Brima M. Younis

Paromomycin for the treatment of visceral leishmaniasis in Sudan: a randomized, open-label, dose-finding study.

Background A recent study has shown that treatment of visceral leishmaniasis (VL) with the standard dose of 15 mg/kg/day of paromomycin sulphate (PM) for 21 days was not efficacious in patients in Sudan. We therefore decided to test the efficacy of paramomycin for a longer treatment duration (15 mg/kg/day for 28 days) and at the higher dose of 20 mg/kg/day for 21 days. Methods This randomized, open-label, dose-finding, phase II study assessed the two above high-dose PM treatment regimens. Patients with clinical features and positive bone-marrow aspirates for VL were enrolled. All patients received their assigned courses of PM intramuscularly and adverse events were monitored. Parasite clearance in bone-marrow aspirates was tested by microscopy at end of treatment (EOT, primary efficacy endpoint), 3 months (in patients who were not clinically well) and 6 months after EOT (secondary efficacy endpoint). Pharmacokinetic data were obtained from a subset of patients weighing over 30 kg. Findings 42 patients (21 per group) aged between 4 and 60 years were enrolled. At EOT, 85% of patients (95% confidence interval [CI]: 63.7% to 97.0%) in the 20 mg/kg/day group and 90% of patients (95% CI: 69.6% to 98.8%) in the 15 mg/kg/day group had parasite clearance. Six months after treatment, efficacy was 80.0% (95% CI: 56.3% to 94.3%) and 81.0% (95% CI: 58.1% to 94.6%) in the 20 mg/kg/day and 15 mg/kg/day groups, respectively. There were no serious adverse events. Pharmacokinetic profiles suggested a difference between the two doses, although numbers of patients recruited were too few to make it significant (n = 3 and n = 6 in the 20 mg/kg/day and 15 mg/kg/day groups, respectively). Conclusion Data suggest that both high dose regimens were more efficacious than the standard 15 mg/kg/day PM for 21 days and could be further evaluated in phase III studies in East Africa. Trial Registration ClinicalTrials.gov NCT00255567

Safety and Efficacy of Single Dose versus Multiple Doses of AmBisome® for Treatment of Visceral Leishmaniasis in Eastern Africa: A Randomised Trial.

Background Anti-leishmanial drug regimens that include a single dose AmBisome® could be suitable for eastern African patients with symptomatic visceral leishmaniasis (VL) but the appropriate single dose is unknown. Methodology A multi-centre, open-label, non-inferiority, randomized controlled trial with an adaptive design, was conducted to compare the efficacy and safety of a single dose and multiple doses of AmBisome® for the treatment of VL in eastern Africa. The primary efficacy endpoint was definitive cure (DC) at 6 months. Symptomatic patients with parasitologically-confirmed, non-severe VL, received a single dose of AmBisome® 7.5 mg/kg body weight or multiple doses, 7 times 3 mg/kg on days 1–5, 14, and 21. If interim analyses, evaluated 30 days after the start of treatment following 40 or 80 patients, showed the single dose gave significantly poorer parasite clearance than multiple doses at the 5% significance level, the single dose was increased by 2·5 mg/kg. In a sub-set of patients, parasite clearance was measured by quantitative reverse transcriptase (qRT) PCR. Principal Findings The trial was terminated after the third interim analysis because of low efficacy of both regimens. Based on the intention-to-treat population, DC was 85% (95%CI 73–93%), 40% (95%CI 19–64%), and 58% (95%CI 41–73%) in patients treated with multiple doses (n = 63), and single doses of 7·5 (n = 21) or 10 mg/kg (n = 40), respectively. qRT-PCR suggested superior parasite clearance with multiple doses as early as day 3. Safety data accorded with the drug label. Conclusions The tested AmBisome® regimens would not be suitable for VL treatment across eastern Africa. An optimal single dose regimen was not identified. Trials Registration www.clinicaltrials.gov NCT00832208

Safety and Efficacy of miltefosine alone and in combination with sodium stibogluconate and liposomal amphotericin B for the treatment of primary visceral leishmaniasis in East Africa: study protocol for a randomized controlled trial

