Identification and characterization of an atypical Gαs-biased β2AR agonist that fails to evoke airway smooth muscle cell tachyphylaxis

Donghwa Kim, Alina Tokmakova, Lauren K. Lujan, Hannah R. Strzelinski, Nicholas Kim, Maliheh Najari Beidokhti, Marc A. Giulianotti, Amirhossein Mafi, Jung A.A. Woo, Steven S. An, William A. Goddard, Stephen B. Liggett

Research output: Contribution to journalArticlepeer-review

11 Scopus citations


G protein–coupled receptors display multifunctional signaling, offering the potential for agonist structures to promote conformational selectivity for biased outputs. For β2-adrenergic receptors (β2AR), unbiased agonists stabilize conformation(s) that evoke coupling to Gαs (cyclic adenosine monophosphate [cAMP] production/human airway smooth muscle [HASM] cell relaxation) and β-arrestin engagement, the latter acting to quench Gαs signaling, contributing to receptor desensitization/tachyphylaxis. We screened a 40-million-compound scaffold ranking library, revealing unanticipated agonists with dihydroimidazolyl-butyl-cyclic urea scaffolds. The S-stereoisomer of compound C1 shows no detectable β-arrestin engagement/signaling by four methods. However, C1-S retained Gαs signaling—a divergence of the outputs favorable for treating asthma. Functional studies with two models confirmed the biasing: β2AR-mediated cAMP signaling underwent desensitization to the unbiased agonist albuterol but not to C1-S, and desensitization of HASM cell relaxation was observed with albuterol but not with C1-S. These HASM results indicate biologically pertinent biasing of C1-S, in the context of the relevant physiologic response, in the human cell type of interest. Thus, C1-S was apparently strongly biased away from β-arrestin, in contrast to albuterol and C5-S. C1-S structural modeling and simulations revealed binding differences compared with unbiased epinephrine at transmembrane (TM) segments 3,5,6,7 and ECL2. C1-S (R2 = cyclohexane) was repositioned in the pocket such that it lost a TM6 interaction and gained a TM7 interaction compared with the analogous unbiased C5-S (R2 = benzene group), which appears to contribute to C1-S biasing away from β-arrestin. Thus, an agnostic large chemical-space library identified agonists with receptor interactions that resulted in relevant signal splitting of β2AR actions favorable for treating obstructive lung disease.

Original languageEnglish (US)
Article numbere2026668118
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number49
StatePublished - Dec 7 2021
Externally publishedYes

Bibliographical note

Funding Information:
ACKNOWLEDGMENTS. We thank Michel Bouvier for the Ala293 construct, Himeshkumar Patel and Christine Tam for assistance in manuscript preparation, and the NIH for support from Grants HL045967, HL155532, and HL114471 (to S.B.L.).

Publisher Copyright:
© 2021 National Academy of Sciences. All rights reserved.


  • Asthma
  • Biasing
  • Gs
  • β-agonist
  • β-arrestin


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