PURPOSE

Operational failures are system-level errors in the supply of information, equipment, and materials to health care personnel. We aimed to review and synthesize the research literature to determine how operational failures in primary care affect the work of primary care physicians.

METHODS

We conducted a critical interpretive synthesis. We searched 7 databases for papers published in English from database inception until October 2017 for primary research of any design that addressed problems interfering with primary care physicians’ work. All potentially eligible titles/abstracts were screened by 1 reviewer; 30% were subject to second screening. We conducted an iterative critique, analysis, and synthesis of included studies.

RESULTS

Our search retrieved 8,544 unique citations. Though no paper explicitly referred to "operational failures," we identified 95 papers that conformed to our general definition. The included studies show a gap between what physicians perceived they should be doing and what they were doing, which was strongly linked to operational failures—including those relating to technology, information, and coordination—over which physicians often had limited control. Operational failures actively configured physicians’ work by requiring significant compensatory labor to deliver the goals of care. This labor was typically unaccounted for in scheduling or reward systems and had adverse consequences for physician and patient experience.

CONCLUSIONS

Primary care physicians’ efforts to compensate for suboptimal work systems are often concealed, risking an incomplete picture of the work they do and problems they routinely face. Future research must identify which operational failures are highest impact and tractable to improvement.

PURPOSE

Most real-world studies on anticoagulants have been based on health insurance databases or performed in secondary care. The aim of this study was to compare safety and effectiveness between patients treated with vitamin K antagonists (VKAs) and patients treated with direct oral anticoagulants (DOACs) in a general practice setting.

METHODS

The CACAO study (Comparison of Accidents and their Circumstances with Oral Anticoagulants) is a multicenter prospective cohort study conducted among ambulatory patients taking an oral anticoagulant. Participants were patients from the study’s cross-sectional phase receiving oral anticoagulants because of nonvalvular atrial fibrillation, for secondary prevention of venous thromboembolism, or both. They were followed as usual for 1 year by their general practitioners, who collected data on changes in therapy, thromboembolic events, bleeding, and deaths. All events were adjudicated by an independent committee. We used a propensity score and a Cox regression model to derive hazard ratios.

RESULTS

Between April and December 2014, a total of 3,082 patients were included. At 1 year, 42 patients (1.7%) had experienced an arterial or venous event; 151 (6.1%) had experienced bleeding, including 47 (1.9%) who experienced major bleeding; and 105 (4.1%) had died. There was no significant difference between the VKA and DOAC groups regarding arterial or venous events, or major bleeding. The VKA group had a lower risk of overall bleeding (hazard ratio = 0.65; 95% CI, 0.43-0.98) but twice the risk of death (hazard ratio = 1.98; 95% CI, 1.15-3.42).

CONCLUSIONS

VKAs and DOACs had fairly similar safety and effectiveness in general practice. The substantially higher incidence of deaths with VKAs is consistent with known data from health insurance databases and calls for further research to understand its cause.

PURPOSE

General practitioners (GPs) are part of the US physician workforce, but little is known about who they are, what they do, and how they differ from family physicians (FPs). We describe self-identified GPs and compare them with board-certified FPs.

METHODS

Analysis of data on 102,604 Doctor of Medicine and Doctor of Osteopathy physicians in direct patient care in the United States in 2016, who identify themselves as GPs or FPs. The study used linking databases (American Medical Association Masterfile, American Board of Family Medicine [ABFM], Area Health Resource File, Medicare Public Use File) to examine personal, professional, and practice characteristics.

RESULTS

Of the physicians identified, 6,661 self-designated as GPs and 95,943 self-designated as FPs. Of the self-designated GPs, 116 had been ABFM certified and were excluded from the study. Of the remaining 102,488 physicians, those who self-designated as GPs but were never ABFM certified constituted the GP group (n = 6,545, 6%). Self-designated FPs that were ABFM certified made up the FP group (n = 79,449, 78%). The remaining self-designated FPs not ABFM certified constituted the uncertified group (n = 16,494, 16%). GPs differed from FPs in every characteristic examined. Compared with FPs, GPs are more likely to be older, male, Doctors of Osteopathy, graduates of non-US medical schools, and have no family medicine residency training. GPs practice location is similar to FPs, but GPs are less likely to participate in Medicare or to work in hospitals.

CONCLUSIONS

GPs in the United States are a varied group that differ from FPs. Researchers, educators, and policy makers should not lump GPs together with FPs in data collection, analysis, and reporting.

PURPOSE

Online programs may help to engage patients in advance care planning in outpatient settings. We sought to implement an online advance care planning program, PREPARE (Prepare for Your Care; http://www.prepareforyourcare.org), at home and evaluate the changes in advance care planning engagement among patients attending outpatient clinics.

METHODS

We undertook a prospective before-and-after study in 15 primary care clinics and 2 outpatient cancer centers in Canada. Patients were aged 50 years or older (primary care) or 18 years or older (cancer care) and free of cognitive impairment. They used the PREPARE website over 6 weeks, with reminders sent at 2 or 4 weeks. We used the 55-item Advance Care Planning Engagement Survey, which measures behavior change processes (knowledge, contemplation, self-efficacy, readiness) on 5-point scales and actions relating to substitute decision makers, quality of life, flexibility for the decision maker, and asking doctors questions on an overall scale from 0 to 21; higher scores indicate greater engagement.

RESULTS

In total, 315 patients were screened and 172 enrolled, of whom 75% completed the study (mean age = 65.6 years, 51% female, 35% had cancer). The mean behavior change process score was 2.9 (SD 0.8) at baseline and 3.5 (SD 0.8) at follow-up (mean change = 0.6; 95% CI, 0.49-0.73); the mean action measure score was 4.0 (SD 4.9) at baseline and 5.2 (SD 5.4) at follow-up (mean change = 1.2; 95% CI, 0.54-1.77). The effect size was moderate (0.75) for the former and small (0.23) for the latter. Findings were similar in both primary care and cancer care populations.

CONCLUSIONS

Implementation of the online PREPARE program in primary care and cancer care clinics increased advance care planning engagement among patients.

T-cell receptor (TCR)–based therapeutic cells and agents have emerged as a new class of effective cancer therapies. These therapies work on cells that express intracellular cancer-associated proteins by targeting peptides displayed on MHC receptors. However, cross-reactivities of these agents to off-target cells and tissues have resulted in serious, sometimes fatal, adverse events. We have developed a high-throughput genetic platform (termed "PresentER") that encodes MHC-I peptide minigenes for functional immunologic assays and determines the reactivities of TCR-like therapeutic agents against large libraries of MHC-I ligands. In this article, we demonstrated that PresentER could be used to identify the on-and-off targets of T cells and TCR-mimic (TCRm) antibodies using in vitro coculture assays or binding assays. We found dozens of MHC-I ligands that were cross-reactive with two TCRm antibodies and two native TCRs and that were not easily predictable by other methods.

Random transposon mutagenesis is a powerful and unbiased genetic approach to answer fundamental biological questions. Here, we introduce an improved mariner-based transposon system with enhanced stability during propagation and versatile applications in mutagenesis. We used a low-copy-number plasmid as a transposon delivery vehicle, which affords a lower frequency of unintended recombination during vector construction and propagation in Escherichia coli. We generated a variety of transposons allowing for gene disruption or artificial overexpression, each in combination with one of four different antibiotic resistance markers. In addition, we provide transposons that will report gene/protein expression due to transcriptional or translational coupling. We believe that the TnFLX system will help enhance the flexibility of future transposon modification and application in Bacillus and other organisms.

IMPORTANCE The stability of transposase-encoding vectors during cloning and propagation is crucial for the reliable application of transposons. Here, we increased the stability of the mariner delivery vehicle in E. coli. Moreover, the TnFLX transposon system will improve the application of forward genetic methods with an increased number of antibiotic resistance markers and the ability to generate unbiased green fluorescent protein (GFP) fusions to report on protein translation and subcellular localization.

