Research Article: Efficacy of Rifampin Plus Clofazimine in a Murine Model of Mycobacterium ulcerans Disease

Date Published: June 4, 2015

Publisher: Public Library of Science

Author(s): Paul J. Converse, Sandeep Tyagi, Yalan Xing, Si-Yang Li, Yoshito Kishi, John Adamson, Eric L. Nuermberger, Jacques H. Grosset, Richard O. Phillips.

Abstract: Treatment of Buruli ulcer, or Mycobacterium ulcerans disease, has shifted from surgical excision and skin grafting to antibiotic therapy usually with 8 weeks of daily rifampin (RIF) and streptomycin (STR). Although the results have been highly favorable, administration of STR requires intramuscular injection and carries the risk of side effects, such as hearing loss. Therefore, an all-oral, potentially less toxic, treatment regimen has been sought and encouraged by the World Health Organization. A combination of RIF plus clarithromycin (CLR) has been successful in patients first administered RIF+STR for 2 or 4 weeks. Based on evidence of efficacy of clofazimine (CFZ) in humans and mice with tuberculosis, we hypothesized that the combination of RIF+CFZ would be effective against M. ulcerans in the mouse footpad model of M. ulcerans disease because CFZ has similar MIC against M. tuberculosis and M. ulcerans. For comparison, mice were also treated with the gold standard of RIF+STR, the proposed RIF+CLR alternative regimen, or CFZ alone. Treatment was initiated after development of footpad swelling, when the bacterial burden was 4.64±0.14log10 CFU. At week 2 of treatment, the CFU counts had increased in untreated mice, remained essentially unchanged in mice treated with CFZ alone, decreased modestly with either RIF+CLR or RIF+CFZ, and decreased substantially with RIF+STR. At week 4, on the basis of footpad CFU counts, the combination regimens were ranked as follows: RIF+STR>RIF+CLR>RIF+CFZ. At weeks 6 and 8, none of the mice treated with these regimens had detectable CFU. Footpad swelling declined comparably with all of the combination regimens, as did the levels of detectable mycolactone A/B. In mice treated for only 6 weeks and followed up for 24 weeks, there were no relapses in RIF+STR treated mice, one (5%) relapse in RIF+CFZ-treated mice, but >50% in RIF+CLR treated mice. On the basis of these results, RIF+CFZ has potential as a continuation phase regimen for treatment of M. ulcerans disease.

Partial Text: Buruli ulcer (BU), or Mycobacterium ulcerans disease, has become a more readily treatable disease since 2004 with the transition from only surgery and skin grafting to antibiotic therapy with the combination of rifampin (RIF) and streptomycin (STR), the WHO preferred regimen or with RIF plus a fluoroquinolone or clarithromycin if STR is contraindicated, complemented by surgery in the case of extensive lesions to prevent or correct functional disabilities [1]. Although initial trials with the RIF+STR regimen included few reports of ototoxicity, objective measurements of hearing loss indicate that it may occur rather frequently [2]. For this reason and also operational issues with STR, an all-oral regimen is sought. Currently, there is hope that RIF plus clarithromycin (CLR) can be an acceptable oral regimen. However, this regimen would still require eight weeks of drug administration and may be complicated by gastrointestinal intolerance and the induction of CLR metabolism by RIF [3–7]. In addition, CLR is administered twice daily in some settings [5,6,8]. To date, a pilot study in Benin [9] has shown that patients with limited lesions can be successfully treated without relapse if treated with RIF+CLR for 8 weeks. Trials in Ghana have shown that treatment with RIF+STR for 4 weeks, followed by RIF+CLR for 4 weeks [10] or RIF+STR for 2 weeks, followed by RIF+CLR for 6 weeks [11] are as efficacious as RIF+STR for 8 weeks. In mice, RIF+CLR was inferior to RIF+STR and also inferior to the combination of the long-acting rifamycin, rifapentine plus CLR whether assessed by bactericidal activity, regression of footpad swelling, or recurrence (relapse) of swelling after treatment completion [12].

Although the initial bactericidal activity of RIF+CFZ was worse than that of RIF+STR and RIF+CLR (as measured by CFU and the number of mice requiring euthanasia), the sterilizing (relapse protection) was comparable to RIF+STR and superior to the currently proposed all-oral regimen of RIF+CLR. The delayed bactericidal activity of CFZ-containing regimens against M. ulcerans, associated with increased sterilizing activity (absence of relapse), parallel what has been observed against M. tuberculosis [17,18]. The consequences of relapse are potentially much more serious in tuberculosis, a fatal disease, than in Buruli ulcer, a disabling disease. However, adequate treatment durations are required to prevent further morbidity and the need for retreatment in either disease. Limited data indicate that relapse is rare in patients treated with RIF+STR whereas paradoxical reactions that may be confused with relapse are more frequent [32,33]. Given the anti-inflammatory properties of CFZ, its incorporation into BU therapy could potentially reduce the frequency of paradoxical reactions in addition to reducing the treatment duration necessary for cure.



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