The effect of antibiotics on the gut microbiome: a metagenomics analysis of microbial shift and gut antibiotic resistance in antibiotic treated mice.

BACKGROUND: Emergence of antibiotic resistance is a global public health concern. The relationships between antibiotic use, the gut community composition, normal physiology and metabolism, and individual and public health are still being defined. Shifts in composition of bacteria, antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) after antibiotic treatment are not well-understood. METHODS: This project used next-generation sequencing, custom-built metagenomics pipeline and differential abundance analysis to study the effect of antibiotic monotherapy on resistome and taxonomic composition in the gut of Balb/c mice infected with E. coli via transurethral catheterization to investigate the evolution and emergence of antibiotic resistance. RESULTS: There is a longitudinal decrease of gut microbiota diversity after antibiotic treatment. Various ARGs are enriched within the gut microbiota despite an overall reduction of the diversity and total amount of bacteria after antibiotic treatment. Sometimes treatment with a specific class of antibiotics selected for ARGs that resist antibiotics of a completely different class (e.g. treatment of ciprofloxacin or fosfomycin selected for cepA that resists ampicillin). Relative abundance of some MGEs increased substantially after antibiotic treatment (e.g. transposases in the ciprofloxacin group). CONCLUSIONS: Antibiotic treatment caused a remarkable reduction in diversity of gut bacterial microbiota but enrichment of certain types of ARGs and MGEs. These results demonstrate an emergence of cross-resistance as well as a profound change in the gut resistome following oral treatment of antibiotics.

Scientific and regulatory reasons for delay and denial of FDA approval of initial applications for new drugs, 2000-2012.

IMPORTANCE: Some new drug applications fail because of inadequate drug performance and others are not approved because the information submitted to the US Food and Drug Administration (FDA) is unsatisfactory to make that determination. Resubmission of failed applications is costly, delaying marketing approval and the availability of new drugs to patients. OBJECTIVE: To identify the reasons that FDA marketing approval for new drugs was delayed or denied. DESIGN, SETTING, AND PARTICIPANTS: A retrospective review of FDA documents and extraction of data were performed. We examined all drug applications first submitted to the FDA between 2000 and 2012 for new molecular entities (NMEs), which are active ingredients never before marketed in the United States in any form. Using FDA correspondence and reviews, we investigated the reasons NMEs failed to obtain FDA approval. MAIN OUTCOMES AND MEASURES: Reasons for delayed FDA approval or nonapproval of NME applications. RESULTS: Of the 302 identified NME applications, 151 (50%) were approved when first submitted and 222 (73.5%) were ultimately approved. Seventy-one applications required 1 or more resubmissions before approval, with a median delay to approval of 435 days following the first unsuccessful submission. Of the unsuccessful first-time applications, 24 (15.9%) included uncertainties related to dose selection, 20 (13.2%) choice of study end points that failed to adequately reflect a clinically meaningful effect, 20 (13.2%) inconsistent results when different end points were tested, 17 (11.3%) inconsistent results when different trials or study sites were compared, and 20 (13.2%) poor efficacy when compared with the standard of care. The frequency of safety deficiencies was similar among never-approved drugs compared with those with delayed approval (43 of 80 never approved [53.8%] vs 37 of 71 eventually approved [52.1%]; difference, 1.7% [95% CI, -14.86% to 18.05%]; P = .87). However, efficacy deficiencies were significantly more frequent among the never-approved drugs than among those with delayed approvals (61 of 80 never approved [76.3%] vs 28 of 71 eventually approved [39.4%]; difference, 36.9% [95% CI, 20.25% to 50.86%]; P < .001). CONCLUSIONS AND RELEVANCE: Several potentially preventable deficiencies, including failure to select optimal drug doses and suitable study end points, accounted for significant delays in the approval of new drugs. Understanding the reasons for previous failures is helpful to improve the efficiency of clinical development for new drugs.

Decentralized Clinical Trials in the Development of Drugs and Biological Products.