Vibralactone, a hybrid compound derived from phenols and a prenyl group, is a strong pancreatic lipase inhibitor with a rare fused bicyclic β-lactone skeleton. Recently, a researcher reported a vibralactone derivative (compound C1) that caused inhibition of pancreatic lipase with a half-maximal inhibitory concentration of 14 nM determined by structure-based optimization, suggesting a potential candidate as a new antiobesity treatment. In the present study, we sought to identify the main gene encoding prenyltransferase in Stereum vibrans, which is responsible for the prenylation of phenol leading to vibralactone synthesis. Two RNA silencing transformants of the identified gene (vib-PT) were obtained through Agrobacterium tumefaciens-mediated transformation. Compared to wild-type strains, the transformants showed a decrease in vib-PT expression ranging from 11.0 to 56.0% at 5, 10, and 15 days in reverse transcription-quantitative PCR analysis, along with a reduction in primary vibralactone production of 37 to 64% at 15 and 21 days, respectively, as determined using ultra-high-performance liquid chromatography-mass spectrometry analysis. A soluble and enzymatically active fusion Vib-PT protein was obtained by expressing vib-PT in Escherichia coli, and the enzyme’s optimal reaction conditions and catalytic efficiency (K_m/k_cat) were determined. In vitro experiments established that Vib-PT catalyzed the C-prenylation at C-3 of 4-hydroxy-benzaldehyde and the O-prenylation at the 4-hydroxy of 4-hydroxy-benzenemethanol in the presence of dimethylallyl diphosphate. Moreover, Vib-PT shows promiscuity toward aromatic compounds and prenyl donors.

IMPORTANCE Vibralactone is a lead compound with a novel skeleton structure that shows strong inhibitory activity against pancreatic lipase. Vibralactone is not encoded by the genome directly but rather is synthesized from phenol, followed by prenylation and other enzyme reactions. Here, we used an RNA silencing approach to identify and characterize a prenyltransferase in a basidiomycete species that is responsible for the synthesis of vibralactone. The identified gene, vib-PT, was expressed in Escherichia coli to obtain a soluble and enzymatically active fusion Vib-PT protein. In vitro characterization of the enzyme demonstrated the catalytic mechanism of prenylation and broad substrate range for different aromatic acceptors and prenyl donors. These characteristics highlight the possibility of Vib-PT to generate prenylated derivatives of aromatics and other compounds as improved bioactive agents or potential prodrugs.

Most freshwater bacterial communities are characterized by a few dominant taxa that are often ubiquitous across freshwater biomes worldwide. Our understanding of the genomic diversity within these taxonomic groups is limited to a subset of taxa. Here, we investigated the genomic diversity that enables Limnohabitans, a freshwater genus key in funneling carbon from primary producers to higher trophic levels, to achieve abundance and ubiquity. We reconstructed eight putative Limnohabitans metagenome-assembled genomes (MAGs) from stations located along broad environmental gradients existing in Lake Michigan, part of Earth’s largest surface freshwater system. De novo strain inference analysis resolved a total of 23 strains from these MAGs, which strongly partitioned into two habitat-specific clusters with cooccurring strains from different lineages. The largest number of strains belonged to the abundant LimB lineage, for which robust in situ strain delineation had not previously been achieved. Our data show that temperature and nutrient levels may be important environmental parameters associated with microdiversification within the Limnohabitans genus. In addition, strains predominant in low- and high-phosphorus conditions had larger genomic divergence than strains abundant under different temperatures. Comparative genomics and gene expression analysis yielded evidence for the ability of LimB populations to exhibit cellular motility and chemotaxis, a phenotype not yet associated with available Limnohabitans isolates. Our findings broaden historical marker gene-based surveys of Limnohabitans microdiversification and provide in situ evidence of genome diversity and its functional implications across freshwater gradients.

IMPORTANCE Limnohabitans is an important bacterial taxonomic group for cycling carbon in freshwater ecosystems worldwide. Here, we examined the genomic diversity of different Limnohabitans lineages. We focused on the LimB lineage of this genus, which is globally distributed and often abundant, and its abundance has shown to be largely invariant to environmental change. Our data show that the LimB lineage is actually comprised of multiple cooccurring populations for which the composition and genomic characteristics are associated with variations in temperature and nutrient levels. The gene expression profiles of this lineage suggest the importance of chemotaxis and motility, traits that had not yet been associated with the Limnohabitans genus, in adapting to environmental conditions.

Background:

Care services have not been sufficiently adapted to meet the comprehensive care needs of women living with HIV. Our study objective was to engage patients and providers in codesigning care recommendations to improve care for this population in the province of Quebec.

Methods:

We conducted a 5-hour deliberative dialogue workshop in April 2019 in Montréal as the final phase of a mixed-methods study investigating comprehensive care for women living with HIV. The study drew on data from the Canadian HIV Women’s Sexual and Reproductive Health Cohort Study (CHIWOS). Recruitment was guided by a purposive maximum-variation sampling strategy to ensure an appropriate mix of participants and was facilitated by our existing CHIWOS networks. Participants included patients (women living with HIV) and HIV care providers (doctors, nurses, pharmacists). The workshop was facilitated professionally and included a synthesis of the evidence, small- and large-group deliberations, and voting on care improvements.

Results:

Eight patients and 8 HIV care providers participated. Drawing on identified care priorities, the participants identified 4 relatively rapid care improvements and 3 longer-term improvements. The rapid care improvements included delegating medical acts to members of multidisciplinary care teams; greater involvement of HIV community members within care settings and health care decision-making; creating a women’s health information booklet; and increasing HIV education among all health care providers and raising awareness of women’s care needs beyond HIV-specific care among HIV care providers. The longer-term care improvements included advocating for complete financial coverage of antiretroviral therapy within the government-sponsored Medicare program, facilitating access to allied care providers (e.g., physiotherapists and psychologists) and launching a population-wide campaign to increase awareness about the Undetectable = Untransmittable (U=U) initiative and other HIV advances.

Interpretation:

The deliberative dialogue workshop yielded evidence-based, stakeholder-driven recommendations to improve the comprehensive care of women living with HIV in Quebec.

The aim of this study was to assess the association of high-sensitivity cardiac troponin (hs-cTnT) and other cardiac, kidney, hyperglycemia, and inflammatory biomarkers with peripheral neuropathy (PN) in a community-based population.We conducted a cross-sectional analysis of 3056 black and white participants in the Atherosclerosis Risk in Communities (ARIC) study who underwent standardized monofilament PN testing and had measures of cardiac function (hs-cTnT, N-terminal pro–B-type natriuretic peptide [NT-proBNP], and growth differentiation factor 15 [GDF15]), kidney function (serum creatinine, cystatin C, β-2 microglobulin, urine albumin-to-creatinine ratio), hyperglycemia (fasting glucose, hemoglobin A_1c [Hb A_1c], fructosamine, glycated albumin, 1,5-anhydroglucitol), and inflammation (C-reactive protein) assessed at visit 6 (2016–2017; age 71–94 years). We used logistic regression to assess the associations of these biomarkers (modeled in diabetes-specific tertiles) with PN in older adults with and without diabetes after adjusting for traditional risk factors.In total, 33.5% of participants had PN (37.3% with diabetes and 31.9% without diabetes). There was an independent association of hs-cTnT with PN regardless of diabetes status (diabetes T3 vs. T1: odds ratio [OR], 2.15 [95% CI, 1.44–3.22]; no diabetes: OR, 2.31 [95%CI, 1.76–3.03]; P = 0.72 for interaction). Among participants without diabetes, there were also significant associations of NT-proBNP (OR, 1.40 [95% CI, 1.08–1.81]) and urine albumin-to-creatinine ratio (OR, 1.55 [95% CI, 1.22–1.97]) with PN. Associations of hyperglycemia biomarkers including Hb A_1c (OR, 1.76 [95% CI, 1.22–2.54]), fructosamine (OR, 1.71 [95% CI, 1.19–2.46]), and glycated albumin (OR, 1.45 [95% CI, 1.03–2.03]) with PN were significant only among participants with diabetes.Overall, hs-cTnT appears to be a global marker of end organ damage, including PN. Laboratory biomarkers may be able to help us identify those individuals with PN.

