Drug–receptor interaction has fascinated researchers since antiquity. It has been evolved from a concept of “Lock and Key” to the use of Furchgott's method (1966) to quantify dissociation constants and relative efficacies of agonist–receptor complexes. In the last 40 years, great strides in developments in molecular pharmacology have resulted in the accumulation of a vast amount of knowledge. This article briefly summarizes the recent developments in drug–receptor interactions with focus on inverse agonism and its potential therapeutic utility.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3195115/
For the RORgamma transcription factor I would definitely choose a functional assay like cytokine release (for example IL-17 release from Th17 cell, etc) or proliferation of appropriate cells, depending on your system studied. You should establish a measurable basal activity in your assay and look for the decrease of this activity due to the treatment with compounds in interest. I think binding assays developed for the ligand binding domain of RORs are not meaningful for inverse agonism.
Cheers, Karoly
Dear Dr. Karoly Liliom,
Thank you so much for your reply.
Regarding functional assay of IL17 inhibition, how can we differentiate the antagonist and inverse agonist. Because both will inhibit the IL17 production.
Please let me know your thoughts.
Regards,
Kolla
Dear Kolla,
I suggest first you establish a dose-response relationship with a known agonist for reference. Then you measure the dose-response with your compounds alone as well as together with the agonist. If a compound is an antagonist, you will see no change of basal activity if measured alone, while observe a parallel right-shift of the agonist dose-response curve when measured together. In case of an inverse agonist you also observe a shift when measured together, but must observe a decline in the dose-response curve with increasing concentration below the basal level. This can be observed only if a measurable basal activity is present. If a potential inverse agonist compound is identified this way, then you might design binding assays to see where the compound binds and how it affects the binding of agonist (to differentiate the functional effect from an allosteric negative co-operativity effect), but this is a more complicated issue and you need purified protein for that.
Regards,
Karoly
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