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BackgroundExcellent adherence to tuberculosis (TB) treatment is critical to cure TB and avoid the emergence of resistance. Wirelessly observed therapy (WOT) is a novel patient self-management system consisting of an edible ingestion sensor (IS), external wearable patch, and paired mobile device that can detect and digitally record medication ingestions. Our study determined the accuracy of ingestion detection in clinical and home settings using WOT and subsequently compared, in a randomized control trial (RCT), confirmed daily adherence to medication in persons using WOT or directly observed therapy (DOT) during TB treatment.Methods and findingsWe evaluated WOT in persons with active Mycobacterium tuberculosis complex disease using IS-enabled combination isoniazid 150 mg/rifampin 300 mg (IS-Rifamate). Seventy-seven participants with drug-susceptible TB in the continuation phase of treatment, prescribed daily isoniazid 300 mg and rifampin 600 mg, used IS-Rifamate. The primary endpoints of the trial were determination of the positive detection accuracy (PDA) of WOT, defined as the percentage of ingestions detected by WOT administered under direct observation, and subsequently the proportion of prescribed doses confirmed by WOT compared to DOT. Initially participants received DOT and WOT simultaneously for 2-3 weeks to allow calculation of WOT PDA, and the 95% confidence interval (CI) was estimated using the bootstrap method with 10,000 samples. Sixty-one participants subsequently participated in an RCT to compare the proportion of prescribed doses confirmed by WOT and DOT. Participants were randomized 2:1 to receive WOT or maximal in-person DOT. In the WOT arm, if ingestions were not remotely confirmed, the participant was contacted within 24 hours by text or cell phone to provide support. The number of doses confirmed was collected, and nonparametric methods were used for group and individual comparisons to estimate the proportions of confirmed doses in each randomized arm with 95% CIs. Sensitivity analyses, not prespecified in the trial registration, were also performed, removing all nonworking (weekend and public holiday) and held-dose days. Participants, recruited from San Diego (SD) and Orange County (OC) Divisions of TB Control and Refugee Health, were 43.1 (range 18-80) years old, 57% male, 42% Asian, and 39% white with 49% Hispanic ethnicity. The PDA of WOT was 99.3% (CI 98.1; 100). Intent-to-treat (ITT) analysis within the RCT showed WOT confirmed 93% versus 63% DOT (p < 0.001) of daily doses prescribed. Secondary analysis removing all nonworking days (weekends and public holidays) and held doses from each arm showed WOT confirmed 95.6% versus 92.7% (p = 0.31); WOT was non-inferior to DOT (difference 2.8% CI [-1.8%, 9.1%]). One hundred percent of participants preferred using WOT. WOT associated adverse events were ConclusionsIn terms of accuracy, WOT was equivalent to DOT. WOT was superior to DOT in supporting confirmed daily adherence to TB medications during the continuation phase of TB treatment and was overwhelmingly preferred by participants. WOT should be tested in high-burden TB settings, where it may substantially support low- and middle-income country (LMIC) TB programs.Trial registrationClinicalTrials.gov NCT01960257.

Objective to develop a resource of systematically collected, longitudinal clinical data and biospecimens for assisting in the investigation into neuromyelitis optica spectrum disorder (NMOSD) epidemiology, pathogenesis, and treatment. Methods to illustrate its research-enabling purpose, epidemiologic patterns and disease phenotypes were assessed among enrolled subjects, including age at disease onset, annualized relapse rate (ARR), and time between the first and second attacks. Results as of December 2017, the Collaborative International Research in Clinical and Longitudinal Experience Study (CIRCLES) had enrolled more than 1,000 participants, of whom 77.5% of the NMOSD cases and 71.7% of the controls continue in active follow-up. Consanguineous relatives of patients with NMOSD represented 43.6% of the control cohort. Of the 599 active cases with complete data, 84% were female, and 76% were anti-AQP4 seropositive. The majority were white/Caucasian (52.6%), whereas blacks/African Americans accounted for 23.5%, Hispanics/Latinos 12.4%, and Asians accounted for 9.0%. The median age at disease onset was 38.4 years, with a median ARR of 0.5. Seropositive cases were older at disease onset, more likely to be black/African American or Hispanic/Latino, and more likely to be female. Conclusions collectively, the CIRCLES experience to date demonstrates this study to be a useful and readily accessible resource to facilitate accelerating solutions for patients with NMOSD., The Guthy-Jackson Charitable Foundation, The CIRCLES Project

ObjectiveTo gain insights into NMOSD disease impact, which may negatively affect QoL of patients, their families, and social network.MethodsThe current study used validated instruments to assess physical, emotional, and socioeconomic burden of NMOSD on QoL among 193 patients.ResultsA majority of patients reported an initial diagnosis of a disease other than NMOSD. Overall, two-thirds of patients reported NMOSD as having a strong negative impact on physical health (Short Form-36 [SF-36] score 27.1 ± 39.1), whereas emotional well-being was relatively unimpaired on average (SF-36 score 54.0 ± 44.9). A subset of patients reported having the highest category of emotional health despite worse physical health or financial burden, suggesting psychological resilience. Pain (r = 0.61) and bowel/bladder dysfunction (r = 0.41) imposed the greatest negative physical impact on overall QoL. In turn, ability to work correlated inversely with worsened health (r = −0.68). Increased pain, reduced sexual function, inability to work, and reduced QoL had greatest negative impacts on emotional well-being. Dissatisfaction with treatment options and economic burden correlated inversely with QoL.ConclusionsCollectively, the current findings advance the understanding of physical, emotional, social, and financial tolls imposed by NMOSD. These insights offer potential ways to enhance QoL by managing pain, enhancing family and social networks, and facilitating active employment.

Data from clinical trials present numerous problems for the data analyst. These include non-compliance with the prescribed dosing regimen and inaccurate recollection of dosing history by patients as well as mistakes in recording data. Several methods have been proposed to address these issues. One such technique by Lu et al. (Selecting reliable pharmacokinetic data for explanatory analyses of clinical trials in the presence of possible noncompliance. J. Pharmacokinet. Pharmacodyn. 28:343-362 (2001)) identifies occasions in pharmacokinetic (PK) data where the preceding dosing history is likely to be unreliable. We used this method, implemented in the software program NONMEM (beta) VI, to clean a dataset containing indinavir (IDV) plasma concentrations from HIV-1 infected patients. The data was also cleaned by inspection in Microsoft Excel using clinical PK criteria. A one-compartment model with first order absorption and elimination was fit to both sets of cleaned data. IDV population PK parameters obtained from these analyses were similar to those reported previously. It is established that IDV nephrotoxicity is related to high IDV exposure. However, no relationships were found between any PK parameters and nephrotoxicity in the "compliance cleaned" dataset. In the "PK cleaned" dataset, the oral clearance and apparent volume were lower by 9.1% and 6.6%, respectively in patients with any type of nephrotoxicity and the maximum IDV concentration (C(max)) was 12.1% higher. In patients suffering from nephrolithiasis in particular, C(max) was 15.5% higher. Accordingly, the use of the non-compliance detection method did not improve the reliability of our dataset over the usual method of applying clinical criteria. In fact, analyses on the compliance-cleaned dataset missed some exposure-toxicity relationships. Thus, automated methods must be tested rigorously with 'real life' datasets, used with caution, and always in conjunction with clinical reasoning to avoid overlooking a signal in noisy data.

The importance of reviewing and studying sex-based differences in pharmacologic parameters is demonstrated by the increasing data on gender variation in drug efficacy and toxicity profiles. Sex-based differences in the four major factors that contribute to interindividual pharmacokinetic variability—bioavailability, distribution, metabolism, and elimination—are theorized to stem from variations between men and women in factors such as body weight, plasma volume, gastric emptying time, plasma protein levels, cytochrome P450 activity, drug transporter function, and excretion activity. Sex-determined variations in pharmacodynamics have traditionally been more difficult to study, but a number of recent studies have explored these differences. This review examines the biologic basis of differences in pharmacokinetics and pharmacodynamics between the sexes and summarizes studies that have addressed these differences. As an example, sex-based variation in the efficacy and toxicity of antiretroviral therapy in human immunodeficiency virus (HIV)-infected patients is explored more thoroughly to illustrate some of the factors underlying sex-based differences in drug therapy.

