Structural basis for the cyclophilin A binding affinity and immunosuppressive potency of E-ISA247 (voclosporin)

Abstract

E-ISA247 (voclosporin) is a cyclosporin A analogue that is in late-stage clinical development for the treatment of autoimmune diseases and the prevention of organ graft rejection. The X-ray crystal structures of E-ISA247 and its stereoisomer Z-ISA247 bound to cyclophilin A have been determined and their binding affinities were measured to be 15 and 61 nM, respectively, by fluorescence spectroscopy. The higher affinity of E-ISA247 can be explained by superior van der Waals contacts between its unique side chain and cyclophilin A. Comparison with the known ternary structure including calcineurin suggests that the higher immunosuppressive efficacy of E-ISA247 relative to cyclosporin A could be a consequence of structural changes in calcineurin induced by the modified E-ISA247 side chain.

Figures

Figure 1

Residues at position 1 of CsA and the ISA247 stereoisomers. Structural formulae are displayed for (a) the cyclosporin residue MeBmt1, (b) the E-ISA247 residue E-MePmt1 and (c) the Z-ISA247 residue Z-MePmt 1. 2F o − F c electron-density maps around (d) E-MePmt1 and (e) Z-MePmt1 contoured at one standard deviation above the mean density are displayed in green. Surface and ball-and-stick representations of the crystal structures of CypA in grey complexed with (f) CsA (PDB entry 2rma; Ke et al., 1994 ▶), (g) E-ISA247 and (h) Z-ISA247 in yellow are shown. The distances between the Ala103 backbone carbonyl of CypA and residue 1 of the cyclic peptides are marked by black arrows.

Figure 2

Fluorescence titration curve of the E-ISA247 isomer. The measured fluorescence intensities after correction for protein dilution (dots) are fitted with a sigmoidal curve (line).

Figure 3

Superposition of the ternary Cn–CypA–CsA complex with the binary CypA–ISA247 complexes. Cn, CypA and the CsA residue MeBmt1 of the ternary complex (PDB entry 1m63; Huai et al., 2002 ▶) are displayed in yellow, grey and pink, respectively. The MePmt1 residues of E-ISA247 and Z-ISA247 are shown in green and cyan, respectively. A close contact between E-ISA247 and Cn (carbon-to-nitrogen distance of 2.4 Å) is marked by a black arrow.