NANOHAT: Development of a safer and more effective sleeping sickness drug.

Sleeping sickness or human African Trypanosomiasis (HAT) is a potentially fatal disease caused by the parasite, which is transferred to humans by the bite of an infected tsetse fly. After being bitten, the infected human will experience two stages of disease: the first stage is the blood-based stage of the disease and the second stage affects the central nervous system (CNS), which occurs once the parasite penetrates the brain from the blood. The infected human will die if Stage 2 HAT is left untreated. The blood-brain barrier (BBB) makes the CNS stage difficult to treat because it prevents 99% of all known drugs from entering the brain from the blood, including some anti-HAT drugs. Those anti-HAT drugs that do enter the brain are toxic compounds in their own right and have serious side effects. Pentamidine is a less toxic blood stage drug, which our research has shown has a limited ability to cross the BBB due to its removal by efflux transporters, which are membrane proteins that remove toxic substances from cells. Furthermore transporters are considered essential in the mode of action of pentamidine against the parasites that cause HAT. The objective of this multi-disciplinary study is to use nanotechnology to both improve the delivery of pentamidine to the brain and into the parasite, whilst reducing its side effects.

In doing so, we will create a novel formulation that will encapsulate pentamidine. Specifically, we will use an amphiphilic polymer (a long molecule composed of many repeating units, bearing water-loving and water-hating blocks), which spontaneously self-assembles into small aggregates called micelles. These micelles can be used to encapsulate the drug, thus prolonging its circulation time and protecting it. In addition, these specific polymers (namely, Pluronics) have been shown to interact with efflux transporters and thus allow passage of the drug across the BBB. This indicates that this improved drug formulation has potential to treat Stage 2 disease.

The optimal formulation will be determined using an iterative screening process that will utilise a series of cutting-edge research techniques from various biophysical, pharmaceutical and biological disciplines. The end product of this research will be a formulation which will be ready to be taken to pre-clinical testing for the treatment of Stage 2 HAT.

Human African trypanosomiasis (HAT or sleeping sickness) is a potentially fatal disease caused by the parasite Trypanosoma brucei sspp. The disease has two stages - a haemolymphatic stage after the bite of an infected tsetse fly, followed by a central nervous system (CNS) stage where the parasite penetrates the brain, causing death if left untreated. The blood-brain barrier (BBB) makes the CNS stage difficult to treat because it prevents 99% of all known compounds from entering the brain, including some anti-HAT drugs. Those that do enter the brain are toxic compounds in their own right and have serious side effects. Pentamidine is a less toxic blood stage drug, which our research has shown has a limited ability to cross the BBB due to its removal by efflux transporters. Furthermore transporters are considered essential in the mode of action of pentamidine against trypanosomes. The objective of this multi-disciplinary study is to use nanotechnology to both improve the delivery of pentamidine to the brain and trypanosomes, whilst reducing its side effects.

WHO MIGHT BENEFIT FROM THIS RESEARCH?
The proposed studies will generate an optimal Pluronic/pentamidine formulation for treating Stage 2 HAT and should also reduce adverse events associated with pentamidine treatment of Stage 1 HAT. Therefore the primary beneficiaries of this work will be the patients being supported by the National Sleeping Sickness Control Programmes (NSSCPs) and the Non-Government Organisations (NGOs) responsible for HAT control and treatment programmes.

The findings of these studies will also provide valuable information to scientists studying the self-assembly, drug encapsulation and shape and size of resulting aggregates of Pluronics, the efficacy of Pluronics on the ability of pentamidine to cross the BBB, the toxicity and efficacy of the formulations (in vitro & in vivo) and the efficacy of these formulations in live animal models, both nationally and internationally. More generally, the results should be of interest to the greater drug delivery community as Pluronics are currently of significant interest as drug delivery vehicles so all of the findings can be used as a basis for scientists considering using them to deliver different drugs or to treat different diseases.

HOW MIGHT THEY BENEFIT FROM THIS RESEARCH?
Although not a true measure of incidence, analysis of treatment rates for stage 2 HAT published by the WHO (17th Expert Committee on the Selection and Use of Essential Medicines (2009)) indicate that the number of patients diagnosed with and treated for Stage 2 HAT caused by T.b. gambiense is decreasing year on year, but remains steady at ~60% of the annual new infection rate. Thus in 2007 (the latest year for which data are available) 10,473 new cases of T.b. gambiense were detected and 6,461 cases of Stage 2 HAT were treated, yielding a current treatment population of approximately 6,500 per annum, around one fifth of total worldwide cases.

The majority of Stage 2 treatment is moving away from the more toxic melarsoprol, towards either eflornithine, or nifurtimox/eflornithine combination therapy (NECT), which requires 7 days of twice daily intravenous, as well as 10 days of 4 times daily oral therapy. Our solution provides the benefit of being able to treat Stage 2 disease with an existing drug that is currently used to treat Stage 1 disease. It will simplify the treatment regimes and replace eflornithine and NECT therapy in Stage 2 disease. Our combined pentamidine-Pluronic formulation has the potential to provide a single therapeutic entity for the safer, simpler and cost-effective treatment of all Stages of HAT.

More than 98% of all drugs discovered for the brain cannot be used, because they do not cross the BBB. Our solution could be used to re-formulate drugs for CNS delivery or other compartments where access is limited by efflux pumps.

Furthermore, the success of pentamidine against HAT has led to other diamidine compounds being developed. Pentamidine is also used in the treatment of American cutaneous leishmaniasis and pentamidine transporters are expressed in certain Leishmania species, including P-gp-like and MRP-like (pentamidine resistance protein 1) transporters. Pluronics could improve efficacy of other drugs (including diamidine analogues) that target trypanosomes or leishmania.