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Molecular Imaging and Contrast Agent Database (MICAD) [Internet].

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Anti-ICAM-1 antibody-conjugated paramagnetic liposomes

Anti-ICAM ACPLs
, PhD
National Center for Biotechnology Information, NLM, NIH, Bethesda, MD, vog.hin.mln.ibcn@dacim

Created: ; Last Update: February 14, 2008.

Chemical name:Anti-ICAM-1 antibody-conjugated paramagnetic liposomes
Abbreviated name:Anti-ICAM ACPLs
Synonym:
Agent category:Antibody, small molecule (nanoparticle)
Target:ICAM-1
Target category:Antigen
Method of detection:Magnetic resonance imaging (MRI), optical fluorescence microscopy
Source of signal/contrast:Gadolinium, Texas Red
Activation:No
Studies:
  • Checkbox In vitro
  • Checkbox Rodents
No structure is currently available in PubChem.

Background

[PubMed]

ICAM-1 (CD54) is a cell-surface glycoprotein belonging to the immunoglobulin superfamily of cell adhesion molecules (CAMs) (1). Upregulated by inflammatory mediators, ICAM-1 is expressed on various cells including leukocytes, epithelial cells, endothelial cells, and fibroblasts (2). The predominant function of ICAM-1 is the recruitment and trafficking of leukocytes through interactions with leukocyte-expressed integrins (2). Therefore, ICAM-1 plays a critical role in the development of the central nervous system (CNS), in immune and inflammatory responses, and in embryonic development (1). Many cerebrovascular and neurological diseases such as multiple sclerosis and its animal model (i.e., experimental allergic encephalomyelitis (EAE)), Alzheimer’s disease, and Parkinson’s disease have neuroinflammatory responses involving the migration of leukocytes into damaged or infected tissues (3). EAE, which is an ascending encephalomyelitis, is characterized by an intense perivascular inflammatory process in the white matter of the CNS, primarily on the spinal cord, brainstem, and cerebellum, which facilitates the attachment of leukocytes to inflamed endothelium (4). For example, a 10-fold increase in ICAM-1 expression has been found on the endothelium of capillaries and venules throughout the CNS at an early stage of EAE (4). Thus, ICAM-1 has been an attractive target for imaging the early and specific upregulation of vascular CAMs in diseases (4).

Amphiphiles are made from molecules containing both a hydrophobic (non-polar tail) and a hydrophilic (polar head) section. Amphiphiles can self-associate into multiple concentric lipid bilayers (called lamellae) in different sizes and geometries (5). Liposomes are generally formed by a variety of modified amphiphiles and can deliver a substantial amount of encapsulated therapeutic or diagnostic agents to targeted sites (6). Traditional liposomes suffer from the drawbacks of fast elimination from blood and easy capture by the reticulo-endothelial system (7). As an alternative to traditional liposomes, paramagnetic polymerized liposomes (PPLs) are composed of a mixture of lipids with different functional groups on the aqueous exposed surface and a polymerizable group (diacetylene) in the lipid tail (8). A typical formulation consists of gadolinium III–diethylenetriamine pentaacetic acid (Gd-DTPA)–based lipids, amphiphilic carrier lipids, and biotinylated amphiphiles (9).

PPLs possess increased physical stability, unique spectroscopic properties, and chemical modification durability (9). The diacetylene triple bonds in the PPLs are located in the fatty acyl chains. The polymerization of the triple bonds between lipids can be carried out by simple ultraviolet (UV) radiation to form a polydiacetylene backbone of alternating eneyne groups (8). A short polyethylene glycol (PEG) segment is covalently linked to the polar head of PPLs (9). The hydrophilicity of PEG allows for sterical stabilization of PPLs and protects against interactions with other molecular and biological components in blood stream, such as undesired lipid exchange or lipid fusion (9). Both polymerization and PEGylization prolong the in vivo circulation time of PPLs in blood. For instance, the half-life time of the PPLs is found to be ~19 h in rats (4). Because easy access to bulky water, the attachment of Gd-DTPA chelates on the surface of PPLs allows for greater relaxivity enhancement compared to Gd chelates entrapped in the traditional liposomes. The multiple Gd chelates on the surface of the liposomes further increase the sensitivity to detection by magnetic resonance imaging (MRI). The biotin groups in the PPLs allow for conjugation of biotinylated antibodies via an avidin-biotin linkage procedure (10). Avidin, a tetrameric protein with a molecular mass of 68 KDa, is capable of strongly binding four biotins (Ka = 1.7 × 1015 M-1) (10). Thus, avidins serve as a bridge between a biotinylated antibody against neural CAM (NCAM-1) and a biotinylated group on PPLs to form a NCAM-1–specific MRI contrast agent, anti-NCAM antibody-conjugated PPLs (ACPLs) (4).

Synthesis

[PubMed]

Storrs et al. reported the details of synthesis of PPLs (9). Pentacosadiynoic acid (PDA) was treated with N-hydroxysuccinimide (NHS) and 1-(3-(dimethylamino)propyl)-3-ethylcarbodiimide hydrochloride (EDC) to form NHS ester, which was treated with a PEG-diamine linker (NH2(CH2CH2O)2CH2CH2NH2) to produce PEG-PDA derivatives. Then, the product was treated with DTPA dianhydride (DTPAA) to form DTPA-bis(PEG-PDA) diamides followed by a complexation with GdCl3 to produce a neutral amphiphilic Gd chelates. The PEG-PDA derivative was treated with biotinamidocaproic acid NHS ester to form biotinylated lipid conjugates. To generate PPLs, a mixture of 60% PEG-PDA, 29.5% Gd chelate lipid , 10% amino-terminated lipid, and 0.5% biotinylated lipid was polymerized at 0°C under irradiation from an ultraviolet lamp (wavelength = 254 nm) to form a dark blue PDA-PPLs. Sipkins et al. reported the details of preparation of anti–NCAM-1 ACPLs from the PPLs (4). ACPLs were formed by the addition of 2.3 μg avidin and 14.9 μg biotinylated antibody in a PPL solution at a concentration of 5.6 mM acyl chains.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

