A Modified Adenovirus Vector-Mediated Antibody Screening Method Identifies EphA2 as a Cancer Target.

BACKGROUND: We constructed a genetically modified adenovirus vector incorporating an IgG Fc-binding motif from staphylococcal protein A, Z33 (Adv-FZ33). Adv-FZ33 allows an antibody to redirect the vector to a target molecule on the cell surface. We attempted to search for target antigen candidates and antibodies that allowed highly selective gene transduction into malignant tumors.

METHODS: Hybridoma libraries producing monoclonal antibodies (mAbs) were screened that increased transduction efficiency in cancer cell lines after cross-linking with Adv-FZ33. Target antigens of the mAbs were identified by immunoprecipitation and mass spectrometry. Of these mAbs, we noted a clone, F2-27, that recognized the receptor tyrosine kinase EphA2. Next, we generated an adenovirus vector, Ax3CMTK-FZ33, that expressed a herpes simplex virus thymidine kinase (HSV-TK). The therapeutic efficacy of F2-27-mediated HSV-TK gene transduction, followed by ganciclovir (GCV) administration, was studied in vitro. The inhibitory effect of F2-27 on cancer cell invasion was investigated by a three-dimensional spheroid formation assay.

RESULTS: In vitro reporter gene expression after Adv-FZ33 infection via F2-27 was 146 times higher than with control mAb in EphA2-expressing cancer cell lines. F2-27-mediated Ax3CMTK-FZ33 infection induced the HSV-TK gene in an F2-27-dependent manner and had a highly effective cytotoxic effect in a GCV-dependent manner. Additionally, F2-27 independently inhibited migration of EphA2-positive breast cancer cell lines in three-dimensional culture.

CONCLUSION: Our modified adenovirus and hybridoma screening system is useful for the development of targeted cancer therapy, and F2-27 has the potential to be an antibody-based therapy for various EphA2-positive cancers.