Blood-CNS transport mechanisms in pathophysiology and drug delivery
The blood-CNS barriers form a highly selective biological firewall; regulating the passage of solutes between the blood and cerebral microenvironment. This thesis modelled the blood-CNS barriers in vivo to determine the transport of nutrients, metabolic products and drugs across these barriers, and was split into two main parts: 1. L-arginine and ADMA transport - Circulating levels of the cationic amino acid L-arginine and its endogenously produced homologue asymmetric dimethylarginine (ADMA) are in delicately poised equilibrium. Dysregulation of this balance has well-documented implications in cardiovascular disease and also more recently conditions of the brain, as ADMA is a potent inhibitor of nitric oxide synthase (NOS) enzymes, including the endothelial (eNOS) and neuronal (nNOS) isoforms. Until now it has been postulated that ADMA competes for transport across membranes via the same cationic transport system as L-arginine (system y+), however this hypothesis has surprisingly never been tested. The major contributors to transport of both amino acids were investigated to determine how their respective intracellular and extracellular concentrations affect transport across the blood-CNS barriers. These data also have strong relevance to other research areas due to the widespread influence of NO in physiological pathways in health and disease. At the time of writing, this study represents the first true characterisation of ADMA transport across any physiological membrane in vivo and reveals a likely mechanism for explaining ‘the L-arginine paradox’ – the clinical observation that NO-deficient patients respond well to oral supplementation with L-arginine even though [arginine]plasma is easily sufficient to saturate eNOS. 2. ‘NanoHAT’ - Efflux transporters expressed at the blood-CNS barriers remain one of the biggest hurdles for the efficient delivery of therapeutic agents to the brain. The anti-trypanosomal drug pentamidine was shown previously by our group to be a substrate for efflux from the blood-CNS barriers and does not ordinarily accumulate to pharmacologically relevant levels in the brain. Pluronic® copolymers are known biological response modifiers that are approved for use in humans and have generated a great deal of interest due to their ability to inhibit efflux transporters and spontaneously form micelles in solution. In this study, a coformulation of pentamidine and Pluronic® P85 was evaluated as a potential future treatment for CNS-stage Human African Trypanosomiasis, using a combination of in silico, in vitro, and in vivo methodologies. Combining our group’s knowledge and expertise in blood-brain barrier research with colleagues in Molecular Biophysics and Materials & Molecular Modelling has resulted in the creation of a ‘mini formulation-development pathway’ that can be utilised to optimise and develop formulations in the future.
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