Key Contributions by the Swiss Tropical and Public Health Institute Towards New and Better Drugs for Tropical Diseases.

Thanks to its expertise in clinical research, epidemiology, infectious diseases, microbiology, parasitology, public health, translational research and tropical medicine, coupled with deeply rooted partnerships with institutions in low- and middle-income countries (LMICs), the Swiss Tropical and Public Health Institute (Swiss TPH) has been a key contributor in many drug research and development consortia involving academia, pharma and product development partnerships. Our know-how of the maintenance of parasites and their life-cycles in the laboratory, plus our strong ties to research centres and disease control programme managers in LMICs with access to field sites and laboratories, have enabled systems for drug efficacy testing in vitro and in vivo, clinical research, and modelling to support the experimental approaches. Thus, Swiss TPH has made fundamental contributions towards the development of new drugs - and the better use of old drugs - for neglected tropical diseases and infectious diseases of poverty, such as Buruli ulcer, Chagas disease, food-borne trematodiasis (e.g. clonorchiasis, fascioliasis and opisthorchiasis), human African trypanosomiasis, leishmaniasis, malaria, schistosomiasis, soil-transmitted helminthiasis and tuberculosis. In this article, we show case the success stories of molecules to which Swiss TPH has made a substantial contribution regarding their use as anti-infective compounds with the ultimate aim to improve people's health and well-being.

Fexinidazole for Human African Trypanosomiasis, the Fruit of a Successful Public-Private Partnership.

After 100 years of chemotherapy with impractical and toxic drugs, an oral cure for human African trypanosomiasis (HAT) is available: Fexinidazole. In this case, we review the history of drug discovery for HAT with special emphasis on the discovery, pre-clinical development, and operational challenges of the clinical trials of fexinidazole. The screening of the Drugs for Neglected Diseases initiative (DNDi) HAT-library by the Swiss TPH had singled out fexinidazole, originally developed by Hoechst (now Sanofi), as the most promising of a series of over 800 nitroimidazoles and related molecules. In cell culture, fexinidazole has an IC50 of around 1 µM against Trypanosoma brucei and is more than 100-fold less toxic to mammalian cells. In the mouse model, fexinidazole cures both the first, haemolymphatic, and the second, meningoencephalitic stage of the infection, the latter at 100 mg/kg twice daily for 5 days. In patients, the clinical trials managed by DNDi and supported by Swiss TPH mainly conducted in the Democratic Republic of the Congo demonstrated that oral fexinidazole is safe and effective for use against first- and early second-stage sleeping sickness. Based on the positive opinion issued by the European Medicines Agency in 2018, the WHO has released new interim guidelines for the treatment of HAT including fexinidazole as the new therapy for first-stage and non-severe second-stage sleeping sickness caused by Trypanosoma brucei gambiense (gHAT). This greatly facilitates the diagnosis and treatment algorithm for gHAT, increasing the attainable coverage and paving the way towards the envisaged goal of zero transmission by 2030.

Safety and efficacy of oral fexinidazole in children with gambiense human African trypanosomiasis: a multicentre, single-arm, open-label, phase 2-3 trial.

