Serum levels of daptomycin in pediatric patients.

BACKGROUND: Limited data are available on the pharmacokinetics and optimal dosage of daptomycin, a lipopeptide compound possessing activity against Gram-positive bacteria, in the pediatric population, particularly in neonates and infants. We determined serum levels of daptomycin in hospitalized pediatric patients treated with various dosages of this agent. METHODS: Blood samples were obtained from pediatric patients of all ages with normal renal function who had received daptomycin between May 2009 and December 2010. Serum levels prior ("trough") and 30 min after end of the infusion ("peak") were determined using an ultra-performance liquid chromatography-UV detection method. RESULTS: A total of four daptomycin dosages and four patients were studied. Three patients were infants (gestational age: 29-38 weeks, age at sampling 26-65 days) and the fourth was a 7-year-old boy. A dosage of 6 mg/kg/12 h of daptomycin to the infants resulted in trough concentrations of <4-8.4 mg/l and peak concentrations of 10.9-17.7 mg/l. Comparable levels were observed after one of the infants received a dosage of 11 mg/kg/12 h, while a further dosage increase to 15 mg/kg/12 h yielded peak concentrations of 35.5 mg/l. The 7-year-old child received a daptomycin dosage of 12 mg/kg once daily; trough and peak levels were 4.2 and 103.4 mg/l, respectively. CONCLUSIONS: A dosage of daptomycin 6 mg/kg/12 h in small infants results in lower peak and similar trough concentrations compared with a dosage of 4 mg/kg/day administered to adults. This results suggests that daptomycin dosages of more than 6 mg/kg/12 h may be needed for this pediatric age group to achieve a similar drug exposure as adults.

Systemic and local interleukin-17 and linked cytokines associated with Sjögren's syndrome immunopathogenesis.

Recently recognized as a distinct CD4(+) T helper (Th) lineage, Th17 cells have been implicated in host responses to infections and in pathogenesis associated with autoimmune diseases. This cytokine is implicated in primary Sjögren's syndrome (pSS) immunopathology because of the increased levels of circulating interleukin (IL)-17 in pSS. Plasma and minor salivary glands (MSGs) from patients with pSS were therefore evaluated for CD4(+) T cells, T regulatory cells, IL-17, and supporting cytokines by immunohistochemistry, RT-PCR, and microbead assays. MSGs from pSS patients contain IL-17-expressing cells as a dominant population within inflammatory lesions. IL-17 protein expression progressively increased with higher biopsy focus scores (P < 0.0001), in parallel with detection by RT-PCR. Transforming growth factor-beta, IL-6 and IL-23, which are requisite promoters of Th17 differentiation, were found in abundance compared with the amounts in control tissues. Although transforming growth factor-beta is also a pivotal differentiation factor for immunosuppressive Foxp3(+) T regulatory cells (Tregs), an increase in Foxp3(+) Tregs was evident in biopsy specimens with mild and moderate inflammation but this increase was disproportionate to escalating pro-inflammatory Th17 populations in advanced disease. Furthermore, the Th17-centric cytokines IL-17, IL-6, IL-23, and IL-12 were significantly elevated in pSS plasma. These data identify a profusion of IL-17-generating cells and supporting cytokines within diseased pSS MSGs without a compensatory increase in immunomodulatory Tregs; this imbalance seems to foster a pathogenic milieu that may be causative and predictive of infiltrative injury and amenable to therapeutic intervention.

T lymphocytes in Sjögren's syndrome: contributors to and regulators of pathophysiology.

Sjögren's syndrome is a chronic autoimmune disorder characterized by lymphocytic infiltration and malfunction of the exocrine glands, resulting in dry mouth and eyes. This multigenic and multifunctional disease can present as primary Sjögren's syndrome or secondary to an underlying connective tissue disease. Immune activation subsequent to activation or apoptosis of glandular epithelial cells in genetically predisposed individuals may expose autoantigens, which engage self-perpetuating T cell dependent autoimmune sequelae. The cellular and molecular context of this immune response may drive proinflammatory (Th1 and Th17) and restrain inhibitory (Treg) pathways. Inability to suppress the immune response results in persistent tissue damage and compromised function of salivary and lacrimal glands. Defining the contributions of participating T cells may unravel strategies for therapeutic intervention.

Antitumor activity of imidazothioxanthones in murine and human tumor models in vitro and in vivo.

