Gd-DOTA-c(Cys-Arg-Gly-Asp-Cys)

Background

[PubMed]

Magnetic resonance imaging (MRI) maps information about tissues spatially and functionally. Protons (hydrogen nuclei) are widely used to create images because of their abundance in water molecules, which comprise >80% of most soft tissues. The contrast of proton MRI images depends mainly on the density of nuclear proton spins, the relaxation times of the nuclear magnetization (T1, longitudinal; T2, transverse), the magnetic environment of the tissues, and the blood flow to the tissues. However, insufficient contrast between normal and diseased tissues requires the use of contrast agents. Most contrast agents affect the T1 and T2 relaxation of the surrounding nuclei, mainly the protons of water. T2* is the spin–spin relaxation time composed of variations from molecular interactions and intrinsic magnetic heterogeneities of tissues in the magnetic field (1). Cross-linked iron oxide (CLIO) and other iron oxide formulations affect T2 primarily and lead to a decreased signal. On the other hand, paramagnetic T1 agents, such as gadolinium (Gd³⁺) and manganese (Mn²⁺), accelerate T1 relaxation and lead to increased contrast images.

Thrombosis plays a major role in many cardiovascular diseases, such as myocardial infarction, pulmonary embolism (PE), deep venous thrombosis (DVT), atherothrombosis, or cerebral venous thrombosis (2, 3). DVT is a significant cause of PE, which is a potentially life-threatening clinical problem. Thrombosis occurs when platelets deposit in regions of low flow in the deep venous system, followed by an activation process of thrombin, which then converts fibrinogen into fibrin. Platelets become activated and bind to fibrinogen, resulting in platelet aggregation via the platelet integrin GPIIb/IIIa (α_IIbβ₃, CD61/CD41). The thrombus may become organized or detached from the vessel wall.

A single-chain antibody (anti-LIBS 145) has been developed to recognize ligand-induced binding sites (LIBS) of GPIIb/IIIa that become exposed only upon binding to fibrinogen (4). Anti-LIBS 145 single-chain antibody does not bind to circulating platelets. Anti-LIBS 145 single-chain antibody was conjugated to microparticles of iron oxide (MPIOs) to form LIBS-MPIOs for T2-weighted MRI imaging of platelet-containing thrombi (5-8). The cyclic peptide P977 (cyclo(Cys-Arg-Gly-Asp-Cys)) was found to bind to platelet glycoprotein GPIIb/IIIa receptor with good affinity (9). P975 is composed of P977-conjugated gadolinium-tetraazacyclododecane-N,N’,N’’,N’’’-tetraacetic acid (Gd-DOTA) (10). P975 is being developed as a non-invasive T1 MRI agent for GPIIb/IIIa expression in thrombi.

Synthesis

[PubMed]

Gd-DOTA was conjugated to P977 via a small linker at a 1:1 ratio (10). P975 exhibited an r₁ relaxivity value of 9 mM^-1s^-1 at 40°C and 60 MHz.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

In vitro competition binding to activated platelets was performed using FITC-fibrinogen with P975 and P977 (10). The 50% inhibition concentrations for P975 and P977 were 2.1 ± 0.3 µM and 1.6 ± 0.2 µM, respectively.

Animal Studies

Human Studies

[PubMed]

No publication is currently available.

NIH Support

R01 HL71021, R01 HL78667, R01 EB009638

References

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Schwarz M., Katagiri Y., Kotani M., Bassler N., Loeffler C., Bode C., Peter K. Reversibility versus persistence of GPIIb/IIIa blocker-induced conformational change of GPIIb/IIIa (alphaIIbbeta3, CD41/CD61). J Pharmacol Exp Ther. 2004;308(3):1002–11. [PubMed: 14617694]

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von zur Muhlen C., Peter K., Ali Z.A., Schneider J.E., McAteer M.A., Neubauer S., Channon K.M., Bode C., Choudhury R.P. Visualization of activated platelets by targeted magnetic resonance imaging utilizing conformation-specific antibodies against glycoprotein IIb/IIIa. J Vasc Res. 2009;46(1):6–14. [PMC free article: PMC2914450] [PubMed: 18515970]

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von Zur Muhlen C., von Elverfeldt D., Choudhury R.P., Ender J., Ahrens I., Schwarz M., Hennig J., Bode C., Peter K. Functionalized magnetic resonance contrast agent selectively binds to glycoprotein IIb/IIIa on activated human platelets under flow conditions and is detectable at clinically relevant field strengths. Mol Imaging. 2008;7(2):59–67. [PMC free article: PMC2912508] [PubMed: 18706288]

8.

von zur Muhlen C., von Elverfeldt D., Moeller J.A., Choudhury R.P., Paul D., Hagemeyer C.E., Olschewski M., Becker A., Neudorfer I., Bassler N., Schwarz M., Bode C., Peter K. Magnetic resonance imaging contrast agent targeted toward activated platelets allows in vivo detection of thrombosis and monitoring of thrombolysis. Circulation. 2008;118(3):258–67. [PubMed: 18574047]

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Yamada T., Kidera A. Tailoring echistatin to possess higher affinity for integrin alpha(IIb)beta(3). FEBS Lett. 1996;387(1):11–15. [PubMed: 8654558]

10.

Klink A., Lancelot E., Ballet S., Vucic E., Fabre J.E., Gonzalez W., Medina C., Corot C., Mulder W.J., Mallat Z., Fayad Z.A. Magnetic resonance molecular imaging of thrombosis in an arachidonic acid mouse model using an activated platelet targeted probe. Arterioscler Thromb Vasc Biol. 2010;30(3):403–10. [PMC free article: PMC2864133] [PubMed: 20139362]