Decentralized clinical trials (DCTs) are trials where some or all of the trial-related activities occur at locations other than traditional clinical trial sites. FDA supports decentralization to improve participation in clinical trials. While there are benefits of DCTs, including convenience for participants, sponsors and investigators should be aware of potential challenges such as coordination of trial activities at locations other than traditional trial sites and supervision of delegated trial-activities performed remotely. Appropriate training, oversight, and up-front risk assessment and management will be key to implementing a DCT successfully.

A metagenomic analysis for combination therapy of multiple classes of antibiotics on the prevention of the spread of antibiotic-resistant genes.

Antibiotics used systemically to treat infections may have off-target effects on the gut microbiome, potentially resulting in the emergence of drug-resistant bacteria or selection of pathogenic species. These organisms may present a risk to the host and spread to the environment with a risk of transmission in the community. To investigate the risk of emergent antibiotic resistance in the gut microbiome following systemic treatment with antibiotics, this metagenomic analysis project used next-generation sequencing, a custom-built metagenomics pipeline, and differential abundance analysis to study the effect of antibiotics (ampicillin, ciprofloxacin, and fosfomycin) in monotherapy and different combinations at high and low doses, to determine the effect on resistome and taxonomic composition in the gut of Balb/c mice. The results showed that low-dose monotherapy treatments showed little change in microbiome composition but did show an increase in expression of many antibiotic-resistant genes (ARGs) posttreatment. Dual combination treatments allowed the emergence of some conditionally pathogenic bacteria and some increase in the abundance of ARGs despite a general decrease in microbiota diversity. Triple combination treatment was the most successful in inhibiting emergence of relevant opportunistic pathogens and completely suppressed all ARGs after 72 h of treatment. The relative abundances of mobile genetic elements that can enhance transmission of antibiotic resistance either decreased or remained the same for combination therapy while increasing for low-dose monotherapy. Combination therapy prevented the emergence of ARGs and decreased bacterial diversity, while low-dose monotherapy treatment increased ARGs and did not greatly change bacterial diversity.

Digital Health Technologies in Pediatric Trials.

BACKGROUND: Advances in the miniaturization of sensors and other technologies provide opportunities to collect physiological and/or functional data directly from patients participating in clinical trials. The use of such technologies in children is particularly promising. Objective, quantifiable measurements made by these technologies, often on a continuous or frequent basis, may provide more robust data than the episodic reports from caregivers that are used in traditional pediatric trials. METHODS: We reviewed the pros and cons of these technologies for use in a variety of pediatric diseases, including seizure and neuromuscular disorders, cardiorespiratory diseases, and metabolic disorders. RESULTS: Correlation between sensor measurements and patient observations or traditional clinical measurements varied depending on the disease being evaluated. There was a notable dearth of reports on the use of digital health technology in pediatric patients. Given the range of sensors and measurements that can be made by DHTs, selection of the design, metrics and types of sensors best suited to disease evaluation presents challenges for adoption of these technologies in clinical trials. CONCLUSION: Traditional measurements of drug effects are often deficient, particularly in the evaluation of infants and young children. The opportunity to make objective, frequent measurements may increase our power to detect and quantify responses to therapy in these populations. Further research and evaluation are needed to realize the full scientific potential of remote monitoring in pediatric clinical trials.

Evaluation of a Sequential Antibiotic Treatment Regimen of Ampicillin, Ciprofloxacin and Fosfomycin against Escherichia coli CFT073 in the Hollow Fiber Infection Model Compared with Simultaneous Combination Treatment.

