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Discovery of a Novel Chemical Class of mGlu(5) Allosteric Ligands with Distinct Modes of Pharmacology.

Hammond AS, Rodriguez AL, Townsend SD, Niswender CM, Gregory KJ, Lindsley CW, Conn PJ - ACS Chem Neurosci (2010)

Bottom Line: We previously discovered a positive allosteric modulator (PAM) of the metabotropic glutamate receptor subtype 5 (mGlu(5)) termed 4 N-{4-chloro-2-[(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl]phenyl}-2-hydroxybenzamide (CPPHA) that elicits receptor activation through a novel allosteric site on mGlu(5), distinct from the classical mGlu(5) negative allosteric modulator (NAM) MPEP allosteric site.An iterative parallel synthesis approach delivered 22 analogues, optimized mGlu(5) PAM activity to afford VU0357121 (EC(50) = 33 nM, 92% Glu(max)), and also revealed the first non-MPEP site neutral allosteric ligand (VU0365396).Moreover, mutagenesis studies indicate that VU0357121 and related analogues bind to a yet uncharacterized allosteric site on mGlu(5), distinct from CPPHA, yet share a functional interaction with the MPEP site.

View Article: PubMed Central - PubMed

ABSTRACT
We previously discovered a positive allosteric modulator (PAM) of the metabotropic glutamate receptor subtype 5 (mGlu(5)) termed 4 N-{4-chloro-2-[(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl]phenyl}-2-hydroxybenzamide (CPPHA) that elicits receptor activation through a novel allosteric site on mGlu(5), distinct from the classical mGlu(5) negative allosteric modulator (NAM) MPEP allosteric site. However, a shallow structure-activity relationship (SAR), poor physiochemical properties, and weak PAM activity at rat mGlu(5) limited the utility of CPPHA to explore allosteric activation of mGlu(5) at a non-MPEP site. Thus, we performed a functional high-throughput screen (HTS) and identified a novel mGlu(5) PAM benzamide scaffold, exemplified by VU0001850 (EC(50) = 1.3 μM, 106% Glu(max)) and VU0040237 (EC(50) = 350 nM, 84% Glu Max). An iterative parallel synthesis approach delivered 22 analogues, optimized mGlu(5) PAM activity to afford VU0357121 (EC(50) = 33 nM, 92% Glu(max)), and also revealed the first non-MPEP site neutral allosteric ligand (VU0365396). Like CPPHA, PAMs within this class do not appear to bind at the MPEP allosteric site based on radioligand binding studies. Moreover, mutagenesis studies indicate that VU0357121 and related analogues bind to a yet uncharacterized allosteric site on mGlu(5), distinct from CPPHA, yet share a functional interaction with the MPEP site.

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Related in: MedlinePlus

Unlike MPEP, no benzamide analogues were able to antagonize a submaximal (EC80) concentration of glutamate. HEK cells were pretreated with a fixed concentration of test compound (10 μM) or vehicle, followed by EC80 glutamate. Data were normalized to the average maximal response obtained from each experiment as determined by 100 μM glutamate. Data were obtained from three separate experiments, each performed in triplicate, and are expressed as the mean ± SEM.
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fig12: Unlike MPEP, no benzamide analogues were able to antagonize a submaximal (EC80) concentration of glutamate. HEK cells were pretreated with a fixed concentration of test compound (10 μM) or vehicle, followed by EC80 glutamate. Data were normalized to the average maximal response obtained from each experiment as determined by 100 μM glutamate. Data were obtained from three separate experiments, each performed in triplicate, and are expressed as the mean ± SEM.

Mentions: Therefore, these compounds were also screened in a manner to identify NAMs or noncompetitive antagonists of mGlu5. For the antagonist screen, we determined whether the test compounds could inhibit the effect of glutamate on mGlu5-expressing HEK293 cells. The calcium mobilization assay was performed as described above, with cells being treated with test compounds at a fixed concentration of 10 μM, except that they were stimulated with a submaximal concentration of glutamate (∼EC80). In contrast to the blockade induced by the mGlu5 NAM MPEP, VU0357121 and analogues did not inhibit the effect of glutamate on mGlu5; in fact, the majority of compounds induced a mild potentiation of the EC80 glutamate response (Figure 12).


