Application of Minicircle Technology of Self-Reproducing Synthetic Protein Drugs in Preventing Skin Allograft Rejection
Sun Woo Lim, Young Kyun Kim, Narae Park, Long Jin, Jian Jin, Kyoung Chan Doh, Ji Hyeon Ju, Chul Woo Yang
(Convergent Research Consortium for Immunologic Disease, Seoul St, Mary’s Hospital, The Catholic University of Korea, Seoul, South Korea)
Ann Transplant 2015; 20:430-440
Recently, it has been reported that minicircle vectors could allow the expression of transgenes using the protein synthesis system of the host. Here, we tested a novel strategy to permit the production of synthetic biologics using minicircle technology and evaluated their feasibility as a therapeutic tool in a skin allograft model.
MATERIAL AND METHODS: We engineered vectors to carry cassette sequences for tocilizumab [anti-soluble interleukin-6 receptor (sIL-6R) antibody] and/or etanercept [tumor necrosis factor receptor 2 (TNFR2)-Fc fusion protein], and then isolated minicircle vectors from the parent vectors. We verified the production of proteins from minicircles and their duration in HEK293T cells and mice. We also evaluated whether these proteins were expressed at levels sufficient to ameliorate skin allograft rejection in mice.
RESULTS: Each minicircle transfected into cells was detectable for at least 30 days. In mice, the drugs were mainly expressed in the liver and were detectable for at least 10 days after a single injection. These drugs were also detected in the blood. Treatment of mice with minicircles prolonged skin allograft survival, which was accompanied by a reduction of the number of interferon-γ+ or interleukin-17+ lymphocytes and an induction of forkhead box P3 expression. These findings suggest that blocking of sIL-6R and/or TNF-α using minicircles encoding tocilizumab and/or etanercept was functionally active and relevant for preventing acute allograft rejection.
CONCLUSIONS: Self-reproducing synthetic protein drugs produced using minicircle technology are potentially powerful tools for preventing acute rejection in transplantation.
Keywords: Graft Rejection, Interleukin-6, Skin Transplantation, Tumor Necrosis Factor-alpha