospasm due to the discovery of adenosine A₃ receptors on the surface of mast cells.²⁷ Thus, WRC-0470 has the potential to provide comparable or enhanced diagnostic accuracy for CAD detection when used as a pharmacological stress imaging agent, but with a markedly reduced side effect profile. Clinical trials will provide further information regarding the potential advantage.

	Selected Abbreviations and Acronyms

 

CAD = coronary artery disease LAD = left anterior descending coronary artery LCx = left circumflex artery LV = left ventricular NECA = N-ethylcarboxamidoadenosine WRC-0470 = 2-cyclohexylmethylidenehydrazino-adenosine

	Footnotes

 
This study was supported in part by Discovery Therapeutics, Inc.

Received September 29, 1995; revision received April 12, 1996; accepted April 15, 1996.

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Mahmarian JJ, Pratt CM, Nishimura S, Abreu A, Verani MS. Quantitative adenosine ²⁰¹Tl single-photon emission computed tomography for the early assessment of patients surviving acute myocardial infarction. Circulation. 1993;87:1197-1210.[Abstract/Free Full Text]

2. Nishimura S, Mahmarian JJ, Boyce TM, Verani MS. Equivalence between adenosine and exercise thallium-201 myocardial tomography: a multicenter, prospective, crossover trial. J Am Coll Cardiol. 1992;20:265-275.[Abstract]

3. Coyne EP, Belvedere DA, Vande Streek PR, Weiland FL, Evans RB, Spaccavento LJ. Thallium-201 scintigraphy after intravenous infusion of adenosine compared with exercise thallium testing in the diagnosis of coronary artery disease. J Am Coll Cardiol. 1991;17:1289-1294.[Abstract]

4. Gupta NC, Esterbrooks DJ, Hilleman DE, Mohiuddin SM. Comparison of adenosine and exercise thallium-201 single-photon emission computed tomography (SPECT) myocardial perfusion imaging. J Am Coll Cardiol. 1992;19:248-257.[Abstract]

5. Cerqueira MD, Verani MS, Schwaiger M, Heo J, Iskandrian AS. Safety profile of adenosine stress perfusion imaging: results from the Adenoscan Multicenter Trial Registry. J Am Coll Cardiol. 1994;23:384-390.[Abstract]

6. Martin PM, Ueeda M, Olsson RA. 2-Phenylethoxy-9-methyladenine: an adenosine receptor antagonist that discriminates between A₂ adenosine receptors in the aorta and the coronary vessels from the guinea pig. J Pharmacol Exp Ther. 1993;265:248-253.[Abstract/Free Full Text]

7. Nagashima S, Moore HJ, Tracy CM, Solomon AJ, Hewett-Maulman JL, Cowley SM, Woosley RL, Barbey JT. Dose ranging study of N-0861, a new selective A₁ adenosine receptor antagonist, in patients receiving adenosine. J Am Coll Cardiol. 1994;23:478A. Abstract.

8. Hill JA, Bertolet B, Franco E, Kerensky R, Nichols W, Cowley S, Miller J, Belardinelli L. Attenuation of adenosine A1 receptor-mediated effects by N-0861, N⁶-endonorbornan-2-yl-9-methyladenine, in humans. Drug Dev Res. 1994;31:278. Abstract.

9. Gaspardone A, Crea F, Iamele M, Tomai F, Versaci F, Pellegrino A, Chiariello L, Gioffre PA. Bamiphylline improves exercise-induced myocardial ischemia through a novel mechanism of action. Circulation. 1993;88:502-508.[Abstract/Free Full Text]

10. Gaspardone A, Crea F, Tomai F, Versaci F, Iamele M, Gioffre G, Chiariello L, Gioffre PA. Muscular and cardiac adenosine-induced pain is mediated by A1 receptors. J Am Coll Cardiol. 1995;25:251-270.[Abstract]

11. Glover DK, Ruiz M, Sansoy V, Barrett RJ, Beller GA. Effect of N-0861, a selective adenosine A1 receptor antagonist, on pharmacologic stress imaging with adenosine. J Nucl Med. 1995;36:270-275.[Abstract/Free Full Text]

12. Ueeda M, Thompson RD, Arroyo LH, Olsson RA. 2-Alkoxyadenosines: potent and selective agonists at the coronary artery A₂ adenosine receptor. J Med Chem. 1991;34:1334-1339.[Medline] [Order article via Infotrieve]

13. Webb RL, Sills MA, Chovan JP, Balwierczak JL, Francis JE. CGS-21680: a potent selective adenosine A₂ receptor agonist. Cardiovasc Drug Rev. 1992;1:26-53.

