TNF-α is one of the most abundant cytokines produced in many inflammatory and autoimmune conditions such as multiple sclerosis, chronic hepatitis C, or neurodegenerative diseases. These pathologies remain difficult to diagnose and consequently difficult to treat. The aim of this work is to offer a new diagnostic tool by seeking new molecular probes for medical imaging. The target-specific part of the probe consists here of heptameric peptides selected by the phage display technology for their affinity for TNF-α. Several affinity tests allowed isolating 2 peptides that showed the best binding capacity to TNF-α. Finally, the best peptide was synthesized in both linear and cyclic forms and tested on the histological sections of concanavalin-A-(ConA-)treated mice liver. In this well-known hepatitis mouse model, the best results were obtained with the cyclic form of peptide 2, which allowed for the staining of inflamed areas in the liver. The cyclic form of peptide 2 (2C) was, thus, covalently linked to iron oxide nanoparticles (magnetic resonance imaging (MRI) contrast agent) and tested in the ConA-induced hepatitis mouse model. The vectorized nanoparticles allowed for the detection of inflammation as well as of the free peptide. These ex vivo results suggest that phage display-selected peptides can direct imaging contrast agents to inflammatory areas. 1. Introduction Tumor necrosis factor alpha (TNF- ) is a proinflammatory cytokine produced in many inflammatory and autoimmune diseases such as rheumatoid arthritis, Crohn’s disease, multiple sclerosis, or chronic hepatitis C [1, 2]. TNF- is produced by different cell types including macrophages, monocytes, T-cells, smooth muscle cells, adipocytes, and fibroblasts. This cytokine is also implicated in the diseases of the central nervous system like Alzheimer’s and Parkinson’s diseases [3], where it can be produced by several cell populations, including microglia, astrocytes, endothelial cells, Th1 lymphocytes and neurons. Mature TNF- is secreted as a 157-amino acid form [4] with a molecular weight of 17？kDa [5]. Before being released from cells, TNF- is anchored in the plasma membrane as a 26？kDa precursor containing both hydrophobic and hydrophilic regions [6]. The 17？kDa form of TNF- is excised from the integral transmembrane precursor by proteolytic cleavage mediated by the tumor necrosis factor alpha converting enzyme (TACE) [7]. Soluble and transmembrane TNF- are produced by cells as homotrimers that bind to two kinds of receptors, TNF-RI and TNF-RII (tumor necrosis factor receptor type I, p55; type II,
References
[1]
H. Knobler and A. Schattner, “TNF-α, chronic hepatitis C and diabetes: a novel triad,” QJMed, vol. 98, no. 1, pp. 1–6, 2005.
[2]
E. Larrea, N. Garcia, C. Qian, M. P. Civeira, and J. Prieto, “Tumor necrosis factor α gene expression and the response to interferon in chronic hepatitis C,” Hepatology, vol. 23, no. 2, pp. 210–217, 1996.
[3]
P. L. McGeer and E. G. McGeer, “Inflammation and the degenerative diseases of aging,” Annals of the New York Academy of Sciences, vol. 1035, pp. 104–116, 2004.
[4]
W. Fiers, “Precursor structures and structure-function analysis of TNF and lymphotoxin,” Immunology Series, vol. 56, pp. 79–92, 1992.
[5]
B. B. Aggarwal, B. Moffat, and R. N. Harkins, “Human lymphotoxin. Production by a lymphoblastoid cell line, purification, and initial characterization,” Journal of Biological Chemistry, vol. 259, no. 1, pp. 686–691, 1984.
[6]
J. Vilcek and T. H. Lee, “Tumor necrosis factor: new insights into the molecular mechanisms of its multiple actions,” Journal of Biological Chemistry, vol. 266, no. 12, pp. 7313–7316, 1991.
[7]
R. A. Black, C. T. Rauch, C. J. Kozlosky et al., “A metalloproteinase disintegrin that releases tumour-necrosis factor-？ from cells,” Nature, vol. 385, no. 6618, pp. 729–733, 1997.
[8]
B. Beutler, I. W. Milsark, and A. C. Cerami, “Passive immunization against cachectin/tumor necrosis factor protects mice from lethal effect of endotoxin,” Science, vol. 229, no. 4716, pp. 869–871, 1985.
[9]
P. Vassalli, “The pathophysiology of tumor necrosis factors,” Annual Review of Cell and Developmental Biology, vol. 10, pp. 411–452, 1992.
[10]
T. J. M. Molenaar, C. C. M. Appeldoorn, S. A. M. De Haas et al., “Specific inhibition of P-selectin-mediated cell adhesion by phage display-derived peptide antagonists,” Blood, vol. 100, no. 10, pp. 3570–3577, 2002.
[11]
C. L. Chirinos-Rojas, M. W. Steward, and C. D. Partidos, “A peptidomimetic antagonist of TNF-α-mediated cytotoxicity identified from a phage-displayed random peptide library,” Journal of Immunology, vol. 161, no. 10, pp. 5621–5626, 1998.
