A pragmatic randomized controlled trial to evaluate the efficacy and safety of an oral short-course regimen including bedaquiline for the treatment of patients with multidrug-resistant tuberculosis in China: study protocol for PROSPECT.

INTRODUCTION: The lack of safe, effective, and simple short-course regimens (SCRs) for multidrug-resistant/rifampicin-resistant tuberculosis (MDR/RR-TB) treatment has significantly impeded TB control efforts in China. METHODS: This phase 4, randomized, open-label, controlled, non-inferiority trial aims to assess the efficacy and safety of a 9-month all-oral SCR containing bedaquiline (BDQ) versus an all-oral SCR without BDQ for adult MDR-TB patients (18-65 years) in China. The trial design mainly mirrors that of the "Evaluation of a Standardized Treatment Regimen of Anti-Tuberculosis Drugs for Patients with MDR-TB" (STREAM) stage 2 study, while also incorporating programmatic data from South Africa and the 2019 consensus recommendations of Chinese MDR/RR-TB treatment experts. Experimental arm participants will receive a modified STREAM regimen C that replaces three group C drugs, ethambutol (EMB), pyrazinamide (PZA), and prothionamide (PTO), with two group B drugs, linezolid (LZD) and cycloserine (CS), while omitting high-dose isoniazid (INH) for confirmed INH-resistant cases. BDQ duration will be extended from 6 to 9 months for participants with Mycobacterium tuberculosis-positive sputum cultures at week 16. The control arm will receive a modified STREAM regimen B without high-dose INH and injectable kanamycin (KM) that incorporates experimental arm LZD and CS dosages, treatment durations, and administration methods. LZD (600 mg) will be given daily for ≥ 24 weeks as guided by observed benefits and harm. The primary outcome measures the proportion of participants with favorable treatment outcomes at treatment completion (week 40), while the same measurement taken at 48 weeks post-treatment completion is the secondary outcome. Assuming an α = 0.025 significance level (one-sided test), 80% power, 15% non-inferiority margin, and 10% lost to follow-up rate, each arm requires 106 participants (212 total) to demonstrate non-inferiority. DISCUSSION: PROSPECT aims to assess the safety and efficacy of a BDQ-containing SCR MDR-TB treatment at seventeen sites across China, while also providing high-quality data to guide SCRs administration under the direction of the China National Tuberculosis Program for MDR-TB. Additionally, PROSPECT will explore the potential benefits of extending the administration of the 9-month BDQ-containing SCR for participants without sputum conversion by week 16. TRIAL REGISTRATION: ClinicalTrials.gov NCT05306223. Prospectively registered on 16 March 2022 at https://clinicaltrials.gov/ct2/show/NCT05306223?term=NCT05306223&draw=1&rank=1 {2}.

Antigen-specific chemokine profiles as biomarkers for detecting Mycobacterium tuberculosis infection.

Background: Latent tuberculosis (TB) infection can progress to active TB, which perpetuates community transmission that undermines global TB control efforts. Clinically, interferon-γ release assays (IGRAs) are commonly used for active TB case detection. However, low IGRA sensitivity rates lead to false-negative results for a high proportion of active TB cases, thus highlighting IGRA ineffectiveness in differentiating MTB-infected individuals from healthy individuals. Methods: Participants enrolled at Beijing Chest Hospital from May 2020-April 2022 were assigned to healthy control (HC), LTBI, IGRA-positive TB, and IGRA-negative TB groups. Screening cohort MTB antigen-specific blood plasma chemokine concentrations were measured using Luminex xMAP assays then were verified via testing of validation cohort samples. Results: A total of 302 individuals meeting study inclusion criteria were assigned to screening and validation cohorts. Testing revealed significant differences in blood plasma levels of CXCL9, CXCL10, CXCL16, CXCL21, CCL1, CCL19, CCL27, TNF-α, and IL-4 between IGRA-negative TB and HC groups. Levels of CXCL9, CXCL10, IL-2, and CCL8 biomarkers were predictive for active TB, as reflected by AUC values of ≥0.9. CXCL9-based enzyme-linked immunosorbent assay sensitivity and specificity rates were 95.9% (95%CI: 91.7-98.3) and 100.0% (92.7-100.0), respectively. Statistically similar AUC values were obtained for CXCL9 and CXCL9-CXCL10 assays, thus demonstrating that combined analysis of CXCL10 and CXCL9 levels did not improve active TB diagnostic performance. Conclusion: The MTB antigen stimulation-based CXCL9 assay may compensate for low IGRA diagnostic accuracy when used to diagnose IGRA-negative active TB cases and thus is an accurate and sensitive alternative to IGRAs for detecting MTB infection.

