N-Nitrosodimethylamine investigations in Muta™Mouse define point-of-departure values and demonstrate less-than-additive somatic mutant frequency accumulations.

The N-nitrosamine, N-nitrosodimethylamine (NDMA), is an environmental mutagen and rodent carcinogen. Small levels of NDMA have been identified as an impurity in some commonly used drugs, resulting in several product recalls. In this study, NDMA was evaluated in an OECD TG-488 compliant Muta™Mouse gene mutation assay (28-day oral dosing across seven daily doses of 0.02-4 mg/kg/day) using an integrated design that assessed mutation at the transgenic lacZ locus in various tissues and at the endogenous Pig-a gene-locus, along with micronucleus frequencies in peripheral blood. Liver pathology was determined together with NDMA exposure in blood and liver. The additivity of mutation induction was assessed by including two acute single-dose treatment groups (i.e. 5 and 10 mg/kg dose on Day 1), which represented the same total dose as two of the repeat dose treatment groups. NDMA did not induce statistically significant increases in mean lacZ mutant frequency (MF) in bone marrow, spleen, bladder, or stomach, nor in peripheral blood (Pig-a mutation or micronucleus induction) when tested up to 4 mg/kg/day. There were dose-dependent increases in mean lacZ MF in the liver, lung, and kidney following 28-day repeat dosing or in the liver and kidney after a single dose (10 mg/kg). No observed genotoxic effect levels (NOGEL) were determined for the positive repeat dose-response relationships. Mutagenicity did not exhibit simple additivity in the liver since there was a reduction in MF following NDMA repeat dosing compared with acute dosing for the same total dose. Benchmark dose modelling was used to estimate point of departure doses for NDMA mutagenicity in Muta™Mouse and rank order target organ tissue sensitivity (liver > kidney or lung). The BMD50 value for liver was 0.32 mg/kg/day following repeat dosing (confidence interval 0.21-0.46 mg/kg/day). In addition, liver toxicity was observed at doses of ≥ 1.1 mg/kg/day NDMA and correlated with systemic and target organ exposure. The integration of these results and their implications for risk assessment are discussed.

A comparison of the lowest effective concentration in culture media for detection of chromosomal damage in vitro and in blood or plasma for detection of micronuclei in vivo.

It is often assumed that genotoxic substances will be detected more easily by using in vitro rather than in vivo genotoxicity tests since higher concentrations, more cytotoxicity and static exposures can be achieved. However, there is a paucity of data demonstrating whether genotoxic substances are detected at lower concentrations in cell culture in vitro than can be reached in the blood of animals treated in vivo. To investigate this issue, we compared the lowest concentration required for induction of chromosomal damage in vitro (lowest observed effective concentration, or LOEC) with the concentration of the test substance in blood at the lowest dose required for biologically relevant induction of micronuclei in vivo (lowest observed effective dose, or LOED). In total, 83 substances were found for which the LOED could be identified or estimated, where concentrations in blood and micronucleus data were available via the same route of administration in the same species, and in vitro chromosomal damage data were available. 39.8 % of substances were positive in vivo at blood concentrations that were lower than the LOEC in vitro, 22.9 % were positive at similar concentrations, and 37.3 % of substances were positive in vivo at higher concentrations. Distribution analysis showed a very wide scatter of > 6 orders of magnitude across these 3 categories. When mode of action was evaluated, the distribution of clastogens and aneugens across the 3 categories was very similar. Thus, the ability to detect induction of micronuclei in bone marrow in vivo regardless of the mechanism for micronucleus induction, is clearly not solely determined by the concentration of test substance which induced chromosomal damage in vitro.

mRNA or ChAd0x1 COVID-19 Vaccination of Adolescents Induces Robust Antibody and Cellular Responses With Continued Recognition of Omicron Following mRNA-1273.

