The importance of male underwear in cases of alleged sexual assault.

The potential evidential value of male underwear in cases of alleged sexual assault is often overlooked. Male underwear can be a critical item in the investigation of alleged sexual assaults. Body fluids/DNA, which may transfer to the penis during sexual contact, may in turn transfer to the inside front of the underwear, and persist for months or years, provided the underwear are not washed. Here, we demonstrate how the case circumstances drive the sampling strategy of male underwear, in order to maximize the effectiveness of the forensic analysis. Sampling considerations including recovery methods and sampling sequence are discussed, and a methodical examination strategy of male underwear is proposed. To highlight the pertinence of male underwear to the investigation of alleged sexual assaults, three real-life cases are discussed, in which male underwear were examined for multiple body fluids/DNA, and the findings obtained proved evidentially significant. The different cases demonstrate the versatility of male underwear examination in situations, where different body fluids and DNA may transfer based on the specific allegation, and emphasize how targeted sampling can allow the scientist to assess the probability of the findings based on two competing propositions. Accurate sampling strategies are imperative for robust probability assignment in evaluative reporting of scientific findings.

Early thymus and activation-regulated chemokine (TARC) reduction and response following panobinostat treatment in patients with relapsed/refractory Hodgkin lymphoma following autologous stem cell transplant.

Abstract In a phase 2 trial of panobinostat in 129 patients with relapsed or refractory Hodgkin lymphoma, exploratory analyses of chemokines and cytokines were prospectively performed in 109 patients to determine their association with clinical outcomes. Patients were categorized into two groups (reductions > median and reductions ≤ median) based on percentage change from baseline of log10 transformed measurements. Thymus and activation-regulated chemokine (TARC) was most strongly associated with clinical outcome. Early reduction of TARC was observed in responding patients, with the greatest reduction at cycle 1, day 15 (C1D15). Of 93 patients with C1D15 samples, there were three complete and 25 partial responses. The group with TARC reductions > median at C1D15 had more responders (18 [39%] vs. 10 [21%]), longer progression-free survival (10.6 vs. 4.9 months), shorter time to response and longer overall survival than the group with reductions ≤ median. This study is registered at www.ClinicalTrials.gov , NCT00742027.

CGS 35601 and its orally active prodrug CGS 37808 as triple inhibitors of endothelin-converting enzyme-1, neutral endopeptidase 24.11, and angiotensin-converting enzyme.

CGS 35601 is a potent triple inhibitor of endothelin-converting enzyme-1, neutral endopeptidase 24.11, and angiotensin-converting enzyme. It inhibited the activities of these three enzymes with IC50 values of 55, 2 and 22 nM, respectively. In conscious rats, CGS 35601 suppressed the big endothelin-1-induced pressor response by 82% and 72% at 30 and 120 minutes, respectively, following injection at a dose of 10 mg/kg, intravenously. At the same dose, CGS 35601 increased plasma atrial natriuretic peptide (ANP) immunoreactivity by 170% for up to 4 hours in conscious rats infused with ANP, and it inhibited the angiotensin I-induced pressor response by 74-94% within the first 2 hours after dosing. Similar in vivo activities were also observed with its orally active prodrug, CGS 37808. This compound blocked the big endothelin-1- induced pressor response by 71% and 67% at 30 and 120 minutes, respectively, after an oral dose of 10 mgEq/kg in conscious rats. It also increased plasma ANP immunoreactivity by 103% for up to 4 hours and inhibited the angiotensin I-induced pressor response by an average of 49% within the first 4 hours after the same dosing regimen. By suppressing the biosyntheses of endothelin-1 and angiotensin II, two potent vasoconstrictors, while simultaneously potentiating the circulating levels of ANP, a vasorelaxant and diuretic, CGS 35601 and CGS 37808 may represent novel agents for the treatment of cardiovascular and renal diseases.

Synthesis and biological activity of potent heterocyclic thiol-based inhibitors of endothelin-converting enzyme-1.

Directed screening of metalloprotease inhibitors identified CGS 30084 (1) as a potent inhibitor of endothelin-converting enzyme-1 (ECE-1) in vitro (IC(50)=77 nM). Herein we report the syntheses and biological activities of analogues containing modified biphenyl moieties, bearing heterocyclic proximal rings. Compound 20, the thioacetate ethyl ester prodrug derivative of compound 19a, was found to be an orally active and potent inhibitor of ECE-1 activity in rats.

Expression, purification and characterization of the monomeric and dimeric forms of soluble bovine endothelin converting enzyme-1a.

