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The use of posaconazole against Chagas disease

Molina, Israel; Salvador, Fernando; Sánchez-Montalvá, Adrián

Current Opinion in Infectious Diseases: October 2015 - Volume 28 - Issue 5 - p 397–407
doi: 10.1097/QCO.0000000000000192
TROPICAL AND TRAVEL-ASSOCIATED DISEASES: Edited by Joseph M. Vinetz and Yukari C. Manabe
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Purpose of review The current therapeutic scenario against Chagas disease has been recently updated with the use of the triazoles in clinical trials and several experimental assays (in-vitro and in-vivo models) which are bringing novel and promising evidence for the treatment of Chagas diseases, mainly in its chronic phase. We pretend to analyze all the evidence extracted from the in-vitro and in-vivo assays, and try to understand the poor outcome of posaconazole (POS) in the clinical experience.

Recent findings POS is the drug with more advanced development in both experimental model and clinical trial. Despite the promising results initially obtained in the animal model, the clinical trial did not meet expectations. Nevertheless, it has documented the activity against Trypanosoma cruzi either in the animal model or in humans. Also new treatment strategies, combination or sequential schemes, have been evaluated in the animal model.

Summary POS has been tested in humans showing activity against T. cruzi, but not enough to reach cure by itself. Those results represent one of the most important breakthroughs in the treatment of Chagas disease, and open a window to new strategies as combination therapies or even sequential treatments.

Department of Infectious Disease, Vall d’Hebron Teaching Hospital, International Health Program of the Catalan Institute of Health (PROSICS) Barcelona, Universitat Autònoma de Barcelona, Barcelona, Spain

Correspondence to Israel Molina, Department of Infectious Disease, Vall d’Hebron Teaching Hospital, P° Vall d’Hebron 119, 08035 Barcelona, Spain. Tel: +34 93 274 6251; e-mail: imolina@vhebron.net

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INTRODUCTION

Chagas disease, a chronic parasitic infection, has become a world public health concern. According to the last WHO report, 6 million people are infected and 13% of the Latin American population is at risk of infection [1].

Notwithstanding the large number of infected people, access to an adequate healthcare system and to an effective therapy are the major challenges those patients have to face. The pathophysiology of the different disease phases, in addition to conditioning the disease's clinical expression, also has therapeutic implications.

Current therapeutic options are bounded to benznidazole (BNZ) and nifurtimox (NFX), two nitroheterocyclic compounds developed more than 40 years ago. Cure rates between 65 and 80% have been documented for the acute phase, reaching nearly 100% in congenitally transmitted cases treated during the first years of life. In cases of chronic infection, cure rates between 15 and 40% have been achieved, although with a far lesser degree of evidence. Around 50% of chronically infected patients who are treated with BNZ and 75% treated with NFX relate some undesired effect, and between 6 and 40% of these patients abandon treatment for this reason [2].

Despite such modest chronic phase cure rates and the toxic drug profiles, current recommendations advocate for offering universal treatment to all patients in the chronic phase [3]. This consensus is based by one hand on the improved long-term clinical progress observed in patients treated with BNZ after a mean follow-up of 10 years, mainly preventing Chagas cardiomyopathy [4]. By the other hand, the persistence of the parasite is an unequivocal condition for the damage involved in the pathogenesis of Chagas disease, thus the elimination of the parasite from the host seems to be mandatory for a good evolution of the disease [5].

In this paradigmatic scenario, the quest for new more effective and safer compounds has pointed out the ergosterol biosynthesis inhibitors as a very promising drug family.

Sterols are crucial components of eukaryotic cells as they contribute to the stability, permeability and fluidity of the membrane. Whereas mammals produce cholesterol, fungi and protozoa produce ergosterol. Mammals, yeasts or any protozoa, as Trypanosoma brucei, can utilize both endogenous and exogenous sources for the corresponding sterol synthesis. Trypanosoma cruzi is mainly dependent on de-novo production for its survival in all the stages of its life cycle [6]. Therefore, because of its reliance to endogenous ergosterol, T. cruzi is extremely vulnerable to the sterol biosynthetic pathway inhibitors. Among all the enzymes involved in this route, inhibitors of the sterol 14α-demethylase (CYP51) are the most efficient antifungal agents in clinical medicine [7].