Children and adolescents generally experience mild COVID-19. However, those with underlying physical health conditions are at a significantly increased risk of severe disease. Here, we present a comprehensive analysis of antibody and cellular responses in adolescents with severe neuro-disabilities who received COVID-19 vaccination with either ChAdOx1 (n=6) or an mRNA vaccine (mRNA-1273, n=8, BNT162b2, n=1). Strong immune responses were observed after vaccination and antibody levels and neutralisation titres were both higher after two doses. Both measures were also higher after mRNA vaccination and were further enhanced by prior natural infection where one vaccine dose was sufficient to generate peak antibody response. Robust T-cell responses were generated after dual vaccination and were also higher following mRNA vaccination. Early T-cells were characterised by a dominant effector-memory CD4+ T-cell population with a type-1 cytokine signature with additional production of IL-10. Antibody levels were well-maintained for at least 3 months after vaccination and 3 of 4 donors showed measurable neutralisation titres against the Omicron variant. T-cell responses also remained robust, with generation of a central/stem cell memory pool and showed strong reactivity against Omicron spike. These data demonstrate that COVID-19 vaccines display strong immunogenicity in adolescents and that dual vaccination, or single vaccination following prior infection, generate higher immune responses than seen after natural infection and develop activity against Omicron. Initial evidence suggests that mRNA vaccination elicits stronger immune responses than adenoviral delivery, although the latter is also higher than seen in adult populations. COVID-19 vaccines are therefore highly immunogenic in high-risk adolescents and dual vaccination might be able to provide relative protection against the Omicron variant that is currently globally dominant.

Inter-laboratory automation of the in vitro micronucleus assay using imaging flow cytometry and deep learning.

The in vitro micronucleus assay is a globally significant method for DNA damage quantification used for regulatory compound safety testing in addition to inter-individual monitoring of environmental, lifestyle and occupational factors. However, it relies on time-consuming and user-subjective manual scoring. Here we show that imaging flow cytometry and deep learning image classification represents a capable platform for automated, inter-laboratory operation. Images were captured for the cytokinesis-block micronucleus (CBMN) assay across three laboratories using methyl methanesulphonate (1.25-5.0 µg/mL) and/or carbendazim (0.8-1.6 µg/mL) exposures to TK6 cells. Human-scored image sets were assembled and used to train and test the classification abilities of the "DeepFlow" neural network in both intra- and inter-laboratory contexts. Harnessing image diversity across laboratories yielded a network able to score unseen data from an entirely new laboratory without any user configuration. Image classification accuracies of 98%, 95%, 82% and 85% were achieved for 'mononucleates', 'binucleates', 'mononucleates with MN' and 'binucleates with MN', respectively. Successful classifications of 'trinucleates' (90%) and 'tetranucleates' (88%) in addition to 'other or unscorable' phenotypes (96%) were also achieved. Attempts to classify extremely rare, tri- and tetranucleated cells with micronuclei into their own categories were less successful (≤ 57%). Benchmark dose analyses of human or automatically scored micronucleus frequency data yielded quantitation of the same equipotent concentration regardless of scoring method. We conclude that this automated approach offers significant potential to broaden the practical utility of the CBMN method across industry, research and clinical domains. We share our strategy using openly-accessible frameworks.

Permitted daily exposure limits for noteworthy N-nitrosamines.

A genotoxic carcinogen, N-nitrosodimethylamine (NDMA), was detected as a synthesis impurity in some valsartan drugs in 2018, and other N-nitrosamines, such as N-nitrosodiethylamine (NDEA), were later detected in other sartan products. N-nitrosamines are pro-mutagens that can react with DNA following metabolism to produce DNA adducts, such as O6 -alkyl-guanine. The adducts can result in DNA replication miscoding errors leading to GC>AT mutations and increased risk of genomic instability and carcinogenesis. Both NDMA and NDEA are known rodent carcinogens in male and female rats. The DNA repair enzyme, methylguanine DNA-methyltransferase can restore DNA integrity via the removal of alkyl groups from guanine in an error-free fashion and this can result in nonlinear dose responses and a point of departure or "practical threshold" for mutation at low doses of exposure. Following International recommendations (ICHM7; ICHQ3C and ICHQ3D), we calculated permissible daily exposures (PDE) for NDMA and NDEA using published rodent cancer bioassay and in vivo mutagenicity data to determine benchmark dose values and define points of departure and adjusted with appropriate uncertainty factors (UFs). PDEs for NDMA were 6.2 and 0.6 µg/person/day for cancer and mutation, respectively, and for NDEA, 2.2 and 0.04 µg/person/day. Both PDEs are higher than the acceptable daily intake values (96 ng for NDMA and 26.5 ng for NDEA) calculated by regulatory authorities using simple linear extrapolation from carcinogenicity data. These PDE calculations using a bench-mark approach provide a more robust assessment of exposure limits compared with simple linear extrapolations and can better inform risk to patients exposed to the contaminated sartans.