Background: There are insufficient data on pediatric antiretroviral therapy (ART) pharmacokinetics (PK), particularly for children in low- and middle-income countries. Methods: We conducted a prospective nevirapine (NVP) PK study among HIV-infected Kenyan children aged 3-13 years initiating an NVP-based ART regimen. NVP dose timing was measured through medication event monitors. Participants underwent 2 inpatient assessments: 1 at 4-8 weeks after ART initiation and 1 at 3-4 months after ART initiation. Allometric scaling of oral clearance (CL)/bioavailability (F) and volume of distribution (Vd)/F values were computed. Nonlinear mixed-effects modeling using the first-order conditional estimation with interaction method was performed with covariates. The impact of adherence on time below minimum effective concentration was assessed in the final PK model using medication event monitors data and model-estimated individual parameters. Results: Among 21 children enrolled, mean age was 5.4 years and 57% were female. CL/F was 1.67 L/h and Vd/F was 3.8 L for a median child weighing 15 kg. Participants' age had a significant impact on CL/F (P < 0.05), with an estimated decrease in CL of 6.2% for each 1-year increase in age. Total body water percentage was significantly associated with Vd/F (P < 0.001). No children had >10% of time below minimum effective concentration when the PK model assumed perfect adherence compared with 10 children when adherence data were used. Conclusions: Age and body composition were significantly associated with children's NVP PK parameters. ART adherence significantly impacted drug exposure over time, revealing subtherapeutic windows that may lead to viral resistance.

Objective To investigate whether heparin produces vasodilation in human veins and to explore the underlying mechanisms. Methods Eleven healthy volunteers were studied with the dorsal hand vein compliance technique. Dose-response curves to heparin and enoxaparin were generated. Dose-response curves to heparin were also constructed before and after heparin was infused with the nitric oxide synthase inhibitor NG-monomethyl-l-arginine (L-NMMA) or combined histamine H1- and H2-receptor blockade. Results Heparin but not enoxaparin caused significant dose-dependent relaxation with an average apparent maximal response (at an infusion rate of 20 IU/min) of 47% ± 23%. L-NMMA attenuated heparin-induced relaxation (P < .001). The combination of H1- and H2-receptor antagonists attenuated heparin-induced relaxation to a lesser extent (P < .05). Heparin-induced relaxation decreased by 52%, 73%, and 35% in the presence of L-NMMA, indomethacin (INN, indometacin) plus L-NMMA, and combined H1- and H2-receptor blockade, respectively. Conclusion Heparin is an endothelium-dependent venodilator in humans. The mechanism of heparin-induced relaxation involves an increased availability of nitric oxide, possibly partially related to local release of histamine. Clinical Pharmacology & Therapeutics (1999) 66, 232–238; doi

The amino acid l-arginine is the precursor of nitric oxide (NO), a powerful vasodilator with antiplatelet properties. The availability of l-arginine has been suggested to be a rate-limiting factor in the production of NO in conditions such as hypercholesterolemia. It was speculated that fluctuations in plasma concentrations of l-arginine during the day may be dependent upon dietary intake of the amino acid, or other variables, and might modify the elaboration of endogenous NO. Over a 24-h period, the plasma concentrations of l-arginine and NO-related compounds (NOx) were measured during an l-arginine and nitrate/nitrite-free diet (diet A) or a nitrate/nitrite-free diet with a fixed amount of l-arginine intake (3.8 g/d) (diet B) in eight healthy volunteers during a 2-day crossover study. Subjects were randomly selected to begin with diet A or diet B and consumed the other diet on the second day. During diet A, plasma l-arginine decreased significantly from 09.00 to 16.00 (21.4 ± 2.0 to 11.9 ± 1.1 mg/ml), rose slightly in the evening (to 16.6 ± 1.7mg/ml) and gradually increased during the night. During diet B, plasma larginine showed a peak after each meal (approximately 23 mg/ml). Plasma NOx concentrations measured by chemiluminescence did not show any circadian variation on either diet. Plasma l-arginine concentrations change during the day and are influenced by dietary intake. Importantly, plasma NOx do not seem to vary with this pattern in healthy individuals.

Conversion of angiotensin I to angiotensin II likely occurs in human veins, supporting the existence of endothelial angiotensin-converting enzyme (ACE) activity in these vessels. Using the dorsal hand vein technique, we investigated the effects of 2 ACE inhibitors, captopril (single oral dose of 6.25 mg) and enalaprilat (local infusion of 1 μg/min), on venous responsiveness in healthy subjects. Orally administered captopril induced a marked decrease in angiotensin I– but not angiotensin II–induced venoconstriction. This blunted response persisted for at least 4 hours. Enalaprilat and captopril increased the sensitivity to bradykinin, decreasing the dose producing half-maximal response (ED50) of bradykinin 18-fold and 5-fold, respectively, without changing the maximal venodilatory response. These results confirm that there is substantial rapid metabolism of angiotensin I in human veins and suggest that a single dose of locally infused angiotensin I can be used with the dorsal hand vein technique to assess the time-course effect of vascular ACE inhibition after oral administration. Our findings also extend previous in vitro observations in human veins by showing that these agents potentiate the venodilatory effects of bradykinin in vivo. Clinical Pharmacology & Therapeutics (1999) 65, 58–65; doi

Aims Recent reports, largely in animal models, have suggested that either inhibition of nitric oxide (NO) synthase or endothelium removal in arteries inhibits the response to isoprenaline, a β-adrenoceptor agonist, and also enhances the response to sodium nitroprusside, a nitrovasodilator. This in vivo study was designed to determine whether NG-monomethyl-l-arginine (l-NMMA), an inhibitor of NO synthesis, influences relaxation of human hand veins mediated by isoprenaline or by sodium nitroprusside. Methods Using the dorsal hand vein technique, full dose-response curves to bradykinin (0.27–278 ng min−1, n=6), isoprenaline (2.12–271 ng min−1, n=8) and sodium nitroprusside (0.01–634 ng min−1, n=7) were generated on separate occasions before and after l-NMMA co-infusion (50 μg min−1 ). Results In veins preconstricted with the α1-adrenoceptor-selective agonist phenylephrine, the three vasodilators induced maximal responses (Emax ) of 119±35, 72±18 and 103±17%, respectively. l-NMMA inhibited relaxation to bradykinin by 64% (P=0.014) but did not influence relaxation induced by isoprenaline. The sensitivity to sodium nitroprusside was significantly enhanced by l-NMMA co-infusion (concentration shift of 2.3, P=0.031). Conclusions We conclude that in human veins, spontaneously released NO does not play a major role in isoprenaline-induced relaxation. Our results also suggest that the effects of sodium nitroprusside in this vascular bed may be attenuated by endothelium-derived NO.

We investigated the pharmacokinetics of zidovudine, zalcitabine, and saquinavir in AIDS Clinical Trial Group protocol 229. Patients received either saquinavir, zalcitabine, or a combination of both, together with zidovudine three times a day. Approximately 100 patients were enrolled in each treatment arm, and intensive pharmacokinetic studies were performed on about 25 patients per arm at weeks 1 and 12. We estimated the pharmacokinetic parameters of all three drugs by using parametric and nonparametric methods. The mean values of the pharmacokinetic parameters of zidovudine (clearance [CL]/bioavailability [F], 168 liters/h; volume of distribution [V]/F, 185 liters; half-life, 0.76 h) and zalcitabine (CL/F, 25 liters/h; V/F, 92.2 liters; half-life, 2.7 h) were similar to those reported previously. For saquinavir, the mean pharmacokinetic parameter estimates using parametric methods were as follows: maximum concentration of drug in serum [Cmax], 70.8 ng/ml; time to Cmax, 3.11 h; area under the curve, 809 ng x h/ml; CL/F, 989 liters/h; V/F, 1,503 liters; half-life, 1.38 h. For all three drugs, clearance decreased with age. Weight did not influence the clearance of zidovudine, but the clearance of zalcitabine and saquinavir increased with weight. There were no differences in pharmacokinetic parameters between study weeks and arms, suggesting that there is no change in kinetics with chronic administration and that there are no significant pharmacokinetic interactions among these three drugs.

Morphine has been extensively used in the treatment of pulmonary edema, and its action is believed to be mediated in part by its ability to produce peripheral venodilation. This study investigated whether opiates produce venodilation in human hand veins and explored the underlying mechanism(s). Fifteen healthy volunteers (11 men and four women) were studied with use of the dorsal hand vein compliance technique. After preconstriction with the selective alpha 1-adrenergic receptor agonist phenylephrine, dose-response curves were constructed to (1) opiate receptor agonists morphine (1 to 30 micrograms/min) or fentanyl (0.07 to 1 microgram/min), (2) a combination of morphine and the mu-opiate receptor antagonist naloxone, and (3) morphine and a combination of histamine (H1 and H2) receptor antagonists. Infusion of morphine caused venodilation in a dose-dependent manner, whereas fentanyl did not produce venodilation. Coinfusion of naloxone and morphine impaired the venodilation only slightly. Coinfusion of the H1- and H2-antagonists completely abolished the venodilatory effect of morphine. These results suggest that the venodilatory effect of morphine is mediated through histamine release and that mu-opiate receptors have little or no involvement in this process.

