公开(公告)号：US12018081B1

公开(公告)日：2024-06-25

申请号：US17006635

申请日：2020-08-28

申请人： The Regents of the University of Colorado, a body

发明人： Andrew P. Goodwin ,  Jennifer N. Cha ,  Shambojit Roy ,  Michael D. Brasino

IPC分类号： C07K16/28 ,  A61K33/16 ,  A61K39/00 ,  C12N9/78

CPC分类号： C07K16/2863 ,  A61K33/16 ,  C12N9/78 ,  A61K2039/505 ,  C12Y305/04001

摘要： A prodrug enzyme covalently photoconjugated to a live cell receptor survives endosomal proteolysis and retains its catalytic activity on the living cell membrane over multiple days. Antibody-directed enzyme prodrug therapy is a promising approach for selective treatment of solid tumors, but methods are needed to preserve enzyme activity on living cell membranes over multiple prodrug dosings. A fusion protein was designed with both an anti-epidermal growth factor receptor (EGFR) affibody and the prodrug enzyme cytosine deaminase, which can convert prodrug 5-fluorocytosine to the anticancer drug 5-fluorouracil. A benzophenone group was added at a site-specific mutation within the affibody portion, and the fusion protein was selectively and irreversibly photoconjugated to EGFR receptors expressed on membranes of live MDA-MB-468 breast cancer cells. Affinity-mediated covalent conjugation of the affibody-enzymes to cell receptors allows for prolonged expression on membranes and retained enzymatic activity without genetic engineering.