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The designer leptin antagonist peptide Allo-aca compensates for short serum half-life with very tight binding to the receptor

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Abstract

The leptin receptor antagonist peptide Allo-aca exhibits picomolar activities in various cellular systems and sub-mg/kg subcutaneous efficacies in animal models making it a prime drug candidate and target validation tool. Here we identified the biochemical basis for its remarkable in vivo activity. Allo-aca decomposed within 30 min in pooled human serum and was undetectable beyond the same time period from mouse plasma during pharmacokinetic measurements. The C max of 8.9 μg/mL at 5 min corresponds to approximately 22 % injected peptide present in the circulation. The half-life was extended to over 2 h in bovine vitreous fluid and 10 h in human tears suggesting potential efficacy in ophthalmic diseases. The peptide retained picomolar anti-proliferation activity against a chronic myeloid leukemia cell line; addition of a C-terminal biotin label increased the IC50 value by approximately 200-fold. In surface plasmon resonance assays with the biotin-labeled peptide immobilized to a NeutrAvidin-coated chip, Allo-aca exhibited exceptionally tight binding to the binding domain of the human leptin receptor with k a = 5 × 105 M−1 s−1 and k diss = 1.5 × 10−4 s−1 values. Peptides excel in terms of high activity and selectivity to their targets, and may activate or inactivate receptor functions considerably longer than molecular turnovers that take place in experimental animals.

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Abbreviations

Aβ:

Amyloid β peptide

CML:

Chronic myeloid leukemia

Dde:

Dimethyl-dioxo-cyclohexylidene

EDTA:

Ethylene diamine tetraacetate

Fmoc:

Fluorenyl-methoxycarbonyl

G-CSF:

Granulocyte-colony stimulating factor

IL-6:

Interleukin-6

ip:

Intraperitoneally

LBD:

Leptin binding domain

LOQ:

Limit of quantification

MALDI TOF–MS:

Matrix-assisted laser ionization/desorption time-of-flight mass spectrometry

ObR:

Leptin receptor

PK:

Pharmacokinetics

RP-HPLC:

Reversed-phase high performance liquid chromatography

TCA:

Trichloroacetic acid

TFA:

Trifluoroacetic acid

TNBC:

Triple negative breast cancer

sc:

Subcutaneously

SPR:

Surface plasmon resonance

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Acknowledgments

This work was partially supported by the Novo Nordisk Diabetes Innovation Award to E. S. Another part of this work was supported by Australian Research Council Discovery grant DP120101963 to J. D. W. who is also an NHMRC (Australia) Principal Research Fellow.

Conflict of interest

The authors declare no competing financial interests.

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Authors and Affiliations

  1. Department of Biology, Temple University, 1900 North 12th Street, Philadelphia, PA, 19122, USA

    Laszlo Otvos Jr.

  2. Department of Pharmaceutical Sciences, North Dakota State University, Fargo, ND, 58102, USA

    Stefan W. Vetter & Mohit Koladia

  3. Institute of Bioanalytical Chemistry, Faculty of Chemistry and Mineralogy, Center for Biotechnology and Biomedicine, Universität Leipzig, 04103, Leipzig, Germany

    Daniel Knappe, Rico Schmidt & Ralf Hoffmann

  4. Department of Dermatology, Venerology and Dermatooncology, Faculty of Medicine, Semmelweis University, 1085, Budapest, Hungary

    Eszter Ostorhazi

  5. 1st Institute of Pathology and Experimental Cancer Research, Faculty of Medicine, Semmelweis University, 1085, Budapest, Hungary

    Ilona Kovalszky

  6. Torrey Pines Institute for Molecular Studies, Port St Lucie, FL, 34987, USA

    Nina Bionda & Predrag Cudic

  7. Sbarro Institute of Cancer Research and Molecular Therapeutics, Temple University, Philadelphia, PA, 19122, USA

    Eva Surmacz

  8. Florey Institute of Neuroscience and Mental Health, Victoria, 3010, Australia

    John D. Wade

Authors
  1. Laszlo Otvos Jr.
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  2. Stefan W. Vetter
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  8. Nina Bionda
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  9. Predrag Cudic
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  10. Eva Surmacz
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  11. John D. Wade
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  12. Ralf Hoffmann
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Correspondence to Laszlo Otvos Jr..

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Otvos, L., Vetter, S.W., Koladia, M. et al. The designer leptin antagonist peptide Allo-aca compensates for short serum half-life with very tight binding to the receptor. Amino Acids 46, 873–882 (2014). https://doi.org/10.1007/s00726-013-1650-6

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  • DOI: https://doi.org/10.1007/s00726-013-1650-6

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