Discovery of a Novel Chemical Class of mGlu(5) Allosteric Ligands with Distinct Modes of Pharmacology.

Hammond AS, Rodriguez AL, Townsend SD, Niswender CM, Gregory KJ, Lindsley CW, Conn PJ - ACS Chem Neurosci (2010)

Unlike MPEP, no benzamide analogues were able to antagonize a submaximal (EC80) concentration of glutamate. HEK cells were pretreated with a fixed concentration of test compound (10 μM) or vehicle, followed by EC80 glutamate. Data were normalized to the average maximal response obtained from each experiment as determined by 100 μM glutamate. Data were obtained from three separate experiments, each performed in triplicate, and are expressed as the mean ± SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2957851&req=5

fig12: Unlike MPEP, no benzamide analogues were able to antagonize a submaximal (EC80) concentration of glutamate. HEK cells were pretreated with a fixed concentration of test compound (10 μM) or vehicle, followed by EC80 glutamate. Data were normalized to the average maximal response obtained from each experiment as determined by 100 μM glutamate. Data were obtained from three separate experiments, each performed in triplicate, and are expressed as the mean ± SEM.
Mentions: Therefore, these compounds were also screened in a manner to identify NAMs or noncompetitive antagonists of mGlu5. For the antagonist screen, we determined whether the test compounds could inhibit the effect of glutamate on mGlu5-expressing HEK293 cells. The calcium mobilization assay was performed as described above, with cells being treated with test compounds at a fixed concentration of 10 μM, except that they were stimulated with a submaximal concentration of glutamate (∼EC80). In contrast to the blockade induced by the mGlu5 NAM MPEP, VU0357121 and analogues did not inhibit the effect of glutamate on mGlu5; in fact, the majority of compounds induced a mild potentiation of the EC80 glutamate response (Figure 12).

Bottom Line: We previously discovered a positive allosteric modulator (PAM) of the metabotropic glutamate receptor subtype 5 (mGlu(5)) termed 4 N-{4-chloro-2-[(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl]phenyl}-2-hydroxybenzamide (CPPHA) that elicits receptor activation through a novel allosteric site on mGlu(5), distinct from the classical mGlu(5) negative allosteric modulator (NAM) MPEP allosteric site.An iterative parallel synthesis approach delivered 22 analogues, optimized mGlu(5) PAM activity to afford VU0357121 (EC(50) = 33 nM, 92% Glu(max)), and also revealed the first non-MPEP site neutral allosteric ligand (VU0365396).Moreover, mutagenesis studies indicate that VU0357121 and related analogues bind to a yet uncharacterized allosteric site on mGlu(5), distinct from CPPHA, yet share a functional interaction with the MPEP site.

View Article: PubMed Central - PubMed

ABSTRACT
We previously discovered a positive allosteric modulator (PAM) of the metabotropic glutamate receptor subtype 5 (mGlu(5)) termed 4 N-{4-chloro-2-[(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl]phenyl}-2-hydroxybenzamide (CPPHA) that elicits receptor activation through a novel allosteric site on mGlu(5), distinct from the classical mGlu(5) negative allosteric modulator (NAM) MPEP allosteric site. However, a shallow structure-activity relationship (SAR), poor physiochemical properties, and weak PAM activity at rat mGlu(5) limited the utility of CPPHA to explore allosteric activation of mGlu(5) at a non-MPEP site. Thus, we performed a functional high-throughput screen (HTS) and identified a novel mGlu(5) PAM benzamide scaffold, exemplified by VU0001850 (EC(50) = 1.3 μM, 106% Glu(max)) and VU0040237 (EC(50) = 350 nM, 84% Glu Max). An iterative parallel synthesis approach delivered 22 analogues, optimized mGlu(5) PAM activity to afford VU0357121 (EC(50) = 33 nM, 92% Glu(max)), and also revealed the first non-MPEP site neutral allosteric ligand (VU0365396). Like CPPHA, PAMs within this class do not appear to bind at the MPEP allosteric site based on radioligand binding studies. Moreover, mutagenesis studies indicate that VU0357121 and related analogues bind to a yet uncharacterized allosteric site on mGlu(5), distinct from CPPHA, yet share a functional interaction with the MPEP site.

No MeSH data available.


Related in: MedlinePlus