BACKGROUND: Fexinidazole has been reported as an effective oral monotherapy against non-severe gambiense human African trypanosomiasis in a recent trial in adults. We aimed to assess the safety and efficacy of fexinidazole in children across all disease stages of gambiense human African trypanosomiasis. METHODS: We did a multicentre, single-arm, open-label, phase 2-3 trial at eight district hospitals in the Democratic Republic of the Congo. We recruited children with a Karnofsky score of more than 50, those aged 6 years to younger than 15 years, weighing 20 kg or more, and with confirmed gambiense human African trypanosomiasis (any stage). Children weighing 20 kg or more and less than 35 kg received oral fexinidazole of 1200 mg (two × 600 mg tablets) once per day for 4 days (days 1-4) followed by 600 mg (one × 600 mg tablet) once per day for 6 days (days 5-10). Children weighing 35 kg or more received oral fexinidazole of 1800 mg (three × 600 mg tablets) once per day for 4 days (days 1-4), followed by 1200 mg (two × 600 mg tablets) once per day for 6 days (days 5-10). The primary endpoint was fexinidazole treatment success rate 12 months after end of treatment. A rate greater than 80% was deemed acceptable and a target value of 92% was aimed for. Safety was assessed through routine monitoring. This study is completed and registered with ClinicalTrials.gov, number NCT02184689. FINDINGS: Between May 3, 2014, and Nov 22, 2016, we screened a total of 130 paediatric patients, of whom 125 (96%) received at least one dose of fexinidazole. All 125 patients (69 [55%] patients with stage 1, 19 [15%] with early stage 2, and 37 [30%] with late stage 2 gambiense human African trypanosomiasis) completed the 10-day treatment. Treatment success rate at 12 months was 97·6% (95% CI 93·1-99·5; 122 of 125 patients). The primary endpoint was met and the targeted value of 92% was exceeded. Treatment success at 12 months was elevated across all disease stages: 98·6% (95% CI 92·2-99·9; 68 of 69 patients) in stage 1, 94·7% (74·0-99·9; 18 of 19 patients) in early stage 2, and 97·3% (85·8-99·9; 36 of 37 patients) in late stage 2 gambiense human African trypanosomiasis. No new safety issues were observed beyond those found in adult trials. Overall, 116 (93%) of 125 patients reported 586 treatment-emergent adverse events, mainly mild or moderate. The most frequently reported treatment-emergent adverse events of interest during hospital admission were vomiting (86 [69%] of 125) and headache (41 [33%]). Seven (6%) of 125 patients had severe malaria, which was often accompanied by anaemia that was unrelated to fexinidazole. One patient died following dyspnoea and injury due to traumatic aggression 172 days after end of treatment, which was considered unrelated to fexinidazole or gambiense human African trypanosomiasis. INTERPRETATION: Oral fexinidazole is a safe and effective first-line treatment option across all gambiense human African trypanosomiasis disease stages in paediatric patients. FUNDING: Through the Drugs for Neglected Diseases initiative: the Bill & Melinda Gates Foundation (USA), the Republic and Canton of Geneva (Switzerland), the Dutch Ministry of Foreign Affairs (Netherlands), the Norwegian Agency for Development Cooperation (Norway), the Federal Ministry of Education and Research through KfW (Germany), the Brian Mercer Charitable Trust (UK), and other private foundations and individuals from the human African trypanosomiasis campaign. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Effectiveness of Nifurtimox Eflornithine Combination Therapy (NECT) in T. b. gambiense second stage sleeping sickness patients in the Democratic Republic of Congo: Report from a field study.

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.

Oral fexinidazole for stage 1 or early stage 2 African Trypanosoma brucei gambiense trypanosomiasis: a prospective, multicentre, open-label, cohort study.

BACKGROUND: 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. METHODS: In this prospective, multicentre, open-label, single-arm cohort study, patients with stage 1 or early stage 2 g-HAT were recruited from eight treatment centres in the Democratic Republic of the Congo. Primary inclusion criteria included being older than 15 years, being able to ingest at least one complete meal per day (or at least one sachet of Plumpy'Nut®), a Karnofsky score higher than 50, evidence of trypanosomes in the blood or lymph but no evidence of trypanosomes in the CSF, willingness to be admitted to hospital to receive treatment, having a permanent address, and being able to comply with the follow-up visit schedule. Exclusion criteria included severe malnutrition, inability to take medication orally, pregnant or breastfeeding women, any clinically important medical condition that could jeopardise patient safety or participation in the study, severely deteriorated general status, any contraindication to imidazole drugs, HAT treatment in the past 2 years, previous enrolment in the study or previous intake of fexinidazole, abnormalities on electrocardiogram that did not return to normal in pretreatment repeated assessments or were considered clinically important, QT interval corrected using Fridericia's formula of at least 450 ms, and patients not tested for malaria or not having received appropriate treatment for malaria or for soil-transmitted helminthiasis. Patients were classified into stage 1 or early stage 2 g-HAT groups following evidence of trypanosomes in the blood, lymph, and absence in CSF, and using white-blood-cell count in CSF. Patients received 1800 mg fexinidazole once per day on days 1-4 then 1200 mg fexinidazole on days 5-10. Patients were observed for approximately 19 months in total. Study participants were followed up on day 5 and day 8 during treatment, at end of treatment on day 11, at end of hospitalisation on days 11-18, at week 9 for a subset of patients, and after 6 months, 12 months, and 18 months. The primary endpoint was treatment success at 12 months. Safety was assessed through routine monitoring. Analyses were done in the intention-to-treat population. The acceptable success rate was defined as treatment efficacy in more than 80% of patients. This study is completed and registered with ClinicalTrials.gov (NCT02169557). FINDINGS: Patients were enrolled between April 30, 2014, and April 25, 2017. 238 patients were recruited: 195 (82%) patients with stage 1 g-HAT and 43 (18%) with early stage 2 g-HAT. 189 (97%) of 195 patients with stage 1 g-HAT and 41 (95%) of 43 patients with early stage 2 g-HAT were finally included and completed the 10 day treatment period. Three patients with stage 1 g-HAT died after the 10 day treatment period and before the 12 month primary follow-up visit, considered as treatment failure and were withdrawn from the study. Treatment was effective at 12 months for 227 (99%) of 230 patients (95% CI 96·2-99·7): 186 (98%) of 189 patients (95·4-99·7) with stage 1 and 41 (100%) of 41 patients (91·4-100·0) with early stage 2, indicating that the primary study endpoint was met. No new safety issues were observed. The most frequent adverse events were headache and vomiting. In total, 214 (93%) of 230 patients had treatment-emergent adverse events, mainly common-terminology criteria for adverse events grades 1 to 3. None led to treatment discontinuation. INTERPRETATION: Fexinidazole is a valuable first-line treatment option in the early stages of g-HAT. FUNDING: Through the Drugs for Neglected Diseases initiative: the Bill & Melinda Gates Foundation, the Republic and Canton of Geneva (Switzerland), the Dutch Ministry of Foreign Affairs (also known as DGIS; Netherlands), the Norwegian Agency for Development Cooperation (also known as Norad; Norway), the Federal Ministry of Education and Research (also known as BMBF) through KfW (Germany), the Brian Mercer Charitable Trust (UK), and other private foundations and individuals from the HAT campaign. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Oral fexinidazole for late-stage African Trypanosoma brucei gambiense trypanosomiasis: a pivotal multicentre, randomised, non-inferiority trial.