BackgroundTreatment options for Visceral Leishmaniasis (VL) in East Africa are far from satisfactory due to cost, toxicity, prolonged treatment duration or emergence of parasite resistance. Hence there is a need to explore alternative treatment protocols such as miltefosine alone or in combinations including miltefosine, sodium stibogluconate (SSG) or liposomal amphotericin B. The aim of this trial is to identify regimen(s) which are sufficiently promising for future trials in East Africa.Methods/DesignA phase II randomized, parallel arm, open-labelled trial is being conducted to assess the efficacy of each of the three regimens: liposomal amphotericin B with SSG, Liposomal amphotericin B with miltefosine and miltefosine alone. The primary endpoint is cure at day 28 with secondary endpoint at day 210 (6 months). Initial cure is a single composite measure based on parasitologic evaluation (bone marrow, spleen or lymph node aspirate) and clinical assessment. Repeated interim analyses have been planned after recruitment of 15 patients in each arm with a maximum sample size of 63 for each. These will follow group-sequential methods (the triangular test) to identify when a regimen is inadequate (<75% efficacy) or adequate (>90% efficacy). We describe a method to ensure consistency of the sequential analysis of day 28 cure with the non-sequential analysis of day 210 cure.DiscussionA regimen with adequate efficacy would be a candidate for treatment of VL with reasonable costs. The design allows repeated testing throughout the trial recruitment period while maintaining good statistical properties (Type I & II error rates) and reducing the expected sample sizes.Trial RegistrationClinicalTrials.gov: NCT01067443

Immunogenicity and immune modulatory effects of in silico predicted L. donovani candidate peptide vaccines

Visceral leishmaniasis (VL) is a serious parasitic disease for which control measures are limited and drug resistance is increasing. First and second generation vaccine candidates have not been successful. The goal of the present study was to select possibly immunogenic L. donovani donovani GP63 peptides using immunoinformatics tools and to test their immunogenicity in vitro. The amino acid sequence of L. donovani donovani GP63 [GenBank accession: ACT31401] was screened using the EpiMatrix algorithm for putative T cell epitopes that would bind to the most common HLA class II alleles (DRB1*1101 and DRB1*0804) among at–risk populations. Four T cell epitopes were selected from nine potential candidates. Stimulation of whole blood from healthy volunteers using the peptides separately produced mean IFN-γ and IL-4 levels that were not significantly different from negative controls, while the pooled peptides produced a moderate IFN-γ increase in some volunteers. However, mean IL-10 levels were significantly reduced for all individuals compared with controls. The immunogenicity of these epitopes may be harnessed most effectively in a vaccine delivered in combination with immune-modulating adjuvants.

The Economic Burden of Visceral Leishmaniasis in Sudan: An Assessment of Provider and Household Costs

Visceral leishmaniasis (VL) is a neglected parasitic disease that is fatal if left untreated and is endemic in eastern Sudan. We estimated the direct and indirect costs of treatment of VL from the perspective of the provider and the household at three public hospitals in Gedaref State. The median total cost for one VL episode was estimated to be US

Blindness Following Visceral Leishmaniasis : A Neglected Post-Kala-Azar Complication

450. Despite the free provision of VL drugs at public hospitals, households bore 53% of the total cost of VL with one episode of VL representing 40% of the annual household income. More than 75% of households incurred catastrophic out-of-pocket expenditures. The length of treatment of 30 days led to important costs for both health providers and households. Alternative treatment regimens that reduce the duration of treatment are urgently needed.

Leishmania Donovani: Genetic Diversity Of Isolates From Sudan Characterized By PCR-Based RAPD

Visceral leishmaniasis (VL) is an important cause of morbidity and mortality that affects multiple organs. Post-kala-azar ocular involvement is a serious complication that can manifest as blepharo-conjuctivitis or pan-uveitis. Failure of prompt diagnosis and treatment can result in blindness. We report five cases with pan-uveitis that followed the successful treatment of VL and consequent post-kala-azar dermal leishmaniasis were presented. Two patients lost their sight permanently but the rest were successfully treated. A high index of suspicion and prompt treatment are of paramount importance in order to avoid blindness following post-kala-azar ocular uveitis.

