Proceedings of The Physiological Society
King's College London (2011) Proc Physiol Soc 22, PC33
The effect of glioblastoma cells on the function of efflux transporters in the blood-brain barrier.
A. Georgian1, N. J. Abbott1, D. J. Begley1, G. Nichols2, J. E. Preston1
1. Institute of Pharmaceutical Science, King's College London, London, United Kingdom. 2. Clinical Pharmacology & DMPK, AstraZeneca, Cheshire, United Kingdom.
Background and Aim: Efflux transporters such as P-glycoprotein (Pgp), breast cancer resistance protein (BCRP), and multi-drug resistance associated proteins (MRPs), are expressed on the blood- facing surface of blood-brain barrier (BBB) endothelia, and help restrict the movement of compounds from blood to brain. This has been a major problem for delivery of therapeutic drugs to the brain and particularly in treatment of brain tumours because chemotherapeutics are substrates for efflux transporters. It is unclear however why brain tumours are so resistant to chemotherapy and whether BBB efflux transporters play a role. Here we investigate whether BBB efflux transporters are affected during tumour progression in the brain using an in vitro assay. Methods: An efflux activity assay was developed, where 3H-MPP+ (a substrate of Pgp, BCRP and MRPs) was loaded into primary porcine brain endothelial cells (PBECs), and 3H-MPP+ efflux into an extracellular buffer was then measured in the presence and absence of Pgp, BCRP and MRPs inhibitors (verapamil 50μM, haloperidol 60μM, prazosin 35μM Ko143 0.2μM MK571 10μM). Efflux studies were conducted on PBECs in mono-culture or in non-contact co-culture with either primary rat astrocytes or C6 rat glioblastoma cells on suspended Transwell® filters. Results and Conclusions: The presence of inhibitors decreased apical efflux of 3H-MPP+ from PBECs co-cultured with primary astrocytes, demonstrating functionally active Pgp, BCRP and MRPs at the apical (blood face) of the endothelia. The % of apical efflux due to activity of each of the transporters was Pgp 29.4±1.8%, BCRP 20.0±0.7%, MRPs 14.9±0.8% (n=3-8) demonstrating the dominance of Pgp in these cells. When BBB endothelial cells are exposed to C6 glioma cells, Pgp and MRP activity was attenuated, but in contrast BCRP activity was up-regulated. MPP+ efflux due to each transporter was; Pgp 17.8±0.9%, BCRP 29.6±1.5%, MRPs 9.2±0.4%. Down-regulation of Pgp and MRPs would be beneficial for delivery of some chemotherapeutics which are substrates for these efflux transporters, however, up-regulation of BCRP may partially compensate for the decrease in other transporter activity. It may, however, be beneficial for treatment strategies to avoid chemotherapeutics that are substrates for BCRP, such as the anthracyclines (doxorubicin, mitoxantrone) used in the treatment of gliomas.
Where applicable, experiments conform with Society ethical requirements