Multiple cytokine analysis based on QuantiFERON-TB gold plus in different tuberculosis infection status: an exploratory study.

BACKGROUND: More efficient and convenient diagnostic method is a desperate need to reduce the burden of tuberculosis (TB). This study explores the multiple cytokines secretion based on QuantiFERON-TB Gold Plus (QFT-Plus), and screens for optimal cytokines with diagnostic potential to differentiate TB infection status. METHODS: Twenty active tuberculosis (ATB) patients, fifteen patients with latent TB infection (LTBI), ten patients with previous TB and ten healthy controls (HC) were enrolled. Whole blood samples were collected and stimulated by QFT-Plus TB1 and TB2 antigens. The levels of IFN-γ, TNF-α, IL-2, IL-6, IL-5, IL-10, IP-10, IL-1Ra, CXCL-1 and MCP-1 in supernatant were measured by Luminex bead-based multiplex assays. The receiver operating characteristic curve was used to evaluate the diagnostic accuracy of cytokine for distinguishing different TB infection status. RESULTS: After stimulation with QFT-Plus TB1 and TB2 antigens, the levels of all cytokines, except IL-5 in TB2 tube, in ATB group were significantly higher than that in HC group. The levels of IL-1Ra concurrently showed the equally highest AUC for distinguishing TB infection from HC, followed by the levels of IP-10 in both TB1 tube and TB2 tube. Moreover, IP-10 levels displayed the largest AUC for distinguishing ATB patients from non-ATB patients. Meanwhile, the levels of IP-10 also demonstrated the largest AUC in both TB1 tube and TB2 tube for distinguishing ATB patients from LTBI. CONCLUSIONS: In addition to conventional detection of IFN-γ, measuring IP-10 and IL-1Ra based on QFT-Plus may have the more tremendous potential to discriminate different TB infection status.

From Bench to Clinic: A Nitroreductase Rv3368c-Responsive Cyanine-Based Probe for the Specific Detection of Live Mycobacterium tuberculosis.

Tuberculosis (TB), characterized by high mortality and low diagnosis, is caused by a single pathogen, Mycobacterium tuberculosis (Mtb). Imaging tools that can be used to track Mtb without pre-labeling and to diagnose live Mtb in clinical samples can shorten the gap between bench and clinic, fuel the development of novel anti-TB drugs, strengthen TB prevention, and improve patient treatment. In this study, we report an unprecedented novel nitroreductase-responsive cyanine-based fluorescent probe (Cy3-NO2-tre) that rapidly and specifically labels Mtb and detects it in clinical samples. Cy3-NO2-tre generated fluorescence after activation by a specific nitroreductase, Rv3368c, which is conserved in the Mycobacteriaceae. Cy3-NO2-tre effectively imaged mycobacteria within infected host cells, tracked the infection process, and visualized Mycobacterium smegmatis being endocytosed by macrophages. Cy3-NO2-tre also detected Mtb in the sputum of patients with TB and exhibited excellent photostability. Furthermore, the Cy3-NO2-tre/auramine O percentage change within 7 ± 2 days post drug treatment in the sputum of inpatients was closely correlated with the reexamination results of the chest computed tomography, strongly demonstrating the clinical application of Cy3-NO2-tre as a prognostic indicator in monitoring the therapeutic efficacy of anti-TB drugs in the early patient care stage.

Integrated bioinformatics analysis of dendritic cells hub genes reveal potential early tuberculosis diagnostic markers.

BACKGROUND: Dendritic cells (DCs) are most potent antigen-processing cells and play key roles in host defense against Mycobacterium tuberculosis (MTB) infection. In this study, hub genes in DCs during MTB infection were first investigated using bioinformatics approaches and further validated in Monocyte-derived DCs. METHODS: Microarray datasets were obtained from Gene Expression Omnibus (GEO) database. Principal component analysis (PCA) and immune infiltration analysis were performed to select suitable samples for further analysis. Differential analysis and functional enrichment analysis were conducted on DC samples, comparing live MTB-infected and non-infected (NI) groups. The CytoHubba plugin in Cytoscape was used to identify hub genes from the differentially expressed genes (DEGs). The expression of the hub genes was validated using two datasets and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) in human monocyte-derived DCs. Enzyme-linked immunosorbent assay (ELISA) was used to validate interferon (IFN) secretion. Transcription factors (TFs) and microRNAs (miRNAs) that interact with the hub genes were predicted using prediction databases. The diagnostic value of the hub genes was evaluated using receiver operating characteristic (ROC) curves and area under the curve (AUC) values. RESULTS: A total of 1835 common DEGs among three comparison groups (18 h, 48 h, 72 h after MTB infection) were identified. Six DEGs (IFIT1, IFIT2, IFIT3, ISG15, MX1, and RSAD2) were determined as hub genes. Functions enrichment analysis revealed that all hub genes all related to IFN response. RT-qPCR showed that the expression levels of six hub genes were significantly increased after DC stimulated by live MTB. According to the results of ELISA, the secretion of IFN-γ, but not IFN-α/ß, was upregulated in MTB-stimulated DCs. AUC values of six hub genes ranged from 84 to 94% and AUC values of 5 joint indicators of two hub genes were higher than the two hub genes alone. CONCLUSION: The study identified 6 hub genes associated with IFN response pathway. These genes may serve as potential diagnostic biomarkers in tuberculosis (TB). The findings provide insights into the molecular mechanisms involved in the host immune response to MTB infection and highlight the diagnostic potential of these hub genes in TB.

