Mechanistic insights into FX-909: a next-generation PPARγ inverse agonist

Posted by daviskd2 on Thursday, December 11, 2025 in News.

By Cameron I. Cohen

Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor transcription factor which regulates the expression of genes involved in differentiation, metabolism, adipogenesis, and insulin sensitization. PPARγ consists of an N-terminal disordered activation domain (NTD), a central DNA-binding domain (DBD), and a C-terminal ligand-binding domain (LBD). The activation and repression of PPARγ is mediated by the recruitment of coactivator and corepressor complexes, whose binding to PPARγ is modulated further by agonist binding. This complex regulatory cycle is necessary for proper cell function, as evidenced by the fact that aberrant PPARγ signaling occurs in luminal muscle-invasive bladder cancer. Therefore, the development of pharmaceuticals which target PPARγ-mediated transcription is an area of popular research.

In 2002, a covalent PPARγ antagonist, T0070907, was developed which effectively blocked agonist binding and reduced PPARγ-mediated transcription. However, T0070907 was not equally effective against all PPARγ ligands. Furthermore, T0070907 was shown to act not solely as an antagonist, but also as an inverse agonist, which stabilized the corepressor-bound repressive conformation of PPARγ over the coactivator-bound active conformation. Based on this knowledge, pharmaceutical companies have pivoted to designing new inverse agonists with the goal of suppressing PPARγ-mediated transcription to an even greater degree.

In this study led by Zane Laughlin, a former postdoctoral fellow in the Kojetin lab, the researchers report the cellular, biochemical, and structural profiling of FX-909, a new PPARγ inverse agonist from Flare Therapeutics.

To assess the effect of various ligands on PPARγ-mediated transcription, a HEK293T cell transcriptional reporter assay was used which linked a PPAR-binding DNA response element to the firefly luciferase gene. FX-909, along with other covalent inverse agonists, resulted in a concentration-dependent decrease in PPARγ transcription with similar potencies. GW9662, an antagonist, resulted in no change in activity, and rosiglitazone, an agonist, led to an increase in transcription.

The researchers next sought to determine the effect of ligand-PPARγ binding on the association of different coregulator peptides using time-resolved fluorescence energy transfer. FX-909 demonstrated a concentration-dependent increase in corepressor peptide interaction, and a decrease in coactivator interaction with similar potencies to the other covalent inverse agonists. However, FX-909 exhibited a higher TR-FRET efficacy in recruiting the corepressor peptide as compared to its parent compound, T0070907. This difference was further enforced by fluorescence polarization assays which demonstrated a stronger binding affinity between PPARγ LBD and the corepressor peptide in the presence of FX-909 as opposed to T0070907.

The researchers then sought to gain a more complete understanding of the structural changes enacted by FX-909 binding and therefore determined a 2.1 Å crystal structure of the PPARγ LBD bound to FX-909 and a corepressor peptide to. In the structure, the PPARγ LBD adopted a transcriptionally repressive conformation, in which helix12 was occluded, leaving a coregulator binding surface open for the corepressor peptide to bind. However, the binding pose of FX-909 in the ligand-binding pocket and the corepressor peptide on the coregulator binding surface were very similar to crystal structures of the PPARγ LBD bound to T0070907, prompting the use of NMR to fully understand the impact of FX-909 on PPARγ structure. When T0070907 is bound to the PPARγ LBD, the population is split roughly equally between active and repressive conformations, but when FX-909 is bound, the population is highly skewed towards the repressive conformation.

Taken together, these data demonstrate that the increased efficacy of FX-909 is a result of the compound’s ability to stabilize the repressive conformation, increasing corepressor binding, and leading to a decreased level of PPARγ-mediated transcription. This represents the first mechanistic characterization of FX-909, which is currently in clinical trials as a cancer drug, and provides a basis for future drug modulations of PPARγ activity.

Check out the Journal of Medicinal Chemistry for the full story!