Trypanosoma brucei (T.b.) gambiense Human African trypanosomiasis (HAT; sleeping sickness) is a fatal disease. Until 2009, available treatments for 2(nd) stage HAT were complicated to use, expensive (eflornithine monotherapy), or toxic, and insufficiently effective in certain areas (melarsoprol). Recently, nifurtimox-eflornithine combination therapy (NECT) demonstrated good safety and efficacy in a randomised controlled trial (RCT) and was added to the World Health Organisation (WHO) essential medicines list (EML). Documentation of its safety profile in field conditions will support its wider use.In a multicentre, open label, single arm, phase IIIb study of the use of NECT for 2(nd) stage T.b. gambiense HAT, all patients admitted to the trial centres who fulfilled inclusion criteria were treated with NECT. The primary outcome was the proportion of patients discharged alive from hospital. Safety was further assessed based on treatment emergent adverse events (AEs) occurring during hospitalisation.629 patients were treated in six HAT treatment facilities in the Democratic Republic of the Congo (DRC), including 100 children under 12, 14 pregnant and 33 breastfeeding women. The proportion of patients discharged alive after treatment completion was 98.4% (619/629; 95%CI [97.1%; 99.1%]). Of the 10 patients who died during hospitalisation, 8 presented in a bad or very bad health condition at baseline; one death was assessed as unlikely related to treatment. No major or unexpected safety concerns arose in any patient group. Most common AEs were gastro-intestinal (61%), general (46%), nervous system (mostly central; 34%) and metabolic disorders (26%). The overall safety profile was similar to previously published findings.In field conditions and in a wider population, including children, NECT displayed a similar tolerability profile to that described in more stringent clinical trial conditions. The in-hospital safety was comparable to published results, and long term efficacy will be confirmed after 24 months follow-up.The trial is registered at ClinicalTrials.gov, number NCT00906880.
Staging and treatment of human African trypanosomiasis caused by Trypanosoma brucei gambiense (g-HAT) required lumbar puncture to assess cerebrospinal fluid (CSF) and intravenous drugs that cross the blood-brain barrier for late-stage infection. These procedures are inconvenient in rural health systems of disease-endemic countries. A pivotal study established fexinidazole as the first oral monotherapy to be effective against non-severe stage 2 g-HAT. We aimed to assess the safety and efficacy of fexinidazole in early g-HAT.
Background Nifurtimox-eflornithine combination therapy (NECT) for the treatment of second stage gambiense human African trypanosomiasis (HAT) was added to the World Health Organization’s Essential Medicines List in 2009 after demonstration of its non-inferior efficacy compared to eflornithine therapy. A study of NECT use in the field showed acceptable safety and high efficacy until hospital discharge in a wide population, including children, pregnant and breastfeeding women, and patients with a HAT treatment history. We present here the effectiveness results after the 24-month follow-up visit. Methodology/Principal findings In a multicenter, open label, single arm phase IIIb study, second stage gambiense HAT patients were treated with NECT in the Democratic Republic of Congo. Clinical cure was defined 24 months after treatment as survival without clinical and/or parasitological signs of HAT. Of the 629 included patients, 619 (98.4%) were discharged alive after treatment and were examined for the presence of trypanosomes, white blood cell count in cerebro-spinal fluid, and disease symptoms. The clinical cure rate of 94.1% was comparable for all subpopulations analyzed at the 24-month follow-up visit. Self-reported adverse events during follow-up were few and concerned mainly nervous system disorders, infections, and gastro-intestinal disorders. Overall, 28 patients (4.3%) died during the course of the trial. The death of 16 of the 18 patients who died during the follow-up period was assessed as unlikely or not related to NECT. Within 24 months, eight patients (1.3%) relapsed and received rescue treatment. Sixteen patients were completely lost to follow-up. Conclusions/Significance NECT treatment administered under field conditions was effective and sufficiently well tolerated, no major concern arose for children or pregnant or breastfeeding women. Patients with a previous HAT treatment history had the same response as those who were naïve. In conclusion, NECT was confirmed as effective and appropriate for use in a broad population, including vulnerable subpopulations. Trial registration The trial is registered at ClinicalTrials.gov, number NCT00906880 .
