1. A “Proof-of-Concept” Study for Evaluating an Anti-Hypertensive in an Awake, Chronically Instrumented, Pharmacologically Hypertensive Monkey
Michael A. Hawk1, Tom Vinci1, Craig Hassler1, and Robert Hamlin2 
1Battelle, Columbus, OH, USA 2The Ohio State University, Columbus, OH, USA
ABSTRACT
RESULTS
Systemic arterial hypertension is of epidemic proportions (~30% of the U.S. population, ~1 billion, Worldwide)1 and results in significant morbidity and mortality. Only ~1/3 of Americans have their hypertension controlled.2 Thus there is enormous interest by, and value to, the pharmaceutical industry and to drug regulatory agencies worldwide to develop safe and efficacious drugs to manage systemic arterial hypertension. The importance of systemic arterial pressure is recognized by S7A and S7B guidelines. Proof of safety and efficacy is relevant to the arena of Safety Pharmacology, and preclinical studies conducted on infrahuman mammals as surrogates for man are of obvious importance. Whereas, most of these studies have been conducted on rodents and dogs, studies on non-human primates are appealing because of anatomical and physiological similarities between them and man. This proof-of-concept study was conducted on awake, chronically instrumented cynomolgus monkeys with systemic arterial hypertension produced by constant rate infusion of angiotensin II. After production and maintenance of systemic arterial hypertension, monkeys were given escalating doses of sodium nitroprusside to demonstrate the usefulness of the model for evaluating a known antihypertensive. Additionally, we established baroreceptor function from the change in heart rate vs. the change in blood pressure in response to sodium nitroprusside.
Monkeys remained calm throughout the duration of surveillance, and no apparent alteration in activity occurred in response to changing flow rates of the constant infusion pumps. Recordings of systemic arterial pressures and heart rates of quality satisfactory for interpretation were obtained continuously for both monkeys.
Data for monkeys B0571 in response to 0.42 ug/kg/minute of angiotensin II is shown (Figure 1A). In response to angiotensin II, mean systemic arterial pressure increased abruptly from
~70 mmHg to ~120 mmHg, then decreased to ~110 mmHg; correspondingly, heart rate decreased precipitously from
~210 beats/minute to ~130 beats/minute, then increased to ~230 beats/minute. Data for monkeys B0142 in response to
0.42 ug/kg/minute of angiotensin II is shown (Figure 1B). In response to angiotensin II, mean systemic arterial pressure increased abruptly from ~100 mmHg to ~160 mmHg, then decreased to ~130 mmHg; correspondingly, heart rate decreased precipitously from ~170 beats/minute to ~150 beats/minute, then increased to ~190 beats/minute.
The abrupt increase in systemic arterial pressure resulted from constriction of smooth muscle in systemic arterioles, an expected response to angiotensin II and to the release and decreased re-uptake of norepinephrine. Other mechanisms (e.g., neuroendocrines, diseases, drugs) participate in either blunting or decreasing the response to angiotensin II. The abrupt decrease in heart rate resulted from the expected high pressure baroreceptor reflex producing increased parasympathetic efferent traffic to the SA node. A later and more sustained increase in heart rate resulted from the release, and prevention of re-uptake, of norepinephrine.
The responses of systemic arterial pressure and heart rate to escalating doses of sodium nitroprusside are shown for monkey B0142 (Figure 2). It is clear that escalating doses of sodium nitroprusside produced escalating decrements in systemic arterial pressure (Figure 3). For the terminal dose of sodium nitroprusside, Tau for the decrease in pressure was 1.8 minutes, and for the return of pressure after cessation of sodium nitroprusside was 2.4 minutes. Tau for the increase in heart rate was 3.0 minutes, and for the return to normal was 7.2 minutes. These durations expressed by Tau are influenced by many factors (e.g., rate of infusion, rate of binding to receptors, components of the baroreceptor reflex arc); therefore, Tau cannot be interpreted in this pilot study. However, this does demonstrate the feasibility of characterizing the time course of events as well as the magnitudes of changes in heart rate and blood pressures.
The responses of systemic arterial pressure and heart rate to angiotensin II demonstrated the clear dose-response relationships between changes in heart rate and changes in pressure (Figure 4) of the “hypertensive” monkeys responding to escalating doses of sodium nitroprusside. This preparation shows a potential to quantify effects of other hypotensive test articles, in awake chronically instrumented, artificially produced hypertensive monkeys. Furthermore, this model demonstrates the ability to quantify baroreceptor sensitivity (Figure 5), an important pharmacological effect, which may translate to morbidity.3,4
B0571 Responses to 0.42 ug/kg/min of Angiotensin II
Figure 1A. B0571 Response to Angiotensin II
B0142 Responses to 0.42 ug/kg/min of Angiotensin II
INTRODUCTION
Figure 1B. B0142 Response to Angiotensin II
There are two major types of hypertension, primary or essential hypertension and secondary hypertension. Additionally there are four less frequently found types of hypertension that, include malignant, isolated systolic, white coat and resistant hypertension.
