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Remodeling the matrix: doxycycline modulates tuberculosis immunopathology
Jason D. Simmons and Thomas R. Hawn
Department of Medicine, University of Washington School of Medicine, Seattle, Washington, USA.
Address correspondence to: Thomas R. Hawn, University of Washington, 1959 NE Pacific Street, Seattle, Washington 98115, USA. Phone: 206.616.4124; Email: thawn@uw.edu.
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Department of Medicine, University of Washington School of Medicine, Seattle, Washington, USA.
Address correspondence to: Thomas R. Hawn, University of Washington, 1959 NE Pacific Street, Seattle, Washington 98115, USA. Phone: 206.616.4124; Email: thawn@uw.edu.
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Published August 2, 2021 - More info
J Clin Invest. 2021;131(15):e151668. https://doi.org/10.1172/JCI151668.
© 2021 American Society for Clinical Investigation
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Abstract
BACKGROUND Matrix metalloproteinases (MMPs) are key regulators of tissue destruction in tuberculosis (TB) and may be targets for host-directed therapy. We conducted a phase II double-blind, randomized, controlled trial investigating doxycycline, a licensed broad-spectrum MMP inhibitor, in patients with pulmonary TB.METHODS Thirty patients with pulmonary TB were enrolled within 7 days of initiating anti-TB treatment and randomly assigned to receive either 100 mg doxycycline or placebo twice a day for 14 days, in addition to standard care.RESULTS Whole blood RNA-sequencing demonstrated that doxycycline accelerated restoration of dysregulated gene expression in TB towards normality, rapidly down-regulating type I and II interferon and innate immune response genes, and up-regulating B-cell modules relative to placebo. The effects persisted for 6 weeks after doxycycline discontinuation, concurrent with suppressed plasma MMP-1. Doxycycline significantly reduced sputum MMP-1, -8, -9, -12 and -13, suppressed type I collagen and elastin destruction, reduced pulmonary cavity volume without altering sputum mycobacterial loads, and was safe.CONCLUSION Adjunctive doxycycline with standard anti-TB treatment suppressed pathological MMPs in PTB patients. Larger studies on adjunctive doxycycline to limit TB immunopathology are merited.TRIAL REGISTRATION ClinicalTrials.gov NCT02774993.FUNDING Singapore National Medical Research Council (NMRC/CNIG/1120/2014, NMRC/Seedfunding/0010/2014, NMRC/CISSP/2015/009a); the Singapore Infectious Diseases Initiative (SIDI/2013/013); National University Health System (PFFR-28 January 14, NUHSRO/2014/039/BSL3-SeedFunding/Jul/01); the Singapore Immunology Network Immunomonitoring platform (BMRC/IAF/311006, H16/99/b0/011, NRF2017_SISFP09); an ExxonMobil Research Fellowship, NUHS Clinician Scientist Program (NMRC/TA/0042/2015, CSAINV17nov014); the UK Medical Research Council (MR/P023754/1, MR/N006631/1); a NUS Postdoctoral Fellowship (NUHSRO/2017/073/PDF/03); The Royal Society Challenge Grant (CHG\R1\170084); the Sir Henry Dale Fellowship, Wellcome Trust (109377/Z/15/Z); and A*STAR.
Authors
Qing Hao Miow, Andres F. Vallejo, Yu Wang, Jia Mei Hong, Chen Bai, Felicia S.W. Teo, Alvin D.Y. Wang, Hong Rong Loh, Tuan Zea Tan, Ying Ding, Hoi Wah She, Suay Hong Gan, Nicholas I. Paton, Josephine Lum, Alicia Tay, Cynthia B.E. Chee, Paul A. Tambyah, Marta E. Polak, Yee Tang Wang, Amit Singhal, Paul T. Elkington, Jon S. Friedland, Catherine W.M. Ong
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Doxycycline modulates the host Mtb response
Doxycycline is broadly recognized to have antiinflammatory properties, accounting for its efficacy in treating dermatologic diseases, such as bullous pemphigus (18) and acne, and granulomatous diseases, such as rosacea and sarcoidosis (19). Doxycycline is effective for these dermatologic diseases at subantimicrobial (less than 50 mg daily) doses that are also used to treat periodontal diseases where efficacy correlates with the inhibition of collagenase MMPs (20). Doxycycline has pleiotropic effects on host inflammatory cytokines, limits MMP-1 and MMP-3 secretion from Mtb-infected human monocyte–derived macrophages, and reduces collagenase activity of Mtb-infected neutrophil supernatants (21). In respiratory epithelial cells, doxycycline inhibits MMP-1 promoter activity (22), suggesting it may modulate host transcription as opposed to direct inhibition of enzymatic function.