Background The long-term treatment outcome of visceral leishmaniasis (VL) patients with HIV co-infection is complicated by a high rate of relapse, especially when the CD4 count is low. Although use of secondary prophylaxis is recommended, it is not routinely practiced and data on its effectiveness and safety are limited. Methods A prospective cohort study was conducted in Northwest Ethiopia from August 2014 to August 2017 (NCT02011958). HIV-VL patients were followed for up to 12 months. Patients with CD4 cell counts below 200/μL at the end of VL treatment received pentamidine prophylaxis starting one month after parasitological cure, while those with CD4 count ≥200 cells/μL were followed without secondary prophylaxis. Compliance, safety and relapse-free survival, using Kaplan-Meier analysis methods to account for variable time at risk, were summarised. Risk factors for relapse or death were analysed. Results Fifty-four HIV patients were followed. The probability of relapse-free survival at one year was 50% (95% confidence interval [CI]: 35–63%): 53% (30–71%) in 22 patients with CD4 ≥200 cells/μL without pentamidine prophylaxis and 46% (26–63%) in 29 with CD4 <200 cells/μL who started pentamidine. Three patients with CD4 <200 cells/μL did not start pentamidine. Amongst those with CD4 ≥200 cells/μL, VL relapse was an independent risk factor for subsequent relapse or death (adjusted rate ratio: 5.42, 95% CI: 1.1–25.8). Except for one case of renal failure which was considered possibly related to pentamidine, there were no drug-related safety concerns. Conclusion The relapse-free survival rate for VL patients with HIV was low. Relapse-free survival of patients with CD4 count <200cells/μL given pentamidine secondary prophylaxis appeared to be comparable to patients with a CD4 count ≥200 cells/μL not given prophylaxis. Patients with relapsed VL are at higher risk for subsequent relapse and should be considered a priority for secondary prophylaxis, irrespective of their CD4 count.
NO is a potent cellular mediator which has been shown to modulate several immune mechanisms. Using human T lymphocytes as responder cells in a primary mixed lymphocyte reaction, we demonstrated that, at the initiation of the culture, exogenously provided NO via sodium nitroprusside, in non-toxic concentrations, inhibited both allogeneic proliferative and primary cytotoxic responses in a dose-dependent manner. In contrast, it had no effect on the cytotoxic activity of established human TCR (alpha)beta and TCR (gamma)delta cytotoxic T lymphocyte (CTL) clones. The NO inhibitory effect on primary cytotoxic T cell response correlates with inhibition of T cell blastogenesis. Furthermore, under our stimulation conditions, NO induced an inhibition of IL-2 production, an alteration of IL-2R(alpha) expression, and a down-regulation of NF-AT translocation in CD4(+) and CD8(+)allostimulated T cells. Furthermore, we demonstrate that the inhibition of allospecific CTL activity by the NO donor was at least in part related to an inhibition of granzyme B and Fas ligand transcription as revealed respectively by RNase protection and RT-PCR analysis. These results suggest that NO may function to fine tune human CD3(+) T cell activation and subsequent CTL generation.
Interleukin (IL)-12 can enhance the development of effective immune responses against tumors as well as against certain infectious agents. It is therefore a potential candidate for therapeutic use in cancer therapy and in design of vaccines against several infectious diseases.The authors have established a specific cytotoxic T-cell line (TIL-Heu) from lymphocytes infiltrating a human large cell carcinoma of the lung (LCC). In the current report, the authors have investigated the in vivo effect of recombinant human IL-12 (rhIL-12) on the adoptive transfer of TIL-Heu cells in autologous tumor (Heu-n) engrafted into severe combined immunodeficiency disease-nonobese diabetic (SCID-NOD) mice.Initial in vitro experiments indicated that rhIL-12 increased the cytotoxic potential of TIL-Heu cells in a dose-dependent manner. Heu-n tumors transplanted into SCID-NOD mice were injected with TIL-Heu cells, resulting in a significant tumor growth inhibition. When low doses of rhIL-12 were injected intratumorally after TIL-Heu transfer, a clear increase in tumor growth suppression was observed. Surprisingly, higher doses of rhIL-12 had no effect on cytotoxic T lymphocyte (CTL)-induced prevention of tumor growth. Further in vitro experiments revealed an inhibition of tumor cell lysis after incubation with supernatant of TIL-Heu cells stimulated with high doses of rhIL-12, strongly suggesting that an immunosuppressive factor secreted by the high dose IL-12-stimulated CTL may be responsible for the tumor escape observed in vivo.The authors' data indicate that IL-10 may play a critical role in the lack of effect of high dose IL-12, by mediating tumor cell resistance to CTL killing. Therefore, understanding the cross-talk between immunoregulatory and immunosuppressive cytokines ultimately may provide new approaches to improve cytokine-mediated cancer immunotherapy.
