Formation and reversibility of S-linked conjugates of N-(1-methyl-3,3-diphenylpropyl)isocyanate, an in vivo metabolite of N-(1-methyl-3,3-diphenylpropyl)formamaide, in rats

The metabolic disposition of N-(1-methyl-3,3-diphenylpropyl) formamide was studied in rats. The water-soluble metabolites, N-acetyl-S-[N-(1-methyl-3,3-diphenylpropylcarbamoyl)]cysteine and S-[N-(1-methyl-3,3-diphenylpropylcarbamoyl)]glutathione, were identified in urine and bile, respectively, of rats doses with the secondary formamide. The structures of these metabolites were confirmed by comparison with synthetic standards and by using liquid chromatography mass spectrometry and fast atom bombardment mass spectrometry. Synthetic standards of these metabolites were obtained by reacting the N-(1-methyl-3,3-diphenylpropyl)isocyanate with glutathione or N-acetylcysteine in methanolic solutions. The isocyanate was obtained in high yield by reacting 1-methyl-3,3-diphenylpropylamine with trichloromethyl chloroformate. The S-linked conjugates released the isocyanate in mild alkali, but were stable under acidic conditions. The released isocyanate was characterized by comparison with the synthetic standard using GC/MS and HPLC. A mechanism is proposed for the base-catalyzed elimination of the isocyanate from the thiol conjugates.     

Carbamoylation of peptides and proteins in vitro by S-(N-methylcarbamoyl)glutathione and S-(N-methylcarbamoyl)cysteine, two electrophilic S-linked conjugates of methyl isocyanate

The reactivity toward peptides and proteins of S-(N-methylcarbamoyl)glutathione (SMG), the glutathione conjugate of methyl isocyanate, and the corresponding cysteine adduct, S-(N-methylcarbamoyl)cysteine (SMC), was investigated with the aid of in vitro model systems. Incubation of SMC or a trideuteriomethyl analogue of SMC with either the reduced or oxidized forms of oxytocin afforded similar mixtures of mono-, bis- and tris-N-methylcarbamoylated peptides. Structure elucidation of the mono and bis adducts by fast atom bombardment tandem mass spectrometry indicated that carbamoylation of oxytocin occurred preferentially at Cys-6 and that Cys-1 and/or Tyr-2 were secondary sites of modification. Upon incubation of S-[N-([14C]methyl)carbamoyl]glutathione (14C-SMG) with native bovine serum albumin (BSA), radioactivity became bound covalently to the protein in a time- and concentration-dependent fashion. "Blocking" of the lone Cys-34 thiol group of BSA in the form of a disulfide prior to exposure of the protein to 14C-SMG failed to decrease significantly the extent or time course of this covalent binding. It is concluded that carbamate thioester conjugates of MIC are reactive, carbamoylating entities which can donate the elements of MIC to nucleophilic functionalities on peptides and proteins. Free thiols appear to be preferred sites for such carbamoylation processes, a phenomenon that may have important toxicological consequences in the pathology of tissue lesions induced by MIC and related isocyanates.     

Severe Combined Immunodeficiency (SCID)—the Irish Experience

Journal of Clinical Immunology -     

Outcome of children requiring intensive care following haematopoietic SCT for primary immunodeficiency and other non-malignant disorders

Haematopoietic SCT (HSCT) is curative for many children with primary immunodeficiencies or other non-malignant conditions. Outcome for those admitted to intensive care following HSCT for oncology diagnoses has historically been very poor. There is no literature available specifically regarding the outcome for children with primary immunodeficiency requiring intensive care following HSCT. We reviewed our post-HSCT admission to intensive care over a 5-year period. A total of 111 children underwent HSCT. Median age at transplant was 1 year 4 months. The most common diagnosis was SCID. In all, 35% had at least one intensive care admission and 44% survived to be discharged from intensive care. Also, 73% of admission episodes requiring invasive ventilation but no inotropes or renal replacement therapy resulted in survival to discharge. Children undergoing HSCT for immunological diagnoses had a high rate of admission to intensive care. No factors were identified that could predict the need for admission. Invasive ventilation alone has a much better outcome than that in historical series. However, the need for multi-organ system support was still associated with a poor outcome. This information is useful when counselling families of children that have deteriorated and been admitted to intensive care during the HSCT procedure.     

