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Altered Peptide Ligands in the immunotherapy of Multiple Sclerosis: the role of citrullination
Deraos Georgios MSc, PhD ELDRUG S.A.
Patras, Greece
Multiple Sclerosis
Epidemiology of MS
Age onset 20 – 50 years old
Women are 2 times more likely to develop MS
500,000 cases in US
Over 2.5 million people around the world
More prevalent whites of northern European ancestry
Genetic Influences
• Vision problems
• Numbness
• Difficulty walking
• Fatigue, Depression
• Coordination and Balance problems,
• Pain
• Spasticity
Symptoms of MS
http://www.chumsweb.org/types.htm
MS Types
Relapsing-remitting MS (RRMS)Attacks followed by partial or completerecovery Symptoms may be inactive for months or years
•Secondary-progressive MS (SPMS)Occasional relapses but symptoms remain constant, no remissionProgressive disability late in disease course
Primary-progressive MS (PPMS)Affects approximately 10% of MS population. Slow onset but continuous worsening condition
Benign MS. small attacks followed by complete recovery.
Immunopathogenesis of MS
Immunodominant epitopes in MS
MBP 83-99 ENPVVHFFKNIVTPRTP
MOG 35-55 MEVGWYRSPFSRVVHLYRNGK
PLP 139-151 HCLGKWLGHPDKF
HLA-DR2 – PEPTIDE COCMPLEX
Val 87
Phe 90
Lys 91
Crystal Structure of HLA-DR2 (DRA*0101, DRB1*1501) Complexed with a Peptide from Human Myelin Basic Protein kathine J. Smith, Jason Pyrdol, Laurent Gauthier, Don C Wiley, Kai W. Wucherpfenning.
Citrullination in Multiple Sclerosis
Medications and MS
Therapies Administration Class
Avonex IM 1x a week Interferon beta-1a
Betaseron SC, every other day Interferon beta-1b
Copaxone SC 1x a day Glatiramer acetate
Rebif SC 3x a week Interferon beta-1a
Gilenya Oral capsule 1x day Fingolimod
Tysabri IV Monthly at Center Natalizumab
1 Myelin Peptide Analogues2 MBP83-99(Tyr91)
3 MBP83-99(Phe91)
4 MBP83-99(Glu91)
5 Cyclo(83-99)MBP83-99(Ala91, 96)
6 MBP87-99(Cit91, 97)
7 CycloMBP87-99(Cit91, 97)
8 MBP87-99(Arg91Ala96)
9 CycloMBP87-99(Arg91Ala96)
10 MBP87-99(Ala91,96)
11 CycloMBP87-99(Ala91,96)
12 MOG35-55 rat mouse
13 MOG35-55 Human
14 CyclicMOG35-55 rat mouse
15 CyclicMOG35-55 human
Synthesis of peptide analogues Solid Phase peptide Synthesis
Conversion of the citrullinated analogues
HPLC analysis of the citrullinated analogues
Biological evaluation
Citrullinated analoguesMBP87-99(Cit91,97)cycloMBP87-99(Cit91,97)
PBMCs of MS patientsCytokine secretion
Mannan conjugated peptides• MBP83-99(Tyr91)• MBP83-99(Phe91)• MBP83-99(Arg91)• MBP83-99(Glu91)
EAE experimentIn SJL/J miceCytokine secretion Cross reaction antibodies assays
Department of Medical school, University of Patras,
Burnet Institute, Australia
Deraos et al. J. Med. Chem. 2008 Dec 25;51(24):7834-42. .
Citrullinated peptides in PBMCs from MS patients
Katsara et al , Mol Immiunol2008 Aug;45(13):3661-70.
Manosylated Peptideanalogs
IFN-γ IL-4 IL-10 Proliferation Abs Cross reaction MBP
MBP83-99 ++ -- +++ +++
MBP83-99(A91) -- ++ + +
MBP83-99(E91) -- ++ +++ +
MBP83-99(F91) ++ ++ ++ +++ +/- -
MBP83-99(Y91) -- ++ ++ +/- +++ -
MBP83-99(R91,A96) -- ++ +++ +++
Manosylation of MBP83-99 peptide analogues
Katsara et al , Mol Immiunol2008 Aug;45(13):3661-70.