Systemic Inflammation Is Associated With Neurologic Involvement in Pediatric Inflammatory Multisystem Syndrome Associated With SARS-CoV-2.

OBJECTIVE: Pediatric inflammatory multisystem syndrome temporally associated with SARS-CoV-2 (PIMS-TS) is a severe immune-mediated disorder. We aim to report the neurologic features of children with PIMS-TS. METHODS: We identified children presenting to a large children's hospital with PIMS-TS from March to June 2020 and performed a retrospective medical note review, identifying clinical and investigative features alongside short-term outcome of children presenting with neurologic symptoms. RESULTS: Seventy-five patients with PIMS-TS were identified, 9 (12%) had neurologic involvement: altered conciseness (3), behavioral changes (3), focal neurology deficits (2), persistent headaches (2), hallucinations (2), excessive sleepiness (1), and new-onset focal seizures (1). Four patients had cranial images abnormalities. At 3-month follow-up, 1 child had died, 1 had hemiparesis, 3 had behavioral changes, and 4 completely recovered. Systemic inflammatory and prothrombotic markers were higher in patients with neurologic involvement (mean highest CRP 267 vs 202 mg/L, p = 0.05; procalcitonin 30.65 vs 13.11 µg/L, p = 0.04; fibrinogen 7.04 vs 6.17 g/L, p = 0.07; d-dimers 19.68 vs 7.35 mg/L, p = 0.005). Among patients with neurologic involvement, these markers were higher in those without full recovery at 3 months (ferritin 2284 vs 283 µg/L, p = 0.05; d-dimers 30.34 vs 6.37 mg/L, p = 0.04). Patients with and without neurologic involvement shared similar risk factors for PIMS-TS (Black, Asian and Minority Ethnic ethnicity 78% vs 70%, obese/overweight 56% vs 42%). CONCLUSIONS: Broad neurologic features were found in 12% patients with PIMS-TS. By 3-month follow-up, half of these surviving children had recovered fully without neurologic impairment. Significantly higher systemic inflammatory markers were identified in children with neurologic involvement and in those who had not recovered fully.

Peripheral immunophenotypes in children with multisystem inflammatory syndrome associated with SARS-CoV-2 infection.

Recent reports highlight a new clinical syndrome in children related to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)1-multisystem inflammatory syndrome in children (MIS-C)-which comprises multiorgan dysfunction and systemic inflammation2-13. We performed peripheral leukocyte phenotyping in 25 children with MIS-C, in the acute (n = 23; worst illness within 72 h of admission), resolution (n = 14; clinical improvement) and convalescent (n = 10; first outpatient visit) phases of the illness and used samples from seven age-matched healthy controls for comparisons. Among the MIS-C cohort, 17 (68%) children were SARS-CoV-2 seropositive, suggesting previous SARS-CoV-2 infections14,15, and these children had more severe disease. In the acute phase of MIS-C, we observed high levels of interleukin-1ß (IL-1ß), IL-6, IL-8, IL-10, IL-17, interferon-γ and differential T and B cell subset lymphopenia. High CD64 expression on neutrophils and monocytes, and high HLA-DR expression on γδ and CD4+CCR7+ T cells in the acute phase, suggested that these immune cell populations were activated. Antigen-presenting cells had low HLA-DR and CD86 expression, potentially indicative of impaired antigen presentation. These features normalized over the resolution and convalescence phases. Overall, MIS-C presents as an immunopathogenic illness1 and appears distinct from Kawasaki disease.

Probable Vertical Transmission of SARS-CoV-2 Infection.

BACKGROUND: To date, although neonatal infections with severe acute respiratory syndrome coronovirus 2 (SARS-CoV-2) have been described, none of these have been proven to be the result of vertical transmission of SARS-CoV-2. METHODS: We describe the probable vertical transmission of SARS-CoV-2 in a neonate born to a mother with coronavirus disease 2019 (COVID-19). RESULTS: Following cesarean section, the neonate was kept in strict isolation. Molecular tests for SARS-CoV-2 on respiratory samples, blood, and meconium were initially negative, but positive on a nasopharyngeal aspirate on the third day of life. On day 5, the neonate developed fever and coryza, which spontaneously resolved. Viral genomic analysis from the mother and neonate showed identical sequences except for 1 nucleotide. CONCLUSION: This report has important implications for infection control and clinical management of pregnant women with COVID-19 and their newborns.