OBJECTIVE: Employ the hollow fiber infection model (HFIM) to study sequential antibiotic administration (ampicillin, ciprofloxacin and fosfomycin) using human pharmacokinetic profiles to measure changes in the rate of antibiotic resistance development and compare this to simultaneous combination therapy with the same antibiotic combinations. METHODS: Escherichia coli CFT073, a clinical uropathogenic strain, was exposed individually to clinically relevant pharmacokinetic concentrations of ampicillin on day 1, ciprofloxacin on day 2 and fosfomycin on day 3. This sequence was continued for 10 days in the HFIM. Bacterial samples were collected at different time points to enumerate total and resistant bacterial populations. The results were compared with the simultaneous combination therapy previously studied. RESULTS: Sequential antibiotic treatment (ampicillin-ciprofloxacin-fosfomycin sequence) resulted in the early emergence of single and multi-antibiotic-resistant subpopulations, while the simultaneous treatment regimen significantly delayed or prevented the emergence of resistant subpopulations. CONCLUSION: Sequential administration of these antibiotic monotherapies did not significantly delay the emergence of resistant subpopulations compared to simultaneous treatment with combinations of the same antibiotics. Further studies are warranted to evaluate different sequences of the same antibiotics in delaying emergent resistance.

Digital Health Technology to Measure Drug Efficacy in Clinical Trials for Parkinson's Disease: A Regulatory Perspective.

Digital health technology (DHT), including wearable and environmental sensors, video cameras and other electronic tools, has provided new opportunities for the measurement of movement and functionality in Parkinson's disease. Compared to current standards for evaluation of the disease (MDS-UPDRS), DHT may offer new possibilities for more frequent objective measurements of the duration, severity and frequency of disease manifestations over time, that may provide more information than periodic clinic visits. However, DHT measurements are only scientifically and medically useful if they are accurate, reliable and clinically meaningful. Verification and validation, also known as analytical validation and clinical validation, of DHT performance is important to ensure the accuracy and precision of measurements, and the specificity of findings. Given the wide range of clinical manifestations associated with Parkinson's disease and the many tools and metrics to assess them, the challenge is to identify those that may represent a standard for use in clinical trials, and to confirm when digital measurements succeed or fall short of capturing meaningful benefits during drug development.

Eligibility criteria and clinical trials: An FDA perspective.

INTRODUCTION: Randomized clinical trials are regarded as the gold standard for evaluating the effectiveness and safety of interventions. The US Food and Drug Administration (FDA) has made efforts to promote inclusive eligibility criteria. The objective of this study is to identify common trends in eligibility criteria and identify patterns of exclusion criteria among different diseases. METHODS: The authors evaluated the inclusion and exclusion criteria of 38 pivotal clinical trials representing 38 novel drugs approved between 2014 and 2017. Additionally, the authors reviewed the demographic characteristics of participants enrolled across 38 trials. RESULTS: Eighty two percent of trials in our study excluded participants based on hepatic related criteria and 79% trials excluded participants based on renal related criteria and specific infectious diseases. For trials in conditions that affected both men and women, there were no exclusions based on gender and race. More than 90% of trials excluded pregnant, lactating women and women not on adequate contraception. Of the 36,644 patients enrolled in trials for which both men and women were eligible, 62% were men. CONCLUSIONS: The most frequent exclusion criteria were pregnancy, lactation/breastfeeding, renal and hepatic abnormalities, and specific infectious diseases. The preponderance of men in our study likely indicates that factors other than exclusion criteria affect enrollment. The lower representation of women may have been influenced by two large cardiovascular trials that included 75% men. Our study marks an important step in the ongoing efforts of the Agency to increase inclusivity in clinical trials by understanding common clinical trial eligibility criteria patterns.

Effect of drug combinations on the kinetics of antibiotic resistance emergence in Escherichia coli CFT073 using an in vitro hollow-fibre infection model.

Antibiotic resistance is one of the major threats to public health today. To address this problem requires an urgent comprehensive approach. Strategic and multitargeted combination therapy has been increasingly used clinically to treat bacterial infections. The hollow-fibre infection model (HFIM) is a well-controlled in vitro bioreactor system that is increasingly being used in the assessment of resistance emergence with monotherapies and combination antibiotic therapies. In this study, the HFIM was evaluated as a reliable in vitro method to quantitatively and reproducibly analyse the emergence of antibiotic resistance using ampicillin, fosfomycin and ciprofloxacin and their simultaneous combinations against Escherichia coli CFT073, a clinical uropathogenic strain. Bacteria were exposed to clinically relevant pharmacokinetic (PK) concentrations of the drugs for 10 days. Drug and bacterial samples were collected at different time points for PK analysis and to enumerate total and resistant bacterial populations, respectively. The results demonstrated that double or triple combinations significantly delayed the emergence of resistant E. coli CFT073 subpopulations. These findings suggest that strategic combinations of antimicrobials may play a role in controlling the emergence of resistance during treatment. Further animal and human trials will be needed to confirm this and to ensure that there is no adverse impact on the host microbiome or unexpected toxicity. The HFIM system could potentially be used to identify clinically relevant combination dosing regimens for use in a clinical trial evaluating the appearance of resistance to antibacterial drugs.