In this study, the catalytic domain of bovine endothelin converting enzyme-1a (ECE-1a) was cloned into a baculovirus transfer vector behind the human alkaline phosphatase signal sequence. The recombinant baculovirus was then used to infect High Five(TM) insect cells in suspension culture. Both the monomeric (85 kDa) and dimeric (170 kDa) forms of soluble ECE-1a were purified to electrophoretic homogeneity from concentrated culture media following sequential concanavalin A, SP-Sepharose, Mono Q and gel filtration column chromatography. Typically, approximately 11 mg of ECE-1a monomer and 6 mg of dimer were obtained from l litre of culture medium. No interconversion of the two forms was detected after purification. Both forms of ECE-1a had a pH optimum of 7.0, were maximally stimulated by NaCl at a concentration of 500 mM, and were inhibited to the same extent by metalloprotease inhibitors such as phosphoramidon and EDTA. However, in kinetic studies using big endothelin-1 (ET-1) as a substrate, the K(m) and k(cat) values for the monomer were 2.2 microM and 1.6 min(-1) respectively, while those of the dimer were 1.4 microM and 4.9 min(-1) respectively. These results show that, although the two forms of ECE-1a behave similarly in many aspects, the dimeric enzyme is more efficient in catalysing the conversion of big ET-1 to ET-1. The present protocol can be utilized to prepare large quantities of both forms of ECE-1a for further biochemical and structural characterization.

CGS 34226, a thiol-based dual inhibitor of endothelin converting enzyme-1 and neutral endopeptidase 24.11.

Endothelins (ETs) are potent vasoconstrictors and have been implicated in the pathogenesis of various cardiovascular and renal diseases. In contrast, atrial natriuretic peptide (ANP) is a potent vasorelaxant and diuretic agent, which is mainly degraded by neutral endopeptidase 24.11 (NEP) in vivo. Thus, compounds that can suppress the biosynthesis of ETs by inhibiting endothelin converting enzymes (ECEs), which catalyse the final step of post-translational processing of the vasoconstrictors, while simultaneously potentiating the levels of ANP by inhibiting NEP may have novel therapeutic utility. Through targeted screening of our compound library and subsequent optimization, CGS 34226 was identified as a potent, dual inhibitor of ECE-1 and NEP, inhibiting the enzymes with respective IC(50) values of 11 and 4.6 nM. In vivo, CGS 34226 suppressed the big endothelin-1 (big ET-1)-induced pressor response dose-dependently. At 15 and 90 min after an intravenous dose of 30 mg/kg in anaesthetized rats, this compound inhibited the big ET-1-induced effect by 79% and 65% respectively. In addition, CGS 34226 increased plasma ANP immunoreactivity by 120% up to 4 h after an intravenous dose of 10 mg/kg in conscious rats infused with ANP at a rate of 450 ng/kg per min, intravenously. These results show that CGS 34226 is a potent dual inhibitor of ECE-1 and NEP in vitro and in vivo and that the compound may represent a novel agent for the treatment of cardiovascular and renal dysfunction.

Effects of the ECE/NEP inhibitor CGS 34225 on the big ET-1-induced pressor response and plasma atrial natriuretic peptide concentration in conscious rats.

CGS 34226 is a thiol-containing, potent dual inhibitor of endothelin converting enzyme-1 (ECE-1) and neutral endopeptidase 24.11 (NEP) with IC(50) values of 11 and 5 nM respectively. The purpose of the present study was to characterize the inhibitory effects of CGS 34225, an orally active prodrug of CGS 34226, on ECE-1 and NEP in vivo. The effects on ECE-1 and NEP were assessed by determining the inhibition of big endothelin-1 (big ET-1)-induced increases in mean arterial pressure (MAP) and increases in plasma atrial natriuretic peptide (ANP) concentrations respectively, in conscious rats. Thirty and 120 min after the administration of vehicle, big ET-1 (0.3 nmol/kg, intravenously; i.v.) produced pressor responses of approximately 800 mmHg.min (area under the curve for change in MAPxtime). Treatment with CGS 34225 at 1 mgEq/kg, per os (p.o.), decreased the pressor effect of big ET-1 by 39 and 53% at 30 and 120 min respectively (P<0.05, both times). Increasing the dose of CGS 34255 to 30 mgEq/kg, p.o., resulted in greater inhibition, 84 and 92% (P<0.05) at 30 and 120 min respectively. Furthermore, at this higher dose, the inhibitory effect on ECE-1 was long-lasting, averaging 86, 75 and 30% (P<0.05, all times) at 4, 8 and 24 h respectively. In rats treated with vehicle, the infusion of ANP at 450 ng/kg per min i.v. resulted in plasma ANP concentrations of 3.9-4.8 ng/ml that remained relatively constant for 4 h. Treatment with CGS 34225 at 10 mgEq/kg, p.o., increased the ANP level to 7.7+/-1.0 and 10.6+/-1.8 ng/ml at 1 and 4 h after dosing (P<0.05, both times). These data demonstrate that CGS 34225 is a potent, orally active and long-acting inhibitor of ECE-1 and NEP in vivo. It is anticipated that compounds with this dual function may be useful in the treatment of cardiovascular diseases where the ET system plays a pathogenic role and the potentiation of ANP elicits therapeutic benefits.

Oral administration of an inhibitor of endothelin-converting enzyme attenuates cerebral vasospasm following experimental subarachnoid haemorrhage in rabbits.