Based on the experience gained during the past decade in clinical practice, its pharmacokinetics properties and potent anti T. cruzi activity, posaconazole (POS) has appeared as a promising triazole to be recruited against Chagas disease.

The objective of this study was to review the in-vitro and in-vivo evidence and human experiences of POS in Chagas disease.

Box 1

Box 1

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IN-VITRO STUDIES

The first in-vitro study evaluating the efficacy of POS against T. cruzi was published in 1998. POS was used in two stages of the T. cruzi parasite (EP and Y strain), epimastigotes (extracellular form) and amastigotes (the clinically relevant intracellular form). Using different POS concentrations, no immediate effects on epimastigote proliferation was seen; but after 200 h, a delayed lytic effect was observed. The minimal concentration of POS required to eradicate the amastigotes from the host cells was 0.3 nmol. Moreover, POS had no effects on the proliferation of the host cells, indicating a very specific antiparasitic activity. The authors concluded that POS was the most potent sterol biosynthesis inhibitor and antiproliferative agent ever tested against both proliferative stages of T. cruzi[8].

A more recent and novel study compares the activity among current antichagasic drugs (including POS) against different T. cruzi strains and clones belonging to all six discrete typing units (DTUs). For this purpose, amastigote cultures were obtained infecting the osteosarcoma-derived human cell line U20S with tissue-derived trypomastigote forms of T. cruzi, and later exposed to the different compounds. As Table 1 shows, POS had IC50 in the range of low nanomolar against some of the T. cruzi tested, but presented high variability depending on the strain, being ARMA13cl1 (DTU III) and 92–80cl2 (DTU V) resistant to POS. Furthermore, a time-kill assay was performed using the Y strain, showing a residual infection levels up to 7% after 144 h of POS exposure, much higher levels than those reached with BNZ [9▪].

Table 1

Table 1

Other studies have evaluated the in-vitro activity of drug combinations including POS against T. cruzi[10–12]. Two of them are focused on POS and amiodarone combination, as amiodarone is the drug most frequently used to treat arrhythmias in patients with chronic Chagas patients with cardiac involvement, and synergistic effects between amiodarone and other azole drugs have been reported in several fungi. The study performed by Benaim et al.[10] was done with amastigotes of the EP strain of T. cruzi. Amiodarone had minimum inhibitory concentration and IC50 values of 8 and 2.7 μmol, respectively, whereas for POS the corresponding values were 3 and 0.25 nmol. When exploring the activity of amiodarone and POS combination, a fractional inhibitory concentration value (FIC) of 0.42 was obtained, indicating strong synergism [3]. Veiga-Santos et al.[11] also obtained a FIC of 0.42 with amiodarone and POS combination against the Y strain of T. cruzi, and reported the cellular and subcellular alterations caused by this combination using microscopy techniques such as transmission electron microscopy: extreme accumulation of vesicles and the appearance of vacuoles in the parasite's cytoplasm (similar to autophagosomes), membrane shedding, presence of myelin-like structures and disruption of the amastigote plasma membrane [11]. Finally, the study by Planer et al.[12] screened the in-vitro activity and synergism of 2000 compounds against the Tulahuen strain of T. cruzi, showing synergistic effect of POS with clemastine (FIC 0.46), and additive effect of POS with amlodipine (FIC 0.64), BNZ (FIC 0.91), allopurinol (FIC 1.17) and amiodarone (FIC 1.61), respectively [12].

Summarizing, POS has demonstrated significant in-vitro activity against T. cruzi; however, this activity highly varies depending on the strain, and the time-kill assay has shown that POS needs longer periods of exposure with the amastigote than current antichagasic drugs (BNZ and NFX) to eradicate the parasite. On the contrary, a few drug combinations including POS have demonstrated synergistic effects against T. cruzi.