Medical students were introduced to issues relating to medication noncompliance in a simulated clinical setting. Compliance with either a twice-a-day or a three-times-a-day regimen was monitored with use of electronic monitoring devices for a 2-week interval. Compliance with the twice-a-day regimen was higher than compliance with the three-times-a-day regimen, although the difference was not significantly different. Overall, 71% of the prescribed doses were taken by the medical student participants; however, only 46.5% of the doses were taken at the prescribed dosing frequency and 28.5% were taken at the prescribed intervals. The majority of students linked dose taking with routine daily activities and reported that their hectic lifestyles adversely influenced compliance. Similar factors might be expected to influence compliance in patient populations. The goal of this exercise was to demonstrate to future physicians the difficulties that patients have with compliance to prescribed medications.

beta-Adrenergic responses have been shown to decline with aging, particularly in the cardiovascular system. We infused terbutaline, a selective beta 2-adrenoceptor agonist, into the brachial artery of 10 young (mean age 25 yr, range 22–31 yr) and 9 elderly (mean age 73 yr, range 68–81 yr) healthy subjects to examine its effects on nutrient flux. Forearm K+, PO4, free fatty acid (FFA), and glycerol uptake were determined by measurement of forearm blood flow (using dye dilution) and brachial arterial and deep venous plasma substrate concentrations. Elderly subjects were less sensitive to terbutaline-mediated increases in forearm blood flow, net fluxes of K+, and glycerol but not net fluxes of FFA or PO4. The mean fitted slopes of each parameter vs. the log of the terbutaline concentration, a measure of forearm beta-adrenergic sensitivity, for young and elderly groups were 4.9 +/- 1.7 (SD) vs. 2.4 +/- 2.3 for forearm blood flow (P

Epoxyeicosatrienoic acids (EETs) produced by cytochrome P450 (CYP)-dependent epoxidation of arachidonic acid (AA) inhibit thrombocyte adhesion to the vascular wall. Upon dietary omega-3 fatty acid supplementation, EETs are partially replaced by eicosapentaenoic acid (EPA)-derived epoxyeicosatetraenoic acids (EEQs) and docosahexaenoic acid (DHA)-derived epoxydocosapentaenoic acids (EDPs). We hypothesized that the omega-3 epoxy-metabolites may exhibit superior anti-thrombogenic properties compared to their AA-derived counterparts. To test this hypothesis, we analyzed the effects of 11,12-EET, 17,18-EEQ and 19,20-EDP on Ristocetin-induced thrombocyte aggregation (RITA), a process that mimics thrombocyte adhesion to the vascular wall. The eicosanoids were added for 5, 30, or 60 minutes to thrombocyte-rich plasma freshly prepared immediately after blood collection from stringently selected apparently healthy subjects. Thrombocyte aggregation was then induced by Ristocetin (0.75 mg/mL) and assessed by turbidimetric measurements. After 60 minutes of preincubation, all three epoxy-metabolites significantly decreased the rate of RITA. 17,18-EEQ and 19,20-EDP were effective already at 1 μM, whereas 5-fold higher concentrations were required with 11,12-EET. Addition of AUDA, an inhibitor of the soluble epoxide hydrolase, potentiated the effect of 17,18-EEQ resulting in a significant further decrease of the velocity as well as amplitude of the aggregation process. In contrast to their profound effects on RITA, none of the epoxy-metabolites was effective in reducing collagen- or ADP-induced thrombocyte aggregation. These results indicate a highly specific role of CYP-eicosanoids in preventing thromboembolic events and suggest that the formation of 17,18-EEQ and 19,20-EDP may contribute to the anti-thrombotic effects of omega-3 fatty acids.

The enzyme nitric oxide synthase mediates synthesis of nitric oxide (NO) from 1-arginine in endothelial cells. NO, also known as endothelium-dependent relaxing factor (EDRF), diffuses to smooth muscle cells where it leads to cGMP production and dilation. We characterized the potency, efficacy and time course of NG-monomethyl-l-arginine (l-NMMA) as an inhibitor of bradykinin-mediated, endothelium-dependent dilation using the human hand-vein compliance technique. We also compared the efficacy of l-NMMA with methylene blue, an inhibitor of guanylate cyclase, in blocking bradykinin-mediated vasodilation. l-NMMA potently inhibited bradykinin-induced venodilation with a log ED50 of 3.74 +/- 0.52 (geometric mean of 5.5 micrograms/min). Responses to bradykinin (0.27-555 ng/min) were tested in veins pre-constricted with the alpha-adrenergic agonist phenylephrine. l-NMMA (25 micrograms/min) decreased bradykinin's maximal venodilatory response from 90 +/- 22% to 39 +/- 15% (p < 0.05). Complete recovery of bradykinin venodilation was obtained within 155 minutes after stopping l-NMMA infusion, indicating that its effects were reversible. In another set of experiments we compared the efficacy of methylene blue to l-NMMA; methylene blue decreased bradykinin-mediated venodilatory response to 53 +/- 17%; when l-NMMA was added, the response was further decreased to 32 +/- 9% (p < 0.002). We conclude that l-NMMA is a very efficacious NO synthase inhibitor in human veins and it is likely functionally reversible.

BACKGROUND Exercising skeletal muscle releases large amounts of potassium into plasma. beta-adrenergic receptors enhance reuptake of potassium into muscle. Since beta-adrenergic responses decline with aging in many tissues, the elderly might be predisposed to hyperkalemia during exercise. METHODS Venous plasma potassium (K+) was measured during graded bicycle exercise (30 W initial workload with 3-minute 30 W increments) in 18 healthy young men (24.0 +/- 2.9 yrs; mean +/- SD), 18 healthy untrained elderly men (70.0 +/- 3.6 yrs), and 7 elderly master athletes (70.0 +/- 4.6 yrs). Subjects exercised to exhaustion. RESULTS Exercise times and maximal oxygen uptake were: young (N = 14; 19.6 +/- 4.4 min, 42.1 +/- 3.8 ml/kg/min), elderly (N = 13; 11.7 +/- 1.9 min, 26.6 +/- 8.0 ml/kg/min), elderly master athletes (N = 6; 16.8 +/- 2.4 min, 40.2 +/- 10.2 ml/kg/min). The rate of increase in K+ was greater in both elderly (89 +/- 59 mumol/l/min) and elderly master athletes (88 +/- 26) compared with healthy young men (60 +/- 30), p < .05. Despite this greater rate of increase, the elderly group did not achieve higher maximal K+ concentrations than the young because of the much shorter duration of exercise in the elderly group. For subjects performing a maximal exercise test, the maximal increase in K+ was similar in the elderly (1.15 +/- 0.91), but significantly greater in elderly master athletes (1.70 +/- 0.31), compared with healthy young men (1.31 +/- 0.45). CONCLUSIONS Changes in K+ during short-lasting exercise are similar in untrained healthy elderly men compared with healthy young men, but elderly master athletes have greater changes in K+ during exercise. The greater rate of increase in potassium in healthy and athletic subjects supports the hypothesis of an age-related impairment of the beta 2-adrenergic process that mediates potassium flux into skeletal muscle.

Chronic administration of alpha 1-receptor antagonists is associated with loss of clinical efficacy, especially in congestive heart failure, although the mechanism is uncertain. To evaluate changes in venous alpha 1-adrenoceptor responsiveness during chronic alpha 1-adrenoceptor blockade, dose-response curves to phenylephrine and angiotensin II were constructed in 10 healthy subjects before, during, and after administration of terazosin 1 mg orally for 28 d. Terazosin initially shifted the dose-response curve of phenylephrine to the right, with a significant increase in ED50 for phenylephrine from a control value of 102 to 759 ng/min on day 1 of terazosin (P < 0.001). However, by day 28, the dose-response curve had shifted back towards baseline with an ED50 of 112 ng/min. After discontinuing terazosin, the ED50 for phenylephrine remained near the baseline value, indicating no evidence of supersensitivity to phenylephrine. There was no change in responsiveness to angiotensin II during the course of treatment with terazosin. Plasma terazosin concentrations were stable throughout the period of drug administration. The mean Kd of terazosin was estimated as 11 +/- 15 nM in the first few days of treatment. This study demonstrates that pharmacological tolerance to the alpha 1-adrenoceptor blocking action of terazosin occurs in man and may be responsible for loss in efficacy with chronic therapy.

The aim of this study was to determine whether there is an age-related decline in vascular responsiveness to bradykinin, whose vasodilatory action is mediated chiefly through endothelium-derived relaxing factor (EDRF). Dose-response curves for bradykinin were constructed using the dorsal hand vein compliance technique in veins preconstricted with phenylephrine in 27 volunteers (16 male, 11 female) aged 18 to 81 years. At the end of the bradykinin study, 12 subjects had a single infusion of a high dose of isoproterenol. There was no correlation between age and the EMAX or the log ED50 for bradykinin, although the same subjects showed a correlation between age and EMAX for isoproterenol, as previously found. There was no significant difference in either the EMAX or the log ED50 between male and female subjects. The results suggest that bradykinin-induced vasodilation is independent of age or gender.

