Interplay among Conformation, Intramolecular Hydrogen Bonds, and Chameleonicity in the Membrane Permeability and Cyclophilin A Binding of Macrocyclic Peptide Cyclosporin O Derivatives

Abstract

A macrocyclic peptide scaffold with well-established structure−property relationship is desirable for tackling undruggable targets. Here, we adopted a natural macrocycle, cyclosporin O (CsO) and its derivatives (CP1−3), and evaluated the impact of conformation on membrane permeability, cyclophilin A (CypA) binding, and the pharmacokinetic (PK) profile. In nonpolar media, CsO showed a similar conformation to cyclosporin A (CsA), a well-known chameleonic macrocycle, but less chameleonic behavior in a polar environment. The weak chameleonicity of CsO resulted in decreased membrane permeability; however, the more rigid conformation of CsO was not detrimental to its PK profile. CsO exhibited a higher plasma concentration than CsA, which resulted from minimal CypA binding and lower accumulation in red blood cells and moderate oral bioavailability (F = 12%). Our study aids understanding of CsO, a macrocyclic peptide that is less explored than CsA but with greater potential for diversity generation and rational design.

Cyclosporin Structure and Permeability: From A to Z and Beyond

“…For example, CsA forms more intramolecular H-bonds in nonpolar solvents and/or more intermolecular H-bonds in polar solvents. ,,, In contrast, the conformation of CsE is somewhat different from that of CsA in the loop region and is relatively rigid with respect to changes in solvent polarity, as discussed in the previous section. CsO and 11 were also recently found to be rigid with respect to increasing solvent polarity . These cyclosporins maintained a single conformation upon addition of DMSO to CDCl 3 , whereas NMR spectroscopy of CsA under these conditions revealed the presence of minor conformations …”

“…Several of the analogues classified as less permeable than CsA (Table B, compounds 7 – 19 ) tend to have conformations similar to that of CsA in the solid state or in nonpolar solvents. For example, the crystal structures of cyclosporin A-acetate ( 8 ) and Valspodar ( 9 ) are similar to the CsA DEKSAN-type conformation. , NMR and IR spectroscopy of cyclosporin O (CsO, 10 ), [MeLeu1,Nva2,MeLeu11]-CsA ( 11 ), cyclosporin B (CsB, 14 ), cyclosporin D (CsD, 15 ), and cyclosporin C (CsC, 16 ) in nonpolar solvents also show an overall backbone conformation similar to that of CsA. − Like CsA, AM4N ( 17 ) also has the DEKSAN-type conformer as its major conformation in nonpolar solvents (but at a reduced population) . Consequently, out of the 13 less permeable analogues in Table B with available information on the conformation in the solid state or in nonpolar solvents, seven or eight show a propensity to form a conformation similar to the DEKSAN-type conformer of CsA under these conditions.…”

“…CsO and 11 were also recently found to be rigid with respect to increasing solvent polarity . These cyclosporins maintained a single conformation upon addition of DMSO to CDCl 3 , whereas NMR spectroscopy of CsA under these conditions revealed the presence of minor conformations …”

“…Permeability measurements for each analogue should be compared to those of other analogues with the same experiment and authors, if possible, because of variabilities in experimental conditions. PAMPA permeability experiments by May, Ahlbach et al, and Lee et al, Caco-2 permeability experiments by Lee et al, and MDCK-LE experiments by Naylor et al are grouped separately. For the Caco-2 experiments, “AtoB” indicates apical to basolateral permeability, and vice versa for “BtoA” .…”

“…PAMPA permeability experiments by May, Ahlbach et al, and Lee et al, Caco-2 permeability experiments by Lee et al, and MDCK-LE experiments by Naylor et al are grouped separately. For the Caco-2 experiments, “AtoB” indicates apical to basolateral permeability, and vice versa for “BtoA” . Another measure of permeability includes “information from cyclophilin competition/binding”, which is a combination of (1) cyclophilin-binding and (2) cDD–mCherry cytosensor array and/or competition/displacement experiments.…”