Synthetic Peptides as Potential Antigens for Cutaneous Leishmaniosis Diagnosis

This work’s goal was to research new candidate antigens for cutaneous leishmaniosis (CL). In order to reach the goal, we used random peptide phage display libraries screened using antibodies fromLeishmania braziliensispatients. After selection, three peptides (P1, P2, and P3) were synthesized using Fmoc chemistry. The peptides individually or a mixture of them (MIX) was subsequently emulsified in complete and incomplete Freund’s adjuvant and injected subcutaneously in golden hamsters. Sera from the hamsters administered with P1 presented antibodies that recognized proteins between 76 and 150 kDa fromL. braziliensis. Sera from hamsters which had peptides P2 and P3, as well as the MIX, administered presented antibodies that recognized proteins between 52 and 76 kDa ofL. braziliensis. The research on the similarity of the peptides’ sequences in protein databases showed that they match a 63 kDa glycoprotein. The three peptides and the MIX were recognized by the sera from CL patients by immunoassay approach (ELISA). The peptides’ MIX showed the best performance (79% sensitivity) followed by the P1 (72% sensitivity), and the AS presented 91% sensitivity. These results show a new route for discovering molecules for diagnosis or for immunoprotection against leishmaniosis.

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Purification of polyclonal anti-conformational antibodies for use in affinity selection from random peptide phage display libraries: A study using the hydatid vaccine EG95

Journal of Chromatography B ◽

10.1016/j.jchromb.2009.03.036 ◽

2009 ◽

Vol 877 (14-15) ◽

pp. 1516-1522 ◽

Cited By ~ 6

Author(s):

A.J. Read ◽

C.G. Gauci ◽

M.W. Lightowlers

Keyword(s):

Phage Display ◽

Affinity Selection ◽

Random Peptide ◽

Phage Display Libraries ◽

Peptide Phage Display

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Mimotopes for lupus-derived anti-DNA and nucleosome-specific autoantibodies selected from random peptide phage display libraries: facts and follies

Journal of Immunological Methods ◽

10.1016/j.jim.2004.10.010 ◽

2005 ◽

Vol 296 (1-2) ◽

pp. 83-93 ◽

Cited By ~ 20

Author(s):

Jürgen W. Dieker ◽

Yong-Jiang Sun ◽

Cor W. Jacobs ◽

Chaim Putterman ◽

Marc Monestier ◽

...

Keyword(s):

Phage Display ◽

Random Peptide ◽

Phage Display Libraries ◽

Peptide Phage Display

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Selection of peptides that bind to plasminogen activator inhibitor 1 (PAI-1) using random peptide phage-display libraries

FEBS Letters ◽

10.1016/s0014-5793(98)00742-x ◽

1998 ◽

Vol 431 (2) ◽

pp. 170-174 ◽

Cited By ~ 11

Author(s):

Henrik Gårdsvoll ◽

Anton-Jan van Zonneveld ◽

Arne Holm ◽

Eric Eldering ◽

Marja van Meijer ◽

...

Keyword(s):

Phage Display ◽

Plasminogen Activator ◽

Plasminogen Activator Inhibitor ◽

Plasminogen Activator Inhibitor 1 ◽

Random Peptide ◽

Phage Display Libraries ◽

Peptide Phage Display ◽

Activator Inhibitor ◽

Pai 1 ◽

Selection Of

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Selection Dynamic ofEscherichia coliHost in M13 Combinatorial Peptide Phage Display Libraries

Bioscience Biotechnology and Biochemistry ◽

10.1271/bbb.110099 ◽

2011 ◽

Vol 75 (4) ◽

pp. 812-815 ◽

Cited By ~ 8

Author(s):

Stefano ZANCONATO ◽

Giovanni MINERVINI ◽

Irene POLI ◽

Davide De LUCREZIA

Keyword(s):

Phage Display ◽

Phage Display Libraries ◽

Peptide Phage Display

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Epitope Mapping of Inhibitors in Acquired Hemophilia by Phage Display.

Blood ◽

10.1182/blood.v106.11.3202.3202 ◽

2005 ◽

Vol 106 (11) ◽

pp. 3202-3202

Author(s):

Christoph Königs ◽

Christoph Kessel ◽

Sabine Scholz ◽

Susanne Stumpf ◽

Manuela Krause ◽

...

