Cardiovascular pathologies represent one of the main causes of death and morbidity in the world[1]. Fucoidans, sulfated polysaccharides extracted from brown algae are versatile biomolecules endowed with numerous biological activities[2]-[7]. In particular, a low molecular weight fucoidan (LMWF), able to strongly bind to P-selectin, allowed the diagnosis of vascular thrombosis and ischemia by SPECT imaging with 99mTc in rat models[8]-[10].
The radiolabeling studies proceeded by straightly mixing a saline solution of LMWF with 99mTcO4- in appropriate conditions without any chemical modification of the polysaccharide[9]. As part of a joint laboratory with the French company Algues & Mer which produces the LMWF, our works aim to obtain a contrast agent kit for the clinical SPECT imaging of atherothrombosis and heart ischemia.
Materials and Methods: LMWF is prepared by an aqueous treatment of Ascophyscient®, a polysaccharidic extract from Ascophyllum nodosum (Algues & Mer, Ouessant, France)[10]. Molecular weights were determined using HPSEC-MALLS-dRI[9]. Colorimetric assays led to fucose[11], uronic acid[12] and sulfate[13] contents and the neutral sugar composition was obtained by acid hydrolysis of LMWF and HPAEC analysis with amperometric detection. The contrast agent was prepared by mixing a solution of ascorbic acid, tin chloride and sodium chloride and another one containing the LMWF and sodium chloride. 99mTcO4- was added just before injection.
Results and Discussion: Fucoidans are natural polymers and their chemical compositions vary depending on the species, water temperature and the harvest period of the algae. For the preparation of a clinical contrast agent, LMWF had to present well-defined and reproducible physicochemical features. Numerous batches of LMWF have been purified from Ascophyscient® and analyzed to provide these parameters.
The average molecular weights were about 8,000 Da. Fucose and sulfate, as part of the key features responsible for the biological activities of fucoidans[14],[15], and uronic acids constitute about 80% (w/w) of the polymer, with a few amounts of xylose, glucose, galactose and mannose as the remaining 20%.
A radiolabelling kit was prepared, all reactants being kept lyophilized until the radiolabelling with a saline solution of 99mTcO4-. Purity was >95%. The biodistribution study in rat of both compounds labeled with 99mTc evidenced a predominant renal elimination of the purified fucoidan, whereas the crude fucoidan (Ascophyscient®) was mainly retained in liver and spleen. SPECT imaging of rat models have shown the effectiveness and the selectivity of 99mTc-LMWF toward pathological myocardium (figure). All this work is part of a large-scale european project (NanoAthero).
Conclusion: This purified sulfated polysaccharide appears promising for the development of molecular imaging in acute coronary syndrome and a clinical trial will start on 2016.
ANR LabCom; ANR FucoThrombo; ANRT for CIFRE fellowship n°2014/0337 (Lucas Chollet); Large-scale integrated project NanoAthero (NMP-2012-309820)
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