Drug discovery and development in the field of cardiovascular disease. An interview with Dr William Greenlee by Alexandra Hemsley.

William Greenlee, the founder and president of MedChem Discovery Consulting LLC, has over three decades of research experience in industry at Merck Research Laboratories and the Schering-Plough Research Institute. He has contributed to the advancement of 20 drug candidates into preclinical development, 11 of which have progressed into clinical studies. Dr Greenlee was a member of the team that discovered the marketed drugs Vasotec™ and Prinivil™, and has led research teams that discovered several drug candidates that are now in clinical trials. His achievements and significant contributions to the field have been recognized with the presentation of a number of local and national awards, including the Alfred Burger Award in Medicinal Chemistry from the American Chemical Society and the Pennsylvania Drug Discovery Institute Drug Discovery Award. He was inducted into the American Chemical Society Medicinal Chemistry Hall of Fame in 2006. Dr Greenlee was elected a Fellow of the American Association for the Advancement of Science in 2007 and an American Chemical Society Fellow in 2009.

Rapid and sensitive liquid chromatography/tandem mass spectrometry method for simultaneous determination of enalapril and its major metabolite enalaprilat, in human plasma: application to a bioequivalence study.

A rapid and most sensitive method for simultaneous determination of enalapril (ENP) and its metabolite, enalaprilat (ENPT), in human plasma using ESI-LC-MS/MS (electrospray ionization liquid chromatography tandem mass spectrometry) positive ion multiple reactions monitoring (MRM) mode, was developed and validated. The procedure involves a simple solid-phase extraction (SPE) followed by evaporation of the sample. Chromatographic separation was carried out on a Hypurity C(18) column (50 mm × 4.6 mm, 5 µm) with an isocratic mobile phase and a total run time of 2.0 min only. The MRM of ENP and ENPT is 377.10 → 234.20 and 349.20 → 206.10 respectively. The standard calibration curves showed excellent linearity within the range of 0.064 to 431.806 ng/mL for ENA and 0.064 to 431.720 ng/mL for ENPT (r ≥ 0.990). This is the only method which can quantitate upto 0.064 ng/mL for both ENP and ENPT in a single run with the shortest analysis time. In matrix effect experiment, this method shows a % CV (% coefficients of variation) of less than 5, which means that the proposed method is free from any kind of irregular ionization process. This method was successfully applied to a pharmacokinetic study after oral administration of enalapril maleate 20 mg tablet in Indian healthy male volunteers.

Assessment of drug-induced QT interval prolongation in conscious rabbits.

INTRODUCTION: Most preclinical trials are designed to identify potential torsadogenicity test only for surrogates of torsade de pointes, most commonly prolongation of the heart rate corrected QT interval (QTc). This study was conducted to determine which correction method best accounts for the effects of changes in the RR interval on the QT interval of conscious rabbits. This study was also conducted to validate the use of conscious, sling-trained rabbits to assess the QTc interval, and to evaluate the reliability and accuracy of this preparation in predicting drug-induced QTc prolongation in humans. METHODS: ECGs were recorded via bipolar transthoracic ECG leads in 7 conscious rabbits previously trained to rest quietly in slings. The heart rate was slowed with 2.0 mg/kg zatebradine to assess the effects of heart rate on the QT interval. The same ECG and sling preparation was used to evaluate the effects in of three drugs known to be torsadogenic in humans (cisapride, dofetilide and haloperidol), two drugs known to be non-torsadogenic in humans (propranolol and enalaprilat) and a control article (vehicle). All of the test articles were administered intravenously to 4 rabbits, and both RR and QT intervals were measured and the corrected QT values were calculated by an investigator blinded to the test article, utilizing our own algorithm (QTc=QT/(RR)(0.72)) which permitted the least dependency of QTc on RR interval. RESULTS: The following regression equations were obtained relating QT to RR: QT=2.4RR(0.72), r(2)=0.79, with RR intervals varying between 210 and 350 ms. QTc lengthened significantly in all conscious rabbits given intravenous cisapride, dofetilide and haloperidol (p<0.05), and QTc did not change with DMSO (vehicle control), propranolol or enalaprilat. DISCUSSION: Results indicate that a bipolar transthoracic ECG recorded in conscious, sling-trained rabbits may provide an easy and economical methodology useful in predicting QTc lengthening of novel pharmacological entities.

Functional testing: ACEI renography.

Angiotensin-converting enzyme inhibition (ACEI) renography is the only imaging examination that tests directly for the presence of renovascular hypertension (RVH); other imaging examinations test only for the presence of renal artery stenosis (RAS). Consensus panels have recommended that ACEI renograms be interpreted as low, intermediate, or high probability for RVH. ACEI renography is highly accurate in patients with normal renal function and suspected RVH. In this patient population, the sensitivity and specificity of ACEI renography for RAS are approximately 90%; as an initial approach, angiography is not cost effective. Data from 10 studies evaluating cure or improvement in blood pressure in 291 patients undergoing revascularization showed the mean positive predictive value of ACEI renography to be 92%. When azotemic patients present with suspected RVH, as many as 50% of patients may have an intermediate probability ACEI renogram and the sensitivity of detecting RVH falls to approximately 80% even when intermediate and high probability tests are combined.

Assessment of aortic line of elasticity using polynomial regression analysis.

