Introduction
Leishmaniasis is a disease caused by a group of intracellular protozoans of the genus
Leishmania that can be transmitted to mammals, especially humans [
1]. The disease is transmitted by the bite of sandflies, especially
Phlebotomus species, in the Old World [
2]. According to the recent World Health Organization (WHO) report, about 700,000 to 1 million new cases are added annually [
1]. Among the types of leishmaniasis, cutaneous leishmaniasis (CL) is the most prevalent disease [
3], with a concerning reporting rate of 20–33% annual new cases reported to WHO [
1]. Since 2010, when the WHO established the Leishmaniasis Special Committee, there has been increased attention to epidemiological and interventional studies on leishmaniasis [
4].
The management of CL differs from region to region and is primarily based on local experience-based evidence. Several treatment modalities have been used for CL, including topical treatments, systemic therapy, cryotherapy, photodynamic therapy, etc. [
5]. Topical and local treatments are the preferred modality in treating most CL patients, comprising the clinically simple Old World Cutaneous Leishmaniasis (OWCL) lesions and the localized New World Cutaneous Leishmaniasis (NWCL) lesions caused by the
Leishmania species that are not associated with mucosal involvement [
6]. Systemic therapy is required for some
Leishmania species that can cause mucocutaneous involvement. The gold standard for systemic therapy remains systemic antimonial [
7]. Cryotherapy, which involves the use of extreme cold to destroy abnormal tissue, is effective in the treatment of OWCL when combined with topical Juniperus excelsa M. Bieb cream [
8].
Among different treatments for CL, the WHO has suggested the use of pentavalent antimonial drugs as the gold standard therapy [
9]. However, pentavalent antimonial drugs may cause various side effects and toxicities, including musculoskeletal pain, headache, nausea, asthenia, cardiotoxicity, hepatotoxicity, nephrotoxicity, and pancreatitis [
10]. In addition, these drugs are contraindicated among patients with pneumonia, myocarditis, hepatitis, nephritis, pregnant or breast-feeding mothers, etc.; hence, as these agents cannot be used in all patients, many studies have been conducted to investigate the efficacy and safety of various topical and systemic therapies. However, reported treatments do not usually provide consistent results and many treatment modalities may fail in a presenting patient. For example, systemic meglumine antimoniate with a standard dose of 20 mg per Kg per day given intramuscularly over 2 weeks shows about an 81% cure rate for
L. major and 48% for
L. tropica. Noticeably, given the lower efficacy obtained for the
L. tropica CL lesions, several studies have implemented the combination therapies, which have been proved to increase the treatment efficacy [
11].
One of the most useful combination regimens to treat CL is weekly intralesional Glucantime® plus cryotherapy. However, since CL is endemic in many regions, exerts a substantial financial burden on the healthcare system for prevention and treatment, and there still exists a large knowledge gap about treatment modalities with high cure rates, there is a demand for identifying more efficacious treatment strategies. As both fluconazole, an oral medication, and liquid nitrogen, a topical therapy, are shown to be effective for CL [
12,
13], we designed and conducted a placebo-controlled randomized clinical trial (RCT) to evaluate the efficacy and safety of adding oral fluconazole to cryotherapy among CL patients.
Discussion
Our study showed that treatment with oral fluconazole and cryotherapy, as well as cryotherapy plus placebo, resulted in a reduction in the surface area of lesions. Notably, the reduction observed in the oral fluconazole and cryotherapy arm was somewhat more pronounced. Also, the frequency of complete and relative recovery in the interventional arm was higher than in the control arm. However, no significant difference was observed between the two arms. Given the limited number of studies addressing the cure rate based on Leishmania species, additional evidence is essential to conclusively determine the efficacy of azoles against each Leishmania species.
Cryotherapy is a local therapeutic modality in the management of CL with variable reported efficacy [
16]. It is accompanied by a painful sensation and has several mild to serious complications including susceptibility of the wounds to infections. At the same time, evidence exists that combination therapy can improve the treatment duration and outcome [
17].