Primary hypertension is also called essential hypertension, it is by far the most common type of hypertension, and it is constitutes about 95% of cases. Many factors can predispose to primary hypertension: obesity, diet, environment, stress, sedentary life-style, high sodium consumption, and family history.
Secondary hypertension is caused by an underlying condition such as hyperthyroidism, Cushing's syndrome, renal disease, sleep apnea, and pregnancy. When treated, secondary hypertension will diminish.
The purpose of this study was to develop an awake, chronically instrumented cynomolgus monkey model of systemic arterial hypertension produced by constant rate infusion of angiotensin II. After production of hypertension, monkeys were given escalating doses (0.75, 1.5, 3.0, 6.0 and 6.0 µg/kg/minute) of a known vasodilator (sodium nitroprusside) to demonstrate the usefulness of the model for evaluating a known antihypertensive. Additionally, we interrogated baroreceptor function described as changes in heart rate vs. changes in blood pressure in response to sodium nitroprusside.
B0142 Responses to Escalating Doses of 1.5, 3 & 6 ug/kg/min of Nitroprusside
Figure 2. Response to Sodium Nitroprusside
Figure 3. Dose Response Curve
METHODS
B0142 Responses to Angiotensin II Followed by Escalating Doses of 0.75, 1.5, 3, 6 & 6 ug/kg/min of Nitroprusside
Two, young-mature cynomolgus monkeys were instrumented previously with telemetry devices (Data Sciences International, St. Paul, Minnesota) to acquire electrocardiogram (ECG) and systemic arterial pressure (important physiologic signals) remotely without perturbation of the monkeys. Additionally, a dual port infusion catheter was placed in peripheral veins, for constant infusions of angiotensin II and/or sodium nitroprusside. Control of the infusion pumps (SAI, Libertyville, Illinois) was performed remotely so physiological signals could be recorded continuously, and the monkeys did not appear to perceive either the onset or termination of the infusion pump activity.
All studies were performed between 10 AM and 3 PM. Baseline recordings were made for >1 hour, after which angiotensin II was infused at 0.42 ug/kg/minute throughout the experiment to elevate mean systemic arterial pressure from a mean of ~100 mmHg to ~180 mmHg. After systemic arterial pressure had been sustained at a plateau for ~30 minutes, escalating doses (0.75, 1.5, 3.0, 6.0 and 6.0 µg/kg/minute) of sodium nitroprusside were infused at 30-minute increments.
Plots were made of heart rates and systemic arterial pressures (systolic, mean, and diastolic) with each point being an average of 10 seconds. Times, between drugs and cessation of drugs, until new steady states were achieved are expressed as Tau (the time required for the variable to reach 63% of the new steady state). A dose-response curve was generated for changes in systemic arterial pressure vs. sodium nitroprusside doses. In addition, baroreceptor function was assessed by changes in heart rate vs. changes in systemic arterial pressure.
CONCLUSIONS
Systemic arterial pressure is determined by a plethora of factors and it is well known that systemic arterial hypertension has many etiologies. If abnormal activation of vascular smooth muscle is operative in the cause of hypertension (i.e., not due to an elevated cardiac output), then this model of hypertension induced by angiotensin II may be useful for exploring test articles of potential use to treat hypertension. The non-human primate has proven useful to predict both safety and efficacy and, although there are known exceptions, it may be presumed that results of studies conducted on non-human primates may be extrapolated more directly to man than results from studies on non-primates. The model presented here may be particularly attractive.
Figure 4. Sodium Nitroprusside Dose Response in a Hypertensive Model
REFERENCES
Kearney PM, Whelton M, Reynolds K, Muntner P, Whelton PK, He J (2005). "Global burden of hypertension: analysis of worldwide data". Lancet 365 (9455): 217–23.
“US Trends in Prevalence, Awareness, Treatment, and Control of Hypertension, ,” Egan, BM, et al., JAMA 2010; 303(20): doi: /jama
“Hypotension.” National Heart, Lung, and Blood Institute. Accessed Feb. 14,
Kaufmann H, et al. “Mechanisms, causes, and evaluation of orthostatic and postprandial hypotension.” Accessed Feb. 14, 2011.
Figure 5. Baroreceptor Sensitivity