Peripheral neuropathy (PN) is a condition affecting up to 20% of the general population. The symptoms range from mild to disabling, depending on the types of nerve fiber affected and the type and severity of damage.

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This study showed that treatment with a therapeutic monoclonal immunoglobulin-G1 antibody against phosphorylcholine on oxidized phospholipids preserves coronary flow reserve and attenuates atherosclerotic inflammation as determined by the uptake of ¹⁸F-fluorodeoxyglucose in atherosclerotic mice. The noninvasive imaging techniques represent translational tools to assess the efficacy of phosphorylcholine-targeted therapy on coronary artery function and atherosclerosis in clinical studies.

Recently, there has been a worldwide resurgence in pneumoconiosis, or pulmonary fibrosis due to occupational mineral dust exposure. In Queensland, Australia, there has been a re-emergence of coal workers' pneumoconiosis and silicosis. Some coal mining communities have experienced a resurgence of progressive massive fibrosis in the USA and a worldwide epidemic is occurring of accelerated silicosis due to exposure to artificial stone.

These diseases are all preventable and should not be occurring in the 21st century. Best practice prevention includes reduction of exposure to mineral dusts or, ideally, prevention of exposure altogether. However, where dust exposure has occurred, respiratory surveillance can provide a strategy for early disease detection. It is important to identify early signs of occupational lung disease at a stage where intervention may be beneficial, though it must be acknowledged that progression may occur even after cessation of exposure to dusts. Respiratory surveillance should be distinguished from population screening and case finding, which are different methods used for disease investigation and control. Designing an ideal respiratory surveillance programme is challenging, as there is no single test that accurately identifies early disease. Several different respiratory disorders may occur related to the same exposure(s). Physicians organising and interpreting tests used in respiratory surveillance must be aware of the broad range of potential work-related respiratory conditions, complexities in diagnosis, and appropriate interpretation of the exposure history, as well as current management options. A working knowledge of the compensation and medicolegal avenues available to workers in individual jurisdictions is also useful.

Key points

Mineral dust exposure causes a number of conditions, including those specific to dust exposures, such as the pneumoconioses (or pulmonary fibroses due to mineral dust exposure), and others that may additionally be related to other causes, such as COPD.

Identification of multiple conditions using respiratory investigations requires expert interpretation and understanding of the range of potential conditions.

The frequency and content of a respiratory surveillance programme will vary according to the relevant occupational exposures, and be affected by both medical and nonmedical factors, including the background prevalence of local diseases. A programme will also need to consider other factors such as local legislation, availability of resources, worker convenience and cost.

Educational aims

To identify the large range of respiratory diseases caused by exposure to mineral dusts and identify the range of tests that may be used in a surveillance programme for occupational respiratory disorders.

To highlight difficulties that might be experienced by medical practitioners in designing and operating an effective surveillance programme, while incorporating rapidly advancing medical technology and practice.

Caseum, the central necrotic material of tuberculous lesions, is a reservoir of drug-recalcitrant persisting mycobacteria. Caseum is found in closed nodules and in open cavities connecting with an airway. Several commonly accepted characteristics of caseum were established during the preantibiotic era, when autopsies of deceased tuberculosis (TB) patients were common but methodologies were limited. These pioneering studies generated concepts such as acidic pH, low oxygen tension, and paucity of nutrients being the drivers of nonreplication and persistence in caseum. Here we review widely accepted beliefs about the caseum-specific stress factors thought to trigger the shift of Mycobacterium tuberculosis to drug tolerance. Our current state of knowledge reveals that M. tuberculosis is faced with a lipid-rich diet rather than nutrient deprivation in caseum. Variable caseum pH is seen across lesions, possibly transiently acidic in young lesions but overall near neutral in most mature lesions. Oxygen tension is low in the avascular caseum of closed nodules and high at the cavity surface, and a gradient of decreasing oxygen tension likely forms toward the cavity wall. Since caseum is largely made of infected and necrotized macrophages filled with lipid droplets, the microenvironmental conditions encountered by M. tuberculosis in foamy macrophages and in caseum bear many similarities. While there remain a few knowledge gaps, these findings constitute a solid starting point to develop high-throughput drug discovery assays that combine the right balance of oxygen tension, pH, lipid abundance, and lipid species to model the profound drug tolerance of M. tuberculosis in caseum.

Intrathecal administration of anti-infectives is indicated in central nervous system infections by multiresistant pathogens when drugs that can reach adequate cerebrospinal fluid (CSF) concentrations by systemic therapy are not available. Antibiotics that readily pass the blood-brain and blood-CSF barriers and/or that have low toxicity allowing an increase in the daily dosage should not be used for intrathecal therapy. Intrathecal therapy is accompanied by systemic treatment. Antibacterials indispensable for intrathecal therapy include aminoglycosides, colistin, daptomycin, tigecycline, and vancomycin. Limited experience suggests the utility of the antifungals amphotericin B and caspofungin. Intraventricular administration ensures distribution throughout the CSF compartment, whereas intralumbar dosing often fails to attain adequate antibiotic concentrations in the ventricles. The individual dose is determined by the estimated size of the CSF space and by the estimated clearance from CSF. For moderately lipophilic anti-infectives with a molecular weight above approximately 1,000 g/mol, as well as for hydrophilic drugs with a molecular weight above approximately 400 g/mol, one daily dose is normally adequate. The ventricular drain should be clamped for 15 to 120 min to facilitate the distribution of the anti-infective in the CSF space. Therapeutic drug monitoring of the trough levels is necessary only in cases of therapeutic failure.

Purpose:

We hypothesized that the combination of a local stimulus for activating tumor-specific T cells and an anti-immunosuppressant would improve treatment of gliomas. Virally encoded IL15Rα-IL15 as the T-cell activating stimulus and a prostaglandin synthesis inhibitor as the anti-immunosuppressant were combined with adoptive transfer of tumor-specific T cells.

Experimental Design:

Two oncolytic poxviruses, vvDD vaccinia virus and myxoma virus, were each engineered to express the fusion protein IL15Rα-IL15 and a fluorescent protein. Viral gene expression (YFP or tdTomato Red) was confirmed in the murine glioma GL261 in vitro and in vivo. GL261 tumors in immunocompetent C57BL/6J mice were treated with vvDD-IL15Rα-YFP vaccinia virus or vMyx-IL15Rα-tdTr combined with other treatments, including vaccination with GARC-1 peptide (a neoantigen for GL261), rapamycin, celecoxib, and adoptive T-cell therapy.

Results:

vvDD-IL15Rα-YFP and vMyx-IL15Rα-tdTr each infected and killed GL261 cells in vitro. In vivo, NK cells and CD8⁺ T cells were increased in the tumor due to the expression of IL15Rα-IL15. Each component of a combination treatment contributed to prolonging survival: an oncolytic virus, the IL15Rα-IL15 expressed by the virus, a source of T cells (whether by prevaccination or adoptive transfer), and prostaglandin inhibition all synergized to produce elimination of gliomas in a majority of mice. vvDD-IL15Rα-YFP occasionally caused ventriculitis-meningitis, but vMyx-IL15Rα-tdTr was safe and effective, causing a strong infiltration of tumor-specific T cells and eliminating gliomas in 83% of treated mice.