Increasing evidence has implicated endothelin-1 (ET-1), a potent vasoconstrictive peptide, in the pathophysiology of cerebral vasospasm after subarachnoid haemorrhage (SAH). Endothelin-converting enzyme-1 (ECE-1), the protease involved in the final step of post-translational processing of ET-1, cleaves the inactive precursor big ET-1 at the Trp(21)-Val (22) peptide bond. In our previous study, we found that an inhibitor of ECE-1, CGS 26303, could prevent and reverse the arterial narrowing after SAH in rabbits. CGS 26393, a prodrug of CGS 26303, is an orally active, long-acting inhibitor of ECE-1. The present study examined the effects of CGS 26393 on the prevention and reversal of cerebral vasospasm after SAH. New Zealand white rabbits were subjected to experimental SAH by injecting autologous blood into the cisterna magna. In the prevention study, the drug was given orally 1 h before the induction of SAH. All drug treatments in the reversal study were initiated at 23 h after induction of SAH. One of three dosages (3, 10 or 30 mg/kg) of CGS 26393 or vehicle was administrated orally twice daily, and all animals were sacrificed by perfusion and fixation 48 h after SAH. Basilar arteries were removed and sectioned, and cross-sectional areas were measured. Cerebrospinal fluid (CSF) was collected prior to perfusion. Oral administration of CGS 26393 attenuated SAH-induced cerebral vasospasm in a dose-dependent manner in both the prevention and reversal groups. These effects achieved statistical significance at all dosages when compared with the SAH-only or SAH plus vehicle groups. Moreover, the attenuation of vasospasm following oral administration of CGS 26393 was more efficacious than that obtained with bolus injections of CGS 26303. The levels of free CGS 26303 in the CSF were increased in a dose-dependent manner in all three CGS 26393-treated groups. This study provides the first evidence that oral administration of an inhibitor of ECE-1, CGS 26393, is capable of preventing and reversing cerebral vasospasm following SAH. These findings also reinforce evidence demonstrating that treatment with an ECE-1 inhibitor is a potentially viable therapeutic approach for reducing cerebral vasospasm after SAH.

Dipeptide sulfonamides as endothelin ETA/ETB receptor antagonists.

Endothelin-1 (ET-1) is a potent mitogen and modulator of vascular tone. It is synthesized and released from endothelial cells and acts upon two receptor subtypes designated as ETA and ETB. In this study, a series of potent dipeptide sulfonamide dual-endothelin ETA/ETB receptor antagonists were prepared to investigate their potential benefit in vascular diseases. CGS 31398 inhibited [125I]ET-1 binding to human ETA and ETB receptors expressed in Chinese hamster ovary (CHO) cells (ETA/CHO, ETB/CHO) with respective IC50 values of 0.26 and 0.12 nM. However, in anesthetized rats, this compound markedly potentiated ET-1-induced renal vascular resistance, a response normally observed with selective ETB receptor antagonists. To determine whether species differences account for these results, a direct comparison was made between binding to rat and rabbit aortic membranes versus functional antagonism in isolated rat aortic rings. It was found that CGS 31398 had potent affinity for the ETA receptor in rat and rabbit aorta with IC50 values of 0.87 and 0.79 nM, respectively. Inhibition of ET-1-induced contractions of rat aorta by the compound was considerably weaker than expected (pKB = 6.4), while that of sarafotoxin S6c induced contraction of dog saphenous vein (100% inhibition at 100 nM) was consistent with corresponding binding data. These results suggest that although CGS 31398 is a potent dual inhibitor of ETA/ETB receptor binding, it surprisingly displays potent ETB and weak ETA receptor antagonism in functional assays.

Upregulation of endothelin-1 binding in tissues of salt-loaded stroke-prone spontaneously hypertensive rats.

Upon maintained on a 1% NaCl drinking solution beginning at 7 weeks of age, the stroke-prone spontaneously hypertensive rat (SHRsp) developed severe hypertension and stroke; most died by 16 weeks. The mechanism by which these diseases evolve remains unclear. Endothelin-1 (ET-1) is a potent, peptidic vasoconstrictor and is implicated in the pathogenesis of various cardiovascular, renal, and central nervous system diseases. The purpose of the present study was to compare the binding of [125I]ET-1 to the brain, heart, kidney, liver, and spleen membrane preparations of 16-week-old SHRsp and age-matched normotensive Wistar-Kyoto rats (WKY). The KD values for [125I]ET-1 binding to the corresponding tissues of the two strains were not significantly different, except in the brain (SHRsp: 17 +/- 1 pM; WKY: 24 +/- 1 pM). In contrast, the Bmax values measured in the brain, heart, kidney, and liver of SHRsp were 1.5- to 2.1-fold greater than those of their WKY counterparts. Competition of [125I]ET-1 binding to the membrane preparations by the specific ETA receptor antagonist BQ-123 or the specific ETB receptor agonist sarafotoxin S6c revealed a similar proportion of ETA and ETB receptor subtypes in the corresponding tissues of the two rat strains. These results indicate that ET-1 binding is upregulated in SHRsp and suggest that ET-1 may play a pathophysiological role in this animal model of genetic hypertension.