Sputum lipoarabinomannan (LAM) as a biomarker to determine sputum mycobacterial load: exploratory and model-based analyses of integrated data from four cohorts.

BACKGROUND: Despite the high global disease burden of tuberculosis (TB), the disease caused by Mycobacterium tuberculosis (Mtb) infection, novel treatments remain an urgent medical need. Development efforts continue to be hampered by the reliance on culture-based methods, which often take weeks to obtain due to the slow growth rate of Mtb. The availability of a "real-time" measure of treatment efficacy could accelerate TB drug development. Sputum lipoarabinomannan (LAM; an Mtb cell wall glycolipid) has promise as a pharmacodynamic biomarker of mycobacterial sputum load. METHODS: The present analysis evaluates LAM as a surrogate for Mtb burden in the sputum samples from 4 cohorts of a total of 776 participants. These include those from 2 cohorts of 558 non-TB and TB participants prior to the initiation of treatment (558 sputum samples), 1 cohort of 178 TB patients under a 14-day bactericidal activity trial with various mono- or multi-TB drug therapies, and 1 cohort of 40 TB patients with data from the first 56-day treatment of a standard 4-drug regimen. RESULTS: Regression analysis demonstrated that LAM was a predictor of colony-forming unit (CFU)/mL values obtained from the 14-day treatment cohort, with well-estimated model parameters (relative standard error ≤ 22.2%). Moreover, no changes in the relationship between LAM and CFU/mL were observed across the different treatments, suggesting that sputum LAM can be used to reasonably estimate the CFU/mL in the presence of treatment. The integrated analysis showed that sputum LAM also appears to be as good a predictor of time to Mycobacteria Growth Incubator Tube (MGIT) positivity as CFU/mL. As a binary readout, sputum LAM positivity is a strong predictor of solid media or MGIT culture positivity with an area-under-the-curve value of 0.979 and 0.976, respectively, from receiver-operator curve analysis. CONCLUSIONS: Our results indicate that sputum LAM performs as a pharmacodynamic biomarker for rapid measurement of Mtb burden in sputum, and thereby may enable more efficient early phase clinical trial designs (e.g., adaptive designs) to compare candidate anti-TB regimens and streamline dose selection for use in pivotal trials. Trial registration NexGen EBA study (NCT02371681).

Optimising the yield from bronchoalveolar lavage on human participants in infectious disease immunology research.

Bronchoalveolar lavage (BAL) is becoming a common procedure for research into infectious disease immunology. Little is known about the clinical factors which influence the main outcomes of the procedure. In research participants who underwent BAL according to guidelines, the BAL volume yield, and cell yield, concentration, viability, pellet colour and differential count were analysed for association with important participant characteristics such as active tuberculosis (TB) disease, TB exposure, HIV infection and recent SARS-CoV-2 infection. In 337 participants, BAL volume and BAL cell count were correlated in those with active TB disease, and current smokers. The right middle lobe yielded the highest volume. BAL cell and volume yields were lower in older participants, who also had more neutrophils. Current smokers yielded lower volumes and higher numbers of all cell types, and usually had a black pellet. Active TB disease was associated with higher cell yields, and higher proportions of granulocytes, but this declined at the end of treatment. HIV infection was associated with lower cell yields and more bloody pellets, and recent SARS-CoV-2 infection with a higher proportion of lymphocytes. These results allow researchers to optimise their participant and end assay selection for projects involving lung immune cells.

Optimising the yield from bronchoalveolar lavage on human participants in infectious disease immunology research.

Bronchoalveolar lavage (BAL) is becoming a common procedure for research into infectious disease immunology. Little is known about the clinical factors which influence the main outcomes of the procedure. In research participants who underwent BAL according to guidelines, the BAL volume yield, and cell yield, concentration, viability, pellet colour and differential count were analysed for association with important participant characteristics such as active tuberculosis (TB) disease, TB exposure, HIV infection and recent SARS-CoV-2 infection. In 337 participants, BAL volume and BAL cell count were correlated in those with active TB disease, and current smokers. The right middle lobe yielded the highest volume. BAL cell and volume yields were lower in older participants, who also had more neutrophils. Current smokers yielded lower volumes and higher numbers of all cell types, and usually had a black pellet. Active TB disease was associated with higher cell yields, but this declined at the end of treatment. HIV infection was associated with more bloody pellets, and recent SARS-CoV-2 infection with a higher proportion of lymphocytes. These results allow researchers to optimise their participant and end assay selection for projects involving lung immune cells.

