A recombinant adenovirus is applied for treating malignancy of over-expressing proto-oncogene neu/erbB2, wherein an expression cassette, which co-expresses the humanized monoclonal antibody variable region gene of anti proto-oncogene neu/erbB2 and the Mda-7/IL-24 gene, is inserted into E1 deletion region of the recombinant adenovirus. The recombinant adenovirus effectively treats the malignancy of overexpressing proto-oncogene neu/erbB2 without damaging normal cells, such that the recombinant adenovirus is able to be used for the gene therapy of malignancy tumors over-expressing proto-oncogene neu/erbB2.