Interest in using azoles for leishmaniasis was revived after the report by Berman, demonstrating the activity of ketoconazole against
Leishmania species in macrophage culture [
18]. The results of the present study showed that no participants had to exit the study due to the adverse effects of fluconazole or cryotherapy. In line with this, Michelerio et al. reported no significant adverse effects in pediatric OWCL treated with oral fluconazole [
19]. Alrajhi et al. evaluated the effect of fluconazole for the treatment of CL caused by
L. major and showed that side effects were mild and similar in both arms (fluconazole arm and placebo arm) [
15]. Prates et al. who investigated the efficacy of fluconazole in the treatment of CL due to
L. brasiliensis, reported the side effects were similar in both arms (fluconazole arm and Glucantime® arm) [
20]. Fluconazole has been successfully used in children with CL caused by
L. major and
L. tropica. Furthermore, no serious side effects were observed [
19,
21]. Fluconazole has a longer half-life and greater concentrations in skin tissues than other azoles with lower toxicity. Hepatotoxicity is the most frequently reported side effect, and it usually manifests as conjugated hyperbilirubinemia or abnormal liver enzymes. Systemic fluconazole is highly tolerable in lactating and pregnant women [
22]. Generally, no specific complications and side effects have been mentioned for the use of topical formulation of the drug, except for itchiness in several studies [
23], although clinical studies do not appear to be enough to conclude. In the present study, no hepatotoxicity was observed in the treatment arm.
The results of repeated measures analysis within each arm indicated a significant decrease in the mean surface area of the lesion. Also, the lesion size (surface) significantly declined after introducing the intervention in both arms (time effect) but the difference was not significant when both arms were compared to each other. However, the reduction rate of the lesion surface area in patients treated with fluconazole was slightly greater than in patients treated with the placebo.
Consistent with the results of the present study, in the study of Parvizi et al., lesion count, duration of lesions, baseline vertical diameter size, baseline horizontal diameter size, and the baseline area of lesions showed no statistically significant differences between both arms [
8]. In Saudi Arabia, Larbi et al. conducted a double-blind RCT comparing clotrimazole 1% and miconazole 2% topical creams for the treatment of CL lesions over 30 days [
24]. The results showed that in the miconazole arm, no lesions had a complete recovery and only 30.5% of the lesions showed a decrease in size. On the other hand, in the clotrimazole arm, 15.7% of lesions were completely improved and 47.2% showed a size reduction, which confirmed that clotrimazole is significantly more effective than miconazole. Topical ketoconazole did not show a significant difference regarding effectiveness and improvement of lesions compared to placebo cream [
23]. The cure rates were similar among the fluconazole, ketoconazole, and itraconazole arms [
25]. Contrary to the results of our study, Mussi et al. compared the effect of topical fluconazole with topical paromomycin in BALB/c mice infected with
L. major, showing a significantly higher effectiveness for the paromomycin arm [
26]. The anti-leishmanial effects of azole antifungals are due to the inhibition of cytochrome P-450 mediated 14α-demethylation of lanosterol in fungi, which blocks ergosterol synthesis leading to the accumulation of 14α-methyl sterols. The inhibition of sterol biosynthesis causes leishmaniasis growth cessation [
27]. In vitro studies regarding the effects of azoles on sterol biosynthesis have reported that, for most
Leishmania species, itraconazole was slightly more inhibitory than ketoconazole, and fluconazole was much less inhibitory than the other azoles [
13].
Based on the results of the present study, as indicated by the observed difference in lesion size, a statistically significant change first appeared in week 6, coinciding with the completion of the 6-week treatment protocol. This suggests that designing this study based on a 6-week treatment protocol was acceptable for conducting RCTs of this nature.
The duration of therapy is a crucial factor to consider. Some investigators have advocated the use of higher doses of fluconazole for a shorter period (6 weeks) [
28,
29]. For instance, in a previously cited study, fluconazole was used for only 28 days [
20]. The results of another study by Veraldi et al. showed that a longer duration of fluconazole treatment did not lead to significant side effects nor laboratory abnormalities. Previous data on the use of fluconazole for 6 weeks to treat OWCL caused by
L. major have shown high cure rates with daily doses of 200 mg (79%) [
15]. In line, a study by Emad et al. reported increased efficacy with a doubled dose of fluconazole (400 mg per day) [
23]. Therefore, the dose of fluconazole chosen for the present study was based on this rationale.
In the study of Alrajhi et al., the median time to healing was 8.5 weeks for the fluconazole treatment arm, which was shorter than that of the control arm [
15]. Also, none of the independent variables, including the size and number of lesions and their locations, had a significant effect on the healing process [
15]. However, in our study, although the frequency of complete and relative recovery was higher in the interventional arm than in the control arm, no significant difference was observed. In addition, the type of treatment did not show any obvious effect on the amount of secretion.