1. Diabetic autonomic neuropathy causes loss of sympathetic cardiovascular control and is associated with increased vascular sensitivity to catecholamines. Supersensitivity to catecholamines could be due to either a postsynaptic increase in vascular sensitivity or to decreased catecholamine uptake into peripheral sympathetic nerve endings. 2. To differentiate between these possible mechanisms we have measured the responsiveness in vivo to noradrenaline and phenylephrine with local infusions into peripheral veins of diabetic patients with and without symptomatic autonomic neuropathy and of healthy control subjects. The dorsal hand vein compliance technique was used. 3. Symptomatic diabetic patients required significantly lower doses of noradrenaline for half-maximal venoconstriction (ED50) (geometric mean 2.14 ng/min) than control subjects (geometric mean 6.61 ng/min, P = 0.032), but there was no difference in the results from the phenylephrine dose-response curves between the groups. Thesre were no differences in venous responsiveness to noradrenaline or phenylephrine between the asymptomatic diabetic group and the control group. However, in the asymptomatic diabetic group, postural blood pressure change (an index of loss of sympathetic control) was correlated with the ED50 for noradrenaline (r = 0.74, P = 0.014), but not with the ED50 for phenylephrine. In the control group the ED50 values for noradrenaline and phenylephrine were correlated with each other (r= 0.81, P= 0.0005). 4. Both vasopressor drugs act on vascular α-adrenoceptors, but only noradrenaline is taken up into peripheral sympathetic nerve endings. Our results suggest that, in diabetic patients, vascular supersensitivity to catecholamines is primarily determined by decreased neuronal catecholamine uptake. A postsynaptic increase in vascular a-adrenoceptor stimulation does not appear to be prominent in diabetic autonomic neuropathy.

Aging is associated with diminished beta-adrenergic responsiveness in a variety of tissues. Desensitization of tissues secondary to the age-associated increase in sympathetic nervous system activity has been proposed as a potential explanation for diminished beta-adrenergic responsiveness. If this hypothesis is correct, then beta-blockade in older people might be expected to reverse the blunted beta-adrenergic responses of tissues having diminished responsiveness. Cardiac chronotropic responses to bolus isoproterenol (ISO) doses and ISO-induced venous smooth muscle dilatation in superficial hand veins were examined in 8 young (26.2 +/- 2.6 yrs) and 9 elderly (68.0 +/- 2.2 yrs) healthy subjects before, during, and 3 and 7 days following 2 weeks of treatment with propranolol. Baseline cardiac chronotropic responsiveness, CD25, was 1.55 +/- 0.77 mcg in the young and 5.97 +/- 2.77 mcg in the elderly subjects (p less than .01), increasing to 84 +/- 56 mcg and 194 +/- 172 mcg during treatment with propranolol. At 3 and 7 days following withdrawal of propranolol, CD25s were respectively 1.24 +/- 0.79 (p = .14) and 1.10 +/- 0.42 (p = .04) in the young and 5.63 +/- 2.34 (p = .31) and 4.85 +/- 2.37 (p = .05) in the elderly subjects. In contrast, there was no decrease in the ED50 or increase in Emax for ISO-induced venodilation of hand veins following propranolol withdrawal. These results demonstrate that both young and elderly subjects have similar increases in cardiac chronotropic responsiveness following discontinuation of beta-blockade and do not support the concept that desensitization is responsible for the diminished beta-adrenergic responsiveness seen with aging.

Essential hypertension is more prevalent among blacks than whites although the explanation is unknown. It is possible that an altered vascular adrenoceptor responsiveness in blacks may play a role in the etiology of this racial difference. To test this hypothesis, we have compared the diameter changes in superficial veins in response to phenylephrine, an alpha-adrenergic agonist, and to isoproterenol, a beta-adrenergic agonist, in black and white young normotensive males using the dorsal hand vein compliance technique. The maximal venoconstriction after phenylephrine (Emax) was 92 +/- 9% (mean +/- SD) in the whites but only 74 +/- 12% in the blacks (p = 0.00046). The ED50 of phenylephrine was 342 ng/min (geometric mean) in whites and 245 ng/min in blacks (p = 0.50). The Emax and ED50 for isoproterenol-mediated venodilation was not significantly different between the racial groups. These results indicate a decreased maximal responsiveness to alpha-adrenergic stimulation in normotensive blacks. How these changes relate to cardiovascular alterations in hypertensive blacks requires further study.

Fluconazole, a potent bis-triazole antimycotic drug, has been demonstrated to inhibit antipyrine metabolism, a cytochrome P-450 dependent process, in vivo in mice. Cyclosporine is metabolized by the cytochrome P-450 enzyme system in both mice and man. We investigated whether fluconazole had any effects on the metabolism of cyclosporine in vivo in mice. The effects of three different doses of fluconazole (1, 5, and 20 mg/kg) on the metabolism of cyclosporine in CD-1 mice were studied in single-dose experiments. Fluconazole produced significant dose-dependent decreases in the elimination rate constant and increases in the terminal half-life of cyclosporine. The 1 mg/kg dose caused a 26% prolongation of the terminal half-life and the 5 and 20 mg/kg dose prolonged the half-life by 72 and 187%, respectively. Fluconazole doses in the 1–5 mg/kg range are effective in mouse models of fungal infections. These results provide further in vivo evidence that fluconazole is a potent inhibitor of the cytochrome P-450 dependent enzyme system in mice. Future experimental studies in animals and humans are needed to evaluate possible metabolic drug–drug interactions involving fluconazole.Key words: cyclosporine, drug metabolism, drug–drug interaction, fluconazole, cytochrome P-450, animal, antifungal.

Objective The aim of this study was to determine whether pharmacokinetic interactions between the protease inhibitors saquinavir soft gel, nelfinavir, and ritonavir are affected by the timing of administration. Study design We used an open-label, 6-period, incomplete Latin square crossover study in 18 human immunodeficiency virus–negative subjects. Each received single oral doses of 2 of the 3 protease inhibitors during each of 6 periods. Single doses were given either simultaneously or separated by 4 hours. The order of the periods was balanced, and periods were separated by 2 days. We measured protease inhibitor concentrations over a 24-hour period by HPLC and estimated pharmacokinetic parameters by noncompartmental methods. Results Median saquinavir area under the curve (AUC) increased by 62-fold when ritonavir was coadministered, by 50-fold when ritonavir was given 4 hours earlier, and by 16-fold when saquinavir preceded ritonavir by 4 hours. Saquinavir AUC increased by 7-fold when nelfinavir was coadministered. Nelfinavir AUC increased by 2.5-fold with coadministration of ritonavir and by 1.8- and 2.1-fold when ritonavir was administered before nelfinavir and after nelfinavir, respectively. Ritonavir AUCs were unaffected by coadministration of the other drugs. The effect of ritonavir on the kinetics of saquinavir persisted for at least 48 hours after a single dose of ritonavir, suggesting the possibility of metabolic intermediates that form inhibitory complexes. Conclusion Except for saquinavir followed by ritonavir, there is little difference in protease inhibitor exposure for simultaneous or staggered doses. The persistent effect of ritonavir suggests the possibility that lower doses and longer dosing intervals might be effective when ritonavir is used to boost concentrations of other protease inhibitors. Clinical Pharmacology & Therapeutics (2003) 73, 406–416; doi: 10.1016/S0009-9236(03)00006-7

Protease inhibitors are very effective in treating patients infected with HIV. However, many drugs in this class penetrate poorly into the central nervous system (CNS) and may permit this site to be a sanctuary from which resistant virus can emerge. Previous studies have shown that the protease inhibitor saquinavir (SQV) interacts with the multidrug transport system, P-glycoprotein (P-gp), expressed in epithelial cells in the gut mucosa and at the blood-brain barrier, and thus might affect both the oral absorption and the penetration of SQV into the CNS. To determine whether SQV is a substrate for P-gp, its uptake was determined in cancer cells, which do (Dx5) and do not (MES-SA) express P-gp. The distribution of SQV between brain tissue and plasma was also investigated in rats and in normal and P-gp-deficient mdr1a(-/-) mice. The distribution ratio of SQV in plasma:brain:cerebrospinal fluid was approximately 100:10:0.2 in rats. The accumulation of SQV was enhanced in MES-SA cells (P-gp-negative) versus Dx5 cells (P-gp-positive). Bolus i.v. injection of [(14)C]SQV (2 and 5 mg/kg) into mdr1a(-/-) and normal mice (n = 3 or 4) resulted in 3-fold higher radioactivity in brains from mdr1a(-/-) mice. Similarly, oral administration of [(14)C]SQV (500 mg/kg) resulted in a 5-fold increase in systemic exposure and a 10-fold increase in brain levels in mdr1a(-/-) mice. These data demonstrate that saquinavir is a substrate for P-gp and that this transport system may play a role in limiting oral absorption and CNS exposure to this protease inhibitor.