Supporting parents in managing drugs for children with cystic fibrosis

A semi-structured interview guide was used to interview 17 parents of children with cystic fibrosis identified through a shared care patient database at a London hospital to examine parents' medication-related roles and problems. Parents described their responsibilities in the home and the problems experienced, including adherence to regiments, involvement in decision-making, and relationships with the hospital teams and other healthcare professionals. The findings confirmed medication management as an integral part of the informal caring role. Many practical problems of ensuring the availability and administration of medication on a regular basis were described. Information gaps and problems with medication adherence increased the stress of providing effective care. The involvement of healthcare professionals across primary, secondary and tertiary care sectors in supporting parents in all aspects of medication management, including the development of strategies for transferring the responsibility for medication to their children, must be improved.     

Identification of phenylisoxazolines as novel and viable antibacterial agents active against Gram-positive pathogens

A new and promising group of antibacterial agents, collectively known as the oxazolidinones and exemplified by linezolid (PNU-100766, marketed as Zyvox), have recently emerged as important new therapeutic agents for the treatment of infections caused by Gram-positive bacteria. Because of their significance, extensive synthetic investigations into the structure-activity relationships of the oxazolidinones have been conducted at Pharmacia. One facet of this research effort has focused on the identification of bioisosteric replacements for the usual oxazolidinone A-ring. In this paper we describe studies leading to the identification of antibacterial agents incorporating a novel isoxazoline A-ring surrogate. In a gratifying result, the initial isoxazoline analogue prepared was found to exhibit in vitro antibacterial activity approaching that of the corresponding oxazolidinone progenitor. The synthesis and antibacterial activity profile of a preliminary series of isoxazoline analogues incorporating either a C-C or N-C linkage between their B- and C-rings will be presented. Many of the analogues exhibited interesting levels of antibacterial activity. The piperazine derivative 54 displayed especially promising in vitro activity and in vivo efficacy comparable to the activity and efficacy of linezolid.     

Profiling the hepatic effects of flutamide in rats: a microarray comparison with classical aryl hydrocarbon receptor ligands and atypical CYP1A inducers.

The antiandrogen flutamide (FLU) is used primarily for prostate cancer and is an idiosyncratic hepatotoxicant that sometimes causes severe liver problems. To investigate FLU's overt hepatic effects, especially on inducible drug clearance-related gene networks, FLU's hepatic gene expression profile was examined in female Sprague-Dawley rats using approximately 22,500 oligonucleotide microarrays. Rats were dosed daily for 3 days with FLU at 500, 250, 62.5, 31.3, and 15.6 mg/kg/day, and hepatic RNA was isolated. FLU resulted in the dose-dependent regulation of approximately 350 genes. Employing a gene-response compendium, FLU was compared with three classical aryl hydrocarbon receptor (AhR) ligands, 3-methylcholanthrene, benzo[a]pyrene, and beta-naphthoflavone, and four atypical CYP1A inducers, indole-3-carbinol (I3C), omeprazole (OME), chlorpromazine (CPZ), and clotrimazole (CLO). The FLU gene response was comparable with classical AhR ligands across a signature AhR ligand gene set that included CYP1A1 and other members of the AhR gene battery. Dose-related responses of CYP1 genes established a maximum response ceiling and discerned potency differences in atypical inducers. FLU had a sharp down-regulation of c-fos that was comparable with all the compounds except CPZ and CLO. FLU absorption, distribution, metabolism, and excretion (ADME) gene expression analysis revealed that FLU, as well as I3C and OME, induced CYP2B and CYP3A, distinguishing them from the classical AhR ligands. By using a compendium of gene expression profiles, FLU was shown to signal in rats similar to an AhR activator with additional CYP2B and CYP3A effects that most resembled the ADME gene expression pattern of the atypical CYP1A inducers I3C and OME.     

The United Kingdom Primary Immune Deficiency (UKPID) registry 2012 to 2017

This is the second report of the United Kingdom Primary Immunodeficiency (UKPID) registry. The registry will be a decade old in 2018 and, as of August 2017, had recruited 4758 patients encompassing 97% of immunology centres within the United Kingdom. This represents a doubling of recruitment into the registry since we reported on 2229 patients included in our first report of 2013. Minimum PID prevalence in the United Kingdom is currently 5·90/100 000 and an average incidence of PID between 1980 and 2000 of 7·6 cases per 100 000 UK live births. Data are presented on the frequency of diseases recorded, disease prevalence, diagnostic delay and treatment modality, including haematopoietic stem cell transplantation (HSCT) and gene therapy. The registry provides valuable information to clinicians, researchers, service commissioners and industry alike on PID within the United Kingdom, which may not otherwise be available without the existence of a well‐established registry.