Ivana Cvetanovic

Comparative Effects of Rapid Bolus Administration of Aqueous Amiodarone Versus 10-Minute Cordarone I.V. Infusion on Mean Arterial Blood Pressure in Conscious Dogs

Objective: This study was designed to test the hypothesis that rapid bolus administration of an aqueous formulation of intravenous amiodarone causes less hypotension than a 10-minute infusion of the standard formulation, Cordarone IV. Hypotension was the most common adverse event reported with Cordarone IV. The hypotension was not dose related, but related to the rate of infusion. Therefore, product labeling calls Cordarone and its generic formulations to be administered over 10 minutes. Cordarone IV contains polysorbate 80 and benzyl alcohol, each causes hypotension. A new aqueous formulation of amiodarone (Amio-Aqueous) does not contain these agents and therefore may cause less hypotension.Methods: Six conscious beagle dogs were instrumented with a telemetric device for blood pressure monitoring. The study was conducted on 5 days. On the first 2 days, a 10-min infusion or a bolus of D5W was administered (placebo). Over the following 3 days, the dogs received (in randomized order, one per day) a 10-min infusion of 2.5 mg/kg Cordarone IV and boluses of 2.5 mg/kg and 5.0 mg/kg Amio-Aqueous injected over 2 to 5 sec. The dogs were monitored for 2 hrs after dosing.Results: Compared to placebo, boluses of aqueous amiodarone produced no significant changes in the mean arterial blood pressure (MABP). In contrast, Cordarone infusion produced significant decreases in MABP that lasted for at least 2 hrs (p < 0.001).Conclusion: Amio-Aqueous had significantly better hemodynamic profile permitting rapid intravenous administration. This is a significant advantage over the standard formulation, because Cordarone cannot be administered by rapid bolus due to excipient-related hypotension.

Gender Differences in Cardiac Repolarization Following Intravenous Sotalol Administration

Background: Females are more susceptible to drug-induced torsade de pointes (TdP), which is associated with excessive prolongation of the heart rate-corrected QT interval (QTc). Sotalol prolongs the cardiac action potential that can be observed as QT prolongation and can induce TdP. The aim of this study was to assess gender differences in sotalol-induced QTc prolongation. Methods: A total of 15 healthy volunteers, 9 female and 6 male (age: 32 ± 8 years) received 75 mg intravenous sotalol over 2.5 hours at a constant infusion rate. A 12-lead electrocardiograph (ECG) was recorded at baseline, 0.5, 1, 2, 3, 4, and 5 hours following the start of the infusion, and blood samples were collected simultaneously. QTc was calculated by the Fridericia and Framingham formulas. The 2 formulas resulted in virtually identical QTc intervals. The data analysis included repeated measures of analysis of variance (ANOVA), univariate analysis, and linear regression analysis. Results: The longest average QTc intervals were observed at 2 hours of sotalol infusion in both genders. Compared to baseline, the increase was very significant in females (411 ± 13 vs 438 ± 13 ms, P < .001), while it was less significant in males (395 ± 23 vs 413 ± 27 ms, P < .05). The magnitude of individual changes from baseline were greater in females than in males (34 ± 8 vs 21 ± 12 ms, P < .05). In each gender, QTc and serum sotalol concentration strongly correlated (r = .93, P < .001). An upward shift of the regression line in females indicates a longer QTc at any concentration level. Males had greater body weight and body surface area than females (P < .05) but neither correlated with QTc or predicted QTc prolongation. The univariate analysis indicated that the single predictor for the greater QTc prolongation was female gender. Conclusion: Females had greater QTc prolongation than males following sotalol administration. This enhanced response to drug action may explain the higher incidence of drug-induced TdP seen in females.

Pharmacology and toxicology of a new aqueous formulation of intravenous amiodarone (Amio-Aqueous) compared with Cordarone IV.