Conclusions:

IL15Rα-IL15–armed oncolytic poxviruses provide potent antitumor effects against brain tumors when combined with adoptive T-cell therapy, rapamycin, and celecoxib.

Purpose:

There are several agents in early clinical trials targeting components of the adenosine pathway including A2AR and CD73. The identification of cancers with a significant adenosine drive is critical to understand the potential for these molecules. However, it is challenging to measure tumor adenosine levels at scale, thus novel, clinically tractable biomarkers are needed.

Experimental Design:

We generated a gene expression signature for the adenosine signaling using regulatory networks derived from the literature and validated this in patients. We applied the signature to large cohorts of disease from The Cancer Genome Atlas (TCGA) and cohorts of immune checkpoint inhibitor–treated patients.

Results:

The signature captures baseline adenosine levels in vivo (r² = 0.92, P = 0.018), is reduced after small-molecule inhibition of A2AR in mice (r² = –0.62, P = 0.001) and humans (reduction in 5 of 7 patients, 70%), and is abrogated after A2AR knockout. Analysis of TCGA confirms a negative association between adenosine and overall survival (OS, HR = 0.6, P < 2.2e^–16) as well as progression-free survival (PFS, HR = 0.77, P = 0.0000006). Further, adenosine signaling is associated with reduced OS (HR = 0.47, P < 2.2e^–16) and PFS (HR = 0.65, P = 0.0000002) in CD8⁺ T-cell–infiltrated tumors. Mutation of TGFβ superfamily members is associated with enhanced adenosine signaling and worse OS (HR = 0.43, P < 2.2e^–16). Finally, adenosine signaling is associated with reduced efficacy of anti-PD1 therapy in published cohorts (HR = 0.29, P = 0.00012).

Conclusions:

These data support the adenosine pathway as a mediator of a successful antitumor immune response, demonstrate the prognostic potential of the signature for immunotherapy, and inform patient selection strategies for adenosine pathway modulators currently in development.

Purpose:

The development of leptomeningeal melanoma metastases (LMM) is a rare and devastating complication of the late-stage disease, for which no effective treatments exist. Here, we performed a multi-omics analysis of the cerebrospinal fluid (CSF) from patients with LMM to determine how the leptomeningeal microenvironment shapes the biology and therapeutic responses of melanoma cells.

Experimental Design:

A total of 45 serial CSF samples were collected from 16 patients, 8 of these with confirmed LMM. Of those with LMM, 7 had poor survival (<4 months) and one was an extraordinary responder (still alive with survival >35 months). CSF samples were analyzed by mass spectrometry and incubated with melanoma cells that were subjected to RNA sequencing (RNA-seq) analysis. Functional assays were performed to validate the pathways identified.

Results:

Mass spectrometry analyses showed the CSF of most patients with LMM to be enriched for pathways involved in innate immunity, protease-mediated damage, and IGF-related signaling. All of these were anticorrelated in the extraordinary responder. RNA-seq analysis showed CSF to induce PI3K/AKT, integrin, B-cell activation, S-phase entry, TNFR2, TGFβ, and oxidative stress responses in the melanoma cells. ELISA assays confirmed that TGFβ expression increased in the CSF of patients progressing with LMM. CSF from poorly responding patients conferred tolerance to BRAF inhibitor therapy in apoptosis assays.

Conclusions:

These analyses identified proteomic/transcriptional signatures in the CSF of patients who succumbed to LMM. We further showed that the CSF from patients with LMM has the potential to modulate BRAF inhibitor responses and may contribute to drug resistance.

See related commentary by Glitza Oliva and Tawbi, p. 2083

Purpose:

Using standard-of-care CT images obtained from patients with a diagnosis of non–small cell lung cancer (NSCLC), we defined radiomics signatures predicting the sensitivity of tumors to nivolumab, docetaxel, and gefitinib.

Experimental Design:

Data were collected prospectively and analyzed retrospectively across multicenter clinical trials [nivolumab, n = 92, CheckMate017 (NCT01642004), CheckMate063 (NCT01721759); docetaxel, n = 50, CheckMate017; gefitinib, n = 46, (NCT00588445)]. Patients were randomized to training or validation cohorts using either a 4:1 ratio (nivolumab: 72T:20V) or a 2:1 ratio (docetaxel: 32T:18V; gefitinib: 31T:15V) to ensure an adequate sample size in the validation set. Radiomics signatures were derived from quantitative analysis of early tumor changes from baseline to first on-treatment assessment. For each patient, 1,160 radiomics features were extracted from the largest measurable lung lesion. Tumors were classified as treatment sensitive or insensitive; reference standard was median progression-free survival (NCT01642004, NCT01721759) or surgery (NCT00588445). Machine learning was implemented to select up to four features to develop a radiomics signature in the training datasets and applied to each patient in the validation datasets to classify treatment sensitivity.

Results:

The radiomics signatures predicted treatment sensitivity in the validation dataset of each study group with AUC (95 confidence interval): nivolumab, 0.77 (0.55–1.00); docetaxel, 0.67 (0.37–0.96); and gefitinib, 0.82 (0.53–0.97). Using serial radiographic measurements, the magnitude of exponential increase in signature features deciphering tumor volume, invasion of tumor boundaries, or tumor spatial heterogeneity was associated with shorter overall survival.

Conclusions:

Radiomics signatures predicted tumor sensitivity to treatment in patients with NSCLC, offering an approach that could enhance clinical decision-making to continue systemic therapies and forecast overall survival.

Purpose:

Radium-223 is approved for metastatic castration-resistant prostate cancer (mCRPC) based on improved overall survival, and delay in skeletal related events. However, it is not associated with PSA or radiographic response, which poses a challenge in real-time assessment of its efficacy. Surrogate markers of treatment outcomes may facilitate tailoring treatment duration with radium-223, by limiting the duration of therapy with radium-223 in these patients. Here, we sought to investigate the utility of bone metabolic markers (BMMs) as surrogate markers of response to radium-223 in mCRPC.

Patients and Methods:

A prospective phase II trial of radium-223 plus enzalutamide (RE) versus enzalutamide alone was designed to assess surrogacy of BMMs with respect to response to radium-223. Enzalutamide was used as a comparator in lieu of placebo due to the progressive disease. Co-primary endpoints were relative change in serum BMM N-telopeptide (NTP) levels from baseline to 6 months between the two arms and safety and feasibility of the combination.

Results:

Thirty-nine men were randomized to RE (n = 27) or enzalutamide (n = 12). Combination was safe and feasible. Primary endpoint was met. A statistically significant relative change to NTP ratios between arms (0.64, 95% confidence interval, 0.51–0.81; P = 0.00048) favored RE versus enzalutamide. Overall, BMMs decreased with the RE therapy compared with enzalutamide. Improved PSA response rate in RE versus enzalutamide (P = 0.024), correlated with decline in BMMs.

Conclusions:

BMMs declined significantly with combination therapy, and were associated with improved outcomes. Upon external validation, BMMs may emerge as surrogate markers to monitor treatment with radium-223 in real-time.

Recent FDA approvals of regimens targeting programmed death 1 (PD-1) in combination with anti-CTLA-4 or with VEGF tyrosine kinase inhibitors are reshaping front-line therapy for metastatic kidney cancer. In parallel, therapeutics specific for programmed death ligand 1 (PD-L1), one of the two major ligands for PD-1, are under continued investigation. Surprisingly, not all PD-1 and PD-L1 agents lead to similar clinical outcomes, potentially due to biological differences in the cellular expression and regulation of these targets. Here, we review current clinical data on combination immune checkpoint inhibitor therapy in metastatic kidney cancer and discuss the relevant biology of PD-1 and PD-L1. The design of future rational combination therapy trials in metastatic renal cell carcinoma will rely upon an understanding of this biology, along with an evolving understanding of immune cell populations and their functional states in the tumor microenvironment.

