Pharmacology Journal

Acute seizures are a frequent clinical feature of severe malaria (SM) and other infections in children. Over 80% of children presenting to hospital with cerebral malaria have a history of seizures and about 60% have clinical seizures after admission. Protracted and multiple convulsions are often refractory to treatment and are associated with an increased risk of death and/or neurological and cognitive deficits among survivors. Therefore, early, prompt and effective termination and prevention of convulsions may improve outcome in children with SM. Furthermore, rapid and sustained seizure control may avert the need for multiple anticonvulsant administration and prolonged hospitalization.

The ideal drug for treating acute seizures and status epilepticus (SE) should: (i) enter the brain rapidly; (ii) have an immediate onset of anticonvulsant action; (iii) have minimal depression of consciousness or cardiorespiratory function; (iv) have a sustained duration of anticonvulsant action to prevent seizure recurrence; and (v) be easily and safely administered at peripheral healthcare facilities. Benzodiazepines are considered the drugs of choice for rapid termination of acute seizures and SE. In resource-poor countries, diazepam (DZ) is frequently used as the standard first-line treatment for acute convulsions and SE, as it is widely available, cheap and rapidly acting. However, it has several disadvantages. Gaining intravenous (i.v.) access is often technically difficult in most peripheral healthcare settings (with limited resources and healthcare personnel), particularly in young children who are having generalized tonic-clonic convulsions. Following i.v. administration, DZ is rapidly redistributed to lipoid tissues, and consequently, plasma concentrations rapidly decline with breakthrough seizures occurring when the concentrations fall below threshold levels. Accordingly, repeated injections or continuous infusions are often required for sustained control of seizures. However, administration of multiple doses of DZ is undesirable, especially in children with SM, due to accumulation and increased risk of fatal respiratory depression. Moreover, rectal administration of DZ (which has been suggested as a practical alternative to i.v. administration under such settings) generally results in variable absorption, leading to rapid recurrence of seizures. The intramuscular (i.m.) route, which is frequently used in such settings, is not suitable for administration of DZ for termination of acute seizures and SE, since it results in incomplete and erratic absorption.

Lorazepam (LZP) has several advantages over DZ. Lorazepam is less lipid-soluble than DZ and, following i.v. administration, it has a longer duration of anticonvulsant action (12–24 h) than predicted from its elimination half-life. It prevents seizure recurrence for between 2 and 72 h and can be used for both acute treatment and prophylaxis of seizures. It has potent anticonvulsant activity and is effective in management of SE in both adults and children, including SE refractory to phenobarbital and phenytoin. I.v. and intranasal LZP has been successfully used in acute seizures and SE in the emergency room for children and has been shown to be more effective than DZ for out-of-hospital treatment of SE in adults. LZP may be associated with less respiratory depression than DZ.

To date, no studies have been reported describing both the pharmacokinetics and clinical efficacy of LZP in African children, particularly those with SM. The pharmacokinetics of LZP may be altered in children, and a few concentration-dependent side-effects such as respiratory depression have been reported in infants and children. Children with SM may have compromised respiratory function and hypotension due to associated metabolic acidosis and intracranial hypertension, which may be aggravated by LZP.

We have undertaken a study on LZP administered by either the i.v. or i.m. route to children with SM and convulsions in order to: (i) describe and compare the pharmacokinetic profiles of LZP following administration via both routes; (ii) determine whether the currently recommended dose of LZP (0.1 mg kg^−1) is effective in terminating convulsions in this group; (iii) correlate LZP plasma concentrations with termination and recurrence of convulsions, and respiratory and cardiovascular parameters; and (iv) determine whether the i.m. route would provide a comparable bioavailability and pharmacokinetic profile with respect to the i.v. route.

Study site and study design

This was an open-label, nonrandomized, uncontrolled study. The Kenya Medical Research Institute (KEMRI) Scientific Steering Committee, KEMRI/National Ethical Review Committee and the Research Ethics Committee (Liverpool School of Tropical Medicine, Liverpool, UK) approved the study, which was conducted at the paediatric ward of the New Nyanza Provincial General Hospital (NNPGH), Kisumu (a town with a population of approximately 320 000 people) located on the shores of Lake Victoria in Western Kenya. The NNPGH, the main government tertiary referral hospital (400 beds), serves the entire western Kenya region. The hospital includes a 44-bed paediatric inpatient ward, which admits an average of 8000 children annually, of whom about 40% have falciparum malaria (B. R. Ogutu, personal communication). Government-employed and medically qualified clinical and nursing personnel provide inpatient paediatric medical care to children admitted to the paediatric unit, including the study participants. In addition, a qualified and experienced research nurse (trained in intensive care unit skills and with broad experience in paediatric care in a research setting) was primarily responsible for blood sample collection and close monitoring of the study participants.