Clinical Characteristics of 58 Children With a Pediatric Inflammatory Multisystem Syndrome Temporally Associated With SARS-CoV-2.

Importance: In communities with high rates of coronavirus disease 2019, reports have emerged of children with an unusual syndrome of fever and inflammation. Objectives: To describe the clinical and laboratory characteristics of hospitalized children who met criteria for the pediatric inflammatory multisystem syndrome temporally associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (PIMS-TS) and compare these characteristics with other pediatric inflammatory disorders. Design, Setting, and Participants: Case series of 58 children from 8 hospitals in England admitted between March 23 and May 16, 2020, with persistent fever and laboratory evidence of inflammation meeting published definitions for PIMS-TS. The final date of follow-up was May 22, 2020. Clinical and laboratory characteristics were abstracted by medical record review, and were compared with clinical characteristics of patients with Kawasaki disease (KD) (n = 1132), KD shock syndrome (n = 45), and toxic shock syndrome (n = 37) who had been admitted to hospitals in Europe and the US from 2002 to 2019. Exposures: Signs and symptoms and laboratory and imaging findings of children who met definitional criteria for PIMS-TS from the UK, the US, and World Health Organization. Main Outcomes and Measures: Clinical, laboratory, and imaging characteristics of children meeting definitional criteria for PIMS-TS, and comparison with the characteristics of other pediatric inflammatory disorders. Results: Fifty-eight children (median age, 9 years [interquartile range {IQR}, 5.7-14]; 20 girls [34%]) were identified who met the criteria for PIMS-TS. Results from SARS-CoV-2 polymerase chain reaction tests were positive in 15 of 58 patients (26%) and SARS-CoV-2 IgG test results were positive in 40 of 46 (87%). In total, 45 of 58 patients (78%) had evidence of current or prior SARS-CoV-2 infection. All children presented with fever and nonspecific symptoms, including vomiting (26/58 [45%]), abdominal pain (31/58 [53%]), and diarrhea (30/58 [52%]). Rash was present in 30 of 58 (52%), and conjunctival injection in 26 of 58 (45%) cases. Laboratory evaluation was consistent with marked inflammation, for example, C-reactive protein (229 mg/L [IQR, 156-338], assessed in 58 of 58) and ferritin (610 µg/L [IQR, 359-1280], assessed in 53 of 58). Of the 58 children, 29 developed shock (with biochemical evidence of myocardial dysfunction) and required inotropic support and fluid resuscitation (including 23/29 [79%] who received mechanical ventilation); 13 met the American Heart Association definition of KD, and 23 had fever and inflammation without features of shock or KD. Eight patients (14%) developed coronary artery dilatation or aneurysm. Comparison of PIMS-TS with KD and with KD shock syndrome showed differences in clinical and laboratory features, including older age (median age, 9 years [IQR, 5.7-14] vs 2.7 years [IQR, 1.4-4.7] and 3.8 years [IQR, 0.2-18], respectively), and greater elevation of inflammatory markers such as C-reactive protein (median, 229 mg/L [IQR 156-338] vs 67 mg/L [IQR, 40-150 mg/L] and 193 mg/L [IQR, 83-237], respectively). Conclusions and Relevance: In this case series of hospitalized children who met criteria for PIMS-TS, there was a wide spectrum of presenting signs and symptoms and disease severity, ranging from fever and inflammation to myocardial injury, shock, and development of coronary artery aneurysms. The comparison with patients with KD and KD shock syndrome provides insights into this syndrome, and suggests this disorder differs from other pediatric inflammatory entities.