BACKGROUND: Few therapeutic options are available to treat the late-stage of human African trypanosomiasis, a neglected tropical disease, caused by Trypanosoma brucei gambiense (g-HAT). The firstline treatment is a combination therapy of oral nifurtimox and intravenous eflornithine that needs to be administered in a hospital setting by trained personnel, which is not optimal given that patients often live in remote areas with few health resources. Therefore, we aimed to assess the safety and efficacy of an oral regimen of fexinidazole (a 2-substituted 5-nitroimidazole with proven trypanocidal activity) versus nifurtimox eflornithine combination therapy in patients with late-stage g-HAT. METHODS: In this randomised, phase 2/3, open-label, non-inferiority trial, we recruited patients aged 15 years and older with late-stage g-HAT from g-HAT treatment centres in the Democratic Republic of the Congo (n=9) and the Central African Republic (n=1). Patients were randomly assigned (2:1) to receive either fexinidazole or nifurtimox eflornithine combination therapy according to a predefined randomisation list (block size six). The funder, data management personnel, and study statisticians were masked to treatment. Oral fexinidazole was given once a day (days 1-4: 1800 mg, days 5-10: 1200 mg). Oral nifurtimox was given three times a day (days 1-10: 15 mg/kg per day) with eflornithine twice a day as 2 h infusions (days 1-7: 400 mg/kg per day). The primary endpoint was success at 18 months (ie, deemed as patients being alive, having no evidence of trypanosomes in any body fluid, not requiring rescue medication, and having a cerebrospinal fluid white blood cell count ≤20 cells per µL). Safety was assessed through routine monitoring. Primary efficacy analysis was done in the modified intention-to-treat population and safety analyses in the intention-to-treat population. The acceptable margin for the difference in success rates was defined as 13%. This study has been completed and is registered with ClinicalTrials.gov, number NCT01685827. FINDINGS: Between October, 2012, and November, 2016, 419 patients were pre-screened. Of the 409 eligible patients, 14 were not included because they did not meet all inclusion criteria (n=12) or for another reason (n=2). Therefore, 394 patients were randomly assigned, 264 to receive fexinidazole and 130 to receive nifurtimox eflornithine combination therapy. Success at 18 months was recorded in 239 (91%) patients given fexinidazole and 124 (98%) patients given nifurtimox eflornithine combination therapy, within the margin of acceptable difference of -6·4% (97·06% CI -11·2 to -1·6; p=0·0029). We noted no difference in the proportion of patients who experienced treatment-related adverse events (215 [81%] in the fexinidazole group vs 102 [79%] in the nifurtimox eflornithine combination therapy group). Treatment discontinuations were unrelated to treatment (n=2 [1%] in the fexinidazole group). Temporary nifurtimox eflornithine combination therapy interruption occurred in three (2%) patients. 11 patients died during the study (nine [3%] in the fexinidazole group vs two [2%] in the nifurtimox eflornithine combination therapy group). INTERPRETATION: Our findings show that oral fexinidazole is effective and safe for the treatment of T b gambiense infection compared with nifurtimox eflornithine combination therapy in late-stage HAT patients. Fexinidazole could be a key asset in the elimination of this fatal neglected disease. FUNDING: Drugs for Neglected Diseases initiative.

Efficacy, Safety, and Dose of Pafuramidine, a New Oral Drug for Treatment of First Stage Sleeping Sickness, in a Phase 2a Clinical Study and Phase 2b Randomized Clinical Studies.