Hypotension is the most frequent adverse event reported with intravenous amiodarone (Cordarone IV). The hypotension has been attributed to the vasoactive solvents of the formulation, polysorbate 80 and benzyl alcohol, both known to exhibit negative inotropy and hypotensive effect. A new aqueous formulation of intravenous amiodarone (Amio-Aqueous) does not contain vasoactive excipients and may be less toxic and cause less hypotension than Cordarone IV. This hypothesis was tested in a series of animal studies with direct comparison of Amio-Aqueous and Cordarone IV. All studies were performed on Sprague-Dawley rats. The acute toxicology study showed that both LD50 and LD100 were 30% greater for Amio-Aqueous than for Cordarone. At the dose at which all animals expired on Cordarone, 50% of animals were still alive on Amio-Aqueous. The study on myocardial contractility showed that Amio-Aqueous was a far less negative inotropic than Cordarone IV (P < 0.001). Amio Aqueous did not have an effect on contractility at 5- and 10-mg/kg dose levels while Cordarone resulted in a 25% (P < 0.01) and 29% (P < 0.002) decrease, respectively. The study on arterial blood pressure showed that Cordarone caused a significant decrease in blood pressure at each of the 3, 5, 10, and 20 mg/kg doses (P < 0.05 to P < 0.001) while Amio-Aqueous did not. The study on the antiarrhythmic effects showed comparable efficacy for both formulations. In conclusion, Cordarone IV was more toxic and caused significant hypotension and negative inotropy while Amio-Aqueous lacked the hypotensive and cardiotoxic properties of Cordarone. Therefore, Amio-Aqueous is a safer alternative than the standard formulation.

The Influence of Extracellular Acidosis on the Effect of IKr Blockers

Background: Myocardial infarction causes the acidification of the cellular environment and the resultant acidosis maybe arrhythmogenic. The effect of acidosis on the action of antiar-rhythmic drugs, an important issue in the antiarrhythmic drug therapy after myocardial infarction, remains to be studied. Methods: To evaluate the effect of acidosis on rectifier potassium current (Ikr) blockers, the human ether-a-go-go-related gene (HERG), which encodes IKr, was expressed in Xenopus laevis oocytes. The two electrodes voltage clamp technique was used and the experiments were performed at room temperature. Results: Quinidine (10 µM) inhibited HERG tail current by 37% ± 5% at pH7.4. The block decreased to 5% ± 2% with extracellular pH at 6.2. Dofetilide (0.3 µM) inhibited HERG tail current by 34% ± 3% and 1% ± 2% at extracellular pH 7.4 and 6.2, respectively. Azimilide (10 µM) inhibited HERG tail current by 59% ± 3% and 17% ± 3% at extracellular pH 7.4 and 6.2. There were significant differences in the HERG inhibition by quinidine, dofetilide, and azimilide between pH 7.4 and pH 6.2 (P < .01). The drug concentration blocking 50% of current (IC50) was 5.8 ± 0.3 µM for azimilide, 9.9 ±1.0 µM for quinidine, and 0.5 ± 0.02 µM for dofetilide at pH 7.4. When extracellular pH was decreased from 7.4 to 6.2, the IC50 increased to 95.5 ± 11.3 µM for azimilide, 203.2 ± 15.7 µM for quinidine, and 12.6 ± 1.2 µM for dofetilide. Unlike quinidine, dofetilide, and azimilide, there was no significant difference in the percentage of current block by amiodarone between pH 6.2 and 7.4. For amiodarone, the IC50 was 38.3 ± 8.5 µM at pH 7.4 and 27.3 ± 1.6 µM at pH 6.2. Conclusion: Our data show that the Ikr blocking effect of azimilide, dofetilide, and quinidine was attenuated at acid pH, whereas this was not the case for amiodarone. These observations may explain the efficacy of amiodarone in reducing arrhythmic death in patients after a myocardial infarction compared with other IKr blockers.