Spontaneous mutational patterns and novel mutations for bedaquiline and clofazimine resistance in Mycobacterium tuberculosis.

The 2022 World Health Organization guidelines recommend use of two core anti-tuberculosis (TB) drugs, bedaquiline (BDQ) and clofazimine (CFZ), for treatment of drug-resistant (DR)-TB. However, several mutated Mycobacterium tuberculosis (MTB) genes, conferring BDQ and CFZ resistance, have been reported that predominantly arose from sporadic mutations that have not been comprehensively characterized. Herein, MTB clinical isolates collected from drug-susceptible (DS)-, multidrug-resistant (MDR)-, and extensively drug-resistant (XDR)-TB patients were cultured in vitro with BDQ or CFZ to generate progeny strains with resistance to these drugs. Progeny strains exposed to CFZ exhibited increased CFZ minimum inhibitory concentrations (MICs) that exceeded MIC increases of BDQ-exposed progeny strains. Notably, mmpR and pepQ mutations accounted for 83% and 17% of BDQ-induced spontaneous gene mutations, respectively, and 86% and 14% of CFZ-induced spontaneous gene mutations, respectively. Analyses of predicted mutation-induced changes in amino acid sequences and structures of MmpR and PepQ mutants revealed several point mutations affected sequence conversation and functionality as an underlying mechanism for observed acquired BDQ/CFZ resistance. Moreover, our results revealed differences in patterns of BDQ- and CFZ-induced acquired spontaneous mutations that may enhance our understanding of MTB BDQ/CFZ-resistance mechanisms. IMPORTANCE This study of MTB drug resistance mechanisms revealed patterns of spontaneous MTB mutations associated with acquired BDQ and CFZ resistance that arose after clinical MTB isolates were cultured in vitro with BDQ or CFZ. Results of protein sequence and structural analyses provided insights into potential mechanisms underlying associations between MTB gene mutations and DR phenotypes. Taken together, these results revealed differences in acquired BDQ and CFZ resistance mechanisms as a new perspective that may enhance our understanding of BDQ/CFZ resistance mechanisms and facilitate the development of new methods for detecting MTB drug resistance genes.

DNA framework-engineered chimeras platform enables selectively targeted protein degradation.

A challenge in developing proteolysis targeting chimeras (PROTACs) is the establishment of a universal platform applicable in multiple scenarios for precise degradation of proteins of interest (POIs). Inspired by the addressability, programmability, and rigidity of DNA frameworks, we develop covalent DNA framework-based PROTACs (DbTACs), which can be synthesized in high-throughput via facile bioorthogonal chemistry and self-assembly. DNA tetrahedra are employed as templates and the spatial position of each atom is defined. Thus, by precisely locating ligands of POI and E3 ligase on the templates, ligand spacings can be controllably manipulated from 8 Å to 57 Å. We show that DbTACs with the optimal linker length between ligands achieve higher degradation rates and enhanced binding affinity. Bispecific DbTACs (bis-DbTACs) with trivalent ligand assembly enable multi-target depletion while maintaining highly selective degradation of protein subtypes. When employing various types of warheads (small molecules, antibodies, and DNA motifs), DbTACs exhibit robust efficacy in degrading diverse targets, including protein kinases and transcription factors located in different cellular compartments. Overall, utilizing modular DNA frameworks to conjugate substrates offers a universal platform that not only provides insight into general degrader design principles but also presents a promising strategy for guiding drug discovery.

A case report about a child with drug-resistant tuberculous meningitis.

