Interaction of PEGylated anti-hypertensive drugs, amlodipine, atenolol and lisinopril with lipid bilayer membrane: A molecular dynamics simulation study.

The interaction of PEGylated anti-hypertensive drugs, amlodipine, atenolol and lisinopril with lipid bilayer membrane dimyristoylphosphatidylcholine (DMPC) has been studied in nine different simulation systems consisting of 128 lipid molecules and appropriate number of water molecules by molecular dynamics method and by utilizing GROMACS software. The influences of PEGylation on the mentioned drugs and the differences in application of two types of spacer molecules on the performance of drugs and DMPC membrane have been evaluated and mass density of the components in the simulation box, mean square displacement (MSD), electrostatic potential, hydrogen bonding, radial distribution function (RDF), area per lipid, order parameter, and angle distribution of the component molecules including drug, DMPC and PEG has been investigated. Furthermore, umbrella sampling analysis indicated that, PEGylation of the drugs made amlodipine to behave more hydrophilic, whereas in case of lisinopril and atenolol, PEGylation made these drugs to behave more hydrophobic. In almost all of the simulated systems, PEGylation increased the diffusion coefficient of the drugs.

Effects of a domain-selective ACE inhibitor in a mouse model of chronic angiotensin II-dependent hypertension.

The somatic isozyme of ACE (angiotensin I-converting enzyme) comprises two distinct zinc-dependent catalytic domains with different substrate specificities for angiotensin I (cleaved selectively by the C-domain) and bradykinin (cleaved equally efficiently by both the N- and C-domains). Classical ACEIs (ACE inhibitors) target both domains, with side effects such as cough and angio-oedema being attributed, in part, to N-domain inhibition, probably through bradykinin accumulation. We questioned whether a novel C-domain-selective ACEI (lisW-S) has anti-hypertensive effects without influencing bradykinin status. AngII (angiotensin II)-dependent hypertension was studied in mice that express active human renin in the liver (TtRhRen). Compared with wild-type littermates, TtRhRen mice displayed cardiac hypertrophy and had significantly elevated SBP [systolic BP (blood pressure)] as determined by tail cuff sphygmomanometry (150±3 compared with 112±5 mmHg; P<0.05) and telemetry (163±3 compared with 112±2 mmHg; P<0.01). Treatment with the non-selective ACEI lisinopril (1 mg/kg of body weight per day via an osmotic mini-pump for 2 weeks) reduced SBP (127±3 compared with. 154±6; P<0.05). Similarly, treatment with the C-domain selective ACEI lisW-S (lisinopril-tryptophan; 3.6 mg/kg of body weight per day via an osmotic mini-pump for 2 weeks) reduced BP. Treatment with lisinopril or lisW-S significantly reduced levels of AngII in kidneys (~4-fold; P<0.001). Ang-(2-8) [angiotensin-2-8)] was significantly reduced by lisinopril, but not by lisW-S. Plasma bradykinin levels were significantly increased only in the lisinopril group. These data suggest that C-domain-selective ACEIs reduce BP and AngII levels similarly to classical ACEIs. C-domain-selective ACEIs have the potential to avoid undesirable effects on the bradykinin system common to classic ACEIs and may represent a novel approach to the treatment of hypertension.

Targeted catalytic inactivation of angiotensin converting enzyme by lisinopril-coupled transition-metal chelates.

A series of compounds that target reactive transition-metal chelates to somatic angiotensin converting enzyme (sACE-1) have been synthesized. Half-maximal inhibitory concentrations (IC(50)) and rate constants for both inactivation and cleavage of full-length sACE-1 have been determined and evaluated in terms of metal chelate size, charge, reduction potential, coordination unsaturation, and coreactant selectivity. Ethylenediaminetetraacetic acid (EDTA), nitrilotriacetic acid (NTA), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), and tripeptide GGH were linked to the lysine side chain of lisinopril by 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride/N-hydroxysuccinimide coupling. The resulting amide-linked chelate-lisinopril (EDTA-lisinopril, NTA-lisinopril, DOTA-lisinopril, and GGH-lisinopril) conjugates were used to form coordination complexes with iron, cobalt, nickel, and copper, such that lisinopril could mediate localization of the reactive metal chelates to sACE-1. ACE activity was assayed by monitoring cleavage of the fluorogenic substrate Mca-RPPGFSAFK(Dnp)-OH, a derivative of bradykinin, following preincubation with metal chelate-lisinopril compounds. Concentration-dependent inhibition of sACE-1 by metal chelate-lisinopril complexes revealed IC(50) values ranging from 44 to 4500 nM for Ni-NTA-lisinopril and Ni-DOTA-lisinopril, respectively, versus 1.9 nM for lisinopril. Stronger inhibition was correlated with smaller size and lower negative charge of the attached metal chelates. Time-dependent inactivation of sACE-1 by metal chelate-lisinopril complexes revealed a remarkable range of catalytic activities, with second-order rate constants as high as 150,000 M(-1) min(-1) (Cu-GGH-lisinopril), while catalyst-mediated cleavage of sACE-1 typically occurred at much lower rates, indicating that inactivation arose primarily from side chain modification. Optimal inactivation of sACE-1 was observed when the reduction potential for the metal center was poised near 1000 mV, reflecting the difficulty of protein oxidation. This class of metal chelate-lisinopril complexes possesses a range of high-affinity binding to ACE, introduces the advantage of irreversible catalytic turnover, and marks an important step toward the development of multiple-turnover drugs for selective inactivation of sACE-1.

