The Interaction of MS-325 with Human Serum Albumin and Its Effect on Proton Relaxation Rates

Abstract

MS-325 is a novel blood pool contrast agent for magnetic resonance imaging currently undergoing clinical trials to assess blockage in arteries. MS-325 functions by binding to human serum albumin (HSA) in plasma. Binding to HSA serves to prolong plasma half-life, retain the agent in the blood pool, and increase the relaxation rate of water protons in plasma. Ultrafiltration studies with a 5 kDa molecular weight cutoff filter show that MS-325 binds to HSA with stepwise stoichiometric affinity constants (mM(-1)) of K(a1) = 11.0 +/- 2.7, K(a2) = 0.84 +/- 0.16, K(a3) = 0.26 +/- 0.14, and K(a4) = 0.43 +/- 0.24. Under the conditions 0.1 mM MS-325, 4.5% HSA, pH 7.4 (phosphate-buffered saline), and 37 degrees C, 88 +/- 2% of MS-325 is bound to albumin. Fluorescent probe displacement studies show that MS-325 can displace dansyl sarcosine and dansyl-L-asparagine from HSA with inhibition constants (K(i)) of 85 +/- 3 microM and 1500 +/- 850 microM, respectively; however, MS-325 is unable to displace warfarin. These results suggest that MS-325 binds primarily to site II on HSA. The relaxivity of MS-325 when bound to HSA is shown to be site dependent. The Eu(III) analogue of MS-325 is shown to contain one inner-sphere water molecule in the presence and in the absence of HSA. The synthesis of an MS-325 analogue, 5, containing no inner-sphere water molecules is described. Compound 5 is used to estimate the contribution to relaxivity from the outer-sphere water molecules surrounding MS-325. The high relaxivity of MS-325 bound to HSA is primarily because of a 60-100-fold increase in the rotational correlation time of the molecule upon binding (tau(R) = 10.1 +/- 2.6 ns bound vs 115 ps free). Analysis of the nuclear magnetic relaxation dispersion (T(1) and T(2)) profiles also suggests a decrease in the electronic relaxation rate (1/T(1e) at 20 MHz = 2.0 x 10(8) s(-1) bound vs 1.1 x 10(9) s(-1) free) and an increase in the inner-sphere water residency time (tau(m) = 170 +/- 40 ns bound vs 69 +/- 20 ns free).

The extraordinary ligand binding properties of human serum albumin

“…agent is located in the subdomain IIIA of HSA. Interestingly, dansyl-L-asparagine is also displaced by the metal complex, but only at much higher concentrations, allowing it to localize in the subdomain IIA, a weaker binding site (66). However, such analysis did not consider the occurrence of allosteric effects, and it is worth noting that a relevant conformational change of the protein structure has been suggested to explain the experimental observation that several substrates, known to have different HSA binding sites, invariantly reduced the affinity of some Gd-DTPA-like chelates (including MS-325) to the protein (63).…”

High‐Relaxivity MRI Contrast Agents: Where Coordination Chemistry Meets Medical Imaging

“…Once bound to HSA, the proton relaxation efficiency of MS-325 increases and the longer in vivo retention times present opportunities for MR angiography. [13][14][15] Common to all the aminocarboxylatebased commercial agents is the octadentate ligand motif, a chelate design that leaves only one open coordination site for a single innersphere water molecule. This low hydration number (q) limits the potential effectiveness of these complexes as relaxation agents (vide infra).…”

Paramagnetic liposomes containing amphiphilic bisamide derivatives of Gd-DTPA with aromatic side chain groups as possible contrast agents for magnetic resonance imaging

“…In order to prolong the intravascular retention of a contrast agent, several different strategies have been proposed so far. Attachment of a protein-binding group such as diphenylcyclohexyl to a gadolinium(III) chelate via a phosphodiester linkage, like in the MS-325 contrast agent, results in reversible binding of MS-325 to human serum albumin in plasma (Caravan et al 2002). The binding to human serum albumin reduces the extravasation of the contrast agent out of the vascular system and also leads to a high increase in relaxivity.…”