title: Impediments to enhancement of CPT-11 anticancer activity by E. coli directed beta-glucuronidase therapy.
abstract: CPT-11 is a camptothecin analog used for the clinical treatment of colorectal adenocarcinoma. CPT-11 is converted into the therapeutic anti-cancer agent SN-38 by liver enzymes and can be further metabolized to a non-toxic glucuronide SN-38G, resulting in low SN-38 but high SN-38G concentrations in the circulation. We previously demonstrated that adenoviral expression of membrane-anchored beta-glucuronidase could promote conversion of SN-38G to SN-38 in tumors and increase the anticancer activity of CPT-11. Here, we identified impediments to effective tumor therapy with E. coli that were engineered to constitutively express highly active E. coli beta-glucuronidase intracellularly to enhance the anticancer activity of CPT-11. The engineered bacteria, E. coli (lux/βG), could hydrolyze SN-38G to SN-38, increased the sensitivity of cultured tumor cells to SN-38G by about 100 fold and selectively accumulated in tumors. However, E. coli (lux/βG) did not more effectively increase CPT-11 anticancer activity in human tumor xenografts as compared to non-engineered E. coli. SN-38G conversion to SN-38 by E. coli (lux/βG) appeared to be limited by slow uptake into bacteria as well as by segregation of E. coli in necrotic regions of tumors that may be relatively inaccessible to systemically-administered drug molecules. Studies using a fluorescent glucuronide probe showed that significantly greater glucuronide hydrolysis could be achieved in mice pretreated with E. coli (lux/βG) by direct intratumoral injection of the glucuronide probe or by intratumoral lysis of bacteria to release intracellular beta-glucuronidase. Our study suggests that the distribution of beta-glucuronidase, and possibly other therapeutic proteins, in the tumor microenvironment might be an important barrier for effective bacterial-based tumor therapy. Expression of secreted therapeutic proteins or induction of therapeutic protein release from bacteria might therefore be a promising strategy to enhance anti-tumor activity.
authors: Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, Taiwan; Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
authors: Yuan-Ting
authors: YT
authors: Hsieh
authors: Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
authors: Kai-Chuan
authors: KC
authors: Chen
authors: Department of Aquaculture, National Kaohsiung Marine University, Kaohsiung, Taiwan.
authors: Chiu-Min
authors: CM
authors: Cheng
authors: Department of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors: Tian-Lu
authors: TL
authors: Cheng
authors: Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
authors: Mi-Hua
authors: MH
authors: Tao
authors: Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
authors: Steve R
authors: SR
authors: Roffler
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology*; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D045744:Cell Line, Tumor; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005456:Fluorescent Dyes / Q000378:metabolism; D005818:Genetic Engineering; D015316:Genetic Therapy; D005966:Glucuronidase / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D000077146:Irinotecan; D051379:Mice; D014170:Transformation, Genetic
chemical_list: D000970:Antineoplastic Agents; D005456:Fluorescent Dyes; D000077146:Irinotecan; D005966:Glucuronidase; D002166:Camptothecin