BACKGROUND: Hematogenous disseminated tuberculosis predisposes to concurrent tuberculous meningitis (TBM), the most devastating and disabling form of tuberculosis. However, children often have atypical clinical symptoms, difficulty in specimen collection, low specimen content, and an increasing incidence of drug-resistant tuberculosis. Thus, the accurate diagnosis and timely treatment of childhood tuberculosis face monumental challenges. CASE PRESENTATION: The 14-year-old female presented to the hospital with intermittent fever, headache, and blurred vision. Her cerebrospinal fluid (CSF) showed a lymphocytic pleocytosis, an elevated protein level, and a decreased chloride level. And her CSF tested positive for TB-RNA. Xpert MTB/RIF detected Mycobacterium tuberculosis in her CSF, but the rifampin resistance test was unknown. Subsequently, her CSF culture was positive for Mycobacterium tuberculosis. The drug sensitivity test (DST) revealed resistance to isoniazid, rifampin, and fluoroquinolones. A computed tomography (CT) of the chest showed diffuse miliary nodules in both lungs. Intracranial enhanced magnetic resonance imaging (MRI) showed "multiple intensified images of the brain parenchyma, cisterns, and part of the meninges." The final diagnosis is miliary pulmonary tuberculosis and pre-extensive drug-resistant TBM. After 19 months of an oral, individualized antituberculosis treatment, she recovered with no significant neurological sequelae. CONCLUSION: For patients with miliary pulmonary tuberculosis, especially children, even if there are no typical clinical symptoms, it is necessary to know whether there is TBM and other conditions. Always look for the relevant aetiological basis to clarify whether it is drug-resistant tuberculosis. Only a rapid and accurate diagnosis and timely and effective treatment can improve the prognosis and reduce mortality and disability rates.

Clinical characteristics and predictive model of pulmonary tuberculosis patients with pulmonary fungal coinfection.

BACKGROUND: In clinical settings, pulmonary tuberculosis (PTB) patients were often found to have pulmonary fungal coinfection. This study aimed to assess the clinical characteristics of patients suffering from coinfection with TB and pulmonary fungal and construct a predictive model for evaluating the probability of pulmonary fungal coinfection in patients with pulmonary tuberculosis. METHODS: The present case-control study retrospectively collected information from 286 patients affected by PTB who received treatment from December 6,2016- December 6,2021 at Beijing Chest Hospital, Capital Medical University. As control subjects, patients with sex and address corresponding to those of the case subjects were included in the study in a ratio of 1:1. These 286 patients were randomly divided into the training and internal validation sets in a ratio of 3:1. Chi-square test and logistic regression analysis were performed for the training set, and a predictive model was developed using the selected predictors. Bootstrapping was performed for internal validation. RESULTS: Seven variables [illness course, pulmonary cavitation, broad-spectrum antibiotics use for at least 1 week, chemotherapy or immunosuppressants, surgery, bacterial pneumonia, and hypoproteinemia] were validated and used to develop a predictive model which showed good discrimination capability for both training set [area under the curve (AUC) = 0.860, 95% confidence interval (CI) = 0.811-0.909] and internal validation set (AUC = 0.884, 95% CI = 0.799-0.970). The calibration curves also showed that the probabilities predicted using the predictive model had satisfactory consistency with the actual probability for both training and internal validation sets. CONCLUSIONS: We developed a predictive model that can predict the probability of pulmonary fungal coinfection in pulmonary tuberculosis patients. It showed potential clinical utility.

Predominance of Escherichia-Shigella in Gut Microbiome and Its Potential Correlation with Elevated Level of Plasma Tumor Necrosis Factor Alpha in Patients with Tuberculous Meningitis.