Background Cigarette smoking is associated with impaired endothelium-dependent dilatation in human veins and arteries. An in vivo study in animals suggests that nicotine may contribute to this abnormality. We tested the hypothesis that local administration of nicotine at a dose reproducing the plasma concentration observed during smoking would impair endothelium-dependent vasodilatation in human veins in vivo. Methods We studied 11 healthy nonsmokers with the dorsal hand vein compliance technique. After 70% to 80% preconstriction with phenylephrine, endothelium-dependent venous relaxation was assessed by infusion of bradykinin (1 to 278 ng/min), a potent vasodilator acting primarily in this model through endothelial release of nitric oxide and prostanoids. Sodium nitroprusside (0.0001 to 3166 ng/min) was used to test endothelium-independent relaxation. Dose-response curves were constructed before and during nicotine coinfusion at a rate of 40 ng/min, reproducing a plasma concentration of 15 ng/mL. Results After a 10-minute preinfusion, nicotine administration was associated with a loss in sensitivity to bradykinin (P < .001). After 30 and 60 minutes of preinfusion with nicotine, the venorelaxant effect of bradykinin was further reduced (P < .001). A similar inhibition of the response to bradykinin by nicotine persisted in the presence of indomethacin (INN, indometacin). Coinfusion of nicotine did not attenuate sodium nitroprusside–induced venodilation. Conclusion The results show that acute local exposure to nicotine in vivo is associated with an impaired response to endothelium-derived nitric oxide in human veins. This finding may provide further insight into the pathophysiology of smoking-induced endothelial dysfunction. (Clin Pharmacol Ther 2000;67:391-7.) Clinical Pharmacology & Therapeutics (2000) 67, 391–397; doi: 10.1067/mcp.2000.105153

Abstract —Asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, is elevated in hypercholesterolemia. This study was designed to determine the role of ADMA in the increased mononuclear cell adhesiveness observed in human hypercholesterolemia. In patient studies, plasma ADMA levels were determined by high-performance liquid chromatography. Functional mononuclear leukocyte adhesion assays were performed in parallel, and flow cytometry was used to characterize bound monocytes and T lymphocytes. Hypercholesterolemic patients were then placed on an oral l -arginine regimen of 14 or 21 g/d and studied over 12 weeks. In cell culture studies, bovine aortic endothelial cells were incubated with varied concentrations of ADMA. Monocytoid cells were cocultured with these bovine aortic endothelial cells, and their adhesiveness was assessed by use of a binding assay. Flow cytometry was used to quantify adhesion molecule expression. Plasma ADMA levels and adhesiveness of mononuclear cells (specifically, monocytes and T lymphocytes) were elevated in hypercholesterolemic patients. Adhesiveness was inversely correlated with the plasma l -arginine/ADMA ratio. Oral administration of l -arginine normalized plasma l -arginine/ADMA ratios and attenuated monocyte and T-lymphocyte adhesiveness. ADMA had no direct effect on the adhesiveness of mononuclear cells. However, monocytes became hyperadhesive when cocultured with ADMA-exposed endothelial cells. In human hypercholesterolemia, the plasma l -arginine/ADMA ratio is inversely correlated with mononuclear cell adhesiveness. Restoration of the l -arginine/ADMA ratio to control levels normalizes mononuclear cell adhesiveness. Our studies suggest that the elaboration of endothelium-derived nitric oxide affects the behavior of circulating T lymphocytes and monocytes.

The obstructive sleep apnea syndrome (OSAS) is associated with cardiovascular disease and systemic hypertension. Because systemic arterial blood pressure is proportional to venodilation and venous return to the heart, we hypothesized that altered vascular responsiveness might exist in the veins of subjects with OSAS. We therefore investigated venodilator responses in awake, normotensive subjects with and without OSAS, using the dorsal hand vein compliance technique. Dose-response curves to bradykinin and nitroglycerin were obtained from 12 subjects with OSAS and 12 matched control subjects. Maximal dilation (E(max)) to bradykinin was significantly lower in the OSAS group (62.1% +/- 26.1%) than in the control group (94.3% +/- 10.7%) (p0.005). Vasodilation to nitroglycerin tended to be lower in the OSAS group (78.6% +/- 31.8%) than the control group (100.3% +/- 12.9%), but this effect did not reach statistical significance. When six of the OSAS subjects were retested after 60 d of treatment with nasal continuous positive airway pressure (CPAP), E(max) to bradykinin rose from 60.3% +/- 20. 3% to 121.4% +/- 26.9% (p0.01). Vasodilation to nitroglycerin also increased, but this effect did not reach statistical significance. These results demonstrate that a blunted venodilatory responsiveness to bradykinin exists in OSAS. This effect appears to be reversible with nasal CPAP therapy.

Objective In arteries and veins smoking is associated with impaired nitric oxide–mediated relaxation to endothelium-dependent agonists such as bradykinin. We investigated whether acute local angiotensinconverting enzyme (ACE) inhibition, achieved by enalaprilat, could influence bradykinin-induced vasodilation in veins of smokers. Methods We studied 7 smokers and 7 nonsmokers with the hand vein technique. After preconstriction with phenylephrine was performed, endothelium-dependent and independent relaxations were assessed by infusing bradykinin (1 to 278 ng/min) and sodium nitroprusside (0.0001 to 3166 ng/min), respectively. Dose-response curves were constructed before and during enalaprilat coinfusion (1 μg/min for 40 minutes). Results Smokers had impaired venodilation to bradykinin compared with nonsmokers (P < .01). Apparent maximal relaxation induced by bradykinin was 78% ± 9% in the control group and 48% ± 9% in smokers (mean ± SD). ACE inhibition shifted the bradykinin dose-response curve to the left in both groups (P < .001) and was associated with a minimal increase in apparent maximal venodilation in nonsmokers (78% ± 9% to 83% ± 18%). In contrast, in smokers ACE inhibition augmented the magnitude of apparent maximal venodilation to values comparable to those observed in the control group (48% ± 9% to 102% ± 21%). In both groups the response to sodium nitroprusside was not affected by enalaprilat. Conclusions This study shows that acute local ACE inhibition restores bradykinin-induced relaxation in smokers to values found in nonsmokers. This observation suggests that increased vascular metabolism of bradykinin exists in veins of smokers and that the vascular renin-angiotensin system may play a key role in smoking-induced endothelial dysfunction. Clinical Pharmacology & Therapeutics (1999) 65, 295–303; doi

Acute administration of L-arginine, the precursor of endothelial nitric oxide, has been shown to improve endothelial function in hypercholesterolaemic rabbits and humans. Animal studies suggest that this beneficial effect, which is thought to be related to the increased availability of nitric oxide, may not be sustained during chronic oral administration. Pharmacokinetic alterations may contribute to this observation. The present study was designed to examine the disposition of L-arginine in hypercholesterolaemic subjects during long-term administration. Plasma L-arginine concentrations were determined by HPLC in 10 patients (eight women and two men; mean age 46±16 years) after an intravenous dose of 10 or 30 g and an oral dose of 5 or 7 g. Pharmacokinetic studies were performed at regular intervals (4 weeks) during a 12-week period of oral L-arginine administration (14–21 g/day). The average plasma L-arginine concentrations before (baseline) and during administration were 16.1±1.2 and 22.5±1.3 μg/ml respectively (P< 0.05). Plasma concentrations of L-arginine remained above baseline throughout weeks 2–12. The L-arginine exposure, expressed as a normalized area-under-the-curve for 8 h (AUC0–8) after oral or intravenous doses during the first visit, was 894.4±118.7 and 1837.8±157.0 units respectively. There were no significant changes in peak plasma L-arginine concentrations or in the AUC0–8 after oral and intravenous doses during subsequent visits (P> 0.05). The mean non-renal clearance of L-arginine during the four visits remained constant. Knowledge of the pharmacokinetics of L-arginine may be useful in the design of clinical trials involving this agent, as well as in the interpretation of the pharmacodynamics of this important precursor of nitric oxide.

Objective To characterize drug-taking behavior using continuous electronic monitoring in an AIDS clinical trial. Setting This was a substudy of AIDS Clinical Trials Group (ACTG) protocol 175, a phase II/III study of dideoxynucleoside monotherapy versus combination therapy in asymptomatic HIV-positive subjects. Participants were required to comply with regimens containing zidovudine, zalcitabine and didanosine, or matching placebos; the total daily pill count was 16. Design For participants at two ACTG 175 sites, electronic devices were used to monitor drug-taking behavior of all study medications over a period of approximately 90 days. Measurements Four indices of drug-taking behavior were calculated and their distributions and relationship to the prescribed regimen were examined. Results Data from 41 subjects were analyzed. Of the prescribed doses of zidovudine, zalcitabine and didanosine, 88, 84 and 82%, respectively, were taken. Of these, 55, 66 and 79%, respectively, were taken at the prescribed dosing frequency. The median percentage of days on which participants failed to take any of the doses was 2-5%. There was a trend towards lower adherence in the combination therapy arms compared with those assigned to receive monotherapy. In this analysis, older patients demonstrated better adherence, although patient characteristics, in general, were poorly predictive of adherence. Conclusion Drug-taking behavior for all three active study medications differed from that prescribed. One result of this erratic adherence was that study participants sustained little antiretroviral effect during more than 25% of the monitoring period.