Study participants

All children admitted to the NNPGH paediatric unit were eligible for the study if: (i) they were aged between 6 months and 13 years; (ii) they had signs of SM (prostration: unable to sit, drink or breast-feed, but conscious; respiratory distress, or deep (acidotic) breathing); (iii) they had a convulsion lasting ≥5 min; and (iv) the child’s parent(s)/guardian(s) gave written informed consent. Informed consent was obtained from the parents/guardians in a language of their choice, and children were recruited with intent to treat. Children were excluded if: (i) they had received DZ prior to admission to NNPGH (documented in referral case notes, or from the history of the current illness); (ii) informed consent could not be obtained from parents/guardians; and (iii) they had compromised cardiorespiratory function (which, in the opinion of the clinician attending to the child, could be exacerbated by a benzodiazepine). Children exited the study if informed consent was withdrawn or were excluded at the data analysis stage if LZP was detected in the baseline (predose) plasma sample.

Clinical care

All children admitted to the ward were assessed by a medically qualified member of the paediatric team. A clinical history was taken and complete physical examination performed on all children on admission. Venous access was obtained by fixing F.E.P. polymer cannulae (Ven-O-Lit^®; MIGADA, Kiryat Shmona, Israel), one for i.v. fluids and drug administration and another (in the opposite arm) for blood sampling. A single blood sample (4 ml) was drawn for quantitative parasite count, blood culture and full haemogram. A portion of the blood (0.5 ml) was centrifuged (1500 g; 5 min) and the plasma stored at −20°C until assayed for baseline LZP concentrations.

All children received antimalarial treatment with quinine dihydrochloride (Lincoln Pharmaceuticals Ltd, Gandhinagar, India) as a loading dose (15 mg kg^−1 diluted with 10 ml kg^−1 of 5% dextrose saline) infused over 4 h, followed by a maintenance dose (10 mg kg^−1 in 10 ml kg^−1 5% dextrose saline) infused over 4 h every 8 hours, until the child could take oral drugs when antimalarial treatment was completed with first line antimalarial drugs. Other antimalarial drugs including artemisinin derivatives (dihydroartemisin) and amodiaquine were prescribed as required (at the discretion of the clinician attending the child). Severe anaemia (Hb ≤50 g l^−1) was corrected with a blood transfusion (20 ml kg^−1) infused over 4 h. All children were treated with broad-spectrum antibiotics (chloramphenicol, 25 mg kg^−1 6-hourly; and benzyl penicillin) as presumptive treatment for bacteraemia or meningitis until cerebrospinal fluid and blood culture results were available. Children with fever (temperature >38.5°C) were treated with paracetamol (15 mg kg^−1) administered orally, exposure, and tepid sponging. Hypoxia was corrected with 100% oxygen administered by nasal prongs.

Children with convulsions lasting ≥5 min were treated with a single dose (0.1 mg kg^−1) of LZP (Ativan^™; 4 mg LZP BP ml^−1; Wyeth Laboratories, Maidenhead, UK) administered either intravenously as a slow bolus over 1–2 min (to avoid inducing respiratory depression) or as a deep i.m. injection into the anterior aspect of the thigh. If the convulsion did not stop within 10–15 min after LZP administration, i.v. DZ (0.3 mg kg^−1) was administered as a slow bolus over 1–2 min. Convulsions that were refractory to treatment with both LZP and DZ, or those that recurred within the 72-h study period following administration of LZP, were treated with a single dose (15 mg kg^−1) of i.m. phenobarbital (100 mg ml^−1; Lab Renaudin, St Claud, France). The child was closely monitored for any signs of adverse effects, such as respiratory depression.

Other second- or third-line anticonvulsants, including paraldehyde, phenytoin and sodium valproate, were not available as part of the hospital’s routine seizure treatment protocol. Ideally, phenytoin should only be given intravenously together with cardiac monitoring (which is lacking in most health facilities where SM malaria is treated in sub-Saharan Africa, including the present study site).