14. Niiya K, Olsson RA, Thompson RD, Silvia SK, Ueeda M. 2-(N'-Alkylidenehydrazino) adenosines: potent and selective coronary vasodilators. J Med Chem. 1992;35:4557-4561.[Medline] [Order article via Infotrieve]

15. Webb RL, Barclay BW, Graybill SC. Cardiovascular effects of adenosine A₂ agonists in the conscious spontaneously hypertensive rat: a comparative study of three structurally distinct ligands. J Pharmacol Exp Ther. 1991;259:1203-1212.[Abstract/Free Full Text]

16. Glover DK, Ruiz M, Simanis JP, Smith WH, Watson DD, Beller GA. Comparison between thallium-201 and Tc-99m sestamibi uptake during adenosine hyperemia in a canine model of either mild or severe coronary stenoses. Circulation. 1995;91:813-820.[Abstract/Free Full Text]

17. Pelleg A, Hurt CM. Effects of N⁶-endonorbornan-2-yl-9-methyladenine, N-0861, on the negative chronotropic and vasodilatory actions of adenosine in the canine heart in vivo. Can J Physiol Pharmacol. 1992;70:1450-1456.[Medline] [Order article via Infotrieve]

18. Heymann MA, Payne BD, Hoffman JIE, Rudolph AM. Blood flow measurement with radionuclide-labeled particles. Prog Cardiovasc Dis. 1987;20:55-79.

19. Fozard JR, Hannon JP. BW-A522 blocks adenosine A3 receptor-mediated hypotensive responses in the rat. Eur J Pharmacol. 1994;252:R5-R6.[Medline] [Order article via Infotrieve]

20. Hannon JP, Pfannkuche HJ, Fozard JR. A role for mast cells in adenosine A3 receptor-mediated hypotension in the rat. Br J Pharmacol. 1995;115:945-952.[Medline] [Order article via Infotrieve]

21. Martin PL. Relative agonist potencies of C²-substituted analogues of adenosine: evidence for adenosine A_2B receptors in the guinea pig aorta. Eur J Pharmacol. 1992;216:235-242.[Medline] [Order article via Infotrieve]

22. Glover DK, Okada RD. Myocardial kinetics of Tc-MIBI in canine myocardium after dipyridamole. Circulation. 1990;81:628-636.[Abstract/Free Full Text]

23. Granato JE, Watson DD, Belardinelli L, Cannon JM, Beller GA. Effects of dipyridamole and aminophylline on hemodynamics, regional myocardial blood flow and thallium-201 washout in the setting of a critical coronary stenosis. J Am Coll Cardiol. 1990;16:1760-1770.[Abstract]

24. Abreu A, Mahmarian JJ, Nishimura S, Boyce TM, Verani MS. Tolerance and safety of pharmacologic coronary vasodilatation with adenosine in association with thallium-201 scintigraphy in patients with suspected coronary artery disease. J Am Coll Cardiol. 1991;18:730-735.[Abstract]

25. O'Keefe JH Jr, Bateman TM, Silvestri R, Barnhart C. Safety and diagnostic accuracy of adenosine thallium-201 scintigraphy in patients unable to exercise and those with left bundle branch block. Am Heart J. 1992;124:614-621.[Medline] [Order article via Infotrieve]

26. Lette J, Tatum JL, Fraser S, Miller DD, Waters DD, Heller G, Stanton EB, Bom HS, Leppo J, Nattel S. Safety of dipyridamole testing in 73,806 patients: the Multicenter Dipyridamole Safety Study. J Nucl Cardiol. 1995;2:3-17.[Medline] [Order article via Infotrieve]

27. Ramkumar V, Stiles GL, Beaven MA, Ali H. The A₃ adenosine receptor is the unique adenosine receptor which facilitates release of allergic mediators in mast cells. J Biol Chem. 1993;268:16887-16890.[Abstract/Free Full Text]

This article has been cited by other articles:

				E. H. Botvinick Current Methods of Pharmacologic Stress Testing and the Potential Advantages of New Agents J. Nucl. Med. Technol., March 1, 2009; 37(1): 14 - 25. [Abstract] [Full Text] [PDF]

				J. E. Udelson, G. V. Heller, F. J.Th. Wackers, A. Chai, D. Hinchman, P. S. Coleman, V. Dilsizian, M. DiCarli, R. Hachamovitch, J. R. Johnson, et al. Randomized, Controlled Dose-Ranging Study of the Selective Adenosine A2A Receptor Agonist Binodenoson for Pharmacological Stress as an Adjunct to Myocardial Perfusion Imaging Circulation, February 3, 2004; 109(4): 457 - 464. [Abstract] [Full Text] [PDF]

				N K Sabharwal and A Lahiri Role of myocardial perfusion imaging for risk stratification in suspected or known coronary artery disease Heart, November 1, 2003; 89(11): 1291 - 1297. [Abstract] [Full Text] [PDF]

				G. Zhao, A. Linke, X. Xu, M. Ochoa, F. Belloni, L. Belardinelli, and T. H. Hintze Comparative Profile of Vasodilation by CVT-3146, a Novel A2A Receptor Agonist, and Adenosine in Conscious Dogs J. Pharmacol. Exp. Ther., October 1, 2003; 307(1): 182 - 189. [Abstract] [Full Text] [PDF]

				L. M. Riou, S. Unger, M.-C. Toufektsian, M. Ruiz, D. D. Watson, G. A. Beller, and D. K. Glover Effects of Increased Lipid Concentration and Hyperemic Blood Flow on the Intrinsic Myocardial Washout Kinetics of 99mTcN-NOET J. Nucl. Med., July 1, 2003; 44(7): 1092 - 1098. [Abstract] [Full Text] [PDF]