Background —Asymmetric dimethylarginine (ADMA) is an endogenous competitive inhibitor of nitric oxide (NO) synthase. Because endothelial NO elaboration is impaired in hypercholesterolemia, we investigated whether plasma concentrations of ADMA are elevated in young, clinically asymptomatic hypercholesterolemic adults. We further studied whether such elevation of ADMA levels was correlated with impaired endothelium-dependent, NO-mediated vasodilation and urinary nitrate excretion. In a randomized, double-blind, placebo-controlled study, we investigated whether these changes could be reversed with exogenous l -arginine. Methods and Results —We measured plasma levels of l -arginine, ADMA, and symmetrical dimethylarginine (SDMA) by high-performance liquid chromatography in 49 hypercholesterolemic (HC) and 31 normocholesterolemic (NC) humans. In 8 HC subjects, endothelium-dependent forearm vasodilation was assessed before and after an intravenous infusion of l -arginine or placebo and compared with 8 NC control subjects. ADMA levels were significantly elevated by >100% (2.17±0.15 versus 1.03±0.09 μmol/L; P l -Arginine levels were similar, resulting in a significantly decreased l -arginine/ADMA ratio in HC subjects (27.7±2.4 versus 55.7±5.4; P l -arginine significantly increased the l -arginine/ADMA ratio and normalized endothelium-dependent vasodilation and urinary nitrate excretion. ADMA levels were inversely correlated with endothelium-mediated vasodilation ( R =0.762, P R =0.534, P Conclusions —We find that ADMA is elevated in young HC individuals. Elevation of ADMA is associated with impaired endothelium-dependent vasodilation and reduced urinary nitrate excretion. This abnormality is reversed by administration of l -arginine. ADMA may be a novel risk factor for endothelial dysfunction in humans.

Cigarette smoking has been shown to impair endothelium-dependent dilation in arteries. We tested the hypothesis that cigarette smoking also impairs endothelium-dependent venodilation and evaluated changes in this response after smoking cessation in a time-course study using the dorsal hand vein technique. Dose-response curves were constructed in smokers and nonsmokers by infusing bradykinin (1–278 ng/min), an endothelium-dependent vasodilator, and nitroglycerin (0.006–1,583 ng/min), an endothelium-independent vasodilator, into hand veins preconstricted with the selective α1-adrenergic agonist phenylephrine. The maximal venodilation induced by bradykinin was 89 ± 5% in controls ( n = 16) and 61 ± 7% in smokers ( n = 18; P = 0.02). No difference in nitroglycerin-induced venodilation was observed between the two groups. Coinfusion of l-arginine (0.33 mg/min) markedly improved the bradykinin-induced venodilation in smokers (52 ± 7 to 90 ± 9%; P < 0.01). After acute smoking cessation ( n = 7), restoration to normal bradykinin-induced venodilation was observed within 24 h, whereas no change in the response to a maximally effective dose of nitroglycerin (1,583 ng/min) was detected. In a human vein model appropriate for testing vascular functional alterations, this study demonstrates that smoking impairs endothelium-dependent venodilation in heavy smokers. Moreover, this endothelial dysfunction appears to be rapidly reversible after smoking cessation. This model may be useful in studies evaluating mechanisms of endothelial dysfunction and interventions to modify it.

The relationship of CD4+ cell response, level of RNA in plasma, and quantitative peripheral blood mononuclear cell (PBMC) titer to apparent drug exposure was investigated by using data from AIDS Clinical Trial Group protocol 229, a multicenter randomized study. Patients received either saquinavir, zalcitabine, or a combination of both, along with open-label zidovudine. Approximately 100 patients were enrolled in each arm, and the primary study duration was 24 weeks. Individual drug exposure, the area under the concentration-time curve, was estimated by using population-based pharmacokinetic methods. Response was defined as the maximum increase in CD4+ cell count or the maximum decrease in RNA in plasma or PBMC titer adjusted for baseline CD4+ cell count, RNA in plasma, and PBMC titer, respectively. Regression of responses on exposure demonstrated an exposure effect for saquinavir which was significant for the maximum increase in CD4+ cell count and the decrease in RNA in plasma. For the PBMC titer, no significant relationship could be demonstrated but the results suggested a trend similar to that of the other response variables. For all three response variables, the slope of the saquinavir exposure response was greater with the triple combination (saquinavir, zidovudine, and zalcitabine) than with the combination of saquinavir and zidovudine, suggesting possible synergism between saquinavir and zalcitabine.

Objective Hypotension induced by parenteral administration of chloroquine is a common and serious adverse effect of this drug. Our aim was to investigate whether chloroquine produces venodilation in vivo and to explore the underlying mechanisms. Methods Vascular effects of chloroquine were studied in healthy volunteers with use of the dorsal hand vein technique at the Geriatric Research Education and Clinical Center, Veterans Affairs Palo Alto Health Care System. We studied 22 healthy volunteers (19 men and three women). Venous responsiveness was determined with the dorsal hand vein technique, which measures the diameter of the vein. Results Chloroquine was found to produce a dose-dependent relaxation of hand veins preconstricted with the α1-receptor selective agonist phenylephrine. The venodilatory response to chloroquine ranged from 15% ± 19% at an infusion rate of 0.75 μg/min to 61% ± 24% at 48 μg/min. Venodilation was attenuated by the nitric-oxide synthase inhibitor NG-monomethyl-L-arginine (L-NMMA) so that the dose of chloroquine required to produce 20% venodilation was increased from 3.7 μg/min to 15 μg/min (p < 0.01). In the presence of a combination of histamine receptor antagonists, there was also a diminution of the vasodilatory response to chloroquine from 72% ± 5% to 44% ± 5% at the infusion rate of 96 μg/min. The response was further reduced to 33% ± 7% by the coinfusion of H1-/H2-receptor antagonists with L-NMMA. Conclusion Chloroquine produces venodilation at infusion rates that achieve local concentrations likely similar to those observed systematically after clinically relevant intravenous doses. The data also suggest a role for nitric oxide and histamine release in mediating this response. Clinical Pharmacology & Therapeutics (1997) 61, 677–683; doi

Purpose: CCNU (lomustine) and other nitrosourea compounds are rapidly metabolized to alkylating moieties, which are active against various malignancies. In humans, CCNU undergoes biotransformation to the geometric isomers of 4′-hydroxyCCNU. The pharmacokinetics of trans- and cis-4′-hydroxyCCNU were determined in five patients with Hodgkin’s or non-Hodgkin’s lymphoma receiving sequential doses of CCNU (15 mg/kg), etoposide (60 mg/kg) and cyclophosphamide (100 mg/kg) prior to autologous bone marrow transplantation. Methods: Plasma concentrations of the isomeric forms of the metabolites were determined by high-performance liquid chromatography. Data were analysed using noncompartmental pharmacokinetic methods. Results: Formation of the trans-isomer predominated over that of the cis-isomer, with an average exposure ratio of 1.4 (range 1.0–2.1). Peak plasma concentrations occurred between 1 and 4.1 h postdosing and averaged 1.56 mg/l for the trans-isomer and 1.10 mg/l for cis isomer. Peak plasma concen- trations and systemic exposures varied approximately six- and ninefold, respectively, reflecting significant interindividual variability in alkylating activity after high doses of CCNU. Plasma half-lives of the metabolites were 3.1 h (range 1.1–4.5 h) for the trans-isomer and 3.5 h (range 1.3–6.4 h) for the cis- isomer, and varied linearly with increasing patient body weight. There was no significant difference in plasma half-lives after high-dose CCNU administration observed in this study and those reported previously after the administration of substantially lower doses of CCNU. Conclusion. Despite linearity in the pharmacokinetics of the isomeric forms of 4′-hydroxyCCNU at high doses, large interindividual variability in exposure to the CCNU metabolites was observed. Potential saturation of metabolic pathways to the isomers at these doses may manifest as toxic symptoms since alkylating moieties formed directly from the parent, CCNU, may be associated with greater toxicity than those formed from the isomeric forms of the 4′-hydroxylated metabolite.