In this issue of the JCI, Hao Miow et al. report results on a randomized, double-blind, placebo-controlled trial measuring doxycycline host-directed activity in 30 patients with pulmonary TB (Figure 1) (23). The majority (63%) of patients had pulmonary cavities and were positive for Mycobacteria in sputum tests (median Acid-Fast Bacillus 2+) at baseline. Doxycycline or placebo was added within 7 days of starting standard ATT and continued for 14 days only. Using coexpression network analysis of whole blood transcriptional profiles, gene clusters that specifically were downregulated with doxycycline treatment included the innate immune response, Type I and Type II interferon signaling. Transcripts from these gene clusters, including IRF1, APOL1, FCGR1A, and GBP6, which are upregulated by Mtb infection, remained elevated during ATT/placebo treatment but were markedly downregulated in the ATT/doxycycline group at day 56. This occurred 42 days after completion of doxycycline and suggests either durable host-directed activity or pronounced early effects that shape the late immune response (23).
Figure 1The effect of doxycycline on pulmonary TB when added to standard therapy. (i) Aerosolized Mtb is initially contained within lung granulomas where it can remain quiescent or cause progressive pulmonary disease with unrestricted bacterial replication. (ii) In some cases, tissue destruction leads to pulmonary cavitation, which is associated with high mycobacterial burden and poor drug penetration, increasing the risk for transmission and emergent drug resistance. Mtb infection induces MMPs, which regulate inflammatory processes that range from tissue remodeling to tissue destruction (25) and are linked to granuloma maintenance and Mtb dissemination (10–12). MMPs also degrade elastin and collagen, contributing to cavitation (13). (iii) Hao Miow et al. explored the effects of doxycycline when added to standard antituberculous therapy in a randomized, placebo-controlled trial of patients with newly diagnosed pulmonary TB (23). Relative to healthy controls, multiple inflammatory gene programs were enriched among pulmonary TB subjects including coexpression modules related to extracellular matrix (ECM) genes. Longitudinal analysis of blood transcriptomes indicated doxycycline reduced transcription of MMP9 and genes related to interferon (IFN) signaling and the innate immune response. These findings mirrored lower levels of MMP proteins in serum (iv) and and lower MMP-1, -8, -9, -12, -13 levels in sputum (ii). This correlated with decreased elastase (potentially including MMP-9 and -12) and collagenase (potentially including MMP-1, -8, -13) enzyme activities in sputum. While the study was underpowered to detect more relevant clinical or microbiologic outcomes, the broad MMP effects of doxycycline correlated with reduced lung cavity volumes (v) at two months, which is notable considering doxycycline was dosed only during the initial two weeks.
To explore early effects of doxycycline that may shape these later transcriptome changes, Hao Miow and colleagues next measured MMP transcript and protein levels in blood and sputum. After 14 days of treatment, decreased MMP9 transcripts and MMP-1 protein levels were measured in participants who received doxycycline, which were similar to the reduced MMP-1 and MMP-8 levels and collagenase activity measured in sputum. Importantly, the reduced MMP levels detected early following doxycycline treatment are unlikely to simply reflect nonspecific transcriptional effects, since they correlated tightly with measures of immunopathology. In the 26 patients who completed follow-up (13 in each arm), doxycycline treatment resulted in a notable reduction in pulmonary cavity volume (23). These doxycycline effects are consistent with other broad MMP inhibitors, such as marimastat and BB-94, that reduced pulmonary pathology in the murine model (10, 24). Together, these experimental medicine data suggest a potential benefit of doxycycline with a host pathway–dependent mechanism.
Degradation of extracellular matrix provides a niche for extracellular Mtb growth where ATT delivery is impaired (25), which may in part explain improved microbiologic outcomes following treatment with MMP inhibitors in preclinical pulmonary TB models (10, 22, 24). However, a direct antimicrobial effect of doxycycline against Mtb cannot be ruled out. In broth culture, doxycycline was bacteriostatic (22) at concentrations (MIC 2.5 μg/mL) that are within peak serum levels measured previously after standard dosing (26). Doxycycline is used as second-line therapy for M. kansasii and M. fortuitum infections, and the related tetracycline minocycline is effective for M. leprae. No statistically significant microbiologic outcomes differed between groups in the trial by Hao Miow et al., although a trend toward lower median sputum bacterial titers was measured in participants who received doxycycline versus placebo (25 versus 100 CFU/mL, P = 0.32) (23).
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