The ex vivo BBB permeability model enables BBB permeability studies of compounds at a constant brain exposure level. In this model setting, whole brains, complete with an intact brain barrier, are exposed to a test drug dissolved in buffer at a pre-defined concentration level. The insect ex vivo BBB permeability model has been tested in an in-house proof of concept study where a number of known drugs were tested.
The chart below shows the results obtained from testing known drugs varying degrees of intrinsic CNS permeability in the Locust ex vivo BBB model at 3 and 10 uM. Green bars represent drugs with high intrinsic CNS permeability while the red bars represent non-CNS drugs.
Figure text: The locust ex vivo whole brain model can discriminate between mammalian CNS and non-CNS drugs after 5 min constant compound exposure at 30 dgr C. Results are mean +/- SD (n=3)
Since P-gp mediated drug efflux is a major hurdle in CNS drug discovery, it is important to identify actively effluxed compounds in the early screen phase. The degree of active efflux can be quantified in the ex vivo Locust BBB model since a P-gp homolog (MDR65) is expressed in the insect brain barrier. This is exemplified with a study where locust brains were exposed to various concentrations of carbamazepine (a non P-gp CNS drug) and quinidine (a well known P-gp substrate) in the presence and absence of the prototypical P-gp inhibitor verapamil. As can be seen in the figures, high and consistent brain exposure was obtained after carbamazepine exposure regardless of verapamil presence. In contrast, quinidine brain concentrations were considerably lower, in particular in the absence of verapamil demonstrating an impact of efflux transporters in this system. Interestingly, the efflux ratio of quinidine (defined as the ratio of brain levels after co-incubation with and without verapamil) is reduced from 5 at 10 uM to 1.7 at 100 uM, suggesting a partial saturation of quinidine efflux, something that would be expected for P-gp mediated efflux in this concentration range.
Figure text: Locust brains were exposed to 10, 30 and 100 uM of carbamazepine and quinidine in the ex vivo locust model in order to assess the initial rate of absorption. The brains were exposed for 5 minutes after which the brains were recovered, homogenized and analysed. High and consistent levels of carbamazepine were detected in a concentration dependent manner, in the absence and presence of the P-gp inhibitor verapamil. The brain levels of quinidine were considerably lower in the absence of P-gp inhibition but the levels were reproducibly increased in the presence of verapamil.
Results are mean amounts drug per locust brain +/- sd (n=3)