Potent and selective nonpeptidic inhibitors of procollagen C-proteinase.

6-Cyclohexyl-N-hydroxy-3-(1,2,4-oxadiazol-5-yl)hexanamides were previously disclosed as inhibitors of procollagen C-proteinase (PCP) culminating in the identification of amide 1. Our objective was to discover a second inhibitor that would have improved affinity for PCP and to optimize properties for transepidermal delivery (TED) to intact skin. Further investigation of this template identified a number of potent PCP inhibitors (IC50 values of 2-6 nM) with improved TED flux. Sulfonamide 56 had excellent PCP enzyme activity when measured with a peptide substrate (Ki 8.7 nM) or with the endogenous substrate procollagen (IC50 3.4 nM) and demonstrates excellent selectivity over MMPs involved in wound healing (>10 000-fold). In the fibroplasia model, 56 inhibited deposition of insoluble collagen by 76 +/- 2% at 10 microM and was very effective at penetrating human skin in vitro with a TED flux of 1.5 microg/cm2/h, which compares favorably with values for agents that are known to penetrate skin well in vivo. Based on this profile, 56 (UK-421,045) was selected as a candidate for further preclinical evaluation as a topically applied, dermal anti-scarring agent.

Selective urokinase-type plasminogen activator inhibitors. 4. 1-(7-sulfonamidoisoquinolinyl)guanidines.

1-isoquinolinylguanidines were previously disclosed as potent and selective inhibitors of urokinase-type plasminogen activator (uPA). Further investigation of this template has revealed that incorporation of a 7-sulfonamide group furnishes a new series of potent and highly selective uPA inhibitors. Potency and selectivity can be achieved with sulfonamides derived from a variety of amines and is further enhanced by the incorporation of sulfonamides derived from amino acids. The binding mode of these 1-isoquinolinylguanidines has been investigated by X-ray cocrystallization studies. uPA inhibitor 26 was selected for further evaluation based on its excellent enzyme potency (Ki 10 nM) and selectivity profile (4000-fold versus tPA and 2700-fold versus plasmin). In vitro, compound 26 is able to inhibit exogenous uPA in human chronic wound fluid (IC50=0.89 microM). In vivo, in a porcine acute excisional wound model, following topical delivery, compound 26 is able to penetrate into pig wounds and inhibit exogenous uPA activity with no adverse effect on wound healing parameters. On the basis of this profile, compound 26 (UK-371,804) was selected as a candidate for further preclinical evaluation for the treatment of chronic dermal ulcers.

Integrative pharmacology and drug discovery--is the tide finally turning?

In vivo animal experiments are essential (and a regulatory requirement) to demonstrate the potential efficacy, safety and pharmacodynamic/pharmacokinetic profile of candidate drugs. However, the number of pharmacologists (and other bioscientists) with integrative (in vivo) pharmacology skills has been in decline for a number of years, as have the opportunities for students to learn such skills. This article reviews some recent initiatives that are underway to rebuild this essential skills base in the United Kingdom and the United States. Partnerships between industry and national funders of research and education have proved to be a particularly effective approach to support this strategically important area of biological science.

Effect of endothelin antagonists on the renal haemodynamic and tubular responses to ischaemia-reperfusion injury in anaesthetised rats.

In this investigation we have evaluated whether blockade of endothelin receptors influenced the renal haemodynamic and excretory responses to a period of ischaemia and reperfusion in the anaesthetised rat. The renal artery was occluded for 30 min and renal haemodynamic and excretory function followed for 90 min of reperfusion while either saline, the non-selective endothelin 1 receptor (ET(A)/ET(B)) antagonist SB209670 or the selective ET(A) receptor antagonist UK-350,926 was infused. In the post-ischaemic period, renal cortical and medullary perfusions were reduced by 40-50 %. When SB209670 was administered (30 micro g kg(-1) min(-1) I.V.) for 30 min before, during and for 90 min after renal artery occlusion, cortical and medullary perfusions returned to baseline levels, responses different from those obtained during saline infusion (both P < 0.05). In the presence of UK-350,926 (30 micro g kg(-1) min(-1) I.V.), perfusion in the medulla returned to baseline on clamp removal whereas that in the cortex remained depressed (P < 0.05). Renal ischaemia for 30 min decreased glomerular filtration rate during reperfusion and increased urine flow and sodium excretion 5- to 15-fold. UK-350,926 (30 micro g kg(-1) min(-1) I.V.) reduced (P < 0.05) fluid excretion prior to ischaemia but during reperfusion, glomerular filtration rate returned to basal levels and there were progressive increases in fluid excretion which were smaller compared to the saline-treated group (all P < 0.05). The ischaemic challenge may cause release of endothelin, which acts on ET(B) receptors in the cortex and ET(A) receptors in the medulla to decrease perfusion. The blunted natriuresis and diuresis during blockade of ET(A) receptors may result from either a vascular or tubular action of endothelin.

The effect of endothelin antagonists on renal ischaemia-reperfusion injury and the development of acute renal failure in the rat.

BACKGROUND: It is recognized that endothelins are released in response to hypoperfusion and anoxia of the kidney and may be responsible for the consequent deterioration in renal function. This study examined the ability of a non-selective (SB209670) and ET(A)-selective (UK-350,926) endothelin antagonist to attenuate ischaemia-induced renal failure in unilaterally nephrectomized rats. METHODS: The animals were anaesthetized, drug infusion commenced, and the renal artery occluded for 30 min. The endothelin antagonists were given for 30 min before, during, and 60 min after the ischaemic period, at 10, 30 and 100 micro g/kg/min or for 60 min after the start of reperfusion. RESULTS: On day 1, following 30 min renal artery occlusion, there was a 95% reduction in glomerular filtration rate, an 8-10-fold increase in plasma creatinine, and 10-15-fold increases in fractional excretions of sodium and potassium, which were partially resolved on day 3 and normalized on day 8. The lowest dose of SB209670 was without effect on the renal functional responses but they were blunted (all P<0.05) by the highest dose. At 30 and 100 micro g/kg/min UK-350,926, the decreases in renal function subsequent to the ischaemic challenge were attenuated. Administration of UK-350,926 at 100 micro g/kg/min for 1 h starting 60 min after the start of reperfusion, had no effect on the magnitude of the renal disturbances over the first 3 days. CONCLUSIONS: The data show that both the ET(A)/ET(B) and selective ET(A)-receptor antagonist ameliorated the ischaemia-reperfusion injury when given in the peri-ischaemic period but not when the ET(A)-receptor antagonist was given for 60 min at 100 micro g/kg/min after the ischaemic period.