Structural Elucidation and Covalent Modulation of the Autorepressed Orphan Nuclear Receptor NR2F6

The orphan nuclear receptor NR2F6 (Nuclear Receptor subfamily
2
group F member 6) is an emerging therapeutic target for cancer immunotherapy.
Upregulation of NR2F6 expression in tumor cells has been linked to
proliferation and metastasis, while in immune cells NR2F6 inhibits
antitumor T-cell responses. Small molecule modulation of NR2F6 activity
might therefore be a novel strategy in cancer treatment, benefiting
from this dual role of NR2F6. However, there are no molecular strategies
available for targeting NR2F6, hampered among others by lack of structural
insights and appropriate biochemical assays. To overcome these challenges,
several noncanonical nuclear receptor coregulator peptide motifs were
identified to be constitutively recruited to the NR2F6 ligand binding
domain (LBD). Co-crystallization of the NR2F6 LBD with a peptide from
the coregulator Nuclear Receptor Binding SET Domain Protein 1 (NSD1)
enabled, for the first time, the structural elucidation of the unliganded
(apo) form of NR2F6. This revealed an autorepressed, homodimeric LBD
conformation in which helix 12 folds over the canonical coregulator
binding site, generating an alternative contact surface for NSD1 binding.
Screening of a focused library of covalent NR probes identified compounds
that preferentially target a cysteine residue near the NSD1 binding
site, inhibiting NR2F6 coregulator recruitment. Combined, these results
provide structural insights into the ligand-independent transcriptional
activity of NR2F6 and may serve as a starting point for the development
of novel NR2F6 modulators.