Tuberculous meningitis (TBM), the most lethal and disabling form of tuberculosis (TB), may be related to gut microbiota composition, warranting further study. Here we systematically compared gut microbiota compositions and blood cytokine profiles of TBM patients, pulmonary TB patients, and healthy controls. Notably, the significant gut microbiota dysbiosis observed in TBM patients was associated with markedly high proportions of Escherichia-Shigella species as well as increased blood levels of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6). Next, we obtained a fecal bacterial isolate from a TBM patient and administered it via oral gavage to mice in order to develop a murine gut microbiota dysbiosis model for use in exploring mechanisms underlying the observed relationship between gut microbial dysbiosis and TBM. Thereafter, cells of commensal Escherichia coli (E. coli) were isolated and administered to model mice by gavage and then mice were inoculated with Mycobacterium tuberculosis (M. tuberculosis). Subsequently, these mice exhibited increased blood TNF-α levels accompanied by downregulated expression of tight junction protein claudin-5, increased brain tissue bacterial burden, and elevated central nervous system inflammation relative to corresponding indicators in controls administered PBS by gavage. Thus, our results demonstrated that a signature dysbiotic gut microbiome profile containing a high proportion of E. coli was potentially associated with an increased circulating TNF-α level in TBM patients. Collectively, these results suggest that modulation of dysbiotic gut microbiota holds promise as a new strategy for preventing or alleviating TBM. IMPORTANCE As the most severe form of tuberculosis, the pathogenesis of tuberculous meningitis (TBM) is still unclear. Gut microbiota dysbiosis plays an important role in a variety of central nervous system diseases. However, the relationship between gut microbiota and TBM has not been identified. In our study, significant dysbiosis in gut microbiota composition with a high proportion of E. coli and increased levels of TNF-α in plasma was noted in TBM patients. A commensal E. coli was isolated and shown to increase the plasma level of TNF-α and downregulate brain tight junction protein claudin-5 in the murine model. Gavage administration of E. coli aggravated the bacterial burden and increased the inflammatory responses in the central nervous system after M. tuberculosis infection. Dysbiosis of gut microbiota may be a promising therapeutic target and biomarker for TBM prevention or treatment.

Delamanid suppresses CXCL10 expression via regulation of JAK/STAT1 signaling and correlates with reduced inflammation in tuberculosis patients.

Background: Apart from bactericidal effects, anti-tuberculosis drugs can interfere with the host's immune system. In this study, we analyzed the role of delamanid (DLM), an inhibitor of mycolic acid synthesis of mycobacterial cell wall, on human macrophages. Methods: Based on a cohort of multidrug-resistant tuberculosis (MDR-TB) patients treated with DLM, the levels of C-reaction protein (CRP) and cytokines in the plasma were monitored using immunoturbidimetric assay and flow cytometry, respectively. We investigated the role of DLM on CXCL10 expression in U937 cell model using the following methods: cell viability assay, reverse transcription-quantitative polymerase chain reaction, enzyme linked immunosorbent assay, immunoblot, and transwell co-culture assay. Results: A total of 23 MDR-TB patients were included, comprising of 13 patients treated with optimized background therapeutic regimen (OBR) plus DLM regimen (OBR+DLM) and 10 patients treated with OBR plus placebo. DLM administration was associated with a significant reduce in circulating CRP level. Correspondingly, after treatment, the level of CXCL10 in patients treated with OBR+DLM was significantly lower than that with control. Using cell model, DLM dramatically suppressed CXCL10 expression, which majorly depended on inhibiting the JAK/STAT pathway, and impaired the migration of PBMCs. Conclusion: Our data firstly demonstrate that DLM suppresses CXCL10 expression via regulation of JAK2/STAT1 signaling and correlates with reduced inflammation in MDR-TB patients. DLM could be used as a potential drug for immunotherapy of patients with overactive immune response due to CXCL10.

Spontaneous Mutational Patterns and Novel Mutations for Delamanid Resistance in Mycobacterium tuberculosis.

Delamanid (DLM) and pretomanid (PTM) are recent additions to the anti-tuberculosis (TB) drug armamentarium, and they offer more effective options for drug-resistant TB treatment. In particular, DLM is included in Group C, which is recommended for use in longer multidrug-resistant (MDR)-TB regimens. Previous studies have shown that resistance to DLM/PTM is caused by mutations in the ddn, fgd1, fbiA, fbiB, fbiC, and fbiD genes, which are related to the F420-dependent bioactivation pathway. Herein, we conduct in vitro selection of DLM-resistant strains using clinical Mycobacterium tuberculosis (MTB) isolates with various drug resistance profiles. The spontaneous resistance frequency of drug-susceptible (DS) MTB (1.14 × 10-6 to 1.04 × 10-4) to DLM was similar to that of H37Rv (8.88 × 10-6 to 9.96 × 10-6) but higher than those of multidrug-resistant MTB (2.03 × 10-7 to 3.18 × 10-6) and extensively drug-resistant (XDR) MTB (4.67 × 10-8 to 3.60 × 10-6). Of the 100 independently selected DLM-resistant MTB mutants, 65% harbored mutations in genes associated with either DLM prodrug activation (ddn, 39.73%; fgd1, 16.44%) or the F420 biosynthetic pathway (fbiA, 16.44%; fbiB, 5.48%; fbiC, 21.92%). Of the 45 mutations we identified, 38 were not previously reported. A structure analysis revealed that several point mutations affected the ligand binding or structural stability of enzymes related to DLM resistance, which would block the enzyme activity required for prodrug activation. Our results elucidate the in vitro spontaneous DLM-resistance patterns of different clinical strains, which could improve the understanding of the causes of DLM resistance in clinical strains and of the effects on drug resistance of different mutations in genes that are related to the DLM activation pathway.

