28 August 2009. The fine balance between potency and selectivity that looms large in drug development has long stymied efforts to target the M1 muscarinic acetylcholine receptor (mAChR) for treatment of Alzheimer disease and other cognitive disorders. However, hopes of bringing M1’s therapeutic potential to the clinic have resurged with recent data hinting at better success by targeting allosteric sites. In this week’s Early Edition of the Proceedings of the National Academy of Sciences, researchers led by William James Ray at Merck & Co. in West Point, Pennsylvania, describe an allosteric compound that potentiates M1 activity with high specificity and relieves some cognitive and behavioral symptoms in mice. Ray presented the gist of these findings at last year’s Keystone conference on AD (see ARF related news story); the paper offers additional proof of selectivity in animal models.
The selectivity data is critical. The acetylcholine binding site is highly conserved among all types of mACh receptor. For this reason, clinically efficacious M1 compounds that target that region have fizzled due to nasty cholinergic side effects, mostly gastrointestinal, stemming from cross-reactivity (see, e.g., Bodick et al., 1997 on xanomeline). More recent efforts have shifted toward identifying compounds that selectively enhance M1 activity by targeting unique sites away from the substrate-binding action. By and large, this approach appears promising. It has produced a number of allosteric compounds that bind M1 with higher specificity than molecules targeting the acetylcholine site—including several M1 agonists (see Jones et al., 2008 and ARF related news story; and Vanover et al., 2008), and a set of positive allosteric modulators (Marlo et al., 2009).
The published Merck compound, benzyl quinolone carboxylic acid (BQCA), falls into the latter category. Unlike an agonist, it does not activate M1 in the absence of acetylcholine. Instead, it functions by reducing the transmitter concentration required to stimulate M1—in this case, by up to 129-fold. First author Lei Ma and colleagues demonstrated this by applying 100 microM BQCA to CHO cells that stably expressed human, rhesus, dog, rat, or mouse M1. BQCA had no effect on M2-M5, though, indicating the compound had more than 100-fold selectivity for M1.
To further confirm the specificity, the researchers treated primary mouse cortical neurons with BQCA or vehicle, then with acetylcholine, and measured levels of the inositol triphosphate (IP3) metabolite IP1 (which rise in response to M1, M3, or M5 activation) using a fluorescence assay. In wild-type mice, acetylcholine increased IP1 levels, and BQCA potentiated this effect about 23-fold. However, in M1 knockout mice, IP1 levels rose only in response to acetylcholine, and showed no effect with BQCA.
Distinguishing itself from other M1-selective compounds, BQCA relieves both cognitive and psychotic-like symptoms in animal models, Ray wrote in an e-mail to ARF. This bears some resemblance to an α7 nicotinic AChR agonist that has shown similar dual benefits in patients with AD and with schizophrenia (see ARF related conference story). In the current study, BQCA, but not M1 allosteric agonists TBPB and AC-42, was able to reverse memory loss induced by the muscarinic antagonist scopolamine in a contextual fear-conditioning regimen. BQCA also differs mechanistically from the other two compounds. In a β-galactosidase assay, only BQCA and its analogs potentiated acetylcholine-induced recruitment of β-arrestin (which mediates G protein-coupled receptor internalization) with reasonable potency.
Furthermore, BQCA and its analogs have drug-like properties. The compounds cross the blood-brain barrier, are readily handled by the gut, have good metabolic properties, and don’t inhibit any other enzymes, transporters, or ion channels out of more than 300 tested, Ray noted. BQCA itself is not a clinical compound, but Ray was unable to state exactly where in the pipeline potential analogs are at this point. (Pharma companies frequently prohibit their researchers from disclosing pipeline advances.)
The new data “represent a fundamental advance and provide further evidence that it will be possible to develop highly selective activators of the M1 muscarinic receptor by targeting allosteric sites,” wrote Jeffrey Conn of Vanderbilt University in Nashville, Tennessee, in an e-mail to ARF. Conn’s lab published a recent paper demonstrating TBPB’s antipsychotic-like activity in rats (Jones et al., 2008 and ARF related news story).
“BQCA is the first M1 allosteric potentiator that has both exquisite selectivity and pharmacokinetic properties needed for use in studies in animal models that are relevant to Alzheimer disease,” Conn wrote. “Thus, when combined with recently described highly selective systemically active M1 allosteric agonists, this provides the first set of tools to allow systematic studies aimed at determining the optimal properties of a drug that should be advanced as a clinical candidate for treatment of AD and other disorders.”(Full comment below; for reviews, see Conn et al., 2009 and
Fisher, 2008).—Esther Landhuis.
Ma L, Seager M, Wittmann M, Jacobson M, Bickel D, Burno M, Jones K, Graufelds VK, Xu G, Pearson M, McCampbell A, Gaspar R, Shughrue P, Danziger A, Regan C, Flick R, Pascarella D, Garson S, Doran S, Kreatsoulas C, Veng L, Lindsley CW, Shipe W, Kuduk S, Sur C, Kinney G, Seabrook GR, Ray WJ. Selective activation of the M1 muscarinic acetylcholine receptor achieved by allosteric potentiation. PNAS Early Edition. 26 August 2009. Abstract