The T1 relaxivity of PPLs was determined to be 8.7 mM-1•s-1 at 2.0 T (8, 9). Gel electrophoresis and immunodetection techniques demonstrated the capability of attaching the monoclonal antibodies to biotinylated PPLs. The binding specificity of anti-ICAM-1 ACPLs was examined with an enzyme-linked immunosorbent assay (ELISA) in which the anti–ICAM-1 ACPLs competed with the inhibitory binding of horseradish peroxidase (HRP)-labeled anti–ICAM-1 antibodies (4). Then, the anti–ICAM-1 ACPL recognition of antigens was further confirmed in cultured endothelial cells with bacterial lipopolysaccharide–stimulated ICAM-1 expression. The multivalent anti–ICAM-1 ACPLs appeared to have significantly higher binding affinities compared to free antibody.

Animal Studies

Rodents

[PubMed]

The in vivo targeting of anti–ICAM-1 ACPLs was examined in mice with EAE (4). Texas Red fluorophore-labeled anti-NCAM-1 ACPLs were administrated intravenously to EAE mice at 1.2 mg Gd/kg and 890 μg antibody/kg in the early phase of ICAM-1 upregulation. At 24 h after injection, brains were harvested for microscopic localization of fluorescent ACPLs. All EAE mice showed positive ACPL binding to CNS microvasculature in the cerebellum, brainstem, and spinal cord (4). The location of anti–ICAM-1 binding appeared to correlate with the immunohistochemically defined patterns for ICAM-1 expressions. For example, the binding of anti–ICAM-1 ACPLs was found in multiple vessels in the cerebellum that were surrounded by inflammatory infiltrates, in microvessels that were not associated with inflammatory infiltrates, and in small vessels but not in the large central arterioles. The complementary halves of the mouse brains were reserved from the fluorescence microscopy experiments and further imaged with high-resolution MRI microscopy techniques. Each brain was fixed in 45% paraformaldehyde, and T1-weighted images were collected on a 9.4 T imager. There were substantial increases in MRI signal intensity in the cerebellar (32%) and cerebral (28%) cortex, with moderate signal intensity increases in the cerebellar white matter (18%). The contrast between gray and white matter was improved particularly in the cerebellum because gray matter was significantly more vascularized than white matter. The PPLs had a blood plasma half-life of 19 h, which was comparable to that of PEG-stabilized liposomes (9).

Other Non-Primate Mammals

[PubMed]

No publication is currently available.

Non-Human Primates

[PubMed]

No publication is currently available.

Human Studies

[PubMed]

No publication is currently available.

NIH Support

GM 07365

References

1.
Hubbard A.K. , Rothlein R. Intercellular adhesion molecule-1 (ICAM-1) expression and cell signaling cascades. Free Radic Biol Med. 2000;28(9):1379–86. [PubMed: 10924857]
2.
Hopkins A.M. , Baird A.W. , Nusrat A. ICAM-1: targeted docking for exogenous as well as endogenous ligands. Adv Drug Deliv Rev. 2004;56(6):763–78. [PubMed: 15063588]
3.
Turowski P. , Adamson P. , Greenwood J. Pharmacological targeting of ICAM-1 signaling in brain endothelial cells: potential for treating neuroinflammation. Cell Mol Neurobiol. 2005;25(1):153–70. [PubMed: 15962512]
4.
Sipkins D.A. , Gijbels K. , Tropper F.D. , Bednarski M. , Li K.C. , Steinman L. ICAM-1 expression in autoimmune encephalitis visualized using magnetic resonance imaging. J Neuroimmunol. 2000;104(1):1–9. [PubMed: 10683508]
5.
Baxter A.G. The origin and application of experimental autoimmune encephalomyelitis. Nat Rev Immunol. 2007;7(11):904–12. [PubMed: 17917672]
6.
Strijkers G.J. , Mulder W.J. , van Heeswijk R.B. , Frederik P.M. , Bomans P. , Magusin P.C. , Nicolay K. Relaxivity of liposomal paramagnetic MRI contrast agents. Magma. 2005;18(4):186–92. [PubMed: 16155762]
7.
Torchilin V.P. Recent advances with liposomes as pharmaceutical carriers. Nat Rev Drug Discov. 2005;4(2):145–60. [PubMed: 15688077]
8.
Storrs R.W. , Tropper F.D. , Li H.Y. , Song C.K. , Sipkins D.A. , Kuniyoshi J.K. , Bednarski M.D. , Strauss H.W. , Li K.C. Paramagnetic polymerized liposomes as new recirculating MR contrast agents. J Magn Reson Imaging. 1995;5(6):719–24. [PubMed: 8748492]
9.
Storrs R.W. , Tropper F.D. , Li H.Y. , Song C.K. , Kuniyoshi J.K. , Sipkins D.A. , Li C.P.K. , Bednarski M. Paramagnetic polymerized lyposomes: synthesis, characterization, and applications for magnetic resonance imaging. J. Am. Chem. Soc. 1995;117:7301.
10.
Mulder W.J. , Strijkers G.J. , Griffioen A.W. , van Bloois L. , Molema G. , Storm G. , Koning G.A. , Nicolay K. A liposomal system for contrast-enhanced magnetic resonance imaging of molecular targets. Bioconjug Chem. 2004;15(4):799–806. [PubMed: 15264867]

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