Interim analysis: A rational approach of decision making in clinical trial.

Interim analysis of especially sizeable trials keeps the decision process free of conflict of interest while considering cost, resources, and meaningfulness of the project. Whenever necessary, such interim analysis can also call for potential termination or appropriate modification in sample size, study design, and even an early declaration of success. Given the extraordinary size and complexity today, this rational approach helps to analyze and predict the outcomes of a clinical trial that incorporate what is learned during the course of a study or a clinical development program. Such approach can also fill the gap by directing the resources toward relevant and optimized clinical trials between unmet medical needs and interventions being tested currently rather than fulfilling only business and profit goals.

Application of critical path analysis in clinical trials.

Clinical research operates in a strictly regulated environment under various management models, but a distinct management model of clinical trial (CT) still needs exploration and research. Critical path analysis (CPA) is a management approach can be used for monitoring, analysis, and prediction of success of its time-bound operational activities. A model CT was compiled with 78 activities, which were further merged into 35 major activities. After performing dependence analysis, the list was finalized with 25 activities which were taken in activity predecessor to create a network diagram and perform CPA considering patients, conduct, and outcome. Activities were inclusive, described the trial entirely with accuracy, and were in chronological and logical sequences. This approach does not replace an understanding of or adherence to the requirements contained in all applicable regulations, guidelines or standard operating procedures governing clinical studies but ensures the proper use of operational and decisional approaches including optimal resource management. As the need to meet deadlines becomes more important and the need to produce good, stable project plans, CPA is very useful for determining activities that can lead to project delay. With this approach, project may be effectively monitored, and realistic schedules can be maintained.

Influence of ionization source design on matrix effects during LC-ESI-MS/MS analysis.

Glycerophosphocholines (GPChos) are known to cause matrix ionization effects during the analysis of biological samples (i.e. plasma, urine, etc.) in LC-MS/MS. In general, such matrix effect is directly related to an insufficient sample clean-up of the biofluids. In addition to GPCho; design of ionization source and/or LC also plays a very important role in matrix effects. In this research paper, different types of matrix effects, i.e. ion suppression or enhancement were observed in differently designed ion sources coupled with different LCs, from the same molecule, acamprosate (ACM), under the same chromatographic conditions. ACM was analyzed in a negative polarity in electrospray ionization interface using Z-spray and orthogonal spray ion source design. The analyte showed almost complete ion suppression in the Z-spray ionization source coupled with UPLC/HPLC, whereas there was very little ion enhancement in the orthogonal spray ionization source coupled with HPLC. In both the cases different GPChos were responsible, as evident from the presence of m/z 815.4 in Z-spray ion source and m/z 759.0 in orthogonal spray ion source. Hence, this approach can be used to evaluate the matrix effects in plasma samples during development and validation of LC-MS/MS method of drugs and their metabolites in different biological matrixes.

Rapid and sensitive liquid chromatography/tandem mass spectrometry method for simultaneous determination of enalapril and its major metabolite enalaprilat, in human plasma: application to a bioequivalence study.

A rapid and most sensitive method for simultaneous determination of enalapril (ENP) and its metabolite, enalaprilat (ENPT), in human plasma using ESI-LC-MS/MS (electrospray ionization liquid chromatography tandem mass spectrometry) positive ion multiple reactions monitoring (MRM) mode, was developed and validated. The procedure involves a simple solid-phase extraction (SPE) followed by evaporation of the sample. Chromatographic separation was carried out on a Hypurity C(18) column (50 mm × 4.6 mm, 5 µm) with an isocratic mobile phase and a total run time of 2.0 min only. The MRM of ENP and ENPT is 377.10 → 234.20 and 349.20 → 206.10 respectively. The standard calibration curves showed excellent linearity within the range of 0.064 to 431.806 ng/mL for ENA and 0.064 to 431.720 ng/mL for ENPT (r ≥ 0.990). This is the only method which can quantitate upto 0.064 ng/mL for both ENP and ENPT in a single run with the shortest analysis time. In matrix effect experiment, this method shows a % CV (% coefficients of variation) of less than 5, which means that the proposed method is free from any kind of irregular ionization process. This method was successfully applied to a pharmacokinetic study after oral administration of enalapril maleate 20 mg tablet in Indian healthy male volunteers.

Clinical trials of antihypertensives: Nature of control and design.

