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A novel approach in potential anticoagulants from peptides epitope 558–565 of A2 subunit of factor VIII

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Abstract

Factor VIII, a human blood plasma protein, plays an important role during the intrinsic pathway of blood coagulation cascade after its activation by thrombin. The activated form of FVIII acts as cofactor to the serine protease Factor IXa, in the conversion of the zymogen Factor X to the active enzyme Factor Xa. The Ser558–Gln565 region of the A2 subunit of Factor VIII has been shown to be crucial for FVIIIa–FIXa interaction. Based on this, a series of linear peptides, analogs of the 558–565 loop of the A2 subunit of the heavy chain of Factor VIII were synthesized using the acid labile 2-chlorotrityl chloride resin and biologically evaluated in vitro by measuring the chronic delay of activated partial thromboplastin time and the inhibition of Factor VIII activity, as potential anticoagulants.

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Abbreviations

AcOH:

Acetic acid

aPTT:

Activated partial thromboplastin time

Boc:

Tert-butoxycarbonyl

Bzl:

Benzyl group

CLTR-Cl:

2-chlorotrityl chloride

DCM:

Dichloromethane

DIC:

N,N′-diisopropylcarbodiimide

DIPEA:

Diisopropylethylamine

DMF:

N,N′-dimethylformamide

dPPP:

Deficient platelet poor plasma

ESI-MS:

Electrospray ionization mass spectrometry

Fmoc:

9-Fluorenylmethyloxycarbonyl group

FVIII:

Factor VIII

FIX:

Factor IX

HOBt:

1-hydroxybenzotriazole

i-PrOH:

2-propanol

Me:

Methyl group

MeCN:

Acetonitrile

MeOH:

Methanol

MBHA:

4-methylbenzhydrylamine

PPP:

Platelet poor plasma

PyBOP:

(Benzotriazol-1-yloxy)-tris(pyrrolidino)phosphonium hexafluorophosphate

PT:

Prothrombin time

rFVIII:

Recombinant Factor VIII

tBu:

Tert-butyl group

RP-HPLC:

Reversed phase high performance liquid chromatography

TES:

Triethylsilane

TFA:

Trifluoroacetic acid

TFE:

2,2,2-trifluoroethanol

TLC:

Thin layer chromatography

Tol:

Toluene

Trt:

Trityl group

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Acknowledgments

Anticoagulant assays were performed under the supervision of Professor of Haematology, Dr Pantelis Makris, School of Medicine, Aristotle University of Thessaloniki, Greece. This Research Project is co-financed: 80 % by European Union—European Social Fund and 20 % by General Secretary of Research & Technology (PENED 03ED569). We also thank SANOFI S.A., Athens, Greece for the financial support.

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Authors and Affiliations

  1. Department of Chemistry, University of Patras, Patras, Greece

    C. Anastasopoulos, Y. Sarigiannis & G. Stavropoulos

Authors
  1. C. Anastasopoulos
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  2. Y. Sarigiannis
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  3. G. Stavropoulos
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Corresponding author

Correspondence to G. Stavropoulos.

Additional information

Abbreviations of common amino acids are in accordance with the recommendations of IUPAC-IUB Joint Commission on Biochemical Nomenclature: Arch Biochem Biophys 206 (1988) v–xxii, J Biol Chem 264 (1989) 668–673, J Peptide Sci 12 (2006) 1–8.

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Anastasopoulos, C., Sarigiannis, Y. & Stavropoulos, G. A novel approach in potential anticoagulants from peptides epitope 558–565 of A2 subunit of factor VIII. Amino Acids 44, 1159–1165 (2013). https://doi.org/10.1007/s00726-012-1448-y

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  • DOI: https://doi.org/10.1007/s00726-012-1448-y

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