Evaluation of a 12-Lead Digital Holter System for 24-Hour QT Interval Assessment

Background: Drug induced QT prolongation may precipitate life threatening cardiac arrhythmias. Evaluation of the QT prolonging effect of new pharmaceutical agents in a ‘thorough QT/QTc study’ is being mandated by FDA. The purpose of this study was to evaluate an automated 12-lead digital Holter system for a thorough QT/QTc study. Methods: Five healthy volunteers underwent 24-hour digital Holter monitoring. Each recording underwent a fully automated QT analysis (AQA) followed by an onscreen complete manual over read (MOR). Each recording was analyzed twice at least 2 weeks apart. The effect of data sampling (5-min segment/hour), the system sensitivity to detect 5-ms increase in QT, and the ability to assess circadian variation were evaluated. Results: The AQA resulted in identical QT for the first and second analyses, but with obvious errors in QT measurements. Compared to the complete onscreen MOR, the mean QT was longer with AQA (416 ± 41 vs. 387 ± 30 ms, p < 0.001), correlation; r = 0.3. The reproducibility of AQA with complete MOR was very good (QT: 387 ± 30 vs. 387 ± 30 ms, coefficient of variation: 0.2%, r = 0.986. The 5-min mean QT intervals correlated well with the hourly mean QT intervals (r = 0.994, p < 0.001, coefficient of variation = 1 ms) and both showed a similar circadian variation. The system was sensitive to detect a 5-ms change in QT intervals (5 ± 2 ms, coefficient of variation = 0.6%, r = 0.998, p < 0.001). Conclusions: The AQA is not an acceptable method, while the automatic analysis with complete MOR is a highly sensitive and reproducible method. Data sampling by analyzing 5-min segments per hour is sensitive and reproducible.

A mechanism for the potential proarrhythmic effect of acidosis, bradycardia, and hypokalemia on the blockade of human ether-a-go-go-related gene (HERG) channels

Many drugs are proarrhythmic by inhibiting the cardiac rapid delayed rectifier potassium channel (IKr). In this study, we use quinidine as an example of highly proarrhythmic agent to investigate the risk factors that may facilitate the proarrhythmic effects of drugs. We studied the influence of pacing, extracellular potassium, and pH on quinidines IKr blocking effect, all potential factors influencing quinidines cardiac toxicity. Since the HERG gene encodes IKr, we studied quinidines effect on HERG expressed in Xenopus oocytes by the 2-electrode voltage clamp technique. When extracellular K+ was 5 mmol/L, quinidine blocked the HERG current dose dependently, with an IC50 of 6.3 ± 0.2 μmol/L. The blockade was much more prominent at more positive membrane potentials. The inhibition of HERG by quinidine was not use dependent. There was no significant difference between block with or without pacing. When extracellular K+ was lowered to 2.5 mmol/L, the current inhibition by quinidine was enhanced, and IC50 decreased to 4.6 ± 0.5 μmol/L. At 10 mmol/L extracellular K+, there was less inhibition by quinidine and the IC50 was 11.2 ± 3.1 μmol/L. Extracellular acidification decreased both steady state and tail currents of HERG. We conclude that the inhibitory effect of quinidine on IKr was decreased with extracellular acidification, which may produce heterogeneity in the repolarization between normal and ischemic cardiac tissue. Thus, the use-independent blockade of IKr by QT-prolonging agents such as quinidine may contribute to cardiac toxicity with bradycardia, hypokalemia, and acidosis further exaggerating the proarrhythmic potential of these agents.

The Effect of High Extracellular Potassium on IKr Inhibition by Anti-Arrhythmic Agents

Background: Hyperkalemia is a potentially life-threatening disorder frequently occurring in hospitalized patients. The ischemic myocardium releases potassium into the extracellular space which can cause regional hyperkalemia. These changes may modify the effects of anti-arrhythmic drugs acting on the rapid component of the delayed rectifier potassium current (IKr). We evaluated the influence of increased extracellular potassium concentration [K+]e on IKr inhibition by amiodarone, azimilide, dofetilide, quinidine and sotalol. Methods and Results: Experiments were performed at room temperature. IKr current was studied by using HERG gene expressed in Xenopus oocytes as a model of cardiac IKr. Two-electrode voltage clamp technique was employed. The recording bath solutions contained either 5 or 10 mmol/l KCl. Amiodarone, azimilide, dofetilide, quinidine and sotalol all produced a dose-dependent inhibition of HERG current. At 5 mmol/l [K+]e, the IC50 was 37.0 ± 12.5 µM for amiodarone, 5.8 ± 0.4 µM for azimilide, 1.5 ± 0. 2 µM for dofetilide, 9.1 ± 1.5 µM for quinidine, and 5.1 ± 0.8 mM for sotalol. Raising the extracellular potassium to 10 mmol/l, HERG block by azimilide, dofetilide, quinidine and sotalol was significantly decreased, while the block by amiodarone was unchanged. The differences in the percentage current block produced by 3 µM drugs at 5 and 10 mmol/l [K+]e were: –0.9% for amiodarone, 13.8% for quinidine, 20.5% for azimilide, and 16.2% for dofetilide. The differences in percentage block between 5 and 10 mmol/l [K+]e by sotalol 10 and 30 mM were 7.1 and 5.6%. At 10 mmol/l [K+]e, the IC50 was increased for azimilide, dofetilide, quinidine and sotalol but not for amiodarone; the IC50 was 24.7 ± 7.4 µM for amiodarone, 29.3 ± 3.9 µM for azimilide, 2.7 ± 0.2 µM for dofetilide, 27.6 ± 4.0 µM for quinidine, and 7.2 ± 1.7 mM for sotalol. Conclusion: Inhibition of IKr by azimilide, quinidine, dofetilide and sotalol was diminished by increasing [K+]e, while the inhibition by amiodarone was unchanged at normal and high [K+]e. The differential effects of azimilide, dofetilide, quinidine and sotalol at normal and high [K+]e could be pro-arrhythmic by favoring re-entry arrhythmias. These results further support the unique electrophysiological effect of amiodarone.

