Potent cytotoxicity of an auristatin-containing antibody-drug conjugate targeting melanoma cells expressing melanotransferrin/p97

• Published in: Molecular cancer therapeutics Vol. 5; no. 6; pp. 1474 - 1482
• Main Authors: Smith, Leia M, Nesterova, Albina, Alley, Stephen C, Torgov, Michael Y, Carter, Paul J
• Format: Journal Article
• Language: English
• Published: United States American Association for Cancer Research 01.06.2006
• Subjects: Animals; Antibodies, Monoclonal - therapeutic use; antibody-drug conjugates; Antigens, Neoplasm - immunology; auristatin; Biomarkers, Tumor - metabolism; Cell Survival - drug effects; Drug Delivery Systems; Humans; Immunoconjugates - therapeutic use; melanoma; Melanoma - drug therapy; Melanoma - immunology; Melanoma-Specific Antigens; melanotransferrin; Mice; Neoplasm Proteins - immunology; Oligopeptides - therapeutic use; p97; Skin Neoplasms - drug therapy; Skin Neoplasms - immunology; therapeutics; Tumor Cells, Cultured
• DOI: 10.1158/1535-7163.MCT-06-0026
• ISSN: 1535-7163

Identifying factors that determine the sensitivity or resistance of cancer cells to cytotoxicity by antibody-drug conjugates is essential in the development of such conjugates for therapy. Here the monoclonal antibody L49 is used to target melanotransferrin, a glycosylphosphatidylinositol-anchored glycoprotein first identified as p97, a cell-surface marker in melanomas. L49 was conjugated via a proteolytically cleavable valine-citrulline linker to the antimitotic drug, monomethylauristatin F (vcMMAF). Effective drug release from L49-vcMMAF likely requires cellular proteases most commonly located in endosomes and lysosomes. Melanoma cell lines with the highest surface p97 expression (80,000–280,000 sites per cell) were sensitive to L49-vcMMAF whereas most other cancer cell lines with lower p97 expression were resistant, as were normal cells with low copy numbers (≤20,000 sites per cell). Cell line sensitivity to L49-vcMMAF was found by immunofluorescence microscopy to correlate with intracellular fate of the conjugate. Specifically, L49-vcMMAF colocalized with the lysosomal marker CD107a within sensitive cell lines such as SK-MEL-5 and A2058. In contrast, in resistant cells expressing lower p97 levels (H3677; 72,000 sites per cell), L49-vcMMAF colocalized with caveolin-1, a protein prominent in caveolae, but not with CD107a. Thus, for antibody-drug conjugates targeting p97, antigen level and trafficking to the lysosomes are important factors for achieving robust in vitro cytotoxicity against cancer cells. Immunohistochemical analysis with L49 revealed that 62% of metastatic melanoma tumors had strong staining for p97. Overexpression of p97 in melanoma as compared with normal tissue, in conjunction with the greater sensitivity of tumor cells to L49-vcMMAF, supports further evaluation of antibody-drug conjugates for targeting p97-overexpressing tumors. [Mol Cancer Ther 2006;5(6):1474–82]