Stable expression of Human B7-H4 in a Mouse Mammary Tumor Model as a Target for Cancer Immunotherapy.

摘要

B7-H4 is a member of the B7 family, which is a costimulatory molecule and negatively regulates the immune response of T cells. Its mechanism of tumoral immune escape and therapeutic targeting is still unknown. B7-H4 is overly expressed in breast cancer, ovarian cancer, and many solid tumors and is being investigated as a good immunotherapy target for antibodies or antibody drug conjugates. To elucidate its immune function as well as investigate newly developed anti-B7-H4 antibodies, the murine mammary tumor cell line, 4T1, was transfected with the human B7-H4 gene to generate a syngeneic tumor model in immuno-competent BALB/c mice. To facilitate the generation of a stabile cell line for in vivo studies, 4T1 a mouse breast cancer cell line with metastatic potential similar to that seen in human breast cancer, was transfected with the pIRES-eGFP plasmid containing an internal ribosomal entry site of encephalomyocarditis virus (CMV), the gene for green fluorescent protein (GFP), and the Genetecin resistant gene for selection. The human B7-H4 gene was first amplified from the SKBR3 human breast cancer cell line known to be a high expressor of B7-H4 and cloned into pIRES-eGFP via the EcoRI and BamHI restriction sites. Tumors grown on BALB/c mice with transfected 4T1 cells were not rejected demonstrating the utility of this approach.