Anti-tumor activity of K1-LysPE38QQR, an immunotoxin targeting mesothelin,a cell-surface antigen overexpressed in ovarian cancer and malignant mesothelioma

Mesothelin, a differentiation antigen, is a 40-kD glycosylphosphatidylinosi tol-linked cell-surface glycoprotein, that is present on the surface of nor mal mesothelium and is overexpressed in many patients with epithelial ovari an cancer and malignant mesotheliomas. Monoclonal antibody K1 is a murine i mmunoglobulin G1 that recognizes mesothelin. LysPE38QQR is a truncated form of Pseudomonas exotoxin that lacks the cell-binding domain, but retains th e translocation and adenosine diphosphate-ribosylation domains. It has a si ngle lysine residue near the amino terminus that is available for conjugati on to antibodies. To prevent chemical conjugation of the antibody to lysine residues at the C-terminus of Pseudomonas exotoxin, the two lysine residue s at positions 590 and 606 were mutated to glutamine, and the lysine residu e at position 613 was mutated to arginine. Monoclonal antibody K1 was chemi cally conjugated with LysPE38QQR, by modifying the antibody with sulfosucci nimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate and coupling it with SPDP N-succinimidyl 3-(2-pyridyldithio)propionate-modified LysPE38QQR. The resulting immunotoxin K1-LysPE38QQR was highly toxic to A431-K5 cells (a h uman epidermoid carcinoma cell line transfected with a mesothelin expressio n plasmid) with a half-maximal inhibitory concentration of 3-6 ng/mL. The i mmunotoxin had negligible activity against A431 cells, which do not express mesothelin (median inhibitory concentration > 100 ng/mL). This immunotoxin also caused complete regression of turners in nude mice that received xeno grafts of mesothelin-positive human carcinomas. These results show that imm unotoxins directed against mesothelin are a therapeutic option that merits further investigation for the treatment of ovarian cancer and malignant mes otheliomas.