Mycobacterium tuberculosis bloodstream infection prevalence, diagnosis, and mortality risk in seriously ill adults with HIV: a systematic review and meta-analysis of individual patient data.

BACKGROUND: The clinical and epidemiological significance of HIV-associated Mycobacterium tuberculosis bloodstream infection (BSI) is incompletely understood. We hypothesised that M tuberculosis BSI prevalence has been underestimated, that it independently predicts death, and that sputum Xpert MTB/RIF has suboptimal diagnostic yield for M tuberculosis BSI. METHODS: We did a systematic review and individual patient data (IPD) meta-analysis of studies performing routine mycobacterial blood culture in a prospectively defined patient population of people with HIV aged 13 years or older. Studies were identified through searching PubMed and Scopus up to Nov 10, 2018, without language or date restrictions and through manual review of reference lists. Risk of bias in the included studies was assessed with an adapted QUADAS-2 framework. IPD were requested for all identified studies and subject to harmonised inclusion criteria: age 13 years or older, HIV positivity, available CD4 cell count, a valid mycobacterial blood culture result (excluding patients with missing data from lost or contaminated blood cultures), and meeting WHO definitions for suspected tuberculosis (presence of screening symptom). Predicted probabilities of M tuberculosis BSI from mixed-effects modelling were used to estimate prevalence. Estimates of diagnostic yield of sputum testing with Xpert (or culture if Xpert was unavailable) and of urine lipoarabinomannan (LAM) testing for M tuberculosis BSI were obtained by two-level random-effect meta-analysis. Estimates of mortality associated with M tuberculosis BSI were obtained by mixed-effect Cox proportional-hazard modelling and of effect of treatment delay on mortality by propensity-score analysis. This study is registered with PROSPERO, number 42016050022. FINDINGS: We identified 23 datasets for inclusion (20 published and three unpublished at time of search) and obtained IPD from 20, representing 96·2% of eligible IPD. Risk of bias for the included studies was assessed to be generally low except for on the patient selection domain, which was moderate in most studies. 5751 patients met harmonised IPD-level inclusion criteria. Technical factors such as number of blood cultures done, timing of blood cultures relative to blood sampling, and patient factors such as inpatient setting and CD4 cell count, explained significant heterogeneity between primary studies. The predicted probability of M tuberculosis BSI in hospital inpatients with HIV-associated tuberculosis, WHO danger signs, and a CD4 count of 76 cells per µL (the median for the cohort) was 45% (95% CI 38-52). The diagnostic yield of sputum in patients with M tuberculosis BSI was 77% (95% CI 63-87), increasing to 89% (80-94) when combined with urine LAM testing. Presence of M tuberculosis BSI compared with its absence in patients with HIV-associated tuberculosis increased risk of death before 30 days (adjusted hazard ratio 2·48, 95% CI 2·05-3·08) but not after 30 days (1·25, 0·84-2·49). In a propensity-score matched cohort of participants with HIV-associated tuberculosis (n=630), mortality increased in patients with M tuberculosis BSI who had a delay in anti-tuberculosis treatment of longer than 4 days compared with those who had no delay (odds ratio 3·15, 95% CI 1·16-8·84). INTERPRETATION: In critically ill adults with HIV-tuberculosis, M tuberculosis BSI is a frequent manifestation of tuberculosis and predicts mortality within 30 days. Improved diagnostic yield in patients with M tuberculosis BSI could be achieved through combined use of sputum Xpert and urine LAM. Anti-tuberculosis treatment delay might increase the risk of mortality in these patients. FUNDING: This study was supported by Wellcome fellowships 109105Z/15/A and 105165/Z/14/A.