Manosylated Peptideanalogs
IFN-γ IL-4 IL-10 Proliferation Abs Cross reaction MBP
MBP83-99 ++ -- +++ +++
MBP83-99(A91) -- ++ + +
MBP83-99(E91) -- ++ +++ +
MBP83-99(F91) ++ ++ ++ +++ +/- -
MBP83-99(Y91) -- ++ ++ +/- +++ -
MBP83-99(R91,A96) -- ++ +++ +++
Manosylation of MBP83-99 peptide analogues
Katsara et al , Mol Immiunol2008 Aug;45(13):3661-70.
Manosylated Peptideanalogs
IFN-γ IL-4 IL-10 Proliferation Abs Cross reaction MBP
MBP83-99 ++ -- +++ +++
MBP83-99(A91) -- ++ + +
MBP83-99(E91) -- ++ +++ +
MBP83-99(F91) ++ ++ ++ +++ +/- -
MBP83-99(Y91) -- ++ ++ +/- +++ -
MBP83-99(R91,A96) -- ++ +++ +++
Manosylation of MBP83-99 peptide analogues
Katsara et al , Mol Immiunol2008 Aug;45(13):3661-70.
Manosylated Peptideanalogs
IFN-γ IL-4 IL-10 Proliferation Abs Cross reaction MBP
MBP83-99 ++ -- +++ +++
MBP83-99(A91) -- ++ + +
MBP83-99(E91) -- ++ +++ +
MBP83-99(F91) ++ ++ ++ +++ +/- -
MBP83-99(Y91) -- ++ ++ +/- +++ -
MBP83-99(R91,A96) -- ++ +++ +++
Manosylation of MBP83-99 peptide analogues
Katsara et al , Mol Immiunol2008 Aug;45(13):3661-70.
Manosylated Peptideanalogs
IFN-γ IL-4 IL-10 Proliferation Abs Cross reaction MBP
MBP83-99 ++ -- +++ +++
MBP83-99(A91) -- ++ + +
MBP83-99(E91) -- ++ +++ +
MBP83-99(F91) ++ ++ ++ +++ +/- -
MBP83-99(Y91) -- ++ ++ +/- +++ -
MBP83-99(R91,A96) -- ++ +++ +++
Manosylation of MBP83-99 peptide analogues
Katsara et al , Mol Immiunol2008 Aug;45(13):3661-70.
Manosylated Peptideanalogs
IFN-γ IL-4 IL-10 Proliferation Abs Cross reaction MBP
MBP83-99 ++ -- +++ +++
MBP83-99(A91) -- ++ + +
MBP83-99(E91) -- ++ +++ +
MBP83-99(F91) ++ ++ ++ +++ +/- -
MBP83-99(Y91) -- ++ ++ +/- +++ -
MBP83-99(R91,A96) -- ++ +++ +++
Manosylation of MBP83-99 peptide analogues
Katsara et al , Mol Immiunol2008 Aug;45(13):3661-70.
Manosylated Peptideanalogs
IFN-γ IL-4 IL-10 Proliferation Abs Cross reaction MBP
MBP83-99 ++ -- +++ +++
MBP83-99(A91) -- ++ + +
MBP83-99(E91) -- ++ +++ +
MBP83-99(F91) ++ ++ ++ +++ +/- -
MBP83-99(Y91) -- ++ ++ +/- +++ -
MBP83-99(R91,A96) -- ++ +++ +++
Manosylation of MBP83-99 peptide analogues
Conclusions
➢Citrullination is essential for antigenic recognition of self epitopes by T cells and
may be a new strategy of citrullinated inhibitors like drugs in the future
➢Mutations to the native MBP83–99 peptide were able to induce IL-4 responses,
although IFN-γ responses were still present.
➢ However, conjugating MBP83–99 peptide analogs to mannan was able to further
divert the immune response from Th1 to Th2, demonstrating the importance of
mannan as conjugates.
➢ APLs linear or cyclic can be used as immunomodulator agents in the MS.
Although linear peptides have the disadvantage of being degradated easily from
proteolytic enzymes and can not be used as oral administered drugs.
ELDRUG S.A COMPANY
PATRAS GREECE
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