Gudrun Würthwein

Monitoring of asparaginase activity and asparagine levels in children on different asparaginase preparations

The antileukaemic enzyme L-asparaginase is used to achieve the greatest possible reduction in blood levels of the amino acid asparagine, an essential factor for the growth of leukaemic blasts. There are two main sources of the enzyme, E. coli and Erwinia. Faced with increasing reports of treatment complications, we established a programme to monitor enzyme activity and asparagine levels in serum, in children receiving treatment for acute lymphoblastic leukaemia (ALL) and non-Hodgkins lymphoma (NHL). Trough asparagine and asparaginase levels were measured in 49 children on induction treatment with different E. coli preparations (Asparaginase medac, Crasnitin) and in 52 children on re-induction (Asparaginase medac, Crasnitin, and, in the event of allergic reactions, Erwinase) just prior to each sequential application of 10000 U/m2 of asparaginase. Measurements were made by an enzyme assay and an HPLC method. During induction, both Escherichia coli preparations induced the desired reduction in asparagine, but the asparaginase activity with Asparaginase medac was significantly higher than with Crasnitin (median of trough levels 475 versus 74 U/l). Under re-induction treatment (median, Asparaginase medac 528 U/l, Crasnitin 49 U/l, and Erwinase < 20 U/l) complete asparagine depletion was recorded on day 3 in more than 90% of Asparaginase medac samples, more than 60% of Crasnitin samples and in 26% of Erwinase samples. The latter two groups included some children with unchanged asparagine levels and no measurable enzyme activity. Different asparaginase preparations are not readily interchangeable. When Asparaginase medac is used instead of Crasnitin, and identical dose will be associated with significantly higher enzyme activity, well above the level required for complete asparagine depletion. Clinical studies will need to specify both the preparation and the dose to be used. When substitution of an alternative drug is mandatory owing to allergic reactions, monitoring is advisable.

Population Pharmacokinetics of Amphotericin B Lipid Complex in Neonates

ABSTRACT The pharmacokinetics of amphotericin B lipid complex (ABLC) were investigated in neonates with invasive candidiasis enrolled in a phase II multicenter trial. Sparse blood (153 samples; 1 to 9 per patient, 1 to 254 h after the dose) and random urine and cerebrospinal fluid (CSF) samples of 28 neonates (median weight [WT], 1.06 kg; range, 0.48 to 4.9 kg; median gestational age, 27 weeks; range, 24 to 41 weeks) were analyzed. Patients received intravenous ABLC at 2.5 (n = 15) or 5 (n = 13) mg/kg of body weight once a day over 1 or 2 h, respectively, for a median of 21 days (range, 4 to 47 days). Concentrations of amphotericin B were quantified as total drug by high-performance liquid chromatography. Blood data for time after dose (TAD) of <24 h fitted best to a one-compartment model with an additive-error model for residual variability, WT0.75 (where 0.75 is an exponent) as a covariate of clearance (CL), and WT as a covariate of volume of distribution (V). Prior amphotericin B, postnatal age, and gestational age did not further improve the model. The final model equations were CL (liters/h) = 0.399 × WT0.75 (interindividual variability, 35%) and V (liters) = 10.5 × WT (interindividual variability, 43%). Noncompartmental analysis of pooled data with a TAD of >24 h revealed a terminal half-life of 395 h. Mean concentrations in the urine after 1, 2, and 3 weeks ranged from 0.082 to 0.430 μg/ml, and those in CSF ranged from undetectable to 0.074 μg/ml. The disposition of ABLC in neonates was similar to that observed in other age groups: weight was the only factor that influenced clearance. Based on these results and previously published safety and efficacy data, we recommend a daily dosage between 2.5 and 5.0 mg/kg for treatment of invasive Candida infections in neonates.