The evaluation of the diuretic action of parenteral formulations of metolazone

Background and ObjectivesThis study was design to compare the diuretic and natriuretic effects of the 2 parenteral formulations of metolazone and the combination of these 2 formulations of metolazone with the parenteral administration of furosemide. Metolazone is an anthracrene acid derivate and manifests a dual diuretic effect on the proximal and distal tubule with a minimal kaluretic effect. It is currently only marketed in an orally administrable formulation, and this has limited its utility in critically ill patients. Metolazone given orally and furosemide given orally or parenterally are frequently administrated together when furosemide alone is clinically inadequate at producing a desired diuresis. MethodsSprague Dawley male rats (400 to 450 g) were divided into groups to receive a parenteral formulation of metolazone or furosemide administrated separately intraperitoneally (IP) or administrated IP in combination with one another. Tris buffer-administered IP was used as a control vehicle comparator. The urine volume voided over the following 24 hours was collected, measured and analyzed for sodium content. ResultsVehicle (Tris buffer) caused 9 ± 1 mL/d output of urine with a sodium [Na+] concentration of 194 ± 41 μmol/L (n = 6 per group). Metolazone 2 mg/kg resulted in 16 ± 3 mL/d urine output and sodium [Na+] of 278 ± 76 μmol/L (n = 6 per group). Furosemide 2, 4, and 6 mg/kg resulted in a volume of urine 9 ± 1, 14 ± 2 and 17 ± 2 mL/d and [Na+] μmol/L of 194 ± 41, 206 ± 108, and 229 ± 91, respectively. Metolazone 4 mg/kg combined with furosemide 4 mg/kg resulted in a urine volume of 21 ± 1 mL/d and [Na+] of 326 ± 108 μmol/L. ConclusionCombining metolazone and furosemide can cause an increase in urine volume and sodium excretion. Metolazone administrated parenterally in combination with the parenteral administration of furosemide appears to have an important clinical potential.

The differential antibacterial and gastrointestinal effects of erythromycin and its chiral isolates.