BACKGROUND: A new series of imidazothioxanthones has recently been synthesized as potential anticancer agents with the aim of overcoming drug resistance. The route of synthesis and DNA-binding properties of the compounds were reported previously. This paper describes the general structure-activity relationships for the class of imidazothioxanthones in panels of human and murine tumor cell lines in vitro, and the in vivo activity against human and murine solid tumors of the most potent compound, N-[3-(Dimethylamino)propylo]-11-oxo-11H-benzothiopyrano [3',2':2,3]pyrido[1,2-a]imidazo-2-carboxamide (10a). In addition, the interaction between compound 10a and DNA is also considered in terms of molecular mechanics methods and flexible docking techniques. MATERIALS AND METHODS: The cytotoxicity of compounds 10a, 11-oxo-N-[2-(pyrrolidino)ethylo]-11H-benzothiopyrano [3',2':2,3]pyrido[1,2-a]imidazo-2-carboxamide, 11-oxo-N-[2-(piperidino)ethylo]-11H-benzothiopyrano [3',2':2,3]pyrido[1,2-a]imidazo-2-carboxamide and N-[2-(morpholino)ethylo]-11-oxo-11H-benzothiopyrano [3',2':2,3]pyrido[1,2-a]imidazo-2-carboxamide (10c-10e) was assessed in human tumor cell lines and xenografts using the sulforhodamine B assay, MTT assay and the clonogenic assay. The human ovarian xenograft, PXN/109TC, two human breast carcinomas, MT-1 and MCF-7, and the murine colon adenocarcinoma, MAC15A were used for the in vivo testing of compound 10a. In addition, the interaction between compound 10a and DNA is also considered in terms of molecular mechanics methods and flexible docking techniques. RESULTS: Two compounds, 10a and 10c, showed cytotoxic activity below 10 mM in the NCI in vitro screen of 60 human tumor cell lines. The IC50 value of compound 10a was 6.8 mM and that of 10c, 8.3 mM. In addition, both compounds possessed differential activity against leukemia, colon and mammary cancer. The activity pattern was confirmed in two further screens using monolayer and clonogenic, assays. In vivo antitumor studies showed that 10a was active against the human mammary carcinoma MT-1 and murine colon cancer MAC15A. Marginal activity was observed in human ovarian cancer model PXN/109T/C and the compound was inactive in human mammary cancer MCF-7. CONCLUSION: The results warrant further in vivo testing of 10a in additional human solid tumor models. The molecular modeling showed that the planarity of the chromophore and the side-chain conformation could assist the insertion of compound 10a between the base pairs of the double helix. On the other hand, docking to the nucleotide sequence GGAATTGCCTCA suggested that the molecule could also act as a minor groove binder.

Predictors of sustained amenorrhea from pulsed intravenous cyclophosphamide in premenopausal women with systemic lupus erythematosus.

OBJECTIVE: To identify predictors of intravenous cyclophosphamide (IC) induced sustained amenorrhea, especially in young premenopausal women with systemic lupus erythematosus (SLE). METHODS: The cumulative dose resulting in sustained amenorrhea in 50 and 90% of the treated women (D50 and D90) and predictors of sustained amenorrhea at various ages were determined with Kaplan-Meier plots and Cox regressions in a consecutively enrolled cohort of 67 premenopausal women with SLE who received a pulsed IC regimen (monthly doses of 0.75-1.00 g/m2) for nephritis (n = 59) or other indications (n = 8). RESULTS: Twenty-one of 67 women developed sustained amenorrhea of > 12 months' duration. Age was the strongest determinant of this adverse event. For women in the upper age tertile (>or= 32 years old), D50 was 8 g/m2 and D90 was 12 g/m2, and no strong protective or predisposing factors were identified. Conversely, only 5 of 44 women

Composition and antimicrobial activity of the essential oils of five taxa of Sideritis from Greece.

The chemical compositions of the essential oils obtained from the aerial parts of five taxa of Sideritis were analyzed using various GC-MS techniques. A total of 99 different compounds was identified, and significant differences (qualitative and quantitative) were observed between the samples. The in vitro antimicrobial activity of the essential oils against six bacteria and three fungi is also reported.

Kinetic study on the acidic hydrolysis of lorazepam by a zero-crossing first-order derivative UV-spectrophotometric technique.

A zero-crossing first-order derivative UV-spectrophotometric technique for monitoring the main degradation product, 6-chloro-4-(2-chlorophenyl)-2-quinazoline carboxaldehyde, was developed to study the acidic hydrolysis of lorazepam in hydrochloric acid solutions of 0.1 M. Due to the complete overlap of the spectral bands of the parent drug and the hydrolysis product (the range between their spectral maxima was only 3 nm), the graphical methods of derivative spectrophotometry were not efficient. The relative standard deviation of the proposed technique was less than 2.4% and the detection limit was 6.6x10(-8) M. Accelerated studies at higher temperatures have been employed that enable rapid prediction of the long-term stability of this drug. Pseudo-first order reaction kinetics was observed. Kinetic parameters, k(obs) and t(1/2), were calculated, which were similar to those estimated by an HPLC method developed in our laboratory.

Kinetic study on the degradation of prazepam in acidic aqueous solutions by high-performance liquid chromatography and fourth-order derivative ultraviolet spectrophotometry.