New drugs or therapeutic combinations are urgently needed against Mycobacterium abscessus. Previously, we demonstrated the potent activity of indole-2-carboxamides 6 and 12 against M. abscessus. We show here that these compounds act synergistically with imipenem and cefoxitin in vitro and increase the bactericidal activity of the β-lactams against M. abscessus. In addition, compound 12 also displays synergism with imipenem and cefoxitin within infected macrophages. The clinical potential of these new drug combinations requires further evaluation.

Plazomicin was active against 97.0% of 8,783 Enterobacterales isolates collected in the United States (2016 and 2017), and only 6 isolates carried 16S rRNA methyltransferases conferring resistance to virtually all aminoglycosides. Plazomicin (89.2% to 95.9% susceptible) displayed greater activity than amikacin (72.5% to 78.6%), gentamicin (30.4% to 45.9%), and tobramycin (7.8% to 22.4%) against carbapenem-resistant and extensively drug-resistant isolates. The discrepancies among the susceptibility rates for these agents was greater when applying breakpoints generated using the same stringent contemporary methods applied to determine plazomicin breakpoints.

Treating malaria in HIV-coinfected individuals should consider potential drug-drug interactions. Artemether-lumefantrine is the most widely recommended treatment for uncomplicated malaria globally. Lumefantrine is metabolized by CYP3A4, an enzyme that commonly used antiretrovirals often induce or inhibit. A population pharmacokinetic meta-analysis was conducted using individual participant data from 10 studies with 6,100 lumefantrine concentrations from 793 nonpregnant adult participants (41% HIV-malaria-coinfected, 36% malaria-infected, 20% HIV-infected, and 3% healthy volunteers). Lumefantrine exposure increased 3.4-fold with coadministration of lopinavir-ritonavir-based antiretroviral therapy (ART), while it decreased by 47% with efavirenz-based ART and by 59% in the patients with rifampin-based antituberculosis treatment. Nevirapine- or dolutegravir-based ART and malaria or HIV infection were not associated with significant effects. Monte Carlo simulations showed that those on concomitant efavirenz or rifampin have 49% and 80% probability of day 7 concentrations <200 ng/ml, respectively, a threshold associated with an increased risk of treatment failure. The risk of achieving subtherapeutic concentrations increases with larger body weight. An extended 5-day and 6-day artemether-lumefantrine regimen is predicted to overcome these drug-drug interactions with efavirenz and rifampin, respectively.

KBP-7072 is a novel third-generation tetracycline (aminomethylcycline) antibacterial that overcomes common efflux and ribosomal protection resistance mechanisms that cause resistance in older-generation tetracyclines. KBP-7072 completed phase 1 clinical development studies for safety, tolerability, and pharmacokinetics (ClinicalTrials.gov identifier NCT02454361) and multiple ascending doses in healthy subjects (ClinicalTrials.gov identifier NCT02654626) in December 2015. Both oral and intravenous formulations of KBP-7072 are being developed. In this study, we evaluated the in vitro activities of KBP-7072 and comparator agents by CLSI document M07 (2018) broth microdilution against 531 recent geographically diverse and/or molecularly characterized Acinetobacter baumannii-A. calcoaceticus species complex (A. baumannii) isolates from the United States, Europe, Asia-Pacific (excluding China), and Latin America. A. baumannii isolates included carbapenem-resistant, colistin-resistant, tetracycline-resistant, and extended-spectrum-β-lactamase (ESBL)- and metallo-β-lactamase (MBL)-producing isolates. Overall, KBP-7072 (MIC_50/90, 0.25/1 mg/liter) was comparable in activity to colistin (92.8%/92.8% susceptible [S] [CLSI/EUCAST]) against A. baumannii isolates, inhibiting 99.2% of isolates at ≤2 mg/liter and 97.6% of isolates at ≤1 mg/liter. KBP-7072 was equally active against A. baumannii isolates, including carbapenem-resistant, colistin-resistant, and tetracycline-resistant isolates, regardless of geographic location, and maintained activity against ESBL- and MBL-producing isolates. KBP-7072 outperformed comparator agents, including ceftazidime (40.3% S [CLSI]), gentamicin (48.2%/48.2% S [CLSI/EUCAST]), levofloxacin (39.5%/37.9% S [CLSI/EUCAST]), meropenem (42.0%/42.0% S [CLSI/EUCAST]), piperacillin-tazobactam (33.3% S [CLSI]), and all tetracycline-class comparator agents, which include doxycycline (67.3% S [CLSI]), minocycline (73.8% S [CLSI]), tetracycline (37.2% S [CLSI]), and tigecycline (79.5% inhibited by ≤2 mg/liter). The potent in vitro activity of KBP-7072 against recent geographically diverse, molecularly characterized, and drug-resistant A. baumannii isolates supports continued clinical development for the treatment of serious infections, including those caused by A. baumannii.

Infections with nontuberculous mycobacteria (NTM) have a poor prognosis in patients with underlying respiratory diseases. Clofazimine (CFZ) showed both experimental and clinical promising results against clinically relevant NTM. However, there are no data on CFZ in combination with the current recommended treatment; therefore, we aimed to study its in vivo activity in an aerosol mouse model of Mycobacterium avium. In an aerosol infection BALB/c mouse model using M. avium strain Chester, we treated 58 mice with four combinations of rifampin (RIF) at 10 mg/kg, CFZ at 25 mg/kg, and clarithromycin (CLR) and ethambutol (EMB) at 100 mg/kg. Treatment efficacy was assessed on the basis of lung CFU counts after 2 (M2) and 4 (M4) months of treatment. At M2, CLR-RIF-EMB was slightly but significantly more efficient than CFZ-RIF-EMB (3.02 ± 0.12 versus 3.55 ± 0.28, respectively, P < 0.01), whereas CLR-CFZ-EMB and CLR-CFZ-RIF-EMB dramatically decreased lung CFU counts by 4.32 and 4.47 log₁₀, respectively, compared to untreated group. At M4, CLR-RIF-EMB was significantly more efficient than CFZ-RIF-EMB (2 ± 0.53 versus 2.66 ± 0.22, respectively, P = 0.01). The addition of CLZ to CLR dramatically decreased the lung CFU count, with CFU counts 5.41 and 5.79 log₁₀ lower in the CLR-CFZ-EMB and CLR-CFZ-RIF-EMB groups, respectively, than in the untreated group. The addition of CFZ to CLR seems to improve the efficacy of CLR as early as M2 and was confirmed at M4. CFZ, in addition to RIF and EMB, on the other hand, is less effective than CLR-RIF-EMB. These results need to be confirmed by similar studies along with CFZ potential for shortening treatment.

Antibiotics revolutionized the treatment of infectious diseases; however, it is now clear that broad-spectrum antibiotics alter the composition and function of the host’s microbiome. The microbiome plays a key role in human health, and its perturbation is increasingly recognized as contributing to many human diseases. Widespread broad-spectrum antibiotic use has also resulted in the emergence of multidrug-resistant pathogens, spurring the development of pathogen-specific strategies such as monoclonal antibodies (MAbs) to combat bacterial infection. Not only are pathogen-specific approaches not expected to induce resistance in nontargeted bacteria, but they are hypothesized to have minimal impact on the gut microbiome. Here, we compare the effects of antibiotics, pathogen-specific MAbs, and their controls (saline or control IgG [c-IgG]) on the gut microbiome of 7-week-old, female, C57BL/6 mice. The magnitude of change in taxonomic abundance, bacterial diversity, and bacterial metabolites, including short-chain fatty acids (SCFA) and bile acids in the fecal pellets from mice treated with pathogen-specific MAbs, was no different from that with animals treated with saline or an IgG control. Conversely, dramatic changes were observed in the relative abundance, as well as alpha and beta diversity, of the fecal microbiome and bacterial metabolites in the feces of all antibiotic-treated mice. Taken together, these results indicate that pathogen-specific MAbs do not alter the fecal microbiome like broad-spectrum antibiotics and may represent a safer, more-targeted approach to antibacterial therapy.

