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Purpose@#Although Mycobacterium bovis Bacillus Calmette-Guérin (BCG) is the most widely used bladder cancer immunotherapy, innate immune responses involving antimicrobial peptides (AMPs) cause BCG failure. Here, we developed genetically modified recombinant BCG (rBCG) strains which escape AMPs and evaluate the efficacy and effects of rBCG. @*Materials and Methods@#We constructed rBCG strains expressing Streptococcal inhibitor of complement (Sic), which confers resistance to human α-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. The efficacy of the Sic and dltA gene electroporation into BCG was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). The internalization rates and anticancer effects of the rBCG strains containing Sic (rBCG-Sic) and dltA (rBCG-dltA) was evaluated by the orthotopic bladder cancer mouse model. @*Results@#The cycle quantification (Cq) values of rBCG-Sic (y=-4.8823x+13.645, R2=0.9996) and rBCG-dltA (y=-5.438x+11.641, R2=0.9995) were inverse correlations to the amount of Sic and dltA genes dose dependently. The mean introduction proportions of Sic and dltA genes into BCG by electroporation were 22.2%, 27.5% and showed constant efficacy. In the orthotopic bladder cancer mouse model, the relative internalization number of rBCG-Sic, and rBCG-dltA into bladder cell in mouse bladder were higher than that of BCG and the tumor volume at rBCG-Sic were lower than at BCG and rBCG-dltA at 11, 14 and 18 days. @*Conclusions@#Our results showed that constructed rBCG-Sic and rBCG-dltA by electroporation and the rBCG-Sic and rBCG-dltA can effectively escape BCG-stimulated AMPs, and significantly improved immunotherapeutic tools to treat bladder cancer in orthotopic bladder cancer mouse model.
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BACKGROUND: Several studies on the frequency and distribution of unexpected antibodies of Koreans have been done by some investgators. But more studies are necessory to grasp the real frequency and distribution. We hoped to investigate the frequency and distribution of unexpected antiboddis of Koreans. METHODS: We performed 188 unexpected antibody identification tests to evaluate the distribution of unexpected antibodies from May 1997 to June 1998. The sera we tested are antibody screening positive ones sent from 11 hospitals and 1 blood center. We also analized the frequecny of unexpected antibodues from the data of Dongsan Medical Center's antibody screening tests. RESULTS: The frequency of unexpected antibody of this study (using tube method) was 1.0%. Both anti-Lea and anti-E (with anti-c) antibodies were the most frequent antibodies up to 31 cases (33.0%) each. Among the identified antibodies from Dongsan Medical Center which used tube method, anti-Lea antibody was the most frequent antibody (41.3%). Otherwise among the identified antibodies from other two hospitals which used gel card method, anti-E (with anti-c) antibody was the most frequent antibody (45.5%). The cause of all four cases of hemolytic disease of newborn was anti-E antibody. About half of the specimens' antibodies were not identified due to the problemes of identification cells, tested sera, test methods. CONCLUSION: Anti-Lea and anti-E (with anti-c) antibodies were the most frequent ones in Korean people. But the prequency was varied according to antibody screenig test methods. Anti-E antibody is the most important unexpected antibidy for hemolytic disease of newborn in Korea. To decrease the missed cases of antibody identification tests both Korean and foreign identification cells, enzyme method should be used.