Biomarkers of Tuberculosis Severity and Treatment Effect: A Directed Screen of 70 Host Markers in a Randomized Clinical Trial

Abstract

More efficacious treatment regimens are needed for tuberculosis, however, drug development is impeded by a lack of reliable biomarkers of disease severity and of treatment effect. We conducted a directed screen of host biomarkers in participants enrolled in a tuberculosis clinical trial to address this need. Serum samples from 319 protocol-correct, culture-confirmed pulmonary tuberculosis patients treated under direct observation as part of an international, phase 2 trial were screened for 70 markers of infection, inflammation, and metabolism. Biomarker assays were specifically developed for this study and quantified using a novel, multiplexed electrochemiluminescence assay. We evaluated the association of biomarkers with baseline characteristics, as well as with detailed microbiologic data, using Bonferroni-adjusted, linear regression models. Across numerous analyses, seven proteins, SAA1, PCT, IL-1β, IL-6, CRP, PTX-3 and MMP-8, showed recurring strong associations with markers of baseline disease severity, smear grade and cavitation; were strongly modulated by tuberculosis treatment; and had responses that were greater for patients who culture-converted at 8weeks. With treatment, all proteins decreased, except for osteocalcin, MCP-1 and MCP-4, which significantly increased. Several previously reported putative tuberculosis-associated biomarkers (HOMX1, neopterin, and cathelicidin) were not significantly associated with treatment response. In conclusion, across a geographically diverse and large population of tuberculosis patients enrolled in a clinical trial, several previously reported putative biomarkers were not significantly associated with treatment response, however, seven proteins had recurring strong associations with baseline radiographic and microbiologic measures of disease severity, as well as with early treatment response, deserving additional study.

Keywords: Biomarkers; Clinical trials; Host immune response; Tuberculosis.

Fig. 1

Box and whisker plots showing the baseline levels of six biomarkers as a function of five indicators of baseline disease state: presence of cavities in chest x-ray, presence of cavities > 4 cm, lung involvement > 50%, smear grade (≤ 1 vs ≥ 2) and time-to-detection (≤ 5 days vs > 5 days) for MGIT cultures. The asterisks indicate that the difference between two groups is significant (p < 0.05/62 assays = 0.0008).

Fig. 2

Effect of TB treatment on biomarker levels. The plot shows the ratio of post-treatment (week 8) to baseline (week 0) biomarker levels for each biomarker in each study subject. The assays are ordered based on the magnitude of the treatment effect. Asterisks indicate the effect is statistically significant (p < 0.05/62 assays = 0.0008).

Fig. 3

Association of biomarker treatment effect with sputum culture conversion status at week 8. The plot shows the ratio of post-treatment (week 8) to baseline (week 0) biomarker levels for the 25 biomarkers with the strongest treatment effects (the top 25 assays in Supplemental Table 2). Points are separated and colored based on sputum culture conversion status at week 8 (blue = converted, red = non-converted). For each biomarker and group, a horizontal line segment indicates the median biomarker value. The biomarkers with a significant association with culture status (p < 0.05/62 biomarkers = 0.0008) are marked with an asterisk.

Fig. 4

Box and whisker plots showing the magnitude of the treatment effect for six biomarkers (the ratio of week 8 and week 0 biomarker levels) after grouping subjects according to three outcome variables: week 8 culture conversion status, week 12 culture conversion status, and presence/absence of cough at 8 weeks. The asterisks indicate that the difference between two groups is significant (p < 0.05/62 assays = 0.0008).