The comparative efficacy of ceftazidime-avibactam and meropenem-vaborbactam for treatment of carbapenem-resistant Enterobacteriaceae (CRE) infections remains unknown. This was a multicenter, retrospective cohort study of adults with CRE infections who received ceftazidime-avibactam or meropenem-vaborbactam for ≥72 hours from February 2015 to October 2018. Patients with a localized urinary tract infection and repeat study drug exposures after the first episode were excluded. The primary endpoint was clinical success compared between treatment groups. Secondary endpoints included 30- and 90-day mortality, adverse events (AE), 90-day CRE infection recurrence, and development of resistance in patients with recurrent infection. A post hoc subgroup analysis was completed comparing patients who received ceftazidime-avibactam monotherapy, ceftazidime-avibactam combination therapy, and meropenem-vaborbactam monotherapy. A total of 131 patients were included (ceftazidime-avibactam, n = 105; meropenem-vaborbactam, n = 26), 40% of whom had bacteremia. No significant difference in clinical success was observed between groups (62% versus 69%; P = 0.49). Patients in the ceftazidime-avibactam arm received combination therapy more often than patients in the meropenem-vaborbactam arm (61% versus 15%; P < 0.01). No difference in 30- and 90-day mortality resulted, and rates of AE were similar between groups. In patients with recurrent infection, development of resistance occurred in three patients that received ceftazidime-avibactam monotherapy and in no patients in the meropenem-vaborbactam arm. Clinical success was similar between patients receiving ceftazidime-avibactam and meropenem-vaborbactam for treatment of CRE infections, despite ceftazidime-avibactam being used more often as a combination therapy. Development of resistance was more common with ceftazidime-avibactam monotherapy.

Tedizolid (TZD) and daptomycin (DAP) were assessed in a rat endocarditis model against Enterococcus faecalis, Enterococcus faecium (resistant to vancomycin and ampicillin), and Staphylococcus aureus. As a monotherapy, TZD for 5 days was not effective in a comparison with no-treatment controls, while DAP for 5 days was significantly effective against these bacteria. Step-down therapy (DAP for 3 days followed by TZD for 2 days) was as effective as DAP for 5 days and was comparable to 3 days of DAP plus ceftriaxone against all bacteria and to 3 days of DAP plus gentamicin against E. faecalis OG1RF.

Carbapenem resistance in Enterobacterales is a public health threat. Klebsiella pneumoniae carbapenemase (encoded by alleles of the bla_KPC family) is one of the most common transmissible carbapenem resistance mechanisms worldwide. The dissemination of bla_KPC historically has been associated with distinct K. pneumoniae lineages (clonal group 258 [CG258]), a particular plasmid family (pKpQIL), and a composite transposon (Tn4401). In the United Kingdom, bla_KPC has represented a large-scale, persistent management challenge for some hospitals, particularly in North West England. The dissemination of bla_KPC has evolved to be polyclonal and polyspecies, but the genetic mechanisms underpinning this evolution have not been elucidated in detail; this study used short-read whole-genome sequencing of 604 bla_KPC-positive isolates (Illumina) and long-read assembly (PacBio)/polishing (Illumina) of 21 isolates for characterization. We observed the dissemination of bla_KPC (predominantly bla_KPC-2; 573/604 [95%] isolates) across eight species and more than 100 known sequence types. Although there was some variation at the transposon level (mostly Tn4401a, 584/604 [97%] isolates; predominantly with ATTGA-ATTGA target site duplications, 465/604 [77%] isolates), bla_KPC spread appears to have been supported by highly fluid, modular exchange of larger genetic segments among plasmid populations dominated by IncFIB (580/604 isolates), IncFII (545/604 isolates), and IncR (252/604 isolates) replicons. The subset of reconstructed plasmid sequences (21 isolates, 77 plasmids) also highlighted modular exchange among non-bla_KPC and bla_KPC plasmids and the common presence of multiple replicons within bla_KPC plasmid structures (>60%). The substantial genomic plasticity observed has important implications for our understanding of the epidemiology of transmissible carbapenem resistance in Enterobacterales for the implementation of adequate surveillance approaches and for control.

Animal models of bacterial infection have been widely used to explore the in vivo activity of antibacterial drugs. These data are often submitted to the U.S. Food and Drug Administration to support human use in an investigational new drug application (IND). To better understand the range and scientific use of animal models in regulatory submissions, a database was created surveying recent pneumonia models submitted as part of IND application packages. The IND studies were compared to animal models of bacterial pneumonia published in the scientific literature over the same period of time. In this review, we analyze the key experimental design elements, such as animal species, immune status, pathogens selected, and route of administration, and study endpoints.

Nacubactam is a novel β-lactamase inhibitor with dual mechanisms of action as an inhibitor of serine β-lactamases (classes A and C and some class D) and an inhibitor of penicillin binding protein 2 in Enterobacteriaceae. The safety, tolerability, and pharmacokinetics of intravenous nacubactam were evaluated in single- and multiple-ascending-dose, placebo-controlled studies. Healthy participants received single ascending doses of nacubactam of 50 to 8,000 mg, multiple ascending doses of nacubactam of 1,000 to 4,000 mg every 8 h (q8h) for up to 7 days, or nacubactam of 2,000 mg plus meropenem of 2,000 mg q8h for 6 days after a 3-day lead-in period. Nacubactam was generally well tolerated, with the most frequently reported adverse events (AEs) being mild to moderate complications associated with intravenous access and headache. There was no apparent relationship between drug dose and the pattern, incidence, or severity of AEs. No clinically relevant dose-related trends were observed in laboratory safety test results. No serious AEs, dose-limiting AEs, or deaths were reported. After single or multiple doses, nacubactam pharmacokinetics appeared linear, and exposure increased in an approximately dose-proportional manner across the dose range investigated. Nacubactam was excreted largely unchanged into urine. Coadministration of nacubactam with meropenem did not significantly alter the pharmacokinetics of either drug. These findings support the continued clinical development of nacubactam and demonstrate the suitability of meropenem as a potential β-lactam partner for nacubactam. (The studies described in this paper have been registered at ClinicalTrials.gov under NCT02134834 [single ascending dose study] and NCT02972255 [multiple ascending dose study].)

Current treatments for Acanthamoeba keratitis rely on a combination of chlorhexidine gluconate, propamidine isethionate, and polyhexamethylene biguanide. These disinfectants are nonspecific and inherently toxic, which limits their effectiveness. Furthermore, in 10% of cases, recurrent infection ensues due to the difficulty in killing both trophozoites and double-walled cysts. Therefore, development of efficient, safe, and target-specific drugs which are capable of preventing recurrent Acanthamoeba infection is a critical unmet need for averting blindness. Since both trophozoites and cysts contain specific sets of membrane sterols, we hypothesized that antifungal drugs targeting sterol 14-demethylase (CYP51), known as conazoles, would have deleterious effects on A. castellanii trophozoites and cysts. To test this hypothesis, we first performed a systematic screen of the FDA-approved conazoles against A. castellanii trophozoites using a bioluminescence-based viability assay adapted and optimized for Acanthamoeba. The most potent drugs were then evaluated against cysts. Isavuconazole and posaconazole demonstrated low nanomolar potency against trophozoites of three clinical strains of A. castellanii. Furthermore, isavuconazole killed trophozoites within 24 h and suppressed excystment of preformed Acanthamoeba cysts into trophozoites. The rapid action of isavuconazole was also evident from the morphological changes at nanomolar drug concentrations causing rounding of trophozoites within 24 h of exposure. Given that isavuconazole has an excellent safety profile, is well tolerated in humans, and blocks A. castellanii excystation, this opens an opportunity for the cost-effective repurposing of isavuconazole for the treatment of primary and recurring Acanthamoeba keratitis.

