mRNA loading into ATP-responsive polyplex micelles with optimal density of phenylboronate ester crosslinking to balance robustness in the biological milieu and intracellular translational efficiency.

Affiliation

Yoshinaga N(1), Uchida S(2), Dirisala A(3), Naito M(4), Osada K(5), Cabral H(1), Kataoka K(6).
Author information:
(1)Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.
(2)Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan; Innovation Center of NanoMedicine
(iCONM), Kawasaki Institute of Industrial Promotion, 3-25-14 Tonomachi, Kawasaki-ku, Kawasaki 210-0821, Japan. Electronic address: [Email]
(3)Innovation Center of NanoMedicine
(iCONM), Kawasaki Institute of Industrial Promotion, 3-25-14 Tonomachi, Kawasaki-ku, Kawasaki 210-0821, Japan.
(4)Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
(5)Quantum Medical Science Directorate, National Institutes for Quantum and Radiological Science and Technology
(QST), 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan.
(6)Innovation Center of NanoMedicine
(iCONM), Kawasaki Institute of Industrial Promotion, 3-25-14 Tonomachi, Kawasaki-ku, Kawasaki 210-0821, Japan; Institute for Future Initiatives, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan. Electronic address: [Email]

Abstract

Carriers for messenger RNA (mRNA) delivery require propensities to protect the mRNA from enzymatic degradation and to selectively release mRNA in the cytosol for smooth mRNA translation. To meet these requirements, we designed mRNA-loaded polyplex micelles (PMs) with ATP-responsive crosslinking in the inner core by complexing mRNA with poly(ethylene glycol)-polycation block copolymers derivatized with phenylboronic acid and polyol groups, which form crosslinking structures via spontaneous phenylboronate ester formation. PMs thus prepared are tolerable against enzymatic attack and, in turn, disintegrate in the cytosol to release mRNA when triggered by the cleavage of phenylboronate ester linkages in response to elevated ATP concentration. Two structural factors of the PM, including (i) the introduction ratios of phenylboronate ester crosslinkers and (ii) the structure and protonation degree of amino groups in the polycation segment, are critical for maximizing protein expression in cultured cells due to the optimized balance between the robustness in the biological milieu and the ATP-responsive mRNA release in the cytosol. The optimal PM formulation was further stabilized by installing cholesterol moieties into both the mRNA and ω-end of the block copolymer to elicit longevity in blood circulation after intravenous injection.