Unexpected side chain effects at residue 8 of cyclosporin a derivatives allow photoswitching of immunosuppression.
To dissect the enzyme inhibitory properties of the immunosuppressive cyclic undecapeptide cyclosporin A (CsA) and gain access to monospecific, non-calcineurin-inhibiting CsA derivatives, [D-Ser8]CsA was subjected to modifications at the D-Ser side chain. Thus, we modified a CsA residue flanking the calcineurin (CaN) and cyclophilin 18 (Cyp18) binding domains of CsA instead of the residues of the CaN binding domain in order to develop a new specificity-determining site within the cyclic peptide. The [O-(NH2 (CH2)5NHC(O)CH2)-D-Ser8]CsA (derivative 9), with an amino group on a tether, exhibits CsA-like inhibition of the peptidyl prolyl cis/trans isomerase activity of Cyp18 with an IC50 value of 3.2 nm, whereas the CaN inhibition by the Cyp18-derivative 9 complex is completely abolished. Consequently, this compound is not able to inhibit the proliferation and cytokine production of activated T cells. Structure-activity relationship studies with a series of [d-Ser(8)]CsA derivatives indicate that the positively charged side chain is an essential requirement for Cyp18-derivative 9 to be ineffective on CaN. Upon protecting the amino group in derivative 9 with the photolabile moiety 2-nitroveratryloxycarbonyl (NVOC), the Cyp18-[O-(NVOC-NH(CH2)5NHC(O)CH2)-D-Ser8]CsA (derivative 11) complex exhibits strong CaN inhibition and shows potent immunosuppressive activity. In stimulated T cells pretreated with derivative 11, a remarkable recovery of transcriptional activation of the nuclear factor of activated T cells (NFAT) has been achieved through light irradiation, as assessed with a NFAT reporter gene assay.
Zhang
Yixin
Y
Max Planck Research Unit for Enzymology of Protein Folding, Weinbergweg 22, D-06120, Halle/Saale, Germany.
Erdmann
Frank
F
Baumgrass
Ria
R
Schutkowski
Mike
M
Fischer
Gunter
G
D019703:Calcineurin / Q000737:chemistry; D002244:Carbon / Q000737:chemistry; D002460:Cell Line; D049109:Cell Proliferation; D018360:Crystallography, X-Ray; D021983:Cyclophilins / Q000737:chemistry; D016572:Cyclosporine / Q000737:chemistry*; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D017930:Genes, Reporter; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D007166:Immunosuppressive Agents / Q000737:chemistry / Q000494:pharmacology; D020128:Inhibitory Concentration 50; D019169:Jurkat Cells; D008027:Light; D008156:Luciferases / Q000378:metabolism; D008956:Models, Chemical; D008958:Models, Molecular; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012694:Serine / Q000737:chemistry; D013601:T-Lymphocytes / Q000378:metabolism; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation
D016207:Cytokines; D004268:DNA-Binding Proteins; D007166:Immunosuppressive Agents; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D010455:Peptides; D014157:Transcription Factors; D012694:Serine; D002244:Carbon; D016572:Cyclosporine; D008156:Luciferases; D005982:Glutathione Transferase; D019703:Calcineurin; D021983:Cyclophilins