[12]
E. Pick, J. Brostoff, J. Krejci, and J. L. Turk, “Interaction between “sensitized lymphocytes” and antigen in vitro. II. Mitogen-induced release of skin reactive and macrophage migration inhibitory factors,” Cellular Immunology, vol. 1, no. 1, pp. 92–109, 1970.
[13]
E. Pick and J. L. Turk, “The biological activities of soluble lymphocyte products,” Clinical and Experimental Immunology, vol. 10, no. 1, pp. 1–23, 1972.
[14]
G. Tiefs, J. Hentschel, and A. Wendel, “A T cell-dependent experimental liver injury in mice inducible by concanavalin A,” Journal of Clinical Investigation, vol. 90, no. 1, pp. 196–203, 1992.
[15]
A. Nonomura, M. Tanino, H. Kurumaya, G. Ohta, Y. Kato, and K. Kobayashi, “Studies of immune functions of patients with chronic hepatitis,” The Tohoku Journal of Experimental Medicine, vol. 137, no. 2, pp. 163–177.
[16]
J. Sambrook, E. F. Fritsch, and T. Maniatis, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, New York, NY, USA, 1989.
[17]
M. Port, C. Corot, I. Raynal, and O. Rousseaux, “Novel compositions magnetic particles covered with gem-bisphosphonate derivatives,” http://www.patentlens.net/patentlens/patents.html?patnums=US_2004/0253181_A1&returnTo=quick.html.
[18]
V. Rerat, S. Laurent, C. Burtéa et al., “Ultrasmall particle of iron oxide-RGD peptidomimetic conjugate: synthesis and characterisation,” Bioorganic and Medicinal Chemistry Letters, vol. 20, no. 6, pp. 1861–1865, 2010.
[19]
K. A. Radermacher, N. Beghein, S. Boutry et al., “In vivo detection of inflammation using pegylated iron oxide particles targeted at e-selectin a multimodal approach using mr imaging and epr spectroscopy,” Investigative Radiology, vol. 44, no. 7, pp. 398–404, 2009.
[20]
A. Roch, R. N. Muller, and P. Gillis, “Theory of proton relaxation induced by superparamagnetic particles,” Journal of Chemical Physics, vol. 110, p. 5403, 1999.
[21]
C. Trautwein, T. Rakemann, N. P. Malek, J. Plümpe, G. Tiegs, and M. P. Manns, “Concanavalin A-induced liver injury triggers hepatocyte proliferation,” Journal of Clinical Investigation, vol. 101, no. 9, pp. 1960–1969, 1998.
[22]
M. Mammen, S. K. Choi, and G. M. Whitesides, “Polyvalent interactions in biological systems: implications for design and use of multivalent ligands and inhibitors,” Angewandte Chemie International Edition, vol. 37, no. 20, pp. 2754–2794, 1998.
[23]
J. E. Gestwicki, C. W. Cairo, L. E. Strong, K. A. Oetjen, and L. L. Kiessling, “Influencing receptor-ligand binding mechanisms with multivalent ligand architecture,” Journal of the American Chemical Society, vol. 124, no. 50, pp. 14922–14933, 2002.
[24]
L. B. Giebel, R. T. Cass, D. L. Milligan, D. C. Young, R. Arze, and C. R. Johnson, “Screening of cyclic peptide phage libraries identifies ligands that bind streptavidin with high affinities,” Biochemistry, vol. 34, no. 47, pp. 15430–15435, 1995.
[25]
A. Chopra, “99？mTc-labeled murine IgM monoclonal antibody, fanolesomab, that targets the CD15 glycoprotein antigen,” in Molecular Imaging and Contrast Agent Database (MICAD), National Library of Medicine (US) NCBI, Bethesda, Md, USA, 2004–2011.
[26]
C. Burtea, S. Laurent, E. Lancelot et al., “Peptidic targeting of phosphatidylserine for the MRI detection of apoptosis in atherosclerotic plaques,” Molecular Pharmaceutics, vol. 6, no. 6, pp. 1903–1919, 2009.
[27]
L. Larbanoix, C. Burtea, E. Ansciaux et al., “Design and evaluation of a 6-mer amyloid-beta protein derived phage display library for molecular targeting of amyloid plaques in Alzheimer's disease: comparison with two cyclic heptapeptides derived from a randomized phage display library,” Peptides, vol. 32, no. 6, pp. 1232–1243, 2011.
[28]
M. Kopf, M. F. Bachmann, and B. J. Marsland, “Averting inflammation by targeting the cytokine environment,” Nature Reviews Drug Discovery, vol. 9, pp. 703–718, 2010.
[29]
E. Bell, “Inflammation: targeting TNF,” Nature Reviews Immunology, vol. 9, pp. 390–391, 2009.
[30]
P. Laverman, C. P. Bleeker-Rovers, F. H. M. Corstens, O. C. Boerman, and W. J. G. Oyen, “Development of infection and inflammation targeting compounds,” Current Radiopharmaceuticals, vol. 1, no. 1, pp. 42–48, 2008.