Introduction Aging is associated with alterations in the responses to several important vasoactive drugs. We have investigated histamine-mediated venodilation across the adult age range using the human hand vein compliance technique. Histamine produces dilation in human veins by activating both H1- and H2-receptors. Methods Full dose-response curves to histamine were constructed in 16 healthy volunteers (mean age, 47 ± 20 years; age range, 21 to 80 years) by infusing histamine (2 to 136 ng/min) into dorsal hand veins preconstricted with the α-adrenergic selective agonist phenylephrine. Results Histamine was an efficacious venodilator across the age range; the average maximal response (Emax) was 122 ± 45% and the geometric mean ED50 (the dose producing half-maximal response) was 16.6 ng/min for all subjects. Dose-response curves to histamine were repeated after infusion of the H2-selective antagonist cimetidine at a dose sufficient to completely block the H2-mediated response (49 μg/min). Cimetidine did not inhibit the Emax in the elderly as much as it did in the young subjects. The Emax to histamine in the presence of cimetidine plotted against age showed a significant relationship (r = 0.62, p = 0.02). Conclusions These results suggest that, although the overall histamine-induced venodilation is conserved in aging, there is a loss of the signal transduction pathway activated by way of H2-receptors but no loss in function of H1-receptors. Consequently, these results suggest differential changes in function of H1-versus H2-histamine receptors with aging. Because H1-receptors are coupled to endothelial-derived relaxing factor release and because H2-receptors activate cyclic adenosine monophosphate in smooth muscle, the results are compatible with hypothesis that there are specific changes in these signal transduction pathways with aging. Clinical Pharmacology & Therapeutics (1995) 58, 73–80; doi: 10.1016/0009-9236(95)90074-8

BACKGROUND Hypercholesterolemia impairs endothelium-dependent dilation in arteries. We tested the hypothesis that hypercholesterolemia impairs endothelium-dependent vasodilation by an interaction between elevated plasma lipoproteins and a presumably normal endothelium using human veins in vivo; veins do not generally develop atherosclerosis and are appropriate for testing functional alterations. METHODS AND RESULTS Full dose-response curves were constructed in 13 hypercholesterolemic and 12 normocholesterolemic subjects by infusing bradykinin (0.25 to 508 ng/min) into hand veins preconstricted with the alpha-adrenergic agonist phenylephrine. The maximal relaxation induced by bradykinin was 80 +/- 38% in the controls and 103 +/- 40% in subjects with hypercholesterolemia (P = .08). Responsiveness to bradykinin was also determined after infusion of indomethacin (5.4 micrograms/min), a cyclooxygenase inhibitor, to block the contribution of prostaglandins; maximal responsiveness was greater in hypercholesterolemic subjects (112 +/- 41%) than in controls (81 +/- 31%) (P = .03). Hypercholesterolemic subjects were more sensitive to bradykinin, with an ED50 of 4.2 ng/min versus 10.9 ng/min in controls (P = .05); a similarly increased sensitivity was found in the presence of indomethacin. The response to a maximally effective dose of nitroglycerin was greater in hypercholesterolemic subjects (142 +/- 31%) versus 106 +/- 28% in controls (P = .007). In five hypercholesterolemic subjects, treated with lovastatin to normalize serum cholesterol concentrations, maximal responsiveness to bradykinin decreased from 103 +/- 52% to 80 +/- 28%. CONCLUSIONS These results demonstrate that hypercholesterolemia in humans does not impair endothelium-derived relaxing factor-mediated venodilation.

Objective: To determine whether the considerable interindividual variability in nitroglycerin-induced venodilation in humans is related to the polymorphic expression of the μ class of glutathione S-transferase (GSTμ). Recently vascular glutathione S-transferase (EC 2.5.1.18) of the μ-class (GSTμ, a polymorphic group of enzymes present in only about 60% of the population, have been identified and shown in vitro to possess high metabolic activity toward nitroglycerin. Their clinical relevance is unknown. Design: Dose-response relationships to nitroglycerin were constructed in vivo measuring changes in compliance of dorsal hand veins in 26 healthy volunteers during local infusion of small amounts of nitroglycerin. Polymerase chain reaction was applied to detect the deoxyribonudeic acid sequence that codes GST μ in whole blood samples. Results: The GSTμ isozyme was present in 15 subjects (58%) and deficient in 11 subjects. Values for mean maximum venodilation (Emax) and dose rates producing 50% of Emax (ED50) were not significantly different between the groups with or without GSTμ. The respective values were 98% and 103% dilation for Emax and 9 and 16 ng/min for ED50. There was no gender difference in the venodilatory response to nitroglycerin. Conclusions: Subjects lacking GSTμ can clearly respond normally to nitroglycerin, and the large interindividual variability in nitroglycerin potency is not related to the expression of this polymorphic enzyme. Intersubject variability is therefore more likely to be the result of differences in the presence or activity of other vascular enzymes or in steps further distal in the venodilatory cascade. Clinical Pharmacology and Therapeutics (1993) 53, 463–468; doi:10.1038/clpt.1993.52

Bradykinin, a nonapeptide, dilates vascular smooth muscle at least in part via endothelial cell-dependent mechanisms. The aim of this study was to examine the action of bradykinin in human veins in vivo. Utilizing the dorsal hand vein technique, dose-response curves to bradykinin (maximum dose = 513 ng/min) were constructed in veins preconstricted with the alpha-adrenergic agonist phenylephrine in healthy young volunteers. Bradykinin almost fully dilated the veins back to their baseline diameter. To determine if desensitization of bradykinin-mediated vasodilation occurs, two bradykinin dose-response curves were constructed with a short (10 min) interval between studies. The second dose-response curve showed a diminished response. The EMAX of the first curve was 97.8 +/- 47.4% dilation and the EMAX of the second curve was 64.3 +/- 28.0% dilation (p < 0.05). However, when the two curves were separated by 40 min, there was no loss of responsiveness. To investigate the mechanism by which bradykinin caused vasodilation, we used methylene blue to antagonize endothelium-derived relaxing factor, and indomethacin to block prostaglandin-dependent effects. Methylene blue partially antagonized the vasodilatory response to bradykinin, decreasing the EMAX by 50%. Indomethacin also partially antagonized the vasodilatory response, but to a lesser extent than did methylene blue. The vasodilatory response to bradykinin was not fully antagonized with concurrent infusion of both methylene blue and indomethacin. The mechanism of bradykinin-induced vasodilation involves both EDRF and prostacyclin, and possibly another, as yet unidentified, mediator as well.(ABSTRACT TRUNCATED AT 250 WORDS)

Independent of the route of nitroglycerin administration, substantial amounts of 1,2-glyceryl dinitrate (1,2-GDN) and 1,3-glyceryl dinitrate (1,3-GDN) metabolites accumulate in humans. Thus far their pharmacologic activity in comparison to nitroglycerin in humans is unknown. To compare the venodilatory potency of nitroglycerin and of 1,2-GDN and 1,3-GDN in vivo, cumulative dose-response curves were established in nine healthy volunteers by use of the dorsal hand vein compliance technique. Separated by a washout period, two of the three venodilators were infused in randomized order after preconstriction with phenylephrine. Values for maximum vasodilation were similar for all compounds: nitroglycerin, 109%; 1,2-GDN, 100%; and 1,3-GDN, 106%. The respective values for the dose rate exerting 50% of maximum vasodilation were 5.1, 43, and 60 ng/min, indicating that the dinitrates were about 10 times less potent than nitroglycerin (p < 0.001) but not significantly different from each other. The findings support the hypothesis that activity of nitroglycerin metabolites is related to the number of nitrate groups in the molecule and are in agreement with lower dinitrate potencies found in animal experiments.

The objective of this study was to determine whether the dorsal hand vein could be used as a model to study tolerance to oral nitrates, and whether oral N-acetylcysteine (NAC) could reverse tolerance if present. Dose-response curves to nitroglycerin were constructed for 11 normotensive volunteers before and during treatment with a sustained-release formulation of isosorbide dinitrate, 80 mg, three times daily for 7 days and followed by concurrent treatment with NAC at a mean dose of 150 mg/kg/day, in divided doses, for 2 days. In separate studies, dose-response curves were constructed for seven normotensive volunteers before and after treatment with oral NAC at the same dose for 2 days. Nitroglycerin's Emax was significantly attenuated from 115 +/- 36 to 77 +/- 22% after treatment with isosorbide dinitrate alone (p < 0.009). Concurrent treatment with NAC reversed this decrease, as nitroglycerin's Emax of 108 +/- 26% during coadministration of isosorbide dinitrate and NAC was not different from its Emax in the control period. There was also no difference in the dose of phenylephrine required to cause 80% of maximal venoconstriction throughout the study. These studies demonstrate that smooth muscle tolerance to nitrates can be demonstrated in medium-sized veins in humans. In addition, high-dose oral NAC can reverse existing tolerance to oral nitrates in human veins. These results indicate that the dorsal hand vein compliance technique is a good model for the clinical investigation of tolerance to nitrates in humans.

To compare the effects of atenolol and nifedipine on mood and cognitive function in elderly hypertensive patients.Randomized, double-blind, crossover trial.Thirty-one elderly volunteers (7 women and 4 men) 60 to 81 years of age with mild to moderate hypertension were recruited from the general community and a Veterans Affairs hospital hypertension clinic. Six volunteers withdrew at early phases of the study for reasons unrelated to adverse drug effects.Participants had 2 weeks of placebo, to 6 weeks of titration with atenolol or nifedipine, and weeks of treatment followed by similar periods with the other drug.Psychometric tests designed to assess mood and cognitive function.In the group first treated with nifedipine, the summed recall score on the Buschke selective reminding test (a test of verbal learning and memory) decreased by 9.3 words (95% CI, 2.8 to 15.6 words), or 0%, during nifedipine treatment compared with placebo (P = 0.031). The group first treated with atenolol showed no improvement in summed recall scores when results seen during atenolol therapy and placebo administration were compared (P = 0.10); however, this group had an improvement of 16.1 words (CI, 5.6 to 26.5 words), or of 16%, when the atenolol score was compared with the nifedipine score (P = 0.026). In the group first treated with nifedipine, 6 of 11 patients 55%) showed a decrease of 5 words or more during nifedipine therapy compared with placebo, whereas only 1 of the 14 patients (7%) in the group first treated with atenolol showed a similar decrease (P less than 0.01). On the digit symbol test (a psychomotor test), patients treated first with atenolol tended to improve, whereas patients treated first with nifedipine tended to decline. The difference between nifedipine and atenolol, in terms of the change from the score seen during placebo, was 4.3 codings (CI, 0.7 to 7.9 codings) or 10% (P = 0.043). No statistically significant differences were seen between nifedipine and atenolol therapy regarding the other measures of psychomotor ability, sustained attention, motor performance, verbal fluency, or abstract reasoning, and no effects of either drug on mood or psychopathologic symptoms were noted.Although atenolol and nifedipine are generally free of gross effects on cognition or mood, nifedipine may subtly impair learning and memory in some elderly hypertensive patients.