BACKGROUND: Sleeping sickness (human African trypanosomiasis [HAT]) is caused by protozoan parasites and characterized by a chronic progressive course, which may last up to several years before death. We conducted two Phase 2 studies to determine the efficacy and safety of oral pafuramidine in African patients with first stage HAT. METHODS: The Phase 2a study was an open-label, non-controlled, proof-of-concept study where 32 patients were treated with 100 mg of pafuramidine orally twice a day (BID) for 5 days at two trypanosomiasis reference centers (Angola and the Democratic Republic of the Congo [DRC]) between August 2001 and November 2004. The Phase 2b study compared pafuramidine in 41 patients versus standard pentamidine therapy in 40 patients. The Phase 2b study was open-label, parallel-group, controlled, randomized, and conducted at two sites in the DRC between April 2003 and February 2007. The Phase 2b study was then amended to add an open-label sequence (Phase 2b-2), where 30 patients received pafuramidine for 10 days. The primary efficacy endpoint was parasitologic cure at 24 hours (Phase 2a) or 3 months (Phase 2b) after treatment completion. The primary safety outcome was the rate of occurrence of World Health Organization Toxicity Scale Grade 3 or higher adverse events. All subjects provided written informed consent. FINDINGS/CONCLUSION: Pafuramidine for the treatment of first stage HAT was comparable in efficacy to pentamidine after 10 days of dosing. The cure rates 3 months post-treatment were 79% in the 5-day pafuramidine, 100% in the 7-day pentamidine, and 93% in the 10-day pafuramidine groups. In Phase 2b, the percentage of patients with at least 1 treatment-emergent adverse event was notably higher after pentamidine treatment (93%) than pafuramidine treatment for 5 days (25%) and 10 days (57%). These results support continuation of the development program for pafuramidine into Phase 3.

Efficacy and Safety of Pafuramidine versus Pentamidine Maleate for Treatment of First Stage Sleeping Sickness in a Randomized, Comparator-Controlled, International Phase 3 Clinical Trial.

BACKGROUND: Sleeping sickness (human African trypanosomiasis [HAT]) is a neglected tropical disease with limited treatment options that currently require parenteral administration. In previous studies, orally administered pafuramidine was well tolerated in healthy patients (for up to 21 days) and stage 1 HAT patients (for up to 10 days), and demonstrated efficacy comparable to pentamidine. METHODS: This was a Phase 3, multi-center, randomized, open-label, parallel-group, active control study where 273 male and female patients with first stage Trypanosoma brucei gambiense HAT were treated at six sites: one trypanosomiasis reference center in Angola, one hospital in South Sudan, and four hospitals in the Democratic Republic of the Congo between August 2005 and September 2009 to support the registration of pafuramidine for treatment of first stage HAT in collaboration with the United States Food and Drug Administration. Patients were treated with either 100 mg of pafuramidine orally twice a day for 10 days or 4 mg/kg pentamidine intramuscularly once daily for 7 days to assess the efficacy and safety of pafuramidine versus pentamidine. Pregnant and lactating women as well as adolescents were included. The primary efficacy endpoint was the combined rate of clinical and parasitological cure at 12 months. The primary safety outcome was the frequency and severity of adverse events. The study was registered on the International Clinical Trials Registry Platform at www.clinicaltrials.gov with the number ISRCTN85534673. FINDINGS/CONCLUSIONS: The overall cure rate at 12 months was 89% in the pafuramidine group and 95% in the pentamidine group; pafuramidine was non-inferior to pentamidine as the upper bound of the 95% confidence interval did not exceed 15%. The safety profile of pafuramidine was superior to pentamidine; however, 3 patients in the pafuramidine group had glomerulonephritis or nephropathy approximately 8 weeks post-treatment. Two of these events were judged as possibly related to pafuramidine. Despite good tolerability observed in preceding studies, the development program for pafuramidine was discontinued due to delayed post-treatment toxicity.

Melarsoprol- and pentamidine-resistant Trypanosoma brucei rhodesiense populations and their cross-resistance.

Resistance to melarsoprol and pentamidine was induced in bloodstream-form Trypanosoma brucei rhodesiense STIB 900 in vitro, and drug sensitivity was determined for melarsoprol, pentamidine and furamidine. The resistant populations were also inoculated into immunosuppressed mice to verify infectivity and to monitor whether rodent passage selects for clones with altered drug sensitivity. After proliferation in the mouse, trypanosomes were isolated and their IC(50) values to the three drugs were determined. To assess the stability of drug-induced resistance, drug pressure was ceased for 2 months and the drug sensitivity was determined again. Resistance was stable, with a few exceptions that are discussed. Drug IC(50)s indicated cross-resistance among all drugs, but to varying extents: resistance of the melarsoprol-selected and pentamidine-selected trypanosomes to pentamidine was the same, but the pentamidine-selected trypanosome population showed lower resistance to melarsoprol than the melarsoprol-selected trypanosomes. Interestingly, both resistant populations revealed the same intermediate cross-resistance to furamidine. Resistant trypanosome populations were characterised by molecular means, referring to the status of the TbAT1 gene. The melarsoprol-selected population apparently had lost TbAT1, whereas in the pentamidine-selected trypanosome population it was still present.