Significant difference in Th1/Th2 paradigm induced by tuberculosis-specific antigens between IGRA-positive and IGRA-negative patients.

False negative interferon-γ release assay (IGRA) results constitute the major dilemma for the diagnosis of tuberculosis (TB) infections. Herein, we conducted a cohort study to compare the host immunological response to TB-specific antigens between active TB patients with positive and negative IGRA results and control groups. A total of 274 laboratory-confirmed TB patients were included in our analysis, consisting of 221 were IGRA positive and 53 were IGRA negative. Patients with the elderly were identified as an independent risk factor for negative IGRA results. In addition, the elevated level of IL-4 and the decreased levels of IFN-γ, IL-2, IL-6, IL-1ß, and IL-12 in IGRA negative TB relative to IGRA positive TB group, demonstrating a significant difference in Th1/Th2 paradigm between two groups. The IFN-γ&IL-2 based assay could correctly identify 247 out of 307 MTB-infected individuals [271 TB patients and 36 individuals with latent TB infection (LTBI)], demonstrating a sensitivity of 80.5%. Then the IFN-γ and IL-4 were applied to distinguish healthy control and IGRA-negative group. When using the stepwise algorithm, the sensitivity for detecting Mycobacterium tuberculosis (MTB) infections was significantly increased from 80.5% to 89.6%. Additionally, patients with negative IGRA results had a conversion to culture-negative status longer than those with positive IGRA results. In conclusion, a stepwise algorithm outperforms IGRA assays to accurately identify MTB infections by the combination IFN-γ, IL-2, and IL-4. Further study is needed to evaluate the accuracy of our diagnostic algorithm in the LTBI population.

Development and Validation of a Nomogram for Prediction of QT Interval Prolongation in Patients Administered Bedaquiline-Containing Regimens in China: a Modeling Study.

Corrected QT duration (QTc) interval prolongation is the most frequent adverse event associated with bedaquiline (BDQ) use. It may affect the safety of antituberculosis therapy, which leads to the consequent demands of needing to monitor during therapy. Our objective was to establish and validate a prediction model for estimating the risk of QTc prolongation after initiation of BDQ-containing regimens to multidrug-resistant tuberculosis (MDR-TB) patients. We constructed an individualized nomogram model based on baseline demographic and clinical characteristics of each patient within a Chinese cohort during BDQ treatment. The generalizability of this model was further validated through use of externally acquired data obtained from Beijing Chest Hospital from 2019 to 2020. Overall, 1,215 and 165 patients were included in training and external validation cohorts, respectively, whereby during anti-TB drug treatment, QTc prolongation was observed in 273 (22.5%) and 29 (17.6%) patients within these respective cohorts, for whom QTc values were >500 ms in 86 (31.5%) and 10 (34.7%) patients, respectively. Next, a total of four Cox proportional hazards models were created and assessed; then, nomograms derived from the models were plotted based on independent predictors of clofazimine, baseline QTc interval, creatinine, extensive drug-resistance (XDR), moxifloxacin, levofloxacin, and sex. Nomogram analysis revealed concordance index values of 0.723 (95% confidence interval [CI], 0.695 to 0.750) for the training cohort and 0.710 (95% CI, 0.627 to 0.821) for the external validation cohort, thus indicating relatively fair agreement between predicted and observed probabilities of QTc prolongation occurrence based on data obtained during 8-week, 16-week, and 24-week anti-TB treatment of both cohorts. Taken together, results obtained using these models demonstrated that coadministration of clofazimine and abnormal baseline QTc interval significantly contributed to QTc prolongation development during MDR-TB patient treatment with a BDQ-containing anti-TB treatment regimen.

Case report: A 9-year systematic treatment failure of a pulmonary tuberculosis patient.

Objective: To explore the reasons of failure in a case of pulmonary tuberculosis (PTB) after 9 years systematic treatment. Methods: We extracted the patients' treatment history, drug susceptibility testing (DST), Computed tomography (CT) images, and sequenced the isolated strains by whole gene sequencing (WGS). Results: Although most results of the phenotypical DSTs were consistent with the genotype DST, the occurrence of gene resistance to amikacin (AMK), capreomycin (CAP), moxifloxacin (MFX) was earlier than the phenotypical DST. Based on the continuously reversed results of phenotypical DSTs, CT images in different stages and WGS, it can be confirmed that the patient was infected with two different strains of Mycobacterium tuberculosis (M.TB). Moreover, severe cavities may be another factor leading to treatment failure. Conclusion: Given the suggestive effect of genotype DST is earlier than the phenotypical DST, so genotype DST can play a better guiding role in patients with MDR-TB. Additionally, for patients who have not been cured for a long time, medication should be more cautious and the role of WGS in drug resistance surveillance should be fully utilized.