Visceral leishmaniasis relapse hazard is linked to reduced miltefosine exposure in patients from Eastern Africa: a population pharmacokinetic/pharmacodynamic study

Drug unresponsiveness in patients with visceral leishmaniasis (VL) is a problem in many endemic areas. This study aimed to determine genetic diversity of Leishmania donovani isolates from a VL endemic area in Sudan as a possible explanation for drug unresponsiveness in some patients. Thirty clinically stibogluconate (SSG)-sensitive isolates were made SSG-unresponsive in vitro by gradually increasing SSG concentrations. The sensitive isolates and their SSG-unresponsive counterparts were typed using mini-circle kDNA and categorized using PCR-RAPD. All the isolates were typed as L. donovani, the resulting PCR-RAPD characterization of the SSG-sensitive isolates gave three distinct primary genotypes while, the SSG-unresponsive isolates showed only a single band. L. donovani isolates from eastern Sudan are diverse; this probably resulted from emergence of new L. donovani strains during epidemics due to the pressure of widespread use of antimonials. In this communication the possible role of isolates diversity in antimonial unresponsiveness and the in vitro changing PCR-RAPD band pattern in SSG-unresponsive strains were discussed.

Expression profiling of Sudanese visceral leishmaniasis patients pre- and post-treatment with sodium stibogluconate

Abstract Background Low efficacy of miltefosine in the treatment of visceral leishmaniasis was recently observed in Eastern Africa. Objectives To describe the pharmacokinetics and establish a pharmacokinetic/pharmacodynamic relationship for miltefosine in Eastern African patients with visceral leishmaniasis, using a time-to-event approach to model relapse of disease. Methods Miltefosine plasma concentrations from 95 patients (48 monotherapy versus 47 combination therapy) were included in the population pharmacokinetic model using non-linear mixed effects modelling. Subsequently a time-to-event model was developed to model the time of clinical relapse. Various summary pharmacokinetic parameters (various AUCs, Time > EC50, Time > EC90), normalized within each treatment arm to allow simultaneous analysis, were evaluated as relapse hazard-changing covariates. Results A two-compartment population model with first-order absorption fitted the miltefosine pharmacokinetic data adequately. Relative bioavailability was reduced (−74%, relative standard error 4.7%) during the first week of treatment of the monotherapy arm but only the first day of the shorter combination regimen. Time to the relapse of infection could be described using a constant baseline hazard (baseline 1.8 relapses/year, relative standard error 72.7%). Miltefosine Time > EC90 improved the model significantly when added in a maximum effect function on the baseline hazard (half maximal effect with Time > EC90 6.97 days for monotherapy). Conclusions Miltefosine drug exposure was found to be decreased in Eastern African patients with visceral leishmaniasis, due to a (transient) initial lower bioavailability. Relapse hazard was inversely linked to miltefosine exposure. Significantly lower miltefosine exposure was observed in children compared with adults, further urging the need for implementation of dose adaptations for children.

Leishmania : Probable genetic hybrids between species in Sudanese isolates

Visceral leishmaniasis (VL) in Sudan caused by Leishmania donovani is fatal in susceptible individuals if untreated. Treatment with sodium stibogluconate (SSG) leads to post‐kala‐azar dermal leishmaniasis (PKDL) in 58% of patients. Here, Affymetrix microarrays were used to identify genes differentially expressed in lymph nodes (N=9 paired samples) pre‐ and post‐treatment with SSG. Using the Bioconductor package limma, 438 genes from 28 869 post‐quality‐control probe sets were differentially expressed (Pnominal≤.02) post‐ vs pretreatment. Canonical pathway analysis using Ingenuity Pathway Analysis™ identified “role of nuclear factor of activated T‐cell in regulation of immune response” (Pnominal=1.35×10−5; PBH‐adjusted=4.79×10−3), “B‐cell development” (Pnominal=2.04×10−4; PBH‐adjusted=.024), “Fcγ receptor‐mediated phagocytosis in macrophages and monocytes” (Pnominal=2.04×10−4; PBH‐adjusted=.024) and “OX40 signalling” (Pnominal=2.82×10−4; PBH‐adjusted=.025) as pathways differentially regulated post‐ vs pretreatment. Major network hub genes included TP53, FN1, MYC, BCL2, JUN, SYK, RUNX2, MMP1 and ACTA2. Top endogenous upstream regulators included IL‐7 (P=2.28×10−6), TNF (P=4.26×10−6), Amyloid Precursor Protein (P=4.23×10−5) and SPI1/PI.1 (P=1.17×10−7). Top predicted chemical drug regulators included the flavonoid genistein (P=4.56×10−7) and the quinoline alkaloid camptothecin (P=5.14×10−5). These results contribute to our understanding of immunopathology associated with VL and response to SSG treatment. Further replication could identify novel therapeutic strategies that improve on SSG treatment and reduce the likelihood of progression to PKDL.