A positron emission tomography imaging study to confirm target engagement in the lungs of patients with idiopathic pulmonary fibrosis following a single dose of a novel inhaled αvß6 integrin inhibitor.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive lung disease with poor prognosis and a significant unmet medical need. This study evaluated the safety, pharmacokinetics (PK) and target engagement in the lungs, of GSK3008348, a novel inhaled alpha-v beta-6 (αvß6) integrin inhibitor, in participants with IPF. METHODS: This was a phase 1b, randomised, double-blind (sponsor unblind) study, conducted in the UK (two clinical sites, one imaging unit) between June 2017 and July 2018 (NCT03069989). Participants with a definite or probable diagnosis of IPF received a single nebulised dose of 1000 mcg GSK3008348 or placebo (ratio 5:2) in two dosing periods. In period 1, safety and PK assessments were performed up to 24 h post-dose; in period 2, after a 7-day to 28-day washout, participants underwent a total of three positron emission tomography (PET) scans: baseline, Day 1 (~ 30 min post-dosing) and Day 2 (~ 24 h post-dosing), using a radiolabelled αvß6-specific ligand, [18F]FB-A20FMDV2. The primary endpoint was whole lung volume of distribution (VT), not corrected for air volume, at ~ 30 min post-dose compared with pre-dose. The study success criterion, determined using Bayesian analysis, was a posterior probability (true % reduction in VT > 0%) of ≥80%. RESULTS: Eight participants with IPF were enrolled and seven completed the study. Adjusted posterior median reduction in uncorrected VT at ~ 30 min after GSK3008348 inhalation was 20% (95% CrI: - 9 to 42%). The posterior probability that the true % reduction in VT > 0% was 93%. GSK3008348 was well tolerated with no reports of serious adverse events or clinically significant abnormalities that were attributable to study treatment. PK was successfully characterised showing rapid absorption followed by a multiphasic elimination. CONCLUSIONS: This study demonstrated engagement of the αvß6 integrin target in the lung following nebulised dosing with GSK3008348 to participants with IPF. To the best of our knowledge this is the first time a target-specific PET radioligand has been used to assess target engagement in the lung, not least for an inhaled drug. TRIAL REGISTRATION: clinicaltrials.gov: NCT03069989; date of registration: 3 March 2017.

In Vivo Mutagenicity Testing of Arylboronic Acids and Esters.

Arylboronic acids and esters (referred to collectively as arylboronic compounds) are commonly used intermediates in the synthesis of pharmaceuticals but pose a challenge for chemical syntheses because they are often positive for bacterial mutagenicity in vitro. As such, arylboronic compounds are then typically controlled to levels that are acceptable for mutagenic impurities, that is, the threshold of toxicological concern (TTC). This study used ICH M7 guidance to design and conduct a testing strategy to investigate the in vivo relevance of the in vitro positive findings of arylboronic compounds. Eight arylboronic compounds representing a variety of chemical scaffolds were tested in Sprague Dawley and/or Wistar rats in the in vivo Pig-a (peripheral blood reticulocytes and mature red blood cells) and/or comet assays (duodenum and/or liver). Five of the eight compounds were also tested in the micronucleus (peripheral blood) assay. The arylboronic compounds tested orally demonstrated high systemic exposure; thus the blood and bone marrow were adequately exposed to test article. One compound was administered intravenously due to formulation stability issues. This investigation showed that arylboronic compounds that were mutagenic in vitro were not found to be mutagenic in the corresponding in vivo assays. Therefore, arylboronic compounds similar to the scaffolds tested in this article may be considered non-mutagenic and managed in accordance with the ICH Q3A/Q3B guidelines. Environ. Mol. Mutagen. 2019. © 2019 Wiley Periodicals, Inc.

Microbial Translocation Does Not Drive Immune Activation in Ugandan Children Infected With HIV.

Objective: Immune activation is associated with morbidity and mortality during human immunodeficiency virus (HIV) infection, despite receipt of antiretroviral therapy (ART). We investigated whether microbial translocation drives immune activation in HIV-infected Ugandan children. Methods: Nineteen markers of immune activation and inflammation were measured over 96 weeks in HIV-infected Ugandan children in the CHAPAS-3 Trial and HIV-uninfected age-matched controls. Microbial translocation was assessed using molecular techniques, including next-generation sequencing. Results: Of 249 children included, 142 were infected with HIV; of these, 120 were ART naive, with a median age of 2.8 years (interquartile range [IQR], 1.7-4.0 years) and a median baseline CD4+ T-cell percentage of 20% (IQR, 14%-24%), and 22 were ART experienced, with a median age of 6.5 years (IQR, 5.9-9.2 years) and a median baseline CD4+ T-cell percentage of 35% (IQR, 31%-39%). The control group comprised 107 children without HIV infection. The median increase in the CD4+ T-cell percentage was 17 percentage points (IQR, 12-22 percentage points) at week 96 among ART-naive children, and the viral load was <100 copies/mL in 76% of ART-naive children and 91% of ART-experienced children. Immune activation decreased with ART use. Children could be divided on the basis of immune activation markers into the following 3 clusters: in cluster 1, the majority of children were HIV uninfected; cluster 2 comprised a mix of HIV-uninfected children and HIV-infected ART-naive or ART-experienced children; and in cluster 3, the majority were ART naive. Immune activation was low in cluster 1, decreased in cluster 3, and persisted in cluster 2. Blood microbial DNA levels were negative or very low across groups, with no difference between clusters except for Enterobacteriaceae organisms (the level was higher in cluster 1; P < .0001). Conclusion: Immune activation decreased with ART use, with marker clustering indicating different activation patterns according to HIV and ART status. Levels of bacterial DNA in blood were low regardless of HIV status, ART status, and immune activation status. Microbial translocation did not drive immune activation in this setting. Clinical Trials Registration: ISRCTN69078957.