DNA Framework-Programmed Ligand Positioning to Modulate the Targeting Performance.

Effective targeting of nanomedicine is still an intricacy since unsatisfactory clinical trial feedback demonstrated their inadequate concentration at the desired area. However, the regulatory effect of ligand-modification patterns on the targeting effect has not been surveyed yet. Based on the superior spatial addressability of DNA frame structures, herein DNA tetrahedrons were used as templates for site-specific modification of targeting ligands. In this work, nanovectors with homogeneous ligand-modification patterns, including various valence of ligands and the precisely controlled distance between ligands at the nanoscale, were established for the first time. In vitro and in vivo targeting performance studies found that merely relying on the augment of the ligand quantity exhibited a confined promotion effect on the targeting efficiency. Notably, the space distance between ligands displayed a more important role in reforming the targeting effect, and the largest ligand distance (approximately 156.55 Å) pattern exhibited an optimal targeting effect and prominently cytostatic activity toward tumor cells. Generally, the survey of ligand-modification patterns on nanovectors provided a valid guidance to direct the optimization of nanomedicine.

A Peptidomic Approach to Identify Novel Antigen Biomarkers for the Diagnosis of Tuberculosis.

Background: Here, we conducted a peptidomic study in murine model to identify novel antigen biomarkers for the diagnosis of tuberculosis (TB) with improved performance. Methods: Four recombinant proteins, including Mycobacterium tuberculosis protein 32 (MPT32), Mycobacterium tuberculosis protein 64 (MPT64), culture filtrate protein 10 (CFP10), and phosphate ABC transporter substrate-binding lipoprotein (PstS1) were expressed and intravenously injected into BALB/c mice. The serum were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The concentrations of candidate peptides in serum of suspected TB patients were determined using competitive enzyme-linked immunosorbent assay. Results: A total of 65 peptides from 4 MTB precursor recombinant proteins were identified in mouse serum by LC-MS/MS, of which 5 peptides were selected as candidates for serological analysis. The concentrations of peptides MPT64-2, CFP10-2 and PstS1-2 in TB patients were significantly higher than those in non-TB patients. MPT64-2 exhibited the most promising sensitivity (81.4%), followed by PstS1-2 and CFP10-2. In addition, PstS1-2 had the highest specificity (93.3%), followed by CFP10-2 and MPT64-2. According to the area under the curve (AUC), MPT64-2 (AUC = 0.863), PstS1-2 (AUC = 0.812) and CFP10-2 (AUC = 0.809) exhibited better diagnostic validity. Conclusion: We develop an effective approach to identify new antigen biomarkers via LC-MS/MS-based peptidomics. Multiple peptides exhibit promising efficacy in diagnosis of active TB patients.

High Incidence of Psychiatric Disorders Associated with Cycloserine Treatment of Multidrug-Resistant Tuberculosis Patients: A Cohort Study in Beijing, China.

Objective: Cycloserine (CS) is a group B anti-tuberculosis (TB) drug endorsed by the World Health Organization (WHO) for the treatment of drug-resistant (DR)-TB. Despite CS widespread acceptance and known efficacy, the high potential of drug-associated psychiatric disorders is a major concern to multidrug-resistant (MDR)-TB patients. In this study, we investigated CS-associated psychiatric disorders in a cohort of MDR-TB patients in Beijing, China. Our aim was to determine psychiatric disorder prevalence rates and associated risk factors in this population. Methods: This MDR-TB cohort study was conducted at Beijing Chest Hospital between February 2018 and February 2021. All patients received individualized treatment regimens that included CS at some point during the full treatment course. Patient psychological status was assessed using the Symptom Checklist (SCL-90) questionnaire during the post-treatment follow-up period. Results: Two hundred and thirty-seven MDR-TB patients were included in the final analysis. Overall, psychiatric disorders were recorded in 22 (9.28%) of the 237 patients in our cohort, with severity grades of 1 or 2 observed for the majority (8.44%) of psychiatric adverse events. As compared to results obtained for control group patients who were ≥40 years of age, patients who were <40 years of age (77.3%, 17/22) were more likely to experience psychiatric disorders [adjusted odds ratio (aOR) = 3.060; 95% CI (1.060-8.828)]. Additionally, patients with body mass index (BMI)s of <18.5 kg/m2 [aOR = 3.824; 95% CI (1.502-9.739)] had significantly higher odds of being afflicted with psychiatric disorders as compared with patients with BMIs that were ≥18.5 kg/m2. Conclusion: Our results demonstrated that approximately one-tenth of Chinese MDR-TB patients experienced psychiatric disorders during CS treatment, with the majority of adverse events of moderate severity. In addition, low BMI and age <40 years were identified as independent risk factors for psychiatric disorders in MDR-TB patients receiving CS therapy.