Characterization of domain-selective inhibitor binding in angiotensin-converting enzyme using a novel derivative of lisinopril.

Human ACE (angiotensin-converting enzyme) (EC 3.4.15.1) is an important drug target because of its role in the regulation of blood pressure via the renin-angiotensin-aldosterone system. Somatic ACE comprises two homologous domains, the differing substrate preferences of which present a new avenue for domain-selective inhibitor design. We have co-crystallized lisW-S, a C-domain-selective derivative of the drug lisinopril, with human testis ACE and determined a structure using X-ray crystallography to a resolution of 2.30 A (1 A=0.1 nm). In this structure, lisW-S is seen to have a similar binding mode to its parent compound lisinopril, but the P2' tryptophan moiety takes a different conformation to that seen in other inhibitors having a tryptophan residue in this position. We have examined further the domain-specific interactions of this inhibitor by mutating C-domain-specific active-site residues to their N domain equivalents, then assessing the effect of the mutation on inhibition by lisW-S using a fluorescence-based assay. Kinetics analysis shows a 258-fold domain-selectivity that is largely due to the co-operative effect of C-domain-specific residues in the S2' subsite. The high affinity and selectivity of this inhibitor make it a good lead candidate for cardiovascular drug development.

Synthesis and evaluation of a series of 99mTc(CO)3+ lisinopril complexes for in vivo imaging of angiotensin-converting enzyme expression.

In animal models of cardiac disease and in human congestive heart failure, expression of angiotensin-converting enzyme (ACE) is upregulated in the failing heart and has been associated with disease progression leading to cardiac failure and fibrosis. To develop probes for imaging ACE expression, a series of di(2-pyridylmethyl)amine (D) chelates capable of binding M(CO)3+ (M = technetium, rhenium) was conjugated to lisinopril by acylation of the epsilon-amine of the lysine residue with a series of di(2-pyridylmethylamino)alkanoic acids where the distance of the chelator from the lisinopril core was investigated by varying the number of methylene spacer groups to produce di(2-pyridylmethyl)amine(Cx)lisinopril analogs: D(C4)lisinopril, D(C5)lisinopril, and D(C8)lisinopril. The inhibitory activity of each rhenium complex was evaluated in vitro against purified rabbit lung ACE and was shown to vary directly with the length of the methylene spacer: Re[D(C8)lisinopril], inhibitory concentration of 50% (IC50) = 3 nM; Re[D(C5)lisinopril], IC50 = 144 nM; and Re[D(C4)lisinopril], IC50 = 1,146 nM, as compared with lisinopril, IC50 = 4 nM. The in vivo specificity for ACE was determined by examining the biodistribution of the 99mTc-[D(C8)lisinopril] analog in rats with and without pretreatment with unlabeled lisinopril. Uptake in the lungs, a tissue that constitutively expresses ACE, was 15.2 percentage injected dose per gram at 10 min after injection and was dramatically reduced by pretreatment with lisinopril, supporting ACE-mediated binding in vivo. Planar anterior imaging analysis of 99mTc-[D(C8)lisinopril] corroborated these data. Thus, high-affinity 99mTc-labeled ACE inhibitor has been designed with potency similar to that of lisinopril and has been demonstrated to specifically localize to tissues that express ACE in vivo. This agent may be useful in monitoring ACE as a function of disease progression in relevant diseases such as heart failure.

