Endoglin Regulation of Oatp-Mediated Drug Transport at the BBB via TGF-beta Signaling in Stroke

Abstract

Stroke is one of the leading causes of death and disability both within the United States and across the globe. Currently, only one pharmacological treatment (i.e., r-tPA) is FDA approved and has limited effectiveness due to its narrow therapeutic window (4.5 hrs) following stroke onset. Therefore, novel therapeutic paradigms for the treatment of ischemic injury are needed to improve patient outcomes and reduce mortality. Drug delivery at therapeutic concentrations is challenging due to the presence of the blood brain barrier (BBB), limiting passage of small and large molecular drugs. Regulated by numerous cells within the neurovascular unit (NVU), alterations to the BBB during stroke pathogenesis further impair adequate and consistent delivery of neuroprotective compounds. Endogenous transporters within endothelial cells allow for transcellular delivery of pharmacotherapeutics and provide a potential mechanism which can be targeted during ischemic stroke to increase CNS drug delivery. One such group of uptake transporters, the organic anion transporting polypeptides (OATPs in humans; Oatps in rodents), has been shown to transport 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (i.e., statins). Clinical studies provide evidence that statins elicit neuroprotective effects which suggests that statins can be utilized as stroke therapeutic agents. An in depth understanding of statin delivery to the brain and their corresponding effects as a stroke treatment are essential. Additionally, manipulation of CNS drug delivery via uptake transporter expression and function within the BBB is a potential method to increase the translational effectiveness of neuroprotective compounds which are endogenous transporter substrates. Previous work from our lab has shown that Oatp manipulation can be achieved through activation of the TGF-?/ALK1 signaling pathway. We demonstrate regional differences in TGF-?/ALK1 signaling response to bone morphogenic protein (BMP), a known ALK1 agonist, resulting in increased Oatp expression and functional uptake of statins within cortical microvessels. Further assessment of reasons behind the regiospecific responses observed, we found the TGF-?/ALK1 co-receptor, endoglin, expressed differentially between the cortex, hippocampus, and cerebellum. Assessing the necessity of endoglin in mediating Oatp1a4 expression and function demonstrated attenuated responses to BMP treatment in endothelial cells which were lacking cell surface endoglin. Utilizing a rodent model for focal ischemia, statin administration was shown to have a Oatp-dependent effect on outcome improvement. Treatment with atorvastatin reduced neuronal damage and edema which was reversed upon inhibition of Oatp1a4. As endoglin is regulated by hypoxia, changes to endoglin were observed following in vitro models of ischemia. Overall, the data demonstrates that adequate CNS delivery of atorvastatin requires functional Oatp transport across the BBB which could potentially be altered by targeting endoglin signaling. Combination therapy by targeting endogenous transporters via TGF-?/ALK1signaling could translate to improved stroke therapeutics.