Eva Klapkova

The importance of therapeutic drug monitoring (TDM) for parenteral busulfan dosing in conditioning regimen for hematopoietic stem cell transplantation (HSCT) in children.

BACKGROUND Series of observations indicate PK/PD variability challenging the accuracy of the body-weight based busulfan (Bu) dosing schedule for (HSCT) conditioning therapy. The purpose of this communication is to describe the frequency of dose changes in initially body-weight-based fixed IV Bu dose and to emphasize the importance of TDM. MATERIAL AND METHODS Sixty-two children (ages 2 months-18 years) were treated with IV busulfan doses based on body weight for myeloablation. TDM utilizing a limited sample strategy (trough concentration immediately before the 5th dose, followed by samples immediately after the end of the 2-h infusion peak, 4 h, and 6 h from initiation of the infusion) was performed in 46 of 62 subjects. Busulfan concentrations were determined by high-performance liquid chromatography (HPLC). AUC was calculated according to the trapezoidal rule. RESULTS We observed trough levels of 25-1244 µg/L, peak levels of 849-4586 µg/L, and AUC of 2225-12818 µg/L·h following body weight-based high-dose busulfan. The doses were changed in 54% of cases. AUC in 5 of 9 patients with VOD were within target, in 3 patients AUS was higher, and in 1 patient AUC was lower. One of the 2 patients with neurotoxicity had higher AUC. Engraftment was 100%, but relapse occurred in 25% of cases. CONCLUSIONS Our results demonstrate that even with IV busulfan, intra-individual PK/PD variability is challenging. Although AUC does not necessarily correspond with outcomes (due to the role of other factors the fact that doses were changed in 54% of cases underlines the importance of TDM.

Re‐evaluation of cord blood arterial and venous reference ranges for pH, pO2, pCO2, according to spontaneous or cesarean delivery

Umbilical cord blood gas analysis (pO2 and pCO2) is now recommended in all high‐risk baby deliveries and in some centers it is performed routinely following all deliveries. The aim of this study was to re‐evaluate cord blood arterial and venous reference ranges for pH, pO2, pCO2 in newborns, delivered by spontaneous vaginal delivery (SVD) and by cesarean section (CS) performed in Faculty Hospital Motol. Two groups of subjects were selected for the study. Group I consisted of 303 newborns with SVD. Group II consisted of 189 newborns delivered by cesarean section. Cord blood samples were analyzed for standard blood gas and pH, using the analytical device Rapid Lab 845 and Rapid Lab 865. We obtained reference values expressed as range (lower and upper reference value expressed as 2.5 and 97.5 percentiles) for cord blood in newborns with SVD: arterial cord blood: pH=7.01–7.39; pCO2=4.12–11.45 kPa; pO2=1.49–5.06 kPa; venous cord blood: pH=7.06–7.44; pCO2=3.33–9.85 kPa; pO2=1.80–6.29 kPa. We also obtained reference values for cord blood in newborns delivered by CS: arterial cord blood: pH=7.05–7.39; pCO2=5.01–10.60 kPa; pO2=1.17–5.94 kPa; venous cord blood: pH=7.10–7.42; pCO2=3.88–9.36 kPa; pO2=1.98–7.23 kPa. Re‐evaluated reference ranges play essential role in monitoring conditions of newborns with spontaneous and caesarean delivery. J. Clin. Lab. Anal. 24:300–304, 2010.

Topotecan vitreous and plasma levels and retinal toxicity after transcorneal intravitreal delivery in healthy albino rabbits: Alternative retinoblastoma treatment

AIM To determine intravitreal and plasma concentrations and retinal toxicity after transcorneal intravitreal injection of 1 μg and 2 μg of topotecan (Hycamtin). METHOD Twelve healthy albino rabbits were included in this in vivo experiment. Six anesthetized albino rabbits received a single transcorneal intravitreal injection of 1 μg (group A) or 2 μg (group B) of topotecan. Vitreous and blood samples were collected until 168 h. Left eyes were treated with the same volume of saline. Plasma and vitreous levels of topotecan were determined by high-performance liquid chromatography. Area under the plasma concentration time curve (AUC) was calculated using trapezoidal rule. Clinical evidence of toxicity was classified into four grades according to anatomical structures. Electroretinograms (ERGs) were recorded. RESULTS Time to maximum concentration was observed up to 2 h after drug injection in group A whereas up to 1 h in group B. Low levels of topotecan were detected in plasma in both groups and in the vitreous humor of the contralateral eye in group B. Topotecan levels (mean vitreous AUC in group A 2.55 μg/mL.h and in group B 5.338 μg/mL.h) were detectable up to 6 h in both groups. We observed following structural changes in rabbit eyes: corneal vascularization, cataract, hemophthalmus, choroidal edema and focal retinal atrophy. Abnormal ERGs were obtained. CONCLUSION Our findings proved that transcorneal intravitreal administration of 1 μg and 2 μg of topotecan results in potentially cytotoxic intraocular concentrations. More studies are needed to establish the safety of topotecan for retinoblastoma in children.