Increased Tuberculosis Incidence Due to Immunotherapy Based on PD-1 and PD-L1 Blockade: A Systematic Review and Meta-Analysis.

Objectives: In this study, we conducted a systematic review to determine tuberculosis (TB) incidence due to immunotherapy with programmed cell death protein-1 (PD-1)/PD ligand (PD-L1) blockade in cancer patients. Methods: We searched PubMed, Cochrance Library, Excerpt Medica Database (Embase), ClinicalTrials.gov, Chinese BioMedical Literature Database (CBM), China National Knowledge Infrastructure Database (CNKI), Wanfang and China Science and Technology Journal Database to identify studies between January 1, 2000 and April 30, 2021, on the reports of TB cases in patients treated with PD-1/PD-L1 blockade. Methodological quality of eligible studies was assessed, and random-effect model meta-analysis was performed to generate the pooled incidence estimate of TB cases in patients undergoing PD-1/PD-L1 therapy. Results: We initially identified 745 records, of which 27 studies ultimately met the inclusion criteria and were included in our meta-analysis. A total of 35 TB cases occurred among patients treated with PD-1/PD-L1 blockade. Nivolumab (51.4%) was the most frequently used PD-1/PD-L1 blockade for cancer treatment. In addition, pulmonary TB was the most common form of tuberculosis seen in 77.1% cases. Clinical outcomes were recorded in 18 patients, of whom 77.8% were cured or achieved remission, and 22.2% were died of TB. Pooled analysis determined that the TB rate in this population was 2,000 cases per 100,000 persons, and the estimated rate for TB associated with PD-1/PD-L1 blockade was 35 times higher than that in the general population. Conclusion: To conclude, our results demonstrate that the clinical use of PD-1/PD-L1 inhibitors significantly increases risk of TB reactivation. An extremely high mortality rate due to TB disease is noted in the patients with PD-1/PD-L1 blockade.

Comparison of diagnostic accuracy of QuantiFERON-TB Gold Plus and T-SPOT.TB in the diagnosis of active tuberculosis in febrile patients.

OBJECTIVE: This study aimed to compare the accuracy of QuantiFERON-TB Gold Plus (QFT-Plus) and T-SPOT.TB for diagnosing active tuberculosis (ATB) in febrile patients, to explore influencing factors of positive results and to verify the potential value of QFT-Plus in the identification of ATB and latent tuberculosis infection (LTBI). METHODS: A total of 240 febrile patients with ATB (n = 80) and non-ATB (n = 160) were recruited to assess the accuracy of QFT-Plus and T-SPOT.TB for diagnosing ATB. Multivariable logistic regression was used to analyze the influencing factors of positive results. RESULTS: The proportion of indeterminate results (ITRS) in QFT-Plus and T-SPOT.TB were 3.3% and 0%, respectively. The consistency between the results of the QFT-Plus and T-SPOT.TB was substantial. The area under the receiver operating characteristic curve (AUROC) of the QFT-Plus and T-SPOT.TB for diagnosing ATB was 0.792 and 0.849 (p = 0.070), respectively. The sensitivity of differentiating ATB from non-ATB was 92.2% in QFT-Plus versus 95.0% in T-SPOT.TB. The influencing factors of T-SPOT.TB positive result were male (odds ratio (OR) = 2.33, 95% confidence interval (CI) 1.27-4.26, p = 0.006), evidence of previous TB (OR 11.36, 95% CI 4.62-27.94, p < 0.001), while male (OR = 3.17, 95% CI 1.73-5.84, p < 0.001), evidence of previous TB (OR = 7.58, 95% CI 3.60-15.98, p <0.001), and use of immunosuppressant (OR = 0.49, 95% CI 0.260.94, p = 0.030) were influencing factors for QFT-Plus positive result. There was no significant difference in QFT-Plus in differentiating ATB from LTBI in febrile patients. CONCLUSION: There was no significant difference between QFT-Plus and T-SPOT.TB for diagnosing ATB in febrile patients. QFT-Plus is prone to ITRS. The influencing factors including males, evidence of the previous TB, and use of immunosuppressant should be considered when interpreting positive results.