ABSTRACT To facilitate future pharmacokinetic studies of combination treatments against leishmaniasis in remote regions in which the disease is endemic, a simple cheap sampling method is required for miltefosine quantification. The aims of this study were to validate a liquid chromatography-tandem mass spectrometry method to quantify miltefosine in dried blood spot (DBS) samples and to validate its use with Ethiopian patients with visceral leishmaniasis (VL). Since hematocrit (Ht) levels are typically severely decreased in VL patients, returning to normal during treatment, the method was evaluated over a range of clinically relevant Ht values. Miltefosine was extracted from DBS samples using a simple method of pretreatment with methanol, resulting in >97% recovery. The method was validated over a calibration range of 10 to 2,000 ng/ml, and accuracy and precision were within ±11.2% and ≤7.0% (≤19.1% at the lower limit of quantification), respectively. The method was accurate and precise for blood spot volumes between 10 and 30 μl and for Ht levels of 20 to 35%, although a linear effect of Ht levels on miltefosine quantification was observed in the bioanalytical validation. DBS samples were stable for at least 162 days at 37°C. Clinical validation of the method using paired DBS and plasma samples from 16 VL patients showed a median observed DBS/plasma miltefosine concentration ratio of 0.99, with good correlation (Pearson's r = 0.946). Correcting for patient-specific Ht levels did not further improve the concordance between the sampling methods. This successfully validated method to quantify miltefosine in DBS samples was demonstrated to be a valid and practical alternative to venous blood sampling that can be applied in future miltefosine pharmacokinetic studies with leishmaniasis patients, without Ht correction.
Research in visceral leishmaniasis in the last decade has been focused on how better to use the existing medicines as monotherapy or in combination. Systematic research by geographical regions has shown that a universal treatment is far from today's reality. Substantial progress has been made in the elimination of kala-azar in South Asia, with a clear strategy on first- and second-line therapy options of single-dose liposomal amphotericin B and a combination of paromomycin and miltefosine, respectively, among other interventions. In Eastern Africa, sodium stibogluconate (SSG) and paromomycin in combination offer an advantage compared to the previous SSG monotherapy, although not exempted of limitations, as this therapy requires 17 days of painful double injections and bears the risk of SSG-related cardiotoxicity. In this region, attempts to improve the combination therapy have been unsuccessful. However, pharmacokinetic studies have led to a better understanding of underlying mechanisms, like the underexposure of children to miltefosine treatment, and an improved regimen using an allometric dosage. Given this global scenario of progress and pitfalls, we here review what steps need to be taken with existing medicines and highlight the urgent need for oral drugs. Furthermore, it should be noted that six candidates belonging to five new chemical classes are reaching phase I, ensuring an optimistic near future.
Background: Visceral leishmaniasis (VL) treatment in HIV patients very often fails and is followed by high relapse and case-fatality rates. Hence, treatment efficacy assessment is imperative but based on invasive organ aspiration for parasite detection. In the search of a less-invasive alternative and because the host immune response is pivotal for treatment outcome in immunocompromised VL patients, we studied changes in the whole blood transcriptional profile of VL-HIV patients during treatment. Methods: Embedded in a clinical trial in Northwest Ethiopia, RNA-Seq was performed on whole blood samples of 28 VL-HIV patients before and after completion of a 29-day treatment regimen of AmBisome or AmBisome/miltefosine. Pathway analyses were combined with a machine learning approach to establish a clinically-useful 4-gene set. Findings: Distinct signatures of differentially expressed genes were identified for patients who failed treatment and were successfully treated. Pathway analyses in the latter highlighted a downregulation of genes associated with host cellular activity and immunity, and upregulation of antimicrobial peptide activity in phagolysosomes. No signs of disease remission nor pathway enrichment were observed in treatment failure patients. Next, we identified a 4-gene signature (HRH4, IL10, PRSS33, SLFN14) that could accurately discriminate treatment outcome at end of treatment, displaying an average area-under-the-ROC-curve of 0.97. Interpretation: A simple blood-based signature thus holds significant promise to facilitate treatment efficacy monitoring and provide an alternative test-of-cure to guide patient management in VL-HIV patients. Funding Statement: Funding was provided by the research and development project, AfricoLeish, supported by the European Union Seventh Framework Programme (EU FP7), under grant agreement n°305178. WA and BC are personally supported by a Research Foundation-Flanders postdoctoral fellowship. PMK is funded by a Wellcome Trust Senior Investigator Award (WT104726). Declaration of Interests: The authors declare that there are no competing interests.Ethics Approval Statement: The study protocol was approved by the University of Gondar Institutional Review Board, the Ethiopian National Research Ethics Review Committee, the Médecins Sans Frontiers Ethics Review Board, the London School of Hygiene and Tropical Medicine Research Ethics Committee, the Antwerp University Hospital Ethics Committee, and the Prince Leopold Institute of Tropical Medicine Institutional Review Board, and carried out in accordance with international guidelines (Helsinki declaration, Good Clinical Practices and local regulations).