The RESTORE-IMI 1 phase 3 trial demonstrated the efficacy and safety of imipenem-cilastatin (IMI) combined with relebactam (REL) for treating imipenem-nonsusceptible infections. The objective of this analysis was to compare the outcomes among patients meeting eligibility requirements based on central laboratory susceptibility versus local laboratory susceptibility. Patients with serious infections caused by imipenem-nonsusceptible, colistin-susceptible, and imipenem-REL-susceptible pathogens were randomized 2:1 to IMI-REL plus placebo or colistin plus IMI for 5 to 21 days. The primary endpoint was a favorable overall response. Key endpoints included the clinical response and all-cause mortality. We compared outcomes between the primary microbiological modified intent-to-treat (mMITT) population, where eligibility was based on central laboratory susceptibility testing, and the supplemental mMITT (SmMITT) population, where eligibility was based on local, site-level testing. The SmMITT (n = 41) and MITT (n = 31) populations had similar baseline characteristics, including sex, age, illness severity, and renal function. In both analysis populations, favorable overall response rates in the IMI-REL treatment group were >70%. Favorable clinical response rates at day 28 were 71.4% for IMI-REL and 40.0% for colistin plus IMI in the mMITT population, whereas they were 75.0% for IMI-REL and 53.8% for colistin plus IMI in the SmMITT population. Day 28 all-cause mortality rates were 9.5% for IMI-REL and 30.0% for colistin plus IMI in the mMITT population, whereas they were 10.7% for IMI-REL and 23.1% for colistin plus IMI in the SmMITT population. The outcomes in the SmMITT population were generally consistent with those in the mMITT population, suggesting that outcomes may be applicable to the real-world use of IMI-REL for treating infections caused by imipenem-nonsusceptible Gram-negative pathogens. (This study has been registered at ClinicalTrials.gov under identifier NCT02452047.)

Helicobacter pylori is an important risk factor for gastric ulcers. However, antibacterial therapies increase the resistance rate and decrease the eradication rate of H. pylori. Inspired by the microaerophilic characteristics of H. pylori, we aimed at effectively establishing an oxygen-enriched environment to eradicate and prevent the recurrence of H. pylori. The effect and the mechanism of an oxygen-enriched environment in eradicating H. pylori and preventing the recurrence were explored in vitro and in vivo. During oral administration and after drug withdrawal, H. pylori counts were evaluated by Giemsa staining in animal cohorts. An oxygen-enriched environment in which H. pylori could not survive was successfully established by adding hydrogen peroxide into several solutions and rabbit gastric juice. Hydrogen peroxide effectively killed H. pylori in Columbia blood agar and special peptone broth. Minimum inhibition concentrations and minimum bactericidal concentrations of hydrogen peroxide were both relatively stable after promotion of resistance for 30 generations, indicating that hydrogen peroxide did not easily promote resistance in H. pylori. In models of Mongolian gerbils and Kunming mice, hydrogen peroxide has been shown to significantly eradicate and effectively prevent the recurrence of H. pylori without toxicity and damage to the gastric mucosa. The mechanism of hydrogen peroxide causing H. pylori death was related to the disruption of bacterial cell membranes. The oxygen-enriched environment achieved by hydrogen peroxide eradicates and prevents the recurrence of H. pylori by damaging bacterial cell membranes. Hydrogen peroxide thus provides an attractive candidate for anti-H. pylori treatment.

The aim of this work was to evaluate the pharmacokinetics of amikacin in Mexican patients with different renal functions receiving once-daily dosing regimens and the influence of clinical and demographical covariates that may influence the optimization of this antibiotic. A prospective study was performed in a total of 63 patients with at least one determination of amikacin plasma concentration. Population pharmacokinetic (PK) parameters were estimated by nonlinear mixed-effects modeling; validations were performed for dosing recommendation purposes based on PK/pharmacodynamic simulations. The concentration-versus-time data were best described by a one-compartment open model with proportional interindividual variability associated with amikacin clearance (CL) and volume of distribution (V); residual error followed a homoscedastic trend. Creatinine clearance (CL_CR) and ideal body weight (IBW) demonstrated significant influence on amikacin CL and V, respectively. The final model [CL (liters/h) = 7.1 x (CL_CR/130)^0.84 and V (liters) = 20.3 x (IBW/68)^2.9] showed a mean prediction error of 0.11 mg/liter (95% confidence interval, –3.34, 3.55) in the validation performed in a different group of patients with similar characteristics. There is a wide variability in amikacin PK parameters in Mexican patients. This leads to inadequate dosing regimens, especially in patients with augmented renal clearance (CL_CR of >130 ml/min). Optimization based on the final population PK model in Mexican patients may be useful, since reliability and clinical applicability have been demonstrated in this study.

Etravirine (ETR) is a nonnucleoside reverse transcriptase inhibitor (NNRTI) used in treatment-experienced individuals. Genotypic resistance test-interpretation systems can predict ETR resistance; however, genotype-based algorithms are derived primarily from HIV-1 subtype B and may not accurately predict resistance in non-B subtypes. The frequency of ETR resistance among recombinant subtype C HIV-1 and the accuracy of genotypic interpretation systems were investigated. HIV-1_LAI containing full-length RT from HIV-1 subtype C-positive individuals experiencing virologic failure (>10,000 copies/ml and >1 NNRTI resistance-associated mutation) were phenotyped for ETR susceptibility. Fold change (FC) was calculated against a composite 50% effective concentration (EC₅₀) from treatment-naive individuals and three classifications were assigned: (i) <2.9-FC, susceptible; (ii) ≥2.9- to 10-FC, partially resistant; and (iii) >10-FC, fully resistant. The Stanford HIVdb-v8.4 was used for genotype predictions merging the susceptible/potential low-level and low-level/intermediate groups for 3 x 3 comparison. Fifty-four of a hundred samples had reduced ETR susceptibility (≥2.9-FC). The FC correlated with HIVdb-v8.4 (Spearman’s rho = 0.62; P < 0.0001); however, 44% of samples were partially (1 resistance classification difference) and 4% completely discordant (2 resistance classification differences). Of the 34 samples with an FC of >10, 26 were HIVdb-v8.4 classified as low-intermediate resistant. Mutations L100I, Y181C, or M230L were present in 27/34 (79%) of samples with an FC of >10 but only in 2/46 (4%) of samples with an FC of <2.9. No other mutations were associated with ETR resistance. Viruses containing the mutation K65R were associated with reduced ETR susceptibility, but 65R reversions did not increase ETR susceptibility. Therefore, genotypic interpretation systems were found to misclassify ETR susceptibility in HIV-1 subtype C samples. Modifications to genotypic algorithms are needed to improve the prediction of ETR resistance for the HIV-1 subtype C.