A reversed-phase HPLC method was developed for the kinetic investigation of the acidic hydrolysis of prazepam which was carried out in hydrochloric acid solutions of 0.01, 0.1 and 1.0 M. In addition, a fourth-order derivative method for monitoring the parent compound itself was proposed and evaluated. One intermediate was observed by HPLC, which should be formed from breakage of the azomethine linkage. Further slow hydrolysis of the amide bond led to the benzophenone product that was isolated and identified. The mechanism of hydrolysis was biphasic, showing a consecutive reaction with a reversible step. Relative standard deviation was less than 2% for HPLC and less than 5% for the derivative method. Detection limits were 1.2 x 10(-7) M for the former method and 6.7 x 10(-7)M for the latter. Accelerated studies at higher temperatures were employed. Results of HPLC and fourth-order derivative methods were statistically the same.

Acidic hydrolysis of bromazepam studied by high performance liquid chromatography. Isolation and identification of its degradation products.

A kinetic study on the acidic hydrolysis of bromazepam was carried out in 0.01 M hydrochloric acid solution at 25 and 95 degrees C. A reversed-phase HPLC method was developed and validated for the determination of bromazepam and its degradation products. Bromazepam degraded by a consecutive reaction with a reversible first step. Two degradation products were isolated and identified by infrared, 1H and 13C nuclear magnetic resonance and mass spectroscopy. Spectroscopic data indicated that N-(4-bromo-2-(2-pyridylcarbonyl)phenyl)-2-aminoacetamide was the intermediate degradation product of this acid hydrolysis, whereas 2-amino-5-bromophenyl-2-pyridylmethanone was the final one. Therefore, the mechanism of this acid-catalysed hydrolysis involved initial cleavage of the 4,5-azomethine bond, followed by slow breakage of the 1,2-amide bond. Statistical evaluation of the HPLC method revealed its good linearity and reproducibility. Detection limits were 3.8 x 10(-7) M for bromazepam, 6.25 x 10(-7) M for the intermediate and 8.16 x 10(-7) M for the benzophenone derivative.

Properties of the mouse alpha-globin HS-26: relationship to HS-40, the major enhancer of human alpha-globin gene expression.

HS-26, the mouse homologue of HS-40, is the major regulatory element of the mouse alpha-globin gene locus. Like HS-40, HS-26 is located within an intron of a house-keeping gene; comparison of the nucleotide sequences of HS-26 and HS-40 reveals conservation of the sequences and positions of several DNA binding motifs in the 5' regions of both elements (3 GATA, 2 NFE-2, and 1 CACCC sites) and the absence in HS-26 of three CACCC sites and one GATA site that are present in the 3' region of HS-40, suggesting that the two elements might not be identical. We report here that when HS-26 is linked to a 1.5 kb Pstl human alpha-globin gene fragment, it has a weak enhancer activity in induced MEL cells and in transgenic embryos, and it does not have any detectable activity in adult transgenic mice. This suggests that HS-26 does not have Locus Control Region (LCR) activity but can act as an enhancer during the embryonic life when integrated at a permissive locus. To further test the importance of HS-26 at its natural locus, we have generated embryonic stem cells and chimeric animals in which 350 bp containing HS-26 have been replaced by a neomycin resistance gene by homologous recombination. The sizes of the chimeras' red cells were then estimated by measuring forward scattering on a FacsScan apparatus in hypotonic conditions. This revealed that a fraction of the chimeric animals' red cells were smaller than normal mouse red cells and were very similar to cells from mice heterozygous for alpha-thalassemia. Density gradient analysis also suggested the presence of thalassemic cells. These results indicated that despite its lack of LCR activity, HS-26 is important for the regulation of the mouse alpha-globin gene locus.

Radioimmunoassay of pulmonary surface-active material in the tracheal fluid of the fetal lamb.

We measured pulmonary surfactant antigen by radioimmunoassay and phospholipid by lipid phosphorus assay in fluid collected during 12-hour periods from the trachea of lambs in utero between 102 and 147 days of gestation. Both phospholipid and antigen were detectable at 114 days and were present at low concentrations until 137 days. The concentrations then increased rapidly and reached 5- to 10-fold higher concentrations before parturition. The ratio of lipid phosphorus to antigen increased approximately 5-fold as the lungs passed from canalicular to alveolar architecture and approximated the adult ratio only during the alveolar stage of development.

Appearance of paoproteins of pulmonary surfactant in human amniotic fluid.

We have isolated and partially characterized the protein found in surface-active material from adult human lungs, and have determined the time of appearance of this protein in amniotic fluid. Fluids were drawn at gestational ages from 12 to 44 wk, and were assayed for their concentration of apoprotein by an agglutination immunoassay. Surfactant apoprotein ((defined as that protein which is reproducibly found in purified preparations of surface active material) was usually first detected from 30 to 32 wk gestation, and its concentration increased almost fivefold to a maximum at 37 wk. The change in apoprotein concentration was approximately paralleled by the change in phospholipid concentration. At all gestational ages there was wide variability in both phospholipid and apoprotein concentrations, and the time of appearance of the apoprotein in amniotic fluid differed among fetuses. The results suggest that the presence of surfactant apoprotein in amniotic fluid is coincident with the biochemical and morphological maturation of the fetal lung, and are additional evidence that this apoprotein is cosecreted with the lipids of surface-active material.