BACKGROUND: The aim of this study was to assess the sigmoid line of elasticity in the human aorta. METHODS AND RESULTS: The pressure-diameter relation was measured in the descending aorta in 120 subjects. In an additional group of 6 subjects, transient vena caval occlusion produced 5 sets of pressure-diameter data. We found that the best fit curve of the pooled pressure-diameter data was a third-order polynomial. A polynomial equation was used to calculate the sigmoid line of elasticity in the entire population and after the administration of diltiazem (15 patients) or enalaprilat (10 patients). The sigmoid line of elasticity was significantly different with respect to age (P<0.001), history of hypertension (P<0.004), and hypercholesterolemia (P<0.02). The difference between the transition point and the peak systolic pressure was increased in normal subjects compared with patients (P<0.0001). The sigmoid line shifted leftward and upward with diltiazem, but it remained unchanged with enalaprilat. During an average of 3 years of follow-up, 19 of 88 patients developed stroke (n=4), unstable angina (n=8), acute myocardial infarction (n=4), or acute pulmonary edema (n=3). CONCLUSIONS: This approach provides a quantitative evaluation of the aortic line of elasticity, which can differentiate the intrinsic from the extrinsic aortic elastic properties. Furthermore, it is a powerful and independent risk factor for cardiovascular events.

Angiotensinergic versus nonangiotensinergic hemodynamic effects of converting enzyme inhibition in patients with chronic heart failure. Assessment by acute renin and converting enzyme inhibition.

BACKGROUND: The contribution of nonangiotensinergic effects of converting enzyme inhibitors to their hemodynamic effects in patients with chronic heart failure is not clear. A comparison of the effects of renin and converting enzyme inhibition should help to clarify this issue. METHODS AND RESULTS: Thirty-six patients with chronic heart failure (New York Heart Association class II or III) were randomly assigned to receive double-blind either intravenous placebo, the renin inhibitor remikiren, or the converting enzyme inhibitor enalaprilat followed by coinfusion of a second placebo infusion, the addition of remikiren to enalaprilat, or the addition of enalaprilat to remikiren, respectively. Systemic hemodynamics (Swan-Ganz and radial artery catheters) were measured before (rest and submaximal recumbent bicycle ergometry), during (rest), and at the end (rest and exercise) of each 45-minute single- or combination-infusion period. Placebo did not change hemodynamics or renin activity. Effective inhibition of the renin-angiotensin system by remikiren and enalaprilat was indicated by increases of plasma immunoreactive renin together with rapid and complete inhibition of renin activity after remikiren and an increase after enalaprilat (all P < or = .05). Remikiren and enalaprilat rapidly and to a similar extent reduced resting blood pressure through a reduction of systemic vascular resistance, and these changes were significantly correlated to baseline plasma renin activity. Both compounds also decreased pulmonary artery, pulmonary capillary wedge, and right atrial pressures to a similar extent (P < .05). During exercise, pulmonary capillary wedge and right atrial pressures were equally reduced and stroke volume index was increased with remikiren and enalaprilat (P < .05) for both). The combination of converting enzyme with renin inhibition or vice versa did not cause additional hemodynamic changes. CONCLUSIONS: Specific renin inhibition in patients with chronic heart failure produces short-term hemodynamic effects that are almost indistinguishable from those of converting enzyme inhibition. This finding and the lack of additional effects of converting enzyme inhibition added to renin inhibition suggest that nonangiotensinergic effects of converting enzyme inhibitors do not play a significant role in their short-term hemodynamic effects in patients with chronic heart failure.

Direct myocardial and coronary effects of enalaprilat in patients with dilated cardiomyopathy: assessment by a bilateral intracoronary infusion technique.

Angiotensin II elicits contractile responses in the coronary arteries and myocardial tissue, which suggests that blockade of the renin-angiotensin system by specific agents should lead to both coronary vasodilation and an alteration of left ventricular inotropism. The present work was designed to delineate--independently from its systemic effects--the intrinsic actions of an angiotensin converting-enzyme inhibitor on the coronary circulation and left ventricular function. To minimize peripheral effects, a bilateral intracoronary infusion of enalaprilat (0.05 mg.min-1, 1 ml.min-1 in each coronary artery) was performed in 16 patients with dilated cardiomyopathy. All patients had normal coronary arteriograms. In 12 patients (group I) the intracoronary infusion of enalaprilat resulted in minimal peripheral changes, with a 5% reduction in the mean aortic pressure (p less than .05) and no significant alteration in indexes of preload, i.e., left ventricular end-diastolic pressure and volume, or of afterload, i.e., left ventricular end-systolic stress and systemic resistances. Myocardial oxygen consumption was also unaffected by the intracoronary infusion of enalaprilat. Coronary vasodilation was demonstrated by a significant elevation of coronary sinus blood flow (+19%, from 181 +/- 73 to 214 +/- 79 ml.min-1, p less than .001) and a reduction of coronary resistance (-18%, from 0.51 +/- 0.17 to 0.41 +/- 0.15 mm Hg.ml-1.min, p less than .001), with a parallel increase in coronary sinus oxygen content and pressure (both p less than .05). Oxygen extraction by the myocardium was reduced (p less than .01).(ABSTRACT TRUNCATED AT 250 WORDS)