Maleimide-functionalized closo-dodecaborate albumin conjugates (MID-AC): Unique ligation at cysteine and lysine residues enables efficient boron delivery to tumor for neutron capture therapy

Development of Albumin‐closo‐Dodecaborate Conjugates as Boron Carriers for Neutron‐Capture Therapy by Ru(bpy)3‐Photocatalyzed Modification of Tyrosine

“…[26] As positive-control experiments, BSA was conjugated to 10 or 100 equivalents of MID at room temperature for 60 min ( Figure 2, lanes 2 and 3). To investigate the conditions for conjugation of 1 or 2 to the tyrosine residues in BSA, we first attempted conjugation to BSA under photocatalytic conditions without ammonium persulfate (APS).…”

“…We found that the modification efficiency increased only slightly, even if more than 10 equivalents of MID was added ( Figure 2, lane 2 vs. 3; see also ref. [26] ). Combined with the current method involving the use of tyrosine residues as the modification scaffold, we expected that we would be able to introduce more boron clusters into one BSA molecule.…”

“…The increase in the band intensity of the doubly modified BSA clearly indicated that MID and 1 were bound to different amino-acid residues in BSA. Subsequently, the effect of the intracellular uptake of the modified BSAs into mouse colon 26 (Figure 3b, left). The band detected with the anti-B 12 cluster antibody, which is visible around 65 kDa, indicated that BSA was taken up by the cells.…”

“…All chemicals and purified proteins for biological experiments were obtained from commercial sources and were used without further purification. [26] Dioxane-linked closo-dodecaborate 3 was synthesized according to the method reported by Bregadze et al [28] To a solution of 3 (1.22 g, 2.58 mmol) in CH 2 Cl 2 (20 mL) was added tetrabutylammonium azide (1.47 g, 5.17 mmol). The mixture was stirred at room temperature for 24 h and was then concentrated under reduced pressure.…”

“…MID albumin conjugates (MID-ACs) showed high and selective in vivo accumulation in tumors, which resulted in significant tumor-growth inhibition by thermal neutron irradiation. [26] On the other hand, we achieved catalytic protein modification of the tyrosine residue by using a ruthenium photocatalyst, such as Ru(bpy) 3 Cl 2 (bpy = 2,2′-bipyridyl) [27] In this method, oxidative radical cross-coupling between a tyrosine residue and a tyrosine radical trapping moiety (TRT), such as N′-acyl-N,Ndimethylphenylenediamine, proceeds under visible-light irradiation in the presence of a ruthenium photocatalyst (Figure 1c). We aimed for the high integration of boron clusters per unit albumin molecule by using tyrosine residues as scaffolds besides a free cysteine residue and lysine residues.…”