Phase II Dose Escalation Study of Caspofungin for Invasive Aspergillosis

ABSTRACT Our objective was to evaluate the maximum tolerated dose of caspofungin for invasive aspergillosis (IA). The safety and pharmacokinetics of escalating dosages of caspofungin were investigated in IA. Eight patients each received caspofungin 70, 100, 150, or 200 mg once a day (QD). Dose-limiting toxicity (DLT) was defined as the same non-hematological treatment-related adverse event of grade ≥4 in 2 of 8 patients or ≥3 in 4 of 8 patients in a cohort. A total of 46 patients (median age, 61 years; 21 female; 89% with hematological malignancies) received caspofungin (9, 8, 9, and 20 patients in the 70-, 100-, 150-, and 200-mg cohorts) for a median of 24.5 days. Plasma pharmacokinetics were linear across the investigated dosages and followed a two-compartment model, with weight as the covariate on clearance and sex as the covariate on central volume of distribution. Simulated peak plasma concentrations at steady state ranged from 14.2 to 40.6 mg/liter (28%), trough concentrations from 4.1 to 11.8 mg/liter (58%), and area under the concentration-time curve from 175 to 500 mg/liter/h (32%) (geometric mean, geometric coefficient of variation). Treatment was well tolerated without dose-limiting toxicity. The rate of complete or partial responses was 54.3%, and the overall mortality at 12-week follow-up was 28.3%. In first-line treatment of invasive aspergillosis, daily doses of up to 200 mg caspofungin were well tolerated and the maximum tolerated dose was not reached. Pharmacokinetics was linear. Response rates were similar to those previously reported for voriconazole and liposomal amphotericin.

Drug monitoring of low-dose PEG-asparaginase (Oncaspartm) in children with relapsed acute lymphoblastic leukaemia

Use of asparaginase (ASNase) in the treatment of relapsed childhood acute lymphoblastic leukaemia (ALL) is associated with a high rate of hypersensitive reactions. ‘Silent’ inactivation may additionally reduce treatment intensity. Therefore, PEG‐ASNase (Oncaspartm), a polyethylene glycol conjugate of the native Escherichia coli‐ASNase, was introduced into the Berlin‐Frankfurt‐Münster (BFM) 96 treatment protocol for relaped ALL under drug monitoring conditions. A single i.v. dose of 500 IU/m2 PEG‐ASNase, substituted for the native ASNases, was administered to supply a plasma activity of 100 IU/l for 1 week. From November 1997 to March 2000, 35 patients from 23 BFM‐associated hospitals, with or without a previous allergic reaction to one or both native preparations, underwent monitoring. After 82 applications, a total of 270 samples were submitted to be tested for ASNase activity. The ASNase activity on the day of the administration and the following day ranged between < 20 and 693 IU/l, with a median of 413 IU/l (53 samples). The median on d 7 ± 1 was 199 IU/l (range <20–421 IU/l; 41 samples) and on d 14 ± 1, 105 IU/l (range <20–188 IU/l; 19 samples). An ASNase activity of > 100 IU/l was seen on d 7 in 36 activity time courses of 52 interpretable applications (69%). Intraindividual variability of activity time courses was low. However, a rapid decrease in ASNase activity after repeated applications was observed in 4 out of 20 children. Previously experienced allergic reactions to native ASNases did not influence PEG‐ASNase pharmacokinetics. PEG‐ASNase is a useful alternative to the native ASNases in children with relapsed ALL. Whenever possible, drug monitoring should be performed to identify patients with ‘silent’ inactivation.

Serum asparaginase activities and asparagine concentrations in the cerebrospinal fluid after a single infusion of 2,500 IU/m2 peg asparaginase in children with ALL treated according to protocol COALL-06-97

Pharmacological surrogate parameters are considered a useful tool in estimating the treatment intensity of asparaginase (ASNase) preparations. When a pegylated ASNase (single infusion of 2,500 IU/m2 polyethylene glycol (PEG)‐ASNase, Oncaspar™) was introduced into the treatment protocols of the German Cooperative Acute Lymphoblastic Leukaemia (COALL) study group, this was accompanied by a drug monitoring programme measuring serum ASNase activity and asparagine (ASN) concentrations in the cerebrospinal fluid (CSF) in 70 children.

Asparagine synthetase activity in paediatric acute leukaemias: AML‐M5 subtype shows lowest activity

Lack of sufficient cellular activity of asparagine synthetase (AS) in blast cells compared with normal tissue is thought to be the basis of the antileukaemic effect of l‐asparaginase in acute lymphoblastic leukaemia (ALL). Although l‐asparaginase is routinely used in ALL, its role and value in the treatment of acute myelogenous leukaemia (AML) is still being discussed. To evaluate the pharmacological basis for l‐asparaginase treatment, we established pretreatment monitoring of the intracellular AS activity in blast cells of patients with AML and ALL. There was no general difference in AS activity between ALL and AML samples. Significantly lower AS activity, however, was found in the B‐lineage ALL subgroups as well as AML‐M5.