This paper reviews the critical issues in the control and design of antihypertension (anti-HT) clinical trials. The international guidelines and current clinical and biostatistical practices were reviewed for relevant clinical, design, end-point assessments and regulatory issues. The results are grouped mainly into ethical, protocol and assessment issues. Ethical issues arise as placebo-controlled trials (PCTs) for HT-lowering agents in patients with moderate to severe HT are undertaken. Patients with organ damage due to HT should not be included in long-term PCT. Active-control trials, however, are suitable for all randomized subsets of patients, including men and women, and different ethnic and age groups. Severity subgroups must be studied separately with consideration to specific study design. Mortality and morbidity outcome studies are not required in anti-HT trials except when significant mortality and cardiovascular morbidity are suspected. Generally, changes in both systolic and diastolic blood pressures (BP) at the end of the dosing interval from the baseline are compared between the active and the control arms as the primary endpoint of anti-HT effect. Onset of the anti-HT effect can be studied as the secondary endpoint. For maintenance of efficacy, long-term studies of ≥6 months need to be undertaken. Error-free measurement of BP is a serious issue as spontaneous changes in BP are large and active drug effect on diastolic BP is often small. Placebo-controlled short-term studies (of ~12 weeks) for dose-response and titration are very useful. Safety studies must be very vigilant on hypotension, orthostatic hypotension and effects on heart. In dose-response studies, at least three doses in addition to placebo should be used to well characterize the benefits and side-effects.

A LC-MS analysis of acamprosate from human plasma: pharmacokinetic application.

A rapid and highly sensitive method for the determination of acamprosate (ACM), in human plasma using ESI-LC-MS/MS (electrospray ionization liquid chromatography tandem mass spectrometry) in negative ionization polarity in multiple reactions monitoring (MRM) mode was developed and validated. The procedure involves a simple protein precipitation step. Chromatographic separation was carried out on a Hypersil BDS C(18) column (150 mm × 4.6 mm, 5 µm) with an isocratic mobile phase and a total run time of 2.5 min. The standard calibration curves were linear within the range of 7.04-702.20 ng/mL for ACM (r ≥ 0.990). This study briefly describes the role of ion source design on matrix effects. ACM shows matrix effects in z-spray ionization source design, whereas it has no matrix effects in orthogonal spray ion source design. This method was successfully applied to a pharmacokinetic study after oral administration of acamprosate 333 mg tablet in Indian healthy male volunteers.

Simultaneous estimation of atorvastatin and its two metabolites from human plasma by ESI-LC-MS/MS.

A selective, sensitive, and fast high performance liquid chromatography (HPLC) method with mass spectrometric (MS) detection mode has been developed and validated completely in human plasma. Atorvastatin (ATO), p-hydroxy atorvastatin (p-HATO), o-hydroxy atorvastatin (o-HATO) and internal standard (IS) are extracted from human plasma via solid phase extraction (SPE) technique. After elution, the solution is evaporated, then reconstituted with 250 µL of Mobile Phase and analyzed using HPLC/MS/MS system. An isocratic mode is used to separate interference peaks using a Symmetry C-18, 75 × 4.6 mm ID, 3.5 µ, column. The m/z of ATO, o-HATO and p-HATO are 559.2/440.2, 575.3/440.4 and 575.0/440.4 respectively. Linearity ranges are 0.05 to 252.92 ng/mL for ATO, p-HATO and o-HATO respectively. Calibration functions, lower limit of quantitation (LLOQ), stability, intra- and inter-day reproducibility, accuracy, and recovery are estimated. This method is free from matrix effects and any abnormal ionization. This method was successfully applied to a single dose 80 mg tablet bioequivalence (BE) study of Atorvastatin. Copyright © 2011 John Wiley & Sons, Ltd.

Effect of a novel antiinflammatory polyherbal preparation (Sudarshanam oil) on hematological parameters in Wistar rats.