Data obtained from neonates receiving zidovudine as part of a phase I study were used to estimate the population pharmacokinetic parameters of this drug and to determine the minimum number of data points necessary to provide accurate estimates of the kinetic parameters and their variability. Analysis was performed with 541 concentrations of zidovudine, obtained from 32 infants and with a variety of reduced data sets using NONMEM (nonlinear mixed effect model). The reduced data sets were derived by randomly reducing the number of sampling time points per dosing interval and/or by randomly reducing the number of available subjects. We determined that accurate estimates of pharmacokinetic parameters and their variability were obtained with the inclusion of all 32 patients using only two concentration-time points per dose interval, provided that one of the points was obtained during the first 2 h after administration of the drug. The parameters themselves were adequately estimated with only 24 subjects and two concentration-time points per dose interval. We suggest that NONMEM should be used in addition to the traditional pharmacokinetic analysis to obtain more precise information directly in the population of interest with a minimum of blood sampling from each patient. This is especially critical in infants whose blood volumes are limited.

delta-Aminolevulinic acid-4-[14C] and [3H]-bilirubin were administered intravenously to five patients with Gilbert's syndrome and four healthy control subjects on two occasions: before and on days 10 through 14 of a course of phenobarbital (2.5 mg/kg/day). The resulting curves of [3H]-bilirubin and [14C]-bilirubin in plasma were analyzed by computer to determine a number of parameters of physiological interest. As expected, phenobarbital produced a highly significant fall in the plasma concentration of unconjugated bilirubin as a result of a significant increase in hepatic bilirubin clearance in all subjects; plasma bilirubin turnover was unaltered. Surprisingly, the drug produced no change in the incorporation of [14C]-delta-aminolevulinic acid into [14C]-early labeled bilirubin. To explain this unexpected finding, the effects of phenobarbital (75 mg/kg/day for 6 days) on incorporation of [14C]-delta-aminolevulinic acid and 2-[14C]-glycine into [14C]-early labeled bilirubin and on the activity of the enzyme delta-aminolevulinic acid synthase were studied in nonfasted, adult, male Sprague-Dawley rats. At the dose and duration of treatment used, phenobarbital administration increased total hepatic delta-aminolevulinic acid synthase activity and produced a significant increase of 70% in the incorporation of [14C]-glycine into early labeled bilirubin. By contrast, no increase in the incorporation of [14C]-delta-aminolevulinic acid into early labeled bilirubin was observed. These data suggest that delta-aminolevulinic acid is an inappropriate precursor for studies of the rate of heme biosynthesis, presumably because it bypasses delta-aminolevulinic acid synthase, the physiological rate-limiting enzyme in the heme biosynthetic pathway.

Vasodilators have varying hemodynamic properties that may be important in determining their efficacy for different disorders. We used the dorsal hand vein technique to measure the effects of several vasodilators infused locally and to measure the action of systemically administered nifedipine. The venodilatory effects of hydralazine, verapamil, diazoxide, and nitroglycerin were determined in peripheral veins of healthy subjects. The potency (ED50, the dose producing half-maximal response) was as follows: nitroglycerin (0.0007 micrograms/min) greater than verapamil (6.5 micrograms/min) greater than diazoxide (75 micrograms/min) greater than hydralazine (660 micrograms/min). The effect of oral nifedipine on alpha-adrenergic responsiveness of hand veins was determined. Nifedipine given in therapeutic doses for the treatment of hypertension was associated with a threefold increase in the ED50 for phenylephrine (92 to 277 ng/min; p less than 0.05) and norepinephrine (2.9 to 11.5 ng/min; p less than 0.05). Therapeutic doses of nifedipine are associated with measurable shifts in the dose-response curves to these two alpha-adrenergic agonists in the hand vein. The hand vein technique can be used not only to compare the potency of locally infused drugs but also to measure venous effects of vasoactive drugs at therapeutic concentrations achieved after systemic administration.

The effect of ranitidine on both induced (phenobarbital) and uninduced cytochrome P450 enzymes was investigated in mice using the [14C]-labeled antipyrine breath test. Ranitidine administration resulted in a decrease in the fraction of the administered dose of antipyrine exhaled as radiolabeled CO2 (CERAUC0-infinity) indicating inhibition in the demethylase pathway (Kdm), and resulted in induction of enzymes in the non-demethylase pathways (Kndm) as well. No change in antipyrine total elimination rate constant (Kel) was seen after ranitidine administration alone. Concurrent administration of ranitidine and phenobarbital resulted in an increase in the (Kel) but the change was less than that seen after phenobarbital alone. A reduction in CERAUC0-infinity was seen after the combination treatment while phenobarbital alone resulted in an increase in this parameter. Ranitidine, therefore, alters the pattern of antipyrine metabolism by inhibition of demethylase enzymes and induction of non-demethylase enzymes, the former activity being more pronounced in induced forms. Because of the simultaneous occurrence of both effects, no change in antipyrine half-life was noted with uninduced P450 isozymes.

The mercapturic acid conjugate of acetaminophen has been proposed as an index of the toxic intermediate of acetaminophen metabolism which is responsible for the increased incidence of liver injury in alcoholics taking the drug. Previous studies which compared alcoholics with normal patients failed to show any significant difference in mercapturic acid production. We undertook a longitudinal study in recovering alcoholics to test the hypothesis that abstinence should lead to a decrease in acetaminophen-mercapturic acid excretion. The patients were given a 1500 mg dose of acetaminophen soon after they stopped drinking then again approximately 2 weeks later. Urine was collected for 24 h and assayed for mercapturic acid conjugate. There was no significant difference in mercapturic acid excretion when the two measurements were compared. If the toxic intermediate hypothesis is correct, either the effect of alcohol is prolonged, or additional factor other than alcohol exposure may influence mercapturic acid excretion.

We evaluated the effects of extracorporeal membrane oxygenation (ECMO) on the pharmacokinetics of gentamicin in 18 infants who underwent ECMO therapy for severe respiratory failure and received gentamicin for possible sepsis. Twelve of these infants continued to receive gentamicin after ECMO had been discontinued. The volume of distribution (Vd) of gentamicin in the newborns receiving ECMO was 0.58 +/- 0.04 liter/kg, compared with a Vd of 0.45 +/- 0.02 liter/kg after ECMO had been discontinued (P = 0.02). The clearance of gentamicin in the patients undergoing ECMO was 42 +/- 3 ml/kg/hour compared with 57 +/- 4 ml/kg/hour in those patients off ECMO (P = 0.003). The elimination half-life in patients receiving ECMO was 10.0 +/- 0.7 hours compared with 5.7 +/- 0.4 hours after ECMO had been discontinued (P less than 0.0001). Neonates undergoing ECMO demonstrate a higher volume of distribution of gentamicin, a lower clearance, and consequently a longer half life for this drug. We conclude that gentamicin and probably other aminoglycosides should be given at dose rates about 25% lower than usual and at longer dosing intervals in patients undergoing ECMO therapy.

1. Blood pressure regulation was studied in 12 healthy elderly subjects after double-blind randomised administration of placebo, 15 mg and 30 mg temazepam at 10.00 h and 22.00 h. 2. Supine and standing heart rate and blood pressure were measured after daytime administration and supine measurements were obtained during sleep. 3. Temazepam caused a fall in systolic blood pressure and an increase in heart rate after morning administration. These changes were greater in the standing position and were dose-dependent; for standing blood pressure and heart rate 1 h after administration there was a 7 mm Hg fall and 6 beats min-1 increase after 15 mg temazepam and a 10 mm Hg fall and 8 beats min-1 increase after 30 mg temazepam. Temazepam magnified the fall in systolic blood pressure and increase in heart rate that occurred with standing. Temazepam enhanced the fall in systolic blood pressure that occurred during sleep (mean +/- s.d.; placebo: -23 +/- 10 mm Hg, 15 mg temazepam: -31 +/- 13 mm Hg, 30 mg temazepam: -36 +/- 14 mm Hg). 4. These changes in blood pressure regulation caused by temazepam may have clinical importance in some elderly individuals.