Genotoxicity Assessment of Nanomaterials: Recommendations on Best Practices, Assays, and Methods.

Nanomaterials (NMs) present unique challenges in safety evaluation. An international working group, the Genetic Toxicology Technical Committee of the International Life Sciences Institute's Health and Environmental Sciences Institute, has addressed issues related to the genotoxicity assessment of NMs. A critical review of published data has been followed by recommendations on methods alterations and best practices for the standard genotoxicity assays: bacterial reverse mutation (Ames); in vitro mammalian assays for mutations, chromosomal aberrations, micronucleus induction, or DNA strand breaks (comet); and in vivo assays for genetic damage (micronucleus, comet and transgenic mutation assays). The analysis found a great diversity of tests and systems used for in vitro assays; many did not meet criteria for a valid test, and/or did not use validated cells and methods in the Organization for Economic Co-operation and Development Test Guidelines, and so these results could not be interpreted. In vivo assays were less common but better performed. It was not possible to develop conclusions on test system agreement, NM activity, or mechanism of action. However, the limited responses observed for most NMs were consistent with indirect genotoxic effects, rather than direct interaction of NMs with DNA. We propose a revised genotoxicity test battery for NMs that includes in vitro mammalian cell mutagenicity and clastogenicity assessments; in vivo assessments would be added only if warranted by information on specific organ exposure or sequestration of NMs. The bacterial assays are generally uninformative for NMs due to limited particle uptake and possible lack of mechanistic relevance, and are thus omitted in our recommended test battery for NM assessment. Recommendations include NM characterization in the test medium, verification of uptake into target cells, and limited assay-specific methods alterations to avoid interference with uptake or endpoint analysis. These recommendations are summarized in a Roadmap guideline for testing.

Assessment of serum bile acid profiles as biomarkers of liver injury and liver disease in humans.

To assess the potential of individual bile acids (IBA) and their profiles as mechanistic biomarkers of liver injury for humans in real world situations, we interrogated samples collected under minimum controlled conditions (ie subjects were not fasted). Total bile acids (TBA) have been considered to be biomarkers of liver injury for decades, and more recently, monitoring of IBA has been proposed for differentiation of variety of etiologies of liver injury. We established a LC-MS/MS methodology to analyze nine IBA, generated reference ranges, and examined effects of age, gender, and ethnicity for each IBA. Furthermore, we evaluated the ability of IBA and their profiles to detect hepatic injury in subjects with a broad range of liver impairments. To date, our study utilized the largest total cohort of samples (N = 645) that were divided into 2 groups, healthy or liver impaired, to evaluate IBA as biomarkers. The TBA serum levels in the Asian ethnic group trended higher when compared to other ethnic groups, and the serum concentrations of IBA, such as glycocholic acid (GCA), glycochenodeoxycholic acid (GCDCA), chenodeoxycholic acid (CDCA), and taurochenoxycholic acid (TCDCA) were significantly increased. To our knowledge, this report is the first to describe ethnic differences in serum concentrations of IBAs. In patients with hepatic impairments, with the exception of deoxycholic acid (DCA), the concentrations of IBAs were significantly elevated when compared with healthy subjects. The conjugated bile acids displayed greater differences between healthy subjects and subjects with hepatic impairments than non-conjugated bile acids. Furthermore, the subjects with hepatic impairments exhibited distinct profiles (signatures) of IBAs that clustered subjects according the nature of their liver impairments. Although additional studies are needed, our data suggested that the analysis of IBA has the potential to become useful for differentiation of various forms of liver injury.

