Presentation Authors: In Ho Chang*, Byung Hoon Chi, Jin Wook Kim, Tae‑Hyoung Kim, Kyung Do Kim, Young Tae Moon, Seoul, Korea, Republic of
Introduction: Although Mycobacterium bovis Bacillus Calmette-GuÃ©rin (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
Methods: We constructed recombinant BCG (rBCG) strains expressing Streptococcal inhibitor of complement (Sic), which confers resistance to human Î±-defensin-1 and cathelicidin, and d-alanyl carrier protein ligase (dltA), which confers resistance to cationic AMPs. Sic and dltA 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 dltA genes in total mRNA of the rBCG strains containing Sic (rBCG-Sic) and dltA (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-Î±, and INF-Î³, resulting in inhibition of bacterial killing and immune modulation via antimicrobial peptides.
Conclusions: rBCG-Sic and rBCG-dltA can effectively evade BCG-stimulated AMPs may be significantly improved immunotherapeutic tools to treat bladder cancer.
Source of Funding: This research was supported by a Chung-Ang University Research Scholarship Grant in 2018 and by the Korea Health Technology R&D Project (HI17C0710 to I.H.C.)