Monitoring of voriconazole plasma concentrations in immunocompromised paediatric patients

OBJECTIVES Voriconazole is approved for management of invasive fungal diseases (IFDs) in paediatric patients. We analysed plasma trough concentrations and explored their association with endpoints of antifungal therapy. PATIENTS AND METHODS The cohort included 74 immunocompromised patients (0.2-18 years of age) who received 101 courses of voriconazole for possible (7) and probable/proven (13) IFDs, as prophylaxis (79) or empirical therapy (2). Voriconazole was given intravenously (4), intravenously and orally (15) and orally (82) at recommended dosages until intolerance or maximum efficacy. IFDs and outcomes were assessed by EORTC/MSG consensus criteria. RESULTS Voriconazole was administered at a median maintenance dosage of 4.8 mg/kg twice daily (range 2.2-17.4) for a median of 40 days (range 6-1002). Trough plasma concentrations at steady state (251 samples; 3.4 ± 4.3/patient) ranged from <0.2 to 14.9 mg/L with high intra- and inter-individual variability and no apparent relationship to dose (P = 0.074, ANOVA). Of the samples 22%, 42% and 58% had voriconazole concentrations <0.2, ≤0.5 and ≤1.0 mg/L, respectively. Adverse events (AEs) occurred in 77/101 (76.2%) courses and were mostly grade I or II. Ten courses (9.9%) were discontinued due to AEs. Treatment success was observed in 8/20 patients (40%) with IFDs, and in 67/81 courses (82.7%) of empirical therapy/prophylaxis. There were no consistent correlations between dose, trough concentrations and laboratory/clinical AEs or treatment response, and proposed threshold values were not discriminative. CONCLUSIONS Voriconazole had acceptable safety and useful efficacy in the management of paediatric IFDs. Pharmacokinetic variability was high and no predictable dose-concentration-effect relationships were observed.

Toxicity and pharmacokinetics of i.v. busulfan in children before stem cell transplantation.

We have introduced a dimethylacetamide-based i.v. formulation of busulfan into pediatrics with a dose intensity [as measured by area under curve (AUC)] comparable to that achieved by oral busulfan while reducing variability. The target AUC was defined at 1600±600 μM·min. The children received 15 doses of i.v. busulfan as 2-h infusions with a dose calculated to be 80% of the oral dose according to the malignancy-related protocol. The first infusion was applied as a double dose over 4 h with the second infusion following 12 h thereafter. Plasma samples were analyzed for busulfan by a validated LC-MS method and toxicity was assessed at least up to day 100+ after transplantation. Nineteen children (median age: 4 years, range: 0.9–17.3) were included. The AUC after the first dose ranged from 570 to 1410 μM·min [geometric mean 1010 μM·min, coefficient of variation (CV)=22%, n=17]. In nine out of 17 patients, the AUC after the first dose was out of the target range. Two patients had neurotoxic symptoms, which were attributable to busulfan in one individual. No case of severe hepatic veno-occlusive disease or other serious toxic events occurred. We conclude that i.v. busulfan displays a smaller interpatient variability in exposure compared to oral busulfan (CV of 24% after i.v. versus CV of 37% after oral busulfan). The equivalent dose to 1 mg/kg oral busulfan with regard to the AUC appears to be higher than 0.8 mg/kg.

Population Pharmacokinetics of Liposomal Amphotericin B and Caspofungin in Allogeneic Hematopoietic Stem Cell Recipients