Blood sampling

Venous blood samples (0.75 ml) for the determination of plasma unconjugated LZP concentrations were collected predose and at 10, 20, 30, 40, 60 min and 2, 4, 6, 8, 12, 24, 36, 48, 60 and 72 h after LZP administration. The cannula was flushed with sterile heparinized normal saline solution (1.0 ml; 20 IU ml^−1). The blood was mixed in lithium-heparinized tubes and centrifuged immediately (1500 g; 5 min) at room temperature. The plasma was separated into polypropylene tubes and stored at −20°C until analysis for unconjugated LZP and paracetamol.

Clinical measurements

All children had respiratory and pulse rates, blood pressure and level of consciousness (Blantyre coma score) recorded at every blood sampling point. In addition, the presence, duration and pattern of convulsions were recorded. Respiratory depression was defined as requiring oxygen by bag and mask, or a poor respiratory effort and reduced respiratory rate following cessation of convulsions (this definition was primarily subjective and did not include routine blood-gas analysis, because facilities for this service were unavailable). Efficacy was assessed using the following measures: (i) the number of children whose convulsions were controlled by a single dose of LZP; (ii) latency (time from initial LZP injection to termination of the seizure); (iii) the use of additional anticonvulsants to control the initial convulsion; (iv) the number of seizures recurring within 72 h after LZP administration; and (v) the duration of seizure control. In a few cases, patients received a second or third anticonvulsant (DZ and/or phenobarbital) after the single dose of LZP. Therefore, the outcome of seizure control was reported descriptively, such as ‘complete control’ or ‘no recurrence’. A successful treatment was defined as one in which the seizure clinically ceased within 15 min after drug administration requiring no further treatment.

Determination of lorazepam and paracetamol concentrations in plasma

The plasma concentrations of unconjugated LZP were measured using sensitive and selective reversed-phase high-performance liquid chromatography with ultraviolet detection. In brief, LZP and the internal standard (oxazepam) were extracted from alkalinized plasma (pH 9.5) into an organic solvent (n-hexane–dichloromethane; 70 : 30%, v/v). After evaporation of the solvent under white spot nitrogen gas, the residue was reconstituted in mobile phase (100 µl) and a 50-µl aliquot was injected. Chromatographic separation was performed on a C18 reversed-phase analytical column (Synergi^™ Max RP; 150 × 4.6 mm i.d.; 4 µm particle size; Phenomenex^®, Macclesfield, UK) using an aqueous mobile phase [10 mm KH₂PO₄ buffer (pH 2.4)–acetonitrile; 65 : 35%, v/v] delivered at a flow rate of 2.5 ml min^−1. The limit of quantification of LZP was 10 ng ml^−1. Calibration curves were linear over the range 10–300 ng (r²Â = 0.99). Lorazepam quality control (QC) samples corresponding to low (LQC), medium (MQC) and high (HQC) concentrations on the calibration curve were 20, 150 and 270 ng ml^−1, respectively. The intra-assay coefficients of variation (CVs) at 20, 150 and 270 ng ml^−1 of LZP were 7.8%, 9.8% (n = 7 in both cases) and 6.6% (n = 8), respectively. The interassay CVs at the above concentrations were 15.9%, 7.7% and 8.4% (n = 7 in all cases), respectively. The method is selective for LZP, with no interference from other commonly coadministered anticonvulsant, antimicrobial, antipyretic or antimalarial drugs.

Both LZP and paracetamol are eliminated via conjugation with glucuronic acid. Some patients administered LZP were also given paracetamol as an antipyretic. Plasma paracetamol concentrations were measured using an Abbott TDx FLx^® fluorescence polarization immunoassay analyser (Abbott Laboratories, Diagnostics Division, Abbott Park, IL, USA). The target (range) concentrations for low (L), medium (M) and high (H) QC samples were 15.0 (12.70–17.30), 35.0 (31.5–38.5) and 150.0 µg ml^−1 (135.0–165.0), respectively. The nominal concentrations for the paracetamol calibrators were 0, 10, 20, 50, 100 and 200 µg ml^−1. The manufacturer supplied the samples for calibration and QC samples. The method is reported to have a sensitivity of 1.0 µg ml^−1.

Filed under: Pharmacodynamics