Plasmodium vivax relapse is one of the major causes of sustained global malaria transmission. Primaquine (PQ) is the only commercial drug available to prevent relapses, and its efficacy is dependent on metabolic activation by cytochrome P450 2D6 (CYP2D6). Impaired CYP2D6 function, caused by allelic polymorphisms, leads to the therapeutic failure of PQ as a radical cure for P. vivax malaria. Here, we hypothesized that the host immune response to malaria parasites modulates susceptibility to P. vivax recurrences in association with CYP2D6 activity. We performed a 10-year retrospective study by genotyping CYP2D6 polymorphisms in 261 malaria-exposed individuals from the Brazilian Amazon. The immune responses against a panel of P. vivax blood-stage antigens were evaluated by serological assays. We confirmed our previous findings, which indicated an association between impaired CYP2D6 activity and a higher risk of multiple episodes of P. vivax recurrence (risk ratio, 1.75; 95% confidence interval [CI], 1.2 to 2.6; P = 0.0035). An important finding was a reduction of 3% in the risk of recurrence (risk ratio, 0.97; 95% CI, 0.96 to 0.98; P < 0.0001) per year of malaria exposure, which was observed for individuals with both reduced and normal CYP2D6 activity. Accordingly, subjects with long-term malaria exposure and persistent antibody responses to various antigens showed fewer episodes of malaria recurrence. Our findings have direct implications for malaria control, since it was shown that nonimmune individuals who do not respond adequately to treatment due to reduced CYP2D6 activity may present a significant challenge for sustainable progress toward P. vivax malaria elimination.

This study was conducted in treatment-naive adults with drug-susceptible pulmonary tuberculosis in Port-au-Prince, Haiti, to assess the safety, bactericidal activity, and pharmacokinetics of nitazoxanide (NTZ). This was a prospective phase II clinical trial in 30 adults with pulmonary tuberculosis. Twenty participants received 1 g of NTZ orally twice daily for 14 days. A control group of 10 participants received standard therapy over 14 days. The primary outcome was the change in time to culture positivity (TTP) in an automated liquid culture system. The most common adverse events seen in the NTZ group were gastrointestinal complaints and headache. The mean change in TTP in sputum over 14 days in the NTZ group was 3.2 h ± 22.6 h and was not statistically significant (P = 0.56). The mean change in TTP in the standard therapy group was significantly increased, at 134 h ± 45.2 h (P < 0.0001). The mean NTZ MIC for Mycobacterium tuberculosis isolates was 12.3 μg/ml; the mean NTZ maximum concentration (C_max) in plasma was 10.2 μg/ml. Negligible NTZ levels were measured in sputum. At the doses used, NTZ did not show bactericidal activity against M. tuberculosis. Plasma concentrations of NTZ were below the MIC, and its negligible accumulation in pulmonary sites may explain the lack of bactericidal activity. (This study has been registered at ClinicalTrials.gov under identifier NCT02684240.)

We report a systematic, cellular phenotype-based antimalarial screening of the Medicines for Malaria Venture Pathogen Box collection, which facilitated the identification of specific blockers of late-stage intraerythrocytic development of Plasmodium falciparum. First, from standard growth inhibition assays, we identified 173 molecules with antimalarial activity (50% effective concentration [EC₅₀] ≤ 10 μM), which included 62 additional molecules over previously known antimalarial candidates from the Pathogen Box. We identified 90 molecules with EC₅₀ of ≤1 μM, which had significant effect on the ring-trophozoite transition, while 9 molecules inhibited the trophozoite-schizont transition and 21 molecules inhibited the schizont-ring transition (with ≥50% parasites failing to proceed to the next stage) at 1 μM. We therefore rescreened all 173 molecules and validated hits in microscopy to prioritize 12 hits as selective blockers of the schizont-ring transition. Seven of these molecules inhibited the calcium ionophore-induced egress of Toxoplasma gondii, a related apicomplexan parasite, suggesting that the inhibitors may be acting via a conserved mechanism which could be further exploited for target identification studies. We demonstrate that two molecules, MMV020670 and MMV026356, identified as schizont inhibitors in our screens, induce the fragmentation of DNA in merozoites, thereby impairing their ability to egress and invade. Further mechanistic studies would facilitate the therapeutic exploitation of these molecules as broadly active inhibitors targeting late-stage development and egress of apicomplexan parasites relevant to human health.

Currently, the expansion of the novel human respiratory coronavirus (known as SARS-CoV-2 [severe acute respiratory syndrome coronavirus 2], COVID-2019 [coronavirus disease 2019], or 2019-nCoV [2019 novel coronavirus]) has stressed the need for therapeutic alternatives to alleviate and stop this new epidemic. The previous epidemics of infections by high-morbidity human coronaviruses, such as SARS-CoV in 2003 and the Middle East respiratory syndrome coronavirus (MERS-CoV) in 2012, prompted the characterization of compounds that could be potentially active against the currently emerging novel coronavirus, SARS-CoV-2. The most promising compound is remdesivir (GS-5734), a nucleotide analog prodrug currently in clinical trials for treating Ebola virus infections. Remdesivir inhibited the replication of SARS-CoV and MERS-CoV in tissue cultures, and it displayed efficacy in nonhuman animal models. In addition, a combination of the human immunodeficiency virus type 1 (HIV-1) protease inhibitors lopinavir/ritonavir and interferon beta (LPV/RTV–IFN-β) was shown to be effective in patients infected with SARS-CoV. LPV/RTV–IFN-β also improved clinical parameters in marmosets and mice infected with MERS-CoV. Remarkably, the therapeutic efficacy of remdesivir appeared to be superior to that of LPV/RTV–IFN-β against MERS-CoV in a transgenic humanized mouse model. The relatively high mortality rates associated with these three novel human coronavirus infections, SARS-CoV, MERS-CoV, and SARS-CoV-2, have suggested that proinflammatory responses might play a role in the pathogenesis. It remains unknown whether the generated inflammatory state should be targeted. Therapeutics that target the coronavirus alone might not be able to reverse highly pathogenic infections. This minireview aims to provide a summary of therapeutic compounds that have shown potential in fighting SARS-CoV-2 infections.

To obtain the optimal dosage regimen in patients receiving extracorporeal membrane oxygenation (ECMO), we developed a population pharmacokinetics model for cefpirome and performed pharmacodynamic analyses. This prospective study included 15 patients treated with cefpirome during ECMO. Blood samples were collected during ECMO (ECMO-ON) and after ECMO (ECMO-OFF) at predose and 0.5 to 1, 2 to 3, 4 to 6, 8 to 10, and 12 h after cefpirome administration. The population pharmacokinetic model was developed using nonlinear mixed effects modeling and stepwise covariate modeling. Monte Carlo simulation was used to assess the probability of target attainment (PTA) and cumulative fraction of response (CFR) according to the MIC distribution. Cefpirome pharmacokinetics were best described by a two-compartment model. Covariate analysis indicated that serum creatinine concentration (SCr) was negatively correlated with clearance, and the presence of ECMO increased clearance and the central volume of distribution. The simulations showed that patients with low SCr during ECMO-ON had lower PTA than patients with high SCr during ECMO-OFF; so, a higher dosage of cefpirome was required. Cefpirome of 2 g every 8 h for intravenous bolus injection or 2 g every 12 h for extended infusion over 4 h was recommended with normal kidney function receiving ECMO. We established a population pharmacokinetic model for cefpirome in patients with ECMO, and appropriate cefpirome dosage regimens were recommended. The impact of ECMO could be due to the change in patient status on consideration of the small population and uncertainty in covariate relationships. Dose optimization of cefpirome may improve treatment success and survival in patients receiving ECMO. (This study has been registered at ClinicalTrials.gov under identifier NCT02581280.)

In India and China, indigenous drug manufacturers market arbitrarily combined parenteral β-lactam and β-lactamase inhibitors (BL-BLIs). In these fixed-dose combinations, sulbactam or tazobactam is indiscriminately combined with parenteral cephalosporins, with BLI doses kept in ratios similar to those for the approved BL-BLIs. Such combinations have been introduced into clinical practice without mandatory drug development studies involving pharmacokinetic/pharmacodynamic, safety, and efficacy assessments being undertaken. Such unorthodox combinations compromise clinical outcomes and also potentially contribute to resistance development.