Safety and immunogenicity of VPM1002 versus BCG in South African newborn babies: a randomised, phase 2 non-inferiority double-blind controlled trial.

BACKGROUND: Tuberculosis is a major public health problem worldwide. Immunisation with Mycobacterium bovis BCG vaccine is partially effective in infants, reducing the incidence of miliary and tuberculosis meningitis, but is less effective against pulmonary tuberculosis. We aimed to compare safety and immunogenicity of VPM1002-a recombinant BCG vaccine developed to address this gap-with BCG in HIV exposed and HIV unexposed newborn babies. METHODS: This double-blind, randomised, active controlled phase 2 study was conducted at four health centres in South Africa. Eligible neonates were aged 12 days or younger with a birthweight of 2·5-4·2 kg, and could be HIV exposed (seropositive mothers) or unexposed (seronegative mothers). Newborn babies were excluded if they had acute or chronic illness, fever, hypothermia, sepsis, cancer, or congenital malformation, or if they received blood products or immunosuppressive therapy. Participants were excluded if their mothers (aged ≥18 years) had active tuberculosis disease, diabetes, a history of immunodeficiency except for HIV, hepatitis B or syphilis seropositivity, received blood products in the preceding 6 months, any acute infectious disease, or any suspected substance abuse. Participants were randomly assigned to VPM1002 or BCG vaccination in a 3:1 ratio, stratified by HIV status using the random number generator function in SAS, using a block size of eight paticipants. The primary outcome was non-inferiority (margin 15%) of VPM1002 to BCG vaccine in terms of incidence of grade 3-4 adverse drug reactions or ipsilateral or generalised lymphadenopathy of 10 mm or greater in diameter by 12 months. The primary outcome was assessed in all vaccinated participants (safety population) at regular follow-up visits until 12 months after vaccination. Secondary immunogenicity outcomes were interferon-γ levels and percentages of multifunctional CD4+ and CD8+ T cells among all lymphocytes across the 12 month study period. The study was registered with ClinicalTrials.gov, NCT02391415. FINDINGS: Between June 4, 2015 and Oct 16, 2017, 416 eligible newborn babies were randomly assigned and received study vaccine. Seven (2%) of 312 participants in the VPM1002 group had a grade 3-4 vaccine-related adverse reaction or lymphadenopathy of 10 mm or greater in diameter compared with 34 (33%) of 104 participants in the BCG group (risk difference -30·45% [95% CI -39·61% to -21·28%]; pnon-inferiority<0·0001); VPM1002 was thus non-inferior to BCG for the primary outcome. Incidence of severe injection site reactions was lower with VPM1002 than BCG: scarring occurred in 65 (21%) participants in the VPM1002 group versus 77 (74%) participants in the BCG group (p<0·0001); ulceration occurred in one (<1%) versus 15 (14%; p<0·0001); and abscess formation occurred in five (2%) versus 23 (22%; p<0·0001). Restimulated IFNγ concentrations were lower in the VPM1002 group than the BCG group at week 6, week 12, month 6, and month 12. The percentage of multifunctional CD4+ T cells was higher in the VPM1002 group than the BCG group at day 14 but lower at week 6, week 12, month 6, and month 12. The percentage of multifunctional CD8+ T cells was lower in the VPM1002 group than the BCG group at week 6, week 12, and month 6, but did not differ at other timepoints. INTERPRETATION: VPM1002 was less reactogenic than BCG and was not associated with any serious safety concern. Both vaccines were immunogenic, although responses were higher with the BCG vaccine. VPM1002 is currently being studied for efficacy and safety in a multicentric phase 3 clinical trial in babies in sub-Saharan Africa. FUNDING: Serum Institute of India.