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IN-VIVO STUDIES

POS has been tested mainly in murine models [8,10,12–19,20▪▪]. The vast majority of experiences have been carried out in the acute infection model, due to the rapidity and ease of the methodology, and as the first step prior to further investigations in the chronic model. One of the major constraints of the murine model of T. cruzi infection is the lack of homogeneity between the published experiences, making comparison difficult. The more recent studies incorporate T. cruzi DNA assessment measured by PCR and immunosuppression with cyclophosphamide to enhance the chance of reactivation by quiescent forms of the parasite.

At this stage and being aware of the difficulty of drawing conclusions, the majority of the studies show that in the murine model, both acute and chronic, POS has at least the same trypanocidal activity as the BNZ against susceptible and resistant T. cruzi strain (Tables 2 and Tables 3). The more optimistic results belong to studies in which the cure criteria did not take into consideration molecular biology techniques. Both drugs increase the survival and cure rates when compared with control groups; moreover, they have a dose-dependent and length-dependent activity, hence a higher dose and longer duration achieve better results. It is worth noting that suboptimal doses of POS and BNZ have a suppressive effect in parasitaemia, but subsequent relapse is often observed when the drug is discontinued.

Table 2

Table 2

Table 2

Table 2

Table 3

Table 3

As happens in other parasitic diseases, the host–immune system activity is essential to remove the microorganism. Chagas disease is not an exception; the immune system plays an important role in the antitrypanosoma activity of POS. In immunosuppressed mice model, POS showed similar trypanocidal activity compared with the immunocompetent mice, and slightly better than obtained with BNZ regardless of the infective T. cruzi strains [13]. Therefore, POS seems to be less dependent of the immune system than the BNZ. More specifically, Ferraz et al. showed that gamma interferon is essential for the activity of BNZ, whereas interleukin-12 is equally important for BNZ and POS [14]. Moreover, CD4+ T lymphocytes are crucial for the antitrypanocidal activity of both BNZ and POS, meanwhile B lymph presence is essential for BNZ activity, but not for POS [15].

Several combination therapies involving POS have been studied. Discrepant results have been published with the combination of POS and BNZ. The association of BNZ and POS does not improve the curative effect of POS when evaluated with Y strain, but in models infected with other T. cruzi strains, the benefit could reach until 100% of cure ratio [12,18,19,20▪▪].

Sequential regimen using POS and BNZ has showed intriguing results. When BNZ was used as the first compound, noneffect or marginal impact have been observed; whereas when POS was used in first place and followed by BNZ, a significant synergistic effect was proved (72.7–100% cure ratio) [19,20▪▪]. Other combinations such as POS and amlodipine or clemastine or amiodarone given simultaneously have shown promising results with cure ratio ranging from 80 to 100% of the treated animals [10,12].

The nonhuman primate model has also been evaluated. Twelve infected baboons were treated with POS. After 37 weeks of follow-up posttreatment, all of the animals had microbiological confirmation of therapeutic failure (PCR and/or haemoculture) [21].

Summarizing, POS has good antitrypanocidal activity against BNZ-susceptible and BNZ-resistant strain in-vivo murine models, increasing survival and parasitological cure in both acute and chronic infections. Although its activity is influenced by the host's immune system, it seems to be less dependent than the activity of BNZ.

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CLINICAL USE OF POSACONAZOLE FOR CHAGAS DISEASE

First in a Chagas disease patient, use of POS was reported by Pinazo et al.[22]. A 44-year-old Argentinean woman living in Spain was diagnosed with systemic lupus erythematosus and Chagas disease. She received corticosteroids and cyclophosphamide because of lupus nephritis. BNZ at a dose of 5 mg/kg/day for 60 days was administered for the treatment of Chagas disease. Unfortunately, T. cruzi blood PCR was positive 6 months after treatment. Because of the required maintenance of immunosuppressive treatment with azathioprine, an off-label treatment with POS was suggested on a compassionate basis. POS treatment of 400 mg per 12 h for 90 days was given to the patient, and T. cruzi blood PCR was consistently negative during a 13-month follow-up period (nine determinations).

