A phase IIb, open-label, randomized controlled dose ranging multi-centre trial to evaluate the safety, tolerability, pharmacokinetics and exposure-response relationship of different doses of delpazolid in combination with bedaquiline delamanid moxifloxacin in adult subjects with newly diagnosed, uncomplicated, smear-positive, drug-sensitive pulmonary tuberculosis.

BACKGROUND: Linezolid is an effective, but toxic anti-tuberculosis drug that is currently recommended for the treatment of drug-resistant tuberculosis. Improved oxazolidinones should have a better safety profile, while preserving efficacy. Delpazolid is a novel oxazolidinone developed by LegoChem Biosciences Inc. that has been evaluated up to phase 2a clinical trials. Since oxazolidinone toxicity can occur late in treatment, LegoChem Biosciences Inc. and the PanACEA Consortium designed DECODE to be an innovative dose-ranging study with long-term follow-up for determining the exposure-response and exposure-toxicity relationship of delpazolid to support dose selection for later studies. Delpazolid is administered in combination with bedaquiline, delamanid and moxifloxacin. METHODS: Seventy-five participants with drug-sensitive, pulmonary tuberculosis will receive bedaquiline, delamanid and moxifloxacin, and will be randomized to delpazolid dosages of 0 mg, 400 mg, 800 mg, 1200 mg once daily, or 800 mg twice daily, for 16 weeks. The primary efficacy endpoint will be the rate of decline of bacterial load on treatment, measured by MGIT liquid culture time to detection from weekly sputum cultures. The primary safety endpoint will be the proportion of oxazolidinone class toxicities; neuropathy, myelosuppression, or tyramine pressor response. Participants who convert to negative liquid media culture by week 8 will stop treatment after the end of their 16-week course and will be observed for relapse until week 52. Participants who do not convert to negative culture will receive continuation phase treatment with rifampicin and isoniazid to complete a six-month treatment course. DISCUSSION: DECODE is an innovative dose-finding trial, designed to support exposure-response modelling for safe and effective dose selection. The trial design allows assessment of occurrence of late toxicities as observed with linezolid, which is necessary in clinical evaluation of novel oxazolidinones. The primary efficacy endpoint is the change in bacterial load, an endpoint conventionally used in shorter dose-finding trials. Long-term follow-up after shortened treatment is possible through a safety rule excluding slow-and non-responders from potentially poorly performing dosages. TRIAL REGISTRATION: DECODE was registered in ClinicalTrials.gov before recruitment start on 22 October 2021 (NCT04550832).

Evaluation of leucocytes from sputum samples of pulmonary tuberculosis patients using flow cytometry.

SETTING: Information about the sputum cells of pulmonary tuberculosis (PTB) patients is scarce. The analysis of sputum cells using optical microscopy (OM) is a well-established method, but it has some serious limitations. OBJECTIVE: To establish a new flow cytometry (FC) protocol for the leucocyte evaluation of sputum samples from PTB patients. DESIGN: A new FC protocol using 0.1% dithiothreitol and 0.5% paraformaldehyde was developed to fluidise sputum samples and kill Mycobacterium tuberculosis, respectively, to allow the analysis of sputum samples collected from TB patients. The protocol was validated by comparing it with OM, and the cellularity of 30 sputum samples from patients with PTB was evaluated. RESULTS: The comparison between leucocyte subsets analysed using OM and FC showed agreement. Immunophenotyping of leucocytes from sputum samples showed that neutrophils (95.7%) comprised the largest proportion of sputum cells, followed by monocytes/macrophages (2.6%) and lymphocytes (1.6%). Among the total T-lymphocytes (100%), 12.3% were T-helper cells, 24.1% were cytotoxic T-cells and 62.9% were gamma/delta T; none of the T lymphocytes had the CD4+/CD8+ phenotype. CONCLUSION: FC is a useful method for evaluating the different subtypes of leucocytes present in the sputum samples of PTB patients.

Alternative sputum preparation to improve polymerase chain reaction assay for Mycobacterium tuberculosis detection.

BACKGROUND: Tuberculosis (TB), one of the major airborne infectious bacterial diseases, remains an important health problem worldwide. It is estimated that there are 1700 new cases per year in Santa Catarina State, Brazil. OBJECTIVE: To improve polymerase chain reaction (PCR) sensitivity in detecting Mycobacterium tuberculosis in sputum samples. METHODS: This study proposed the use of glass beads as a modification of the routine protocol for sputum preparation used in the Laboratory of Molecular Biology and Mycobacteria at the Federal University of Santa Catarina, Florianópolis, Brazil. The study comprised 120 sputum samples, 60 of which were treated with the routine protocol, while 60 were treated with the modified protocol using glass beads. RESULTS: Samples treated with the routine protocol had a sensitivity of 56.7% (95%CI 44.1-69.2) in 16S rRNA PCR and 81.7% (95%CI 71.9-91.5) in insertion sequence (IS) 6110 PCR, compared with culture. Samples treated with the modified protocol had a sensitivity of 73.3% (95%CI 62.1-84.5) in 16S rRNA PCR and 100% in IS6110 PCR. CONCLUSION: The modified protocol using glass beads greatly improved mycobacterial detection in sputum samples compared with the routine protocol.