Synthesis and characterization of bivalent P-glycoprotein inhibitors to enhance brain penetration of CNS therapies
Abstract
P-glycoprotein (P-gp), a member of the ATP-binding cassette (ABC) family of active transport proteins, prevents the access of numerous therapies to the brain due to its presence at the blood brain barrier (BBB). Our research has focused on investigating ways to increase the brain bioavailability of agents targeted against malaria, Alzheimer's disease, HIV, bipolar disorder and schizophrenia. We have found that dimerization of P-gp substrates generates potent inhibitors of P-gp transport. Thus, these novel prodrug dimeric inhibitors have a dual role: inhibition of P-gp and reversion to therapeutic monomer once inside the cell. We hypothesize that administration of the dimeric prodrug inhibitor would serve to increase the level of the therapeutic agent in the brain and potentially lower the overall patient dose level. We generated a reversibly linked quinine homodimer, Q2, as a proof of principle. The reversibility of the tether was confirmed when Q2 was incubated in the presence of esterase in vitro (t½ ∼ 20 h). Further studies revealed that Q2 was a particularly potent P-gp inhibitor, as revealed by substrate accumulation and binding competition assays. This method has since been utilized to generate additional homodimeric prodrug P-gp inhibitors. For instance, dimerization of the antipsychotic quetiapine generated a library of powerful P-gp inhibitors. The shortest tethered homodimer, Qt-C2 was not plasma stable (t½ ∼ 3 h), prompting the synthesis of a hindered prodrug, Qt-C2-Me2 (t ½ ∼ 38 h). Qt-C2-Me2 was an equally potent inhibitor as Qt-C2 as revealed by substrate accumulation assays (IC50 values from 1-3 μM for both Qt-C2 and Qt-C2-Me2). Substrate binding competition assays revealed a very strong interaction with the P-gp substrate binding sites (∼200 nM for both Qt-C2 and Qt-C2-Me2). Finally, P-gp in rat brain capillaries was inhibited by both homodimers. While Qt-C 2 brought luminal fluorescence to control levels (5 μM PSC833) at 5 μM, only 1 μM Qt-C2-Me2 was required to elicit the same response. Overall, these findings show that quetiapine homodimers strongly inhibit P-gp transport activity not only in cultured cells, but also at the BBB.
Degree
Ph.D.
Advisors
Hrycyna, Purdue University.
Subject Area
Biochemistry
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