The present investigation was an attempt to evaluate the anti-inflammatory effect of a polyherbal preparation (Sudarshanam Oil) on hematological parameters in Wistar rats. This polyherbal formula contains main ingredients of Tinaspora Cordifolia, Curcuma longa, Terminalia chebula, Emblica officinalis, Andrographis paniculata and Terminalia belerica. The active phytochemicals such as tannins, glycosides, flavonoids and triterpenoids are extracted from these herbal plants. And extract was prepared in corn oil. Young adult Wistar rats of either sex were divided into 4 groups and each group having 6 males and 6 females were dosed for 28 days. First group was the control group which was dosed with corn oil as vehicle (15 mL/kg body weight). Groups II, III and IV were treated with different doses of Sudarshanam oil, viz. as 5 mL/kg, 10 mL/kg and 15 mL/kg body weight respectively. On 29th day, after overnight fasting, the blood samples were collected through cardiac puncture under CO2 anaesthesia. The blood samples were collected and transferred into prelabelled vaccutainer coated with EDTA for hematological parameters investigation by using Advia-120 hematology analyser. The results revealed that no treatment related adverse effects in any of the hematological parameters. Thus, Sudarshanam oil proves to be highly potent, novel anti-inflammatory preparation which can be a challenge against allopathic anti-inflammatory drug.

A rapid and most sensitive liquid chromatography/tandem mass spectrometry method for simultaneous determination of alverine and its major metabolite, para hydroxy alverine, in human plasma: application to a pharmacokinetic and bioequivalence study.

A rapid and highly sensitive method for the determination of alverine (ALV) and its metabolite, para hydroxy alverine (PHA), in human plasma using LC-MS/MS in positive ion electrospray ionization (ESI) in multiple reactions monitoring (MRM) mode was developed and validated. The procedure involves a simple solid phase extraction (SPE). Chromatographic separation was carried out on a Hypersil GOLD C(18) column (50 mm x 4.6 mm, 5 microm) with an isocratic mobile phase and a total run time of 1.5 min. The standard calibration curves showed excellent linearity within the range of 0.060-10.051 ng/mL for ALV and 0.059-10.017 ng/mL for PHA (r > or = 0.990). This method was successfully applied to a pharmacokinetic study after oral administration of alverine citrate 120 mg capsule in Indian healthy male volunteers.

Preservation of bioavailability of ingredients and lack of drug-drug interactions in a novel five-ingredient polypill (polycap): a five-arm phase I crossover trial in healthy volunteers.

BACKGROUND: The Polycap (polypill; aspirin [acetylsalicylic acid], ramipril, simvastatin, atenolol, and hydrochlorothiazide) was found to be safe and effective for reducing multiple cardiovascular risk factors in The Indian Polycap Study (TIPS). OBJECTIVE: We evaluated the bioavailability of each ingredient of the Polycap and determined any drug-drug interactions relative to single component reference preparations. METHODS: The bioavailability of the ingredients of the Polycap (T; test) when formulated as a single capsule was compared with that of identical capsules with each of its ingredients administered separately (R; reference) in a five-arm, randomized, single-dose, two-period, two-treatment, two-sequence, crossover trial with at least a 2-week washout period in a total of 195 healthy volunteers. Plasma concentrations of each drug and, where applicable, its active metabolite were measured using validated liquid chromatography-tandem mass spectrometry and ultra-performance liquid chromatography. Mean pharmacokinetic parameters and their standard deviations were computed for each analyte. RESULTS: Comparative bioavailability was computed and no drug-drug interactions and no difference in comparative bioavailability were concluded for each ingredient based on point estimates of the T/R ratio of the geometric means falling within 80-125% for peak plasma concentration (C(max)), area under the plasma concentration-time curve from time zero to the last measurable concentration (AUC(t)), and AUC from time zero to infinity (AUC(infinity)). The T/R ratio for C(max), AUC(t) and AUC(infinity) was within 80-125% for atenolol, hydrochlorothiazide, ramipril, ramiprilat and dose-normalized salicylic acid. However, for simvastatin, the T/R point estimates for C(max), AUC(t) and AUC(infinity) for Ln-transformed data were significantly lower ( approximately 3-4%) than the lower bound of 80%. For its active metabolite, simvastatin acid, these estimates were significantly higher ( approximately 25-35%) than the higher bound of 125%. Thus, the increased bioavailability of active simvastatin acid appeared to compensate for the loss of bioavailability of simvastatin. CONCLUSION: The Polycap was found to be effective and safe in the previously published TIPS trial. The present study in healthy volunteers establishes that Polycap is safe (no serious adverse events) and well tolerated, and that there is no indication of pharmacokinetic drug-drug interactions for any of the ingredients, with their bioavailabilities being well preserved.

Therapeutic class-specific signal detection of bradycardia associated with propranolol hydrochloride.