Taking into account all in-vitro and in-vivo information about the efficacy of POS in Chagas disease, a prospective, open-label, randomized clinical trial was performed by Molina et al.[23▪▪] to assess the efficacy and safety of POS in adults with chronic T. cruzi infection: the CHAGASAZOL trial [23▪▪]. Seventy-eight patients were randomly assigned, in a 1 : 1 : 1 ratio, to receive POS at a dose of 400 mg twice daily (high-dose POS), POS at a dose of 100 mg twice daily (low-dose POS), or BNZ at a dose of 150 mg twice daily, administered for 60 days. Efficacy was evaluated as treatment failure, defined as positive T. cruzi blood PCR at any point of the 10-month follow-up period after treatment. In the intention-to-treat analysis, 92.3% of the patients in the low-dose POS group and 80.7% in the high-dose POS group, as compared with 38.4% in the BNZ group, tested positive on PCR during the follow-up period (P < 0.01). Differences between the groups were more marked in the per-protocol analysis: 90% in the low-dose POS group, 80% in the high-dose POS group, and 5.9% in the BNZ group (P < 0.001). POS was well tolerated, and no serious adverse events were reported in the POS groups. A pharmacokinetic analysis of POS was performed in all patients at day 14 of treatment, showing a mean POS serum concentration of 0.909 (SD ± 0.384) μg/ml in the low-dose POS group, and 1.666 (SD ± 0.935) μg/ml in the high-dose POS group. The study concluded that, in patients with chronic Chagas disease, treatment with POS resulted in a significantly larger percentage of treatment failures than did treatment with BNZ.

There is an ongoing randomized, double blind, placebo-controlled phase 2 trial (the STOP Chagas study, ClinicalTrials.gov identifier NCT01377480) that evaluates the safety and efficacy of POS (400 mg twice daily for 60 days), BNZ (200 mg twice daily for 60 days) and a combination of both drugs (POS 400 mg twice daily and BNZ 200 mg twice daily for 60 days) in patients with Chagas disease in the indeterminate stage. Results of the study are expected to be available by the end of 2015.

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DISCUSSION

Despite being difficult to deduce any objective conclusion from the experimental model, due to the high heterogeneity in the assay design, POS seems to be a compelling drug against T. cruzi. After the results from the in-vivo murine models, the next logical step was to evaluate the drug in a clinical trial.

Surprisingly, the results obtained from the first clinical trial did not meet expectations. Patients treated with POS, regardless of the dose, had a high treatment failure ratio (80–92.3% of treatment failure) [23▪▪]. These results are consistent with those from the nonhuman primate model wherein all the animals treated with POS had treatment failure during the follow-up [21].

The natural question would be to try to understand why clinical outcome was so far from the experimental model.

One of the lessons learnt from the in-vivo and in-vitro models is that cure ratio is related to the dose used [13]. Another interesting issue is that, even though ergosterol biosynthesis inhibitors are more potent than nitroheterocyclic compounds, the time needed to reach the same efficacy against T. cruzi is much longer (slow trypanocidal compounds) [9▪]. Bearing in mind those facts, it makes sense to state that the efficacy of POS is directly related to drug exposure, contrary to what happens with BNZ, wherein its activity might be concentration-dependent [9▪,20▪▪]. This fact could be extrapolated from the CHAGASAZOL trial. Although both POS regimens showed similar treatment failure proportion, there were significant differences between the time-to-treatment failure curves. Patients treated with lower dose (100 mg/12 h) had earlier treatment failure compared with those treated with higher dose (400 mg/12 h) [23▪▪]. Furthermore, the pharmacokinetic studies conducted in the study also revealed a lower drug exposure compared with the animal models. Whereas the Cmax and area under the curve obtained from the patients treated with POS were in the range of what has been previously published in patients with invasive mycoses, drug exposure was 5–10-fold lower than in the animal model [24,25]. New POS formulations with higher plasma exposure would offer new possibilities to explore in future clinical trials [26].

