Purpose: Although Mycobacterium bovis Bacillus Calmette-Guerin (BCG) is the most widely used bladder cancer immunotherapy, innate immune responses involving antimicrobial peptides (AMPS) cause BCG failure and unwanted side effects. Here, we generated genetically modified BCG strains with improved immunotherapeutic effects by adding genes that confer evasion of AMPs. Materials and methods: We constructed recombinant BCG (rBCG) strains expressing Streptococcal inhibitor of complement (Sic), which confers resistance to human alpha-defensin-1 and cathelicidin, and dalanyl carrier protein ligase (dltA), which confers resistance to cationic AMPs. Sic and dItA were separately cloned into the pMV306 plasmid and introduced into BCG via electroporation. Then, the efficacy of the rBCGs was tested in a growth inhibition assay using two bladder cancer cell lines (5637, T24). Results: We confirmed the presence of cDNA segments corresponding to the Sic and dItA genes in total mRNA of the rBCG strains containing Sic (rBCG-Sic) and dItA (rBCG-dltA), and these rBCGs showed higher survival against AMPs. The growth inhibitory effects of rBCGs on bladder cancer cells were significantly enhanced compared to those of the parent BCG, and THP-1 migration also increased. After 8 h of infection, the levels of internalization were higher in rBCG-infected bladder cancer cells than in BCG-infected cells, and cells infected with rBCGs showed increased release of antitumor cytokines, such as IL-6/12, TNF-alpha, and INF-gamma, resulting in inhibition of bacterial killing and immune modulation via antimicrobial peptides. Conclusions: rBCG-Sic and rBCG-dltA can effectively evade BCG-stimulated AMPs, and may be significantly improved immunotherapeutic tools to treat bladder cancer. (C) 2018 Elsevier Inc. All rights reserved.