BACKGROUND: Propranolol hydrochloride, one of the most widely used beta-blocker in the treatment of hypertension since 1960s, shows a number of serious and non-serious adverse events. OBJECTIVE: Major objectives of this study were to extract the Canadian Adverse Drug Reaction Monitoring Program (CADRMP) database for possible toxic signal detection (SD) of propranolol hydrochloride, evaluate the frequency of the bradycardia associated with it in different stratified groups for a putative signal, and generate awareness in healthcare professionals regarding usefulness of SD. MATERIALS AND METHODS: Appropriate statistical methods were used for adverse drug reaction (ADR) signal detection such as, proportional reporting ratio (PRR); reporting odds ratio (ROR); the Chi-square (chi(2)) statistic method; the 95% confidence interval (CI); the observed to expected ratio (O/E); and Du Mouchel method were used to calculate the possible signals. Significance of chi(2) and other calculated statistics, e.g., PRR and ROR, was based on a composite criterion of regulatory guidelines and not on any particular statistical level of significance. RESULTS: Calculated statistics by different methods were compared with the regulatory criteria of a statistic value >/=4.0 for chi(2), and >/=3.0 for the rest for SD to be declared significant. The PRR statistic was found to be 2.5054; by the ROR method it was 2.5820; the chi(2) statistic was 3.2598, whereas the lower and upper limits of 95% CI of PRR were found to be 0.0778 and 1.9104, respectively, by the O/E ratio was found to be 2.3978, and PRR with the help of Du Mouchel was found to be 2.3979. Thus, the bradycardia-propranolol signals calculated in this study were not significant. CONCLUSIONS: The therapeutic class specific signal of bradycardia associated with propranolol hydrochloride was not found potent enough to cause bradycardia. However, since the calculated statistics were very high albeit not significant, the possibility of bradycardia-propranolol pairing should still be analyzed from larger databases.

The effect of dyslipidemic drugs on mortality: A meta-analysis.

Most of the previously published meta-analyses include studies exploring the effect of statins, rather than all dyslipidemic drugs, on mortality. We explored the overall effect of all dyslipidemic drugs on coronary artery disease mortality, cardio-vascular disease mortality and all-cause mortality. A meta-analysis of all randomized controlled trials that were published before February 2006 was carried out. Data sources were published articles in bibliographic electronic databases and medical journals. The article selection criteria included all randomized placebo-controlled trials of at least one year duration and those which measured at least one of the following clinical endpoints: coronary artery disease mortality, cardio-vascular mortality or all-cause mortality. Information on sample size, follow up period, drug used, and clinical outcomes was abstracted independently by two authors. Disagreements were resolved by consensus. The meta-analysis (19 trials, 59033 patients) showed a significant relative risk reduction of coronary artery disease mortality of 23% (P<0.00001), cardiovascular disease mortality of 19% (P<0.00001) and all-cause mortality of 14% (P<0.0001), without any significant heterogeneity and inconsistency between the trials. It was concluded from this meta-analyses that dyslipidemic drugs are indeed highly effective medicines and confer benefit to patients, in terms of primary and secondary prevention of coronary artery disease.

Curcumin ameliorates aflatoxin-induced changes in SDH and ATPase activities in liver and kidney of mice.

The present investigation was an attempt to evaluate the ameliorative effect of curcumin on aflatoxin-induced changes in activities of succinate dehydrogenase (SDH) and adenosine triphosphatase (ATPase) in liver and kidney of mice. Aflatoxin was obtained by growing Aspergillus parasiticus in SMKY liquid medium. Pure curcumin (97% purity) was purchased from Hi-Media Laboratories Pvt. Ltd., Mumbai, India. Young adult male albino mice were orally administered with low dose and high dose (750 and 1500 microg/kg body weight) aflatoxin with and without curcumin (2 mg/0.2 mLolive oil/animal/day) for 45 days. On 46th day the animals were sacrificed by cervical dislocation and organs were removed to prepare homogenates for measuring changes in enzyme activities such as succinate dehydrogenase and adenosine triphosphatase. The results showed that in liver and kidney of mice activities of both the enzymes succinate dehydrogenase and adenosine triphosphatase were found to be reduced in the groups treated with low dose and high dose of aflatoxin, which were ameliorated by the treatment of curcumin along with aflatoxin in other groups. Thus, curcumin along with aflatoxin ameliorates aflatoxin-induced changes in succinate dehydrogenase and adenosine triphosphatase activities in liver and kidney of mice.