Another reason that would justify such discrepancies between the experimental results and the clinical outcome might be the animal model itself. The acute model has been the most used to evaluate the candidates prior to testing them in the chronic model. The period for initiating therapy after the infection in the chronic model ranges from 45 to 130 days. Considering the life expectancy of mice (Mus musculus), this period of infection is still far from the stage of infection we found in the clinical practice, wherein the median age of the patients treated is near 40 years old [23▪▪,27]. Thus, the in-vivo model might overestimate the effect of the drugs.

Finally, another argument would rely on the mechanism of action of POS. The activity of ergosterol biosynthesis inhibitors is time-dependent and results from the depletion of ergosterol pools, and T. cruzi is completely dependent on the endogenous synthetized sterols which are crucial for cell division and growth [28]. Those parasitic forms with reduced metabolism activity (quiescent forms) or nondividing stages as trypomastigotes could have a reduced susceptibility to such drugs [29,30].

Beyond these results from experimental and clinical studies, it has been proved that POS has shown antitrypanocidal activity, which represents an encouraging fact, although according to the current evidence, it cannot be considered a valid option against Chagas disease solely, at least with a dose of 800 mg per day and given for 60 days. Combination therapy or even sequential therapeutic schemes open a new window in the treatment of the chronic stage of Chagas disease, and POS (or at least ergosterol biosynthesis inhibitors) could play an interesting role, as it may allow lowering the doses of the compounds, reducing the toxicity and shortening the duration of the treatment.

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CONCLUSION

POS has demonstrated significant in-vitro and in-vivo activity against T. cruzi. This activity highly varies according to the strain and is dependent on the drug exposure. Clinical experience has failed to show sustained trypanocidal activity in humans; however, its safety profile and data from experimental models suggest potential use in combination or in sequential therapies.

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Acknowledgements

None.

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Financial support and sponsorship

None.

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Conflicts of interest

There are no conflicts of interest.

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REFERENCES AND RECOMMENDED READING

Papers of particular interest, published within the annual period of review, have been highlighted as:

  • ▪ of special interest
  • ▪▪ of outstanding interest
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REFERENCES

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2. Bern C. Antitrypanosomal therapy for chronic Chagas’ disease. N Engl J Med 2011; 364:2527–2534.
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9▪. Moraes CB, Giardini MA, Kim H, et al. Nitroheterocyclic compounds are more efficacious than CYP51 inhibitors against Trypanosoma cruzi: implications for Chagas disease drug discovery and development. Sci Rep 2014; 4:4703.

This is the study wherein the activity and exposure duration of current drugs and drug candidates against intracellular T. cruzi amastigotes belonging to all current phylogenetic subdivisions are compared.

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In this study, new therapeutic strategies with POS and benznidazole are evaluated. The evidence from those experiments will inspire future clinical trials in humans.

21. John L. VandeBerg. Treatment trials and efficacy determination in nonhuman primates with chronic T. cruzi infections. https://sites.google.com/site/chagasddc/.
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23▪▪. Molina I, Gómez i Prat J, Salvador F, et al. Randomized trial of posaconazole and benznidazole for chronic Chagas’ disease. N Engl J Med 2014; 370:1899–1908.

This is the first clinical trial in humans with POS compared with BNZ in the chronic phase of Chagas disease. The antitrypanocidal activity was demonstrated, but patients treated with POS had more treatment failure ratio than those treated with BNZ.

24. Ullmann AJ, Cornely OA, Burchardt A, et al. Pharmacokinetics, safety, and efficacy of posaconazole in patients with persistent febrile neutropenia or refractory invasive fungal infection. Antimicrob Agents Chemother 2006; 50:658–666.
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Keywords:

Chagas disease; posaconazole; Trypanosoma cruzi

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