Keyword(s):

Phage Display ◽

Synthetic Peptides ◽

Coagulation Factor ◽

Coagulation Cascade ◽

Acquired Hemophilia ◽

Random Peptide ◽

Functional Studies ◽

Acquired Haemophilia ◽

Phage Libraries ◽

A1 Domain

Abstract In acquired hemophilia polyclonal autoantibodies (inhibitors) to coagulation factor VIII are generated. Inhibitors functionally interfere with interaction of molecules involved in the coagulation cascade for example the interaction of FVIII and the von Willebrand factor. Inhibitors in acquired hemophilia have been reported to bind either to the A2 or to C2 domain of FVIII. Random peptide phage display libraries were screened with patient plasma to identify small peptides mimicking inhibitor epitopes. Peptide sequences were used to map the inhibitor epitope to the surface of FVIII. The phage libraries included libraries with linear or cyclic 7mer or 12mer inserts. Biopanning was performed including positive selections and also negative selections to deplete irrelevant binders. Phages bound to inhibitors were eluted by pH shift or by competition with FVIII. Single phage clones were tested for their specificity for inhibitor positive plasma by ELISA. For specific binders, the sequence of the inserts was identified by DNA sequencing. For one patient 33 phage clones were analysed from three libraries and 28 peptide sequences mimicking the inhibitor epitopemimotopes - were determined. The peptide sequences map to two conformational epitopes on the A2 and A1 domain of FVIII. Clusters of six and four amino acids are the proposed binding regions on the A2 and A1 domain respectively. The matching conformational motifs contain PPLRQ and PPLS. Synthetic peptides corresponding to the sequence displayed on phage were generated and binding of inhibitors to peptides was confirmed by different ELISA formats. Currently, functional studies with synthetic peptides are being performed to further analyse the epitopes. Additionally, more patient samples are being screened to analyse further epitopes and understand immuno dominant regions of FVIII in acquired haemophilia and compare them to the immune response in hemophilia A. The mimotopes isolated could be a basis for the development of ligand effector conjugates to target inhibitor specific B cells.

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Response to Anti CD20 Monoclonal Antibody Rituximab® and Epitope Mapping of Inhibitory Antibodies in Patients with Acquired Haemophilia.

Blood ◽

10.1182/blood.v108.11.1044.1044 ◽

2006 ◽

Vol 108 (11) ◽

pp. 1044-1044

Author(s):

Manuela Krause ◽

Christoph Koenigs ◽

Christoph Kessel ◽

Inge Scharrer ◽

Ralf Grossmann ◽

...

Keyword(s):

Immunomodulatory Therapy ◽

C2 Domain ◽

Random Peptide ◽

Acquired Haemophilia ◽

Phage Display Libraries ◽

Peptide Phage Display ◽

Initiation Of Therapy ◽

Anti Cd20 ◽

A2 Domain

Abstract Introduction: Acquired haemophilia (AcH) is associated with the development of polyclonal autoantibodies against FVIII, which affect directly the A2 or C2 domain of the FVIII molecule. Immunomodulatory therapy regimes to normalize FVIII levels and to eliminate the inhibitor are essential options in the treatment of patients (pts) with AcH. The aims of the present study were to investigate the response to Rituximab® and to localize the inhibitor epitopes on the FVIII domains. Patients and Methods: In 5 pts with AcH (2 females,3 males; age: 64–81 yrs) the inhibitor titers ranged from 9 to 156 BU and the FVIII activities from <1 % to 6 %. Rituximab® was administered once weekly for 4 weeks (dosage: 375 mg/m2) in combination with a prednisone therapy (1–1.5 mg/kg). Random peptide phage display libraries were used to identify peptide sequences specific for inhibitors. The short peptide sequences allow the mapping of conformational epitopes on the surface of the FVIII molecule. Results: In all pts (median follow up of 24 months, range: 2–49 months) we documented complete remission (CR). The median time to achieve CR was 13 weeks (range: 2–32 weeks). The time to CR was much longer (23/32 weeks) in two pts, who showed high inhibitor titers (50/188 BU) compared to pts (2/4/5 weeks) with low inhibitor titers (9/22/29 BU). One pt with a high inhibitor titer (50 BU) relapsed 19 months after CR, an other pt died in septic shock 8 weeks after the initiation of therapy with Rituximab®. No other relevant side effects were observed. In the pt with a relapse we identified target epitopes in the A1 and A2 domain of FVIII, and in the other pts (n=3) the peptide sequences mapped to epitopes on the C2 domain of FVIII. Conclusion: The therapy with Rituximab®shows high response rates and seems to be effective in the elimination of high and low inhibitors against FVIII in pts with AcH. Limited by the small number of pts, further studies are needed to confirm whether the different target epitopes found in patients with (A1/A2) or without a relapse (C2) might have a relevance in the specific immunomodulatory therapy.

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