ABSTRACT Liposomal amphotericin B (LAMB) and caspofungin (CAS) are important antifungal agents in allogeneic hematopoietic stem cell transplant (aHSCT) recipients. Little is known, however, about the pharmacokinetics (PK) of both agents and their combination in this population. The PK of LAMB and CAS and the potential for PK interactions between both agents were investigated within a risk-stratified, randomized phase II clinical trial in 53 adult aHSCT recipients with granulocytopenia and refractory fever. Patients received either LAMB (n = 17; 3 mg/kg once a day [QD]), CAS (n = 19; 50 mg QD; day 1, 70 mg), or the combination of both (CAS-LAMB; n = 17) for a median duration of 10 to 13 days (range, 4 to 28 days) until defervescence and granulocyte recovery. PK sampling was performed on days 1 and 4. Drug concentrations in plasma (LAMB, 405 samples; CAS, 458 samples) were quantified by high-pressure liquid chromatography and were analyzed using population pharmacokinetic modeling. CAS concentration data best fitted a two-compartment model with a proportional error model and interindividual variability (IIV) for clearance (CL) and central volume of distribution (V1) (CL, 0.462 liter/h ± 25%; V1, 8.33 liters ± 29%; intercompartmental clearance [Q], 1.25 liters/h; peripheral volume of distribution [V2], 3.59 liters). Concentration data for LAMB best fitted a two-compartment model with a proportional error model and IIV for all parameters (CL, 1.22 liters/h ± 64%; V1, 19.2 liters ± 38%; Q, 2.18 liters/h ± 47%; V2, 52.8 liters ± 84%). Internal model validation showed predictability and robustness of both models. None of the covariates tested (LAMB or CAS comedication, gender, body weight, age, body surface area, serum bilirubin, and creatinine clearance) further improved the models. In summary, the disposition of LAMB and CAS was best described by two-compartment models. Drug exposures in aHSCT patients were comparable to those in other populations, and no PK interactions were observed between the two compounds.

Population Pharmacokinetics of Escalating Doses of Caspofungin in a Phase II Study of Patients with Invasive Aspergillosis

ABSTRACT Caspofungin (CAS) is approved for second-line management of proven or probable invasive aspergillosis at a dose of 50 mg once daily (QD). Preclinical and limited clinical data support the concept of the dose-dependent antifungal efficacy of CAS with preservation of its favorable safety profile. Little is known, however, about the pharmacokinetics (PKs) of higher doses of CAS in patients. In a formal multicenter phase II dose-escalation study, CAS was administered as a 2-h infusion at doses ranging from 70 to 200 mg QD. CAS PK sampling (n = 468 samples) was performed on day 1 and at peak and trough time points on days 4, 7, 14, and 28 (70 mg, n = 9 patients; 100 mg, n = 8 patients; 150 mg, n = 9 patients; 200 mg, n = 20 patients; total, n = 46 patients). Drug concentrations in plasma were measured by liquid chromatography tandem mass spectroscopy. Population pharmacokinetic analysis (PopPK) was performed using NONMEM (version 7) software. Model evaluation was performed using bootstrap analysis, prediction-corrected visual predictive check (pcVPC), as well as standardized visual predictive check (SVPC). The four investigated dose levels showed no difference in log-transformed dose-normalized trough levels of CAS (analysis of variance). CAS concentration data fitted best to a two-compartment model with a proportional-error model, interindividual variability (IIV) fitted best on clearance (CL), central and peripheral volume of distribution (V1 and V2, respectively) covariance fitted best on CL and V1, interoccasion variability (IOV) fitted best on CL, and body weight fitted best as a covariate on CL and V1 (CL, 0.411 liters/h ± 29% IIV; IOV on CL, 16%; V1, 5.785 liters ± 29% IIV; intercompartmental clearance, 0.843 liters/h; V2, 6.53 liters ± 67% IIV). None of the other examined covariates (dose level, gender, age, serum bilirubin concentration, creatinine clearance) improved the model further. Bootstrap results showed the robustness of the final PopPK model. pcVPC and SVPC showed the predictability of the model and further confirmed the linear PKs of CAS over the dosage range of 70 to 200 mg QD. On the basis of the final model, geometric mean simulated peak plasma levels at steady state ranged from 13.8 to 39.4 mg/liter (geometric coefficient of variation, 31%), geometric mean trough levels ranged from 4.2 to 12.0 mg/liter (49%), and geometric mean areas under the concentration-time curves ranged from 170 to 487 mg · h/liter (34%) for the dosage range of 70 to 200 mg QD. CAS showed linear PKs across the investigated dosage range of 70 to 200 mg QD. Drug exposure in the present study population was comparable to that in other populations. (This study has been registered with the European Union Drug Regulating Authorities Clinical Trials website under registration no. 2006-001936-30 and at ClinicalTrials.gov under registration no. NCT00404092.)