				M. T. Kearney, K. A. A. Fox, A. J. Lee, R. J. Prescott, A. M. Shah, P. D. Batin, W. Baig, S. Lindsay, T. S. Callahan, W. E. Shell, et al. Predicting death due to progressive heart failure in patients with mild-to-moderate chronic heart failure J. Am. Coll. Cardiol., November 20, 2002; 40(10): 1801 - 1808. [Abstract] [Full Text] [PDF]

				L. M. Riou, M. Ruiz, G. W. Sullivan, J. Linden, H. Leong-Poi, J. R. Lindner, T. D. Harris, G. A. Beller, and D. K. Glover Assessment of Myocardial Inflammation Produced by Experimental Coronary Occlusion and Reperfusion With 99mTc-RP517, a New Leukotriene B4 Receptor Antagonist That Preferentially Labels Neutrophils In Vivo Circulation, July 30, 2002; 106(5): 592 - 598. [Abstract] [Full Text] [PDF]

				D. K. Glover, M. Ruiz, K. Takehana, F. D. Petruzella, L. M. Riou, J. M. Rieger, T. L. Macdonald, D. D. Watson, J. Linden, and G. A. Beller Pharmacological Stress Myocardial Perfusion Imaging With the Potent and Selective A2A Adenosine Receptor Agonists ATL193 and ATL146e Administered by Either Intravenous Infusion or Bolus Injection Circulation, September 4, 2001; 104(10): 1181 - 1187. [Abstract] [Full Text] [PDF]

				Z. Gao, Z. Li, S. P. Baker, R. D. Lasley, S. Meyer, E. Elzein, V. Palle, J. A. Zablocki, B. Blackburn, and L. Belardinelli Novel Short-Acting A2A Adenosine Receptor Agonists for Coronary Vasodilation: Inverse Relationship between Affinity and Duration of Action of A2A Agonists J. Pharmacol. Exp. Ther., July 1, 2001; 298(1): 209 - 218. [Abstract] [Full Text]

				A. K. Hinschen, R. B. Rose'Meyer, and J. P. Headrick Age-related changes in adenosine-mediated relaxation of coronary and aortic smooth muscle Am J Physiol Heart Circ Physiol, May 1, 2001; 280(5): H2380 - H2389. [Abstract] [Full Text] [PDF]

				E. Leistad, K. Ohmori, T. A. Peterson, G. Christensen, and A. N. DeMaria Quantitative assessment of myocardial perfusion during graded coronary artery stenoses by intravenous myocardial contrast echocardiography J. Am. Coll. Cardiol., February 1, 2001; 37(2): 624 - 631. [Abstract] [Full Text] [PDF]

				Z.-X. He, E. Cwajg, W. Hwang, C. J. Hartley, E. Funk, L. H. Michael, and M. S. Verani Myocardial Blood Flow and Myocardial Uptake of 201Tl and 99mTc-Sestamibi During Coronary Vasodilation Induced by CGS-21680, a Selective Adenosine A2A Receptor Agonist Circulation, July 25, 2000; 102(4): 438 - 444. [Abstract] [Full Text] [PDF]

				G. A. Beller and B. L. Zaret Contributions of Nuclear Cardiology to Diagnosis and Prognosis of Patients With Coronary Artery Disease Circulation, March 28, 2000; 101(12): 1465 - 1478. [Full Text] [PDF]

				A. R. Jayaweera, K. Wei, M. Coggins, J. P. Bin, C. Goodman, and S. Kaul Role of capillaries in determining CBF reserve: new insights using myocardial contrast echocardiography Am J Physiol Heart Circ Physiol, December 1, 1999; 277(6): H2363 - H2372. [Abstract] [Full Text] [PDF]

				A. Z. Linka, J. Sklenar, K. Wei, A. R. Jayaweera, D. M. Skyba, and S. Kaul Assessment of Transmural Distribution of Myocardial Perfusion With Contrast Echocardiography Circulation, November 3, 1998; 98(18): 1912 - 1920. [Abstract] [Full Text] [PDF]

				J. C. Shryock, S. Snowdy, P. G. Baraldi, B. Cacciari, G. Spalluto, A. Monopoli, E. Ongini, S. P. Baker, and L. Belardinelli A2A-Adenosine Receptor Reserve for Coronary Vasodilation Circulation, August 18, 1998; 98(7): 711 - 718. [Abstract] [Full Text] [PDF]

				K. Wei, A. R. Jayaweera, S. Firoozan, A. Linka, D. M. Skyba, and S. Kaul Quantification of Myocardial Blood Flow With Ultrasound-Induced Destruction of Microbubbles Administered as a Constant Venous Infusion Circulation, February 10, 1998; 97(5): 473 - 483. [Abstract] [Full Text] [PDF]

This Article Abstract Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Glover, D. K. Articles by Beller, G. A. Search for Related Content PubMed PubMed Citation Articles by Glover, D. K. Articles by Beller, G. A.