The release kinetics, antimicrobial activity and cytocompatibility of differently prepared collagen/hydroxyapatite/vancomycin layers: Microstructure vs. nanostructure

&NA; The aim of this study was to develop an osteo‐inductive resorbable layer allowing the controlled elution of antibiotics to be used as a bone/implant bioactive interface particularly in the case of prosthetic joint infections, or as a preventative procedure with respect to primary joint replacement at a potentially infected site. An evaluation was performed of the vancomycin release kinetics, antimicrobial efficiency and cytocompatibility of collagen/hydroxyapatite layers containing vancomycin prepared employing different hydroxyapatite concentrations. Collagen layers with various levels of porosity and structure were prepared using three different methods: by means of the lyophilisation and electrospinning of dispersions with 0, 5 and 15 wt% of hydroxyapatite and 10 wt% of vancomycin, and by means of the electrospinning of dispersions with 0, 5 and 15 wt% of hydroxyapatite followed by impregnation with 10 wt% of vancomycin. The maximum concentration of the released active form of vancomycin characterised by means of HPLC was achieved via the vancomycin impregnation of the electrospun layers, whereas the lowest concentration was determined for those layers electrospun directly from a collagen solution containing vancomycin. Agar diffusion testing revealed that the electrospun impregnated layers exhibited the highest level of activity. It was determined that modification using hydroxyapatite exerts no strong effect on vancomycin evolution. All the tested samples exhibited sufficient cytocompatibility with no indication of cytotoxic effects using human osteoblastic cells in direct contact with the layers or in 24‐hour infusions thereof. The results herein suggest that nano‐structured collagen‐hydroxyapatite layers impregnated with vancomycin following cross‐linking provide suitable candidates for use as local drug delivery carriers. Graphical abstract Figure. No caption available.

Determination of vitamins K1, MK-4, and MK-7 in human serum of postmenopausal women by HPLC with fluorescence detection

New high‐performance liquid chromatography (HPLC) method was developed for the determination of vitamin K1 and two forms of vitamin K2 (MK‐4 and MK‐7) in human serum, and the levels of vitamin K were determined in 350 samples of postmenopausal women.

A Comparison of Four Methods (Immunochemistry and HPLC) for Determination of 25-(OH)-Vitamin D in Postmenopausal Women.

BACKGROUND Three immunochemical methods for the determination of 25-(OH)-vitamin D and validated HPLC method for the determination of 25-(OH)-vitamin D3 and 25-(OH)-vitamin D2 were compared. 62 patient samples from postmenopausal women were measured and the results obtained by all these methods were compared. METHODS We used three chemiluminescent assays for determination of 25-(OH)-vitamin D. 25-(OH)-vitamin D3 and 25-(OH)-vitamin D2 were determined by HPLC with UV detection (Agilent 1200). The chemiluminescent assays were performed using the Abbott Architect i4000SR analyzer (Abbott Laboratories, Germany), the ADVIA Centaur (Siemens, USA), and the Liaison XL (DiaSorin Inc, USA). The statistical evaluation was done using GraphPad Prism 6.0. RESULTS The data were tested by Tukeys multiple comparison test. All methods showed significant differences in comparison with the immunochemical method from DiaSorin (p < 0.001 for Abbott, p < 0.05 for Siemens, and p < 0.0001 for HPLC). The comparison of the immunochemical method from Siemens with HPLC was also significant, p < 0.05. The mean of DiaSorin measurements was 38% lower than the mean of HPLC measurements. The non-significant difference was shown by the comparison of Abbott with HPLC and also Abbott with Siemens. Means for the 25-(OH)-vitamin D methods used were: Abbott 70.2 ± 24.2 nmol/L, Siemens 67.6 ± 27.9 nmol/L, DiaSorin 53.5 ± 17.1, and HPLC 82.4 ± 40.0 nmol/L. CONCLUSIONS The comparison of the DiaSorin immunochemical assay with other tested methods showed the greatest deviation. The mean of DiaSorin measurements was 38% lower than the mean of HPLC measurements. According to the results of the DiaSorin method, most patients treated with vitamin D would not achieve the optimal level of 25-(OH)-vitamin D and this could negatively affect the clinical decision.