The role and regulation of cardiac angiotensin-converting enzyme for noninvasive molecular imaging in heart failure.

Congestive heart failure is a pathologic condition characterized by progressive decrease in left ventricular contractility and consequent decline of cardiac output. There is convincing clinical and experimental evidence that the renin-angiotensin system (RAS) and its primary effector peptide, angiotensin II, are linked to the pathophysiology of interstitial fibrosis, cardiac remodeling, and heart failure. In addition to the traditional endocrine or circulating RAS, an active tissue RAS has been characterized. Tissue angiotensin-converting enzyme and locally synthesized angiotensin II, for example, by chymase, exert local trophic effects that modulate gene expression, which regulates growth and proliferation in both myocytes and nonmyocytes. The existence of the tissue RAS offers an opportunity for targeted imaging, which may be of considerable value for guiding medical therapy.

Evidence for tissue angiotensin-converting enzyme in explanted hearts of ischemic cardiomyopathy using targeted radiotracer technique.

UNLABELLED: This study aimed to determine the magnitude and distribution of tissue angiotensin-converting enzyme (ACE), mast-cell chymase, and angiotensin II, type 1, plasma membrane receptor (AT1R), in relation to collagen replacement in infarcted and noninfarcted left ventricular myocardial segments. A new radiotracer, 18F-fluorobenzoyl-lisinopril (FBL), was synthesized without compromising its affinity for tissue ACE. METHODS: Five- to 10-microm contiguous short-axis slices of explanted hearts from 3 patients with ischemic cardiomyopathy were incubated in vitro with FBL, with and without 10(-6) M lisinopril. Tissue radioactivity was recorded as a function of position in photostimulating luminescence units (PSL). Immunohistochemistry studies were performed with mouse monoclonal antibody against ACE, anti-mast cell chymase, and polyclonal antibody against the human AT1R. RESULTS: There was specific binding of FBL to ACE; mean FBL binding was 6.6 +/- 5.2 PSL/mm2, compared with 3.4 +/- 2.5 PSL/mm2 in segments incubated in solution containing cold, 10(-6) M lisinopril (P < 0.0001). Mean FBL binding was 6.3 +/- 4.5 PSL/mm2 in infarcted, 7.6 +/- 4.7 PSL/mm2 in periinfarcted, and 5.0 +/- 1.0 PSL/mm2 in remote, noninfarcted (P < 0.02 vs. periinfarcted) segments. The autoradiographic observations concerning FBL binding were confirmed by ACE and AT1R immunoreactivity. Distribution of mast cell chymase differed from ACE, as a higher number of mast cells was present in the remote, noninfarcted myocardium than in the periinfarcted myocardium (5.1 +/- 3.2 vs. 3.2 +/- 2.2 mast cells per field, P < 0.001). The number of mast cells in ischemic hearts exceeded that in normal hearts (4.2 +/- 2.7 vs. 1.5 +/- 1.2 mast cells per field, x200, P < 0.001). CONCLUSION: FBL binds specifically to ACE. The binding is nonuniform in infarcted, periinfarcted, and remote, noninfarcted segments, and there is apparently increased ACE activity in the juxtaposed areas of replacement fibrosis. On the other hand, the distribution of mast cell chymase appears nonuniform and disparate from ACE.

[A new approach to treating secondary pulmonary hypertension].

OBJECTIVE: to study the clinical and hemodynamic effects and safety of the ACE inhibitor dirotone and AT1-receptor antagonist losartan in secondary pulmonary hypertension (PH) in patients with chronic obstructive bronchitis (COB). SUBJECTS AND METHODS: The time course of changes in hemodynamic parameters and diurnal BP profile in 48 patients with COB concurrent with Functional Class (FC) III-IV secondary PH, receiving 4-week therapy including dirotone and losartan. RESULTS: The inclusion of dirotone or losartan into the combined therapy of patients with secondary pulmonary hypertension resulted in alleviated PH, caused positive changes in the right cardiac cavities, and normalized diurnal BP variables. Therapy with dirotone was effective in COB patients with the clinical signs of FC III PH only during its course use, that with losartan was beneficial in FC III and IV PH. The long-term outpatient use of losartan in individually adjusted doses produced pronounced beneficial clinical effects and led to significantly improved hemodynamic parameters. CONCLUSION: The ACE inhibitor dirotone and the AT1-receptor antagonist losartan may be recommended for the correction of hemodynamic disorders in secondary PH in patients with COB.