A Microdose PET Study of the Safety, Immunogenicity, Biodistribution, and Radiation Dosimetry of 18F-FB-A20FMDV2 for Imaging the Integrin αvß6.

The αvß6 integrin is involved in the pathogenesis of cancer and fibrosis. A radiolabeled 20-amino-acid αvß6-binding peptide, derived from the foot and mouth virus (NAVPNLRGDLQVLAQKVART [A20FMDV2]), has been developed to image αvß6 levels preclinically. This study was designed to translate these findings into a clinical PET imaging protocol to measure the expression of αvß6 in humans. Methods: Preclinical toxicology was undertaken, and a direct immunoassay was developed for 4-fluorobenzamide (FB)-A20FMDV2. Four healthy human subjects (2 male and 2 female) received a single microdose of 18F-FB-A20FMDV2 followed by a multibed PET scan of the whole body over more than 3 h. Results: There were no findings in the preclinical toxicology assessments, and no anti-A20FMDV2 antibodies were detected before or after dosing with the PET ligand. The mean and SD of the administered mass of 18F-FB-A20FMDV2 was 8.7 ± 4.4 µg (range, 2.7-13.0 µg). The mean administered activity was 124 ± 20 MBq (range, 98-145 MBq). There were no adverse or clinically detectable pharmacologic effects in any of the subjects. No significant changes in vital signs, laboratory study results, or electrocardiography results were observed. Uptake of radioactivity was observed in the thyroid, salivary glands, liver, stomach wall, spleen, kidneys, ureters, and bladder. Time-activity curves indicated that the highest activity was in the bladder content, followed by the kidneys, small intestine, stomach, liver, spleen, thyroid, and gallbladder. The largest component of the residence times was the voided urine, followed by muscle, bladder, and liver. Using the mean residence time over all subjects as input to OLINDA/EXM, the effective dose was determined to be 0.0217 mSv/MBq; using residence times from single subjects gave an SD of 0.0020 mSv/MBq from the mean. The critical organ was the urinary bladder, with an absorbed dose of 0.18 mGy/MBq. Conclusion:18F-FB-A20FMDV2 successfully passed toxicology criteria, showed no adverse effects in this first-in-humans study, and has an effective dose that enables multiple scans in a single subject.

Presentation of life-threatening invasive nontyphoidal Salmonella disease in Malawian children: A prospective observational study.

Nontyphoidal Salmonellae commonly cause invasive disease in African children that is often fatal. The clinical diagnosis of these infections is hampered by the absence of a clear clinical syndrome. Drug resistance means that empirical antibiotic therapy is often ineffective and currently no vaccine is available. The study objective was to identify risk factors for mortality among children presenting to hospital with invasive Salmonella disease in Africa. We conducted a prospective study enrolling consecutive children with microbiologically-confirmed invasive Salmonella disease admitted to Queen Elizabeth Central Hospital, Blantyre, in 2006. Data on clinical presentation, co-morbidities and outcome were used to identify children at risk of inpatient mortality through logistic-regression modeling. Over one calendar year, 263 consecutive children presented with invasive Salmonella disease. Median age was 16 months (range 0-15 years) and 52/256 children (20%; 95%CI 15-25%) died. Nontyphoidal serovars caused 248/263 (94%) of cases. 211/259 (81%) of isolates were multi-drug resistant. 251/263 children presented with bacteremia, 6 with meningitis and 6 with both. Respiratory symptoms were present in 184/240 (77%; 95%CI 71-82%), 123/240 (51%; 95%CI 45-58%) had gastrointestinal symptoms and 101/240 (42%; 95%CI 36-49%) had an overlapping clinical syndrome. Presentation at <7 months (OR 10.0; 95%CI 2.8-35.1), dyspnea (OR 4.2; 95%CI 1.5-12.0) and HIV infection (OR 3.3; 95%CI 1.1-10.2) were independent risk factors for inpatient mortality. Invasive Salmonella disease in Malawi is characterized by high mortality and prevalence of multi-drug resistant isolates, along with non-specific presentation. Young infants, children with dyspnea and HIV-infected children bear a disproportionate burden of the Salmonella-associated mortality in Malawi. Strategies to improve prevention, diagnosis and management of invasive Salmonella disease should be targeted at these children.