Several studies have demonstrated the in vitro and in vivo efficacy of azole antifungals in the treatment of CL [
30,
31]. The first clinical use of fluconazole in leishmaniasis was against kala-azar, with 0% definite cure rate [
32]. Consistent with the results of our study, Sousa et al. reported a cure rate of 89% among 28 participants treated with fluconazole [
33]. Similarly, Alrajhi et al. found that fluconazole resulted in complete healing for 79% of participants with CL caused by
L. major, compared to 34% in the placebo arm [
15]. In Brazil, a case series involving 28 participants with confirmed leishmaniasis caused by
L. braziliensis, showed varying cure rates (75 to 100%) depending on the fluconazole dosage regimen: Eight participants received 5 mg/Kg/day with a cure rate of 75%, 14 participants received 6.5 mg/Kg/day with a cure rate of 92.8% and six participants received 8 mg/Kg/day with a cure rate of 100% [
27]. However, in Prates et al. study, fluconazole administered orally at a dose of 6.5–8 mg/kg/d for 28 days was found to be ineffective in a high-transmission area for
L. braziliensis [
34].
Regarding imported leishmaniasis cases, oral fluconazole has shown varying and not always satisfying cure rate in RCTs involving
L. braziliensis, which is responsible for mucocutaneous disease in the NWCL [
20]. The cure rates for
L. major infection, the cure rate varied between studies, ranging from 44.4% [
25] to 79% [
15]. The observed differences in cure rates among studies might be attributed to factors such as the dose regimen used, the Leishmania species involved, and variations in study methodology. Specifically, the final efficacy rate for
L. braziliensis was 49%, based on an analysis involving only 138 participants. Based on only one or two studies with efficacy data for each of the other
Leishmania species, the cure rate ranged from 15% for participants with
L. tropica to 89% for
L. mexicana [
19]. These observations reinforce the need for further research to determine the actual efficacy of azole treatments and caution against relying solely on non-comparative and methodologically fragile studies to assess the usefulness of this class of drugs.
A study conducted by Frajzadeh et al. is interesting in this regard. They compared the combination of cryotherapy with oral terbinafine and cryotherapy with systemic meglumine antimoniate in the treatment of leishmaniasis. In their study, although the trend of healing was more slowly in the terbinafine arm, there was no significant difference between the two arms at the end. They used terbinafine at a double dose i.e., 125 mg/kg/d (for less than 20 kg body weight), 250 mg/kg/d (20–40 kg body weight), 500 mg/kg/d (for more than 40 kg body weight) [
35]. Therefore, doubling the dose of antifungals may offer a clue to increase the efficacy of these regimens in treating leishmaniasis and could open new avenues for future research.
A study by Asilian et al. evaluated the effect of intralesional meglumine antimoniate and cryotherapy on leishmaniasis in three arms: intralesional meglumine plus cryotherapy, intralesional meglumine alone, and cryotherapy alone. The results showed that the combination arm achieved a 90.9% cure rate, while both monotherapy arms showed approximately a 55% cure rate. Thus, they stated the combination of Intralesional meglumine antimoniate plus cryotherapy is more effective than each of meglumine antimoniate or cryotherapy alone [
36]. Another study by Noor et al. compared a combination of meglumine antimoniate plus cryotherapy to cryotherapy alone and showed similar better efficacy in the combination arm.
Jowkar et al. evaluated the added benefit of topical nitric oxide 3% to cryotherapy on CL and they reported no significant increase in the efficacy of treatment [
37].
Fekri et al. conducted a study to compare the efficacy of co-administration of topical niosomal dapsone gel and intralesional injection of Glucantime® with cryotherapy plus intralesional injection of Glucantime® in CL. They had two arms of intralesional Glucantime® plus cryotherapy and intralesional Glucantime® plus niosomal dapson gel. They showed that there is no significant difference between the two arms. However, one cannot compare cryotherapy to dapson gel, because the effect of Glucantime® might be the dominant factor and compensate for the difference between cryotherapy and niosomal dapson gel [
38].
This study had limitations. Firstly, lesion characteristics were not equally distributed in the interventional and control arms. Secondly, use of different doses of fluconazole was not evaluated, as varying doses of the medicine may yield different effects in the treatment of the disease. Future studies should consider evaluating the effects of different doses of fluconazole in treating the disease. It is also suggested to evaluate the effect of other azoles in the treatment of leishmaniasis. Thirdly, another aspect that should be highlighted is the effect of leishmaniasis ulcer sizes at baseline on the outcomes. In this study, we did not initially consider this point in study design; nonetheless, we compensated this drawback with an adjustment at the time of analysis, as it stood out as depicted in the figures.
Our study highlights the importance of encouraging research for new effective therapies with oral drugs for CL treatment, preferably in RCTs, stratified by geographic region of study and by Leishmania species. Also, it is prudent to think of multimodal or multi-drug therapy in the treatment of such disease in future research to develop more effective treatments with fewer side effects that could potentially replace antimoniate drugs.