Kinetics of parasite survival and cytokine secretion in response to 4 μM miltefosine and 8 μg Sb^V/mL as meglumine antimoniate in the <i>ex vivo</i> PBMC model.

<p>(A) Parasite survival and (B to E) cytokine secretion (IL-13, TNF-α, IL-10, IFN-γ) over 96 h. Mean values ± SEM for PBMCs from 5 patients. ** p ≤ 0.01, control vs both drugs.</p

Kinetics of parasite and immunologic responses in the absence of drugs.

<p>(A) Kinetics of infection evaluating the parasites: monocyte ratios of 10:1 and 20:1 (left panel), and the parasite burden (right panel) in PBMCs vs macrophages alone (infection ratio 10:1) are shown. Parasite survival is expressed as bioluminescence produced by luciferase activity in relative light units (RLU). Mean signal of uninfected cells: 135.3 ± 23.7 RLU. (B) Kinetics of cytokine secretion (TNF-α, IL-10, IL-13, IFN-γ) over the 72 hours of treatment. Data are based on at least 4 patients and expressed as means ± SEM.</p

Dose-response of cytokine secretion by PBMCs to pentoxifylline and anti-leishmanial-pentoxifylline combinations.

<p>Cells from 10 patients were infected at a parasite: monocyte ratio of 10:1. Cytokine levels (IL-10, IL-13, TNF-α, IFN-γ) were determined after exposure for 96 h to different concentrations of pentoxifylline (PTX), (A) alone, combined with (B) 8 μg Sb^V/ml meglumine antimoniate or (C) 4 μM miltefosine. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.0001, **** p ≤ 0.0001.</p

Dose-response of cytokine secretion by PBMCs to CpG, and anti-leishmanial- CpG combinations.

<p>Cells from 10 patients were infected at a parasite: monocyte ratio of 10:1. Cytokine levels (IL-10, IL-13, TNF-α, IFN-γ) were determined after exposure for 96 h to different concentrations of CpG (A) alone, combined with (B) 8 μg Sb^V/ml meglumine antimoniate or (C) 4 μM miltefosine. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01.</p

Concentration-dependent effect of miltefosine and meglumine antimoniate on parasite survival and cytokine secretion.

<p>(A) Parasite survival and (B) Cytokine secretion after 96 hours of exposure to increasing concentrations of miltefosine (HePC) and meglumine antimoniate (Sb^V). TNFα, IL-10, IFNγ and IL-13 were evaluated in supernatants of PBMCs infected with <i>L</i>. <i>(V) panamensis</i>. Data are based on at least 6 patients and presented as mean ± SEM of the parasite burden or cytokine secretion compared to infected control without drug. ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001.</p

Effect of anti-leishmanial drugs, immunomodulators and their combinations on parasite burden.

<p>(A, C and E) Dose response for pentoxifylline alone, combined with 8 μg Sb^V/ml meglumine antimoniate or 4 μM miltefosine. (B, D and F) Dose response for CpG alone, combined with 8 μg Sb^V/ml meglumine antimoniate or 4 μM miltefosine. Data are presented as means ± SEM of the parasite burden compared to infected control cultures without drugs for PBMCs from 10 patients. * p< 0.05.</p

Relative expression of <i>IDO</i> decreases in CDL patients after treatment and is correlated with <i>IFNG</i>.

<p>RNA was isolated from biopsies of lesions from CDL patients (n = 11) before and after treatment and the expression of <i>FOXP3</i> (<b>A</b>), <i>IFNG</i> (<b>B</b>), <i>IL10</i> (<b>C</b>) and <i>IDO</i> (<b>D</b>) was measured by qRT-PCR. The expression of each gene relative to healthy skin of four normal controls was calculated using the ΔΔCT method. <b>E</b>. Correlation between the relative expression of <i>IDO</i> and <i>FOXP3</i> (left panel) or <i>IFNG</i> (right panel). *p<0.05, Wilcoxon signed-rank test.</p

Tornado diagram of one-way sensitivity analyses of effect on miltefosine cDOT/MA cost ratio of parameters from survey data.

<p>Model parameters are listed on the vertical axis, with the range examined in sensitivity analyses in parentheses. The length of the horizontal bar demonstrates the impact of the changes in the parameter values on the cost ratio of miltefosine cDOT to MA. The solid vertical line indicates the estimated cost ratio of the base case. For example, a cost of $0 was estimated for the base case for miltefosine supply costs. The cost ratio favored miltefosine cDOT for all simulated scenarios. <b>MA:</b> meglumine antimoniate, <b>cDOT:</b> caregiver directly observed therapy.</p

IL-10 does not mediate suppression of effector functions by CD4⁺CD25⁺ cells.

<p>IL-10 was measured in supernatants from the CD4⁺CD25⁻-CD4⁺CD25⁺ co-cultures by ELISA. The level of IL-10 in the absence (1∶0 ratio) or presence (1∶1 ratio) of CD4⁺CD25⁺ cells is shown for AI (n = 6), CDL patients before treatment (n = 14), CDL patients after treatment (n = 11) and all co-cultures combined (n = 31). **p<0.01, Wilcoxon signed-rank test. Means with SEM are shown.</p

Cost-effectiveness of meglumine antimoniate versus miltefosine caregiver DOT for the treatment of pediatric cutaneous leishmaniasis

<div><p>Background</p><p>Oral miltefosine has been shown to be non-inferior to first-line, injectable meglumine antimoniate (MA) for the treatment of cutaneous leishmaniasis (CL) in children. Miltefosine may be administered via in-home caregiver Directly Observed Therapy (cDOT), while patients must travel to clinics to receive MA. We performed a cost-effectiveness analysis comparing miltefosine by cDOT versus MA for pediatric CL in southwest Colombia.</p><p>Methodology/Principle findings</p><p>We developed a Monte Carlo model comparing the cost-per-cure of miltefosine by cDOT compared to MA from patient, government payer, and societal perspectives (societal = sum of patient and government payer perspective costs). Drug effectiveness and adverse events were estimated from clinical trials. Healthcare utilization and costs of travel were obtained from surveys of providers and published sources. The primary outcome was cost-per-cure reported in 2015 USD. Treatment efficacy, costs, and adherence were varied in sensitivity analysis to assess robustness of results. Treatment with miltefosine resulted in substantially lower cost-per-cure from a societal and patient perspective, and slightly higher cost-per-cure from a government payer perspective compared to MA. Mean societal cost-per-cure were $531 (SD±$239) for MA and $188 (SD±$100) for miltefosine, a mean cost-per-cure difference of +$343. Mean cost-per-cure from a patient perspective were$442 (SD ±$233) for MA and$30 (SD±$16) for miltefosine, a mean difference of +$412. Mean cost-per-cure from a government perspective were $89 (SD±$55) for MA and $158 (SD±$98) for miltefosine, with a mean difference of -$69. Results were robust across a variety of assumptions in univariate and multi-way analysis.</p><p>Conclusions/Significance</p><p>Treatment of pediatric cutaneous leishmaniasis with miltefosine via cDOT is cost saving from patient and societal perspectives, and moderately more costly from the government payer perspective compared to treatment with MA. Results were robust over a range of sensitivity analyses. Lower drug price for miltefosine could result in cost saving from a government perspective.</p></div