Evaluation of Immunocompetent Urinary Tract Infected Balb/C Mouse Model For the Study of Antibiotic Resistance Development Using Escherichia Coli CFT073 Infection.

Urinary tract infections (UTI) are common worldwide and are becoming increasingly difficult to treat because of the development of antibiotic resistance. Immunocompetent murine models of human UTI have been used to study pathogenesis and treatment but not for investigating resistance development after treatment with antibiotics. In this study, intravesical inoculation of uropathogenic Escherichia coli CFT073 in immunocompetent Balb/c mice was used as a model of human UTI. The value of the model in investigating antibiotic exposure on in vivo emergence of antibiotic resistance was examined. Experimentally infected mice were treated with 20 or 200 mg/kg ampicillin, 5 or 50 mg/kg ciprofloxacin, or 100 or 1000 mg/kg of fosfomycin. Ampicillin and ciprofloxacin were given twice daily at 8 h intervals, and fosfomycin was given once daily. Antibiotic treatment began 24 h after bacterial inoculation and ended after 72 h following the initial treatment. Although minimum inhibitory concentrations (MIC) for the experimental strain of E. coli were exceeded at peak concentrations in tissues and consistently in urine, low levels of bacteria persisted in tissues in all experiments. E. coli from bladder tissue, kidney, and urine grew on plates containing 1× MIC of antibiotic, but none grew at 3× MIC. This model is not suitable for studying emergent resistance but might serve to examine bacterial persistence.

Mobile health: the power of wearables, sensors, and apps to transform clinical trials.

Mobile technology has become a ubiquitous part of everyday life, and the practical utility of mobile devices for improving human health is only now being realized. Wireless medical sensors, or mobile biosensors, are one such technology that is allowing the accumulation of real-time biometric data that may hold valuable clues for treating even some of the most devastating human diseases. From wearable gadgets to sophisticated implantable medical devices, the information retrieved from mobile technology has the potential to revolutionize how clinical research is conducted and how disease therapies are delivered in the coming years. Encompassing the fields of science and engineering, analytics, health care, business, and government, this report explores the promise that wearable biosensors, along with integrated mobile apps, hold for improving the quality of patient care and clinical outcomes. The discussion focuses on groundbreaking device innovation, data optimization and validation, commercial platform integration, clinical implementation and regulation, and the broad societal implications of using mobile health technologies.

Varicella pneumonia in patients with HIV/AIDS.

OBJECTIVE: To determine the potential role of steroid therapy combined with early antiviral and supportive care in patients infected with human immunodeficiency virus (HIV) with varicella pneumonia. MATERIALS AND METHODS: A retrospective review was conducted of the incidence, clinical course, and outcome of varicella pneumonia in patients with HIV or acquired immunodeficiency syndrome (AIDS). RESULTS: Seven of 12 patients (58%) who were hospitalized with chickenpox developed clinically severe varicella pneumonia. All patients had advanced immunosuppression and all developed diffuse reticulonodular radiographic abnormalities, although two patients had normal chest radiographs on admission. All patients received antiviral therapy within 12 hours of hospital admission. The overall mortality rate was 43%. Six patients were treated with systemic corticosteroids in addition to antiviral agents, including all four of the survivors. CONCLUSIONS: Hospitalized patients with HIV or AIDS with chickenpox are at high risk for developing varicella pneumonia. There is a potentially high rate of death despite prompt initiation of appropriate antiviral therapy. Intensive care management and adjunctive use of systemic corticosteroids may improve outcome.

Models and approaches for anti-TB drug testing.

