Macrocyclic Peptides Uncover a Novel Binding Mode for Reversible Inhibitors of LSD1

Lysine-specific demethylase
1 (LSD1) is an epigenetic enzyme which
regulates the methylation of Lys4 of histone 3 (H3) and is overexpressed
in certain cancers. We used structures of H3 substrate analogues bound
to LSD1 to design macrocyclic peptide inhibitors of LSD1. A linear,
Lys4 to Met-substituted, 11-mer (4) was identified as
the shortest peptide distinctly interacting with LSD1. It was evolved
into macrocycle 31, which was >40 fold more potent (Ki = 2.3 μM) than 4. Linear and macrocyclic
peptides exhibited unexpected differences in structure–activity
relationships for interactions with LSD1, indicating that they bind
LSD1 differently. This was confirmed by the crystal structure of 31 in complex with LSD1-CoREST1, which revealed a novel binding
mode at the outer rim of the LSD1 active site and without a direct
interaction with FAD. NMR spectroscopy of 31 suggests
that macrocyclization restricts its solution ensemble to conformations
that include the one in the crystalline complex. Our results provide
a solid basis for the design of optimized reversible LSD1 inhibitors.