The use of erythromycin has been limited by the gastrointestinal side effect properties, which include abdominal distress and diarrhea. To evaluate the possibility of reducing the toxicity of erythromycin, studies were undertaken to separate erythromycin into chiral isolates and then to test the activity of these chiral isolates on gastrointestinal contractility and bacteriostatic actions. Gastrointestinal contractility was obtained by the use of isolated strips of a rat colon. Antibacterial activity was used by obtaining the MICs of erythromycin and isolated agents against Enterococcus faecalis ATCC 29212. ANOVA was performed using the SPSS v.10 to determine statistical differences in the MICs and the amplitude and frequency of spike bursts. Results were expressed as mean ± SE (N = 5). The MICs (μg/mL) of erythromycin (racemate), chiral isolate X, and chiral isolate Y were 0.45 ± 0.29, 0.53 ± 0.24 (n.s.), and 0.2 ± 0.07 (P ≤ 0.001), respectively. Erythromycin (racemate) at 10−8 mol/L, 10−7 mol/L, 5 × 10−7 mol/L, 10−6 mol/L, and 10−5 mol/L concentrations caused the amplitude of spike bursts to increase by 18 ± 7% (P = n.s.), 43 ± 10% (P ≤ 0.05), 55 ± 12% (P ≤ 0.001), 121 ± 23% (P ≤ 0.001), and 163 ± 16% (P ≤ 0.001), respectively. The chiral isolate Y increased the amplitude of spike bursts at the same concentrations as tested above: 32 ± 11% (P ≤ 0.05), 48 ± 14% (P ≤ 0.001), 84 ± 13% (P ≤ 0.001), 112 ± 18% (P ≤ 0.001), and 121 ± 13% (P ≤ 0.001), respectively. Chiral isolate X caused much reduced effect on the amplitude of spike bursts: 9 ± 6% (P = n.s.), 27 ± 12% (P = n.s.), 27 ± 12% (P = n.s.), 30 ± 11% (P = n.s.), and 30 ± 11.2% (P = n.s.), respectively. EC50 for erythromycin (mixture) was 0.4 × 10−6 mol/L, and for erythromycin Y, it was 0.8 × 10−6 mol/L. The addition of erythromycin at 10−8 mol/L caused the frequency of spike bursts to increase 11 ± 7% at 10−7 mol/L, 5 × 10−7 mol/L, 10−6 mol/L, and 10−5 mol/L; the changes were 13 ± 10% (P = n.s.), 13 ± 10% (P = n.s.), 22 ± 13% (P = ns), and 39 ± 30% (P ≤ 0.05), respectively. Chiral isolate Y of erythromycin, changed the frequency of spike bursts by 26 ± 21% (P = n.s.); 35 ± 20% (P = n.s.), 39 ± 30% (P = n.s.), 41 ± 37% (P = n.s.), and 44 ± 36% (P = n.s.) at the respective concentrations as discussed above. Chiral isolate X altered the frequency of spike bursts at the same concentrations as 40 ± 30% (P = n.s.), 45 ± 30% (P = n.s.), 62 ± 41% (P = n.s.), 62 ± 41% (P = n.s.), and 52 ± 35% (P = n.s.), respectively. Data indicate that erythromycin (racemate) and chiral isolates X and Y possess similar antibacterial activity. It was also shown that erythromycin and chiral isolate Y increase significantly the amplitude of spike bursts compared with baseline. Isolate X does not increase the amplitude of spike bursts in a dose-dependent manner. The frequency of spike bursts is not significantly changed in the presence of erythromycin or the 2 chiral isolates.

The effects of quinidine and its chiral isolates on erg-1sm potassium current and correlation with gastrointestinal augmentation

Smooth-muscle erg 1 (erg1-sm) potassium channel has been recently reported to participate in the modulation of gastrointestinal contractility. Because quinidine inhibits cardiac potassium channel and as a result augments gastrointestinal contractility, it was thought that quinidine may affect erg1-sm. Studies were undertaken to evaluate the effects of quinidine and its chiral isolates on gastrointestinal erg1-sm potassium current and correlate these effects with colon contractility. Chiral separation (high-performance liquid chromatography technique), mass spectrometry, and optical rotation determination were performed to obtain chiral isolates needed for experiments. The erg1-sm potassium channel was expressed in Xenopus oocytes, and the two-electrode patch clamp technique was employed for recording. An isolated rat colon preparation was employed to measure changes in contractility. As a result of chiral separation, two peaks were obtained with elution times of 8.31 and 8.66 minutes, both with a molecular weight of 324; the optical rotations of racemate isolates X and Y were: +258°, ±0°; and +217°, respectively. The percentage changes in amplitudes of colon contraction (from baseline) were determined at different concentrations of quinidine and for the two isolates in five experiments in each group. Quinidine 0.1, 1, and 10 μM increased contractility by 79 ± 34, 125 ± 42, and 217 ± 51 (P ≤ 0.05); for isolate X, the values were 70 ± 20, 115 ± 32, and 272 ± 32 (P ≤ 0.05), and for isolate Y the values were 22 ± 12, 46 ± 17, and 59 ± 22. The inhibition of erg1-sm currents by quinidine was 19 ± 4, 21 ± 5, and 48 ± 6 (P ≤ 0.05), respectively; that by isolate X was 20 ± 4, 23 ± 5, and 39 ± 7 (P ≤ 0.05), and that by isolate Y was 22 ± 4, 21 ± 4, and 31 ± 6. One chiral isolate and quinidine markedly augmented contractility, whereas quinidine and the two chiral isolates inhibited the erg1-sm potassium currents to a similar extent. These results suggest that erg1-sm inhibition does not explain gastrointestinal contractile augmentation caused by the quinidine racemate and its chiral isolates.