Unique challenges remain in the development of new drugs for the treatment of TB. While existing multidrug treatment regimens are prolonged and difficult for patients to adhere to, they are highly efficacious, setting a high bar for the performance of new agents. Complicating matters more, regulatory standards have changed since the first drugs for TB were introduced, with a rigorous characterization of the effect of a new drug within a combination regimen expected. If these demands are to be satisfied, innovative models will be needed to demonstrate drug efficacy. In the past, mycobacterial cultures performed on solid media at the end of treatment have been used as critical biomarkers of drug efficacy, but their inability to predict long-term outcomes with precision has limited their utility. This article reviews a range of nonclinical and clinical models to characterize the bactericidal and/or sterilizing activity of new compounds. Novel approaches, using in vitro and animal models, sensitive biomarkers, as well as creative new clinical trial designs, are discussed. These promise a timely expansion of our TB treatment armamentarium to include potent new drugs and shorter, simpler treatment regimens.

Challenges, successes and hopes in the development of novel TB therapeutics.

Despite efficacious drugs for treatment, TB continues to affect enormous numbers of patients throughout the world. Failure to control TB may be related to the biological characteristics of Mycobacterium tuberculosis, the nature of susceptible hosts often impoverished and poorly supported by healthcare infrastructure and the complex treatment regimens that must be used. Challenges to anti-TB drug development include the organism's slow replication, the ability of M. tuberculosis to survive in a dormant state and to persist despite therapy, its impregnable cell wall and its capacity to develop resistance to drugs. The need for extended therapy using combinations of drugs remains a practical obstacle to effective control in poor, malnourished and diseased communities most susceptible to TB. High-throughput screening of candidate agents and investigation of drugs already in use for other infections are yielding promising new candidates for TB treatment. New families of drugs entering clinical trials include 5-nitroimidazoles, diarylquinolines and ethylene diamines. Increasing funding initiatives, advances in the biology of TB and strategies for drug discovery have rejuvenated the pipeline of new drugs for TB, promising an expanding armamentarium of effective drugs with improved tolerability and potential to treat drug-resistant cases.

Developing new drugs for the treatment of drug-resistant tuberculosis: a regulatory perspective.

Simplifying and shortening treatment for drug-sensitive tuberculosis and providing new treatment options for drug-resistant tuberculosis constitute two principal goals in the development of novel drugs for tuberculosis. Demonstration of clinical efficacy in drug-sensitive tuberculosis is challenging, given high success rates for existing regimens, concerns about substituting an investigational agent for the most effective agents in a regimen and difficulties in determining the effect size of the components of a combination regimen. Large and prolonged studies would be needed either to show superiority over existing regimens or statistically defensible non-inferiority compared to existing regimens. In contrast, exploring efficacy of novel treatments in the setting of drug-resistant disease may present certain opportunities. In drug-resistant disease, the efficacy of existing regimens is comparatively poor, and companion drugs used to treat drug-resistant disease are weak or ineffective, enabling demonstration of the effect of the new drug. Other advantages of this approach, which has been used successfully in the development of antiretroviral agents, include the possibility of demonstrating drug efficacy using smaller studies, the possibility of accelerated approval based on a surrogate endpoint and the opportunity to address an urgent public health need. Experience with the activity and the safety of new agents in drug-resistant disease may provide a platform from which their indication can be broadened to include drug-sensitive disease.

Human immunodeficiency virus-1 RNA levels and CD4 lymphocyte counts, during treatment for active tuberculosis, in South African patients.

During 6 months of treatment, we measured human immunodeficiency virus (HIV)-1 virus loads, CD4 T cell counts, and immune activation markers, in 111 HIV-1-infected patients with active tuberculosis (TB). The median virus load (baseline, 5.58 log(10) copies/mL) significantly increased at 1 month (5.71 log(10) copies/mL), then returned to near-baseline levels at 3 months (5.40 log(10) copies/mL) and at 6 months (5.36 log(10) copies/mL). In contrast, the median CD4 counts increased at 1 month (186/mm(3)), at 3 months (238/mm(3)), and at 6 months (239/mm(3)). CD4 counts and virus loads did not change during therapy. Expression of CD38 and HLA-DR remained high throughout treatment, whereas plasma levels of interleukin-6 decreased over time.