627. Enhanced Transduction and Antitumor Efficiency of Fiber-Modified, CD133-Targeted Oncolytic Adenovirus in Colon Cancer

Abstract

CD133 (Prominin-1), a member of the transmembrane glycoprotein family, is a cell surface marker of cancer stem cells (CSCs) in many malignancies including colon cancer. In colorectal cancer, CD133 expression was correlated with recurrence, metastases and chemotherapy resistance, as well as poor survival. It is thus reasonable to develop novel therapeutic strategies to target CD133 in order to attack colon cancer stem cells. We have been developing therapeutic approaches based on the oncolytic adenovirus, and have recently established a method for isolating transductionally-targeted adenovirus which selectively binds to cell surface molecule by high-throughput screening of the fiber-modified adenovirus library. In this study, we established the CD133-specific oncolytic adenovirus (OAd) and its oncolytic activity was tested in vitro and in vivo, in order to development for new CSCs-targeted therapeutic strategies.Following sequential rounds of screening one clone, TYMLSRN, was identified that could replicate selectively in CD133-expressing 293 cells (293-CD133). CD133-targeted OAd (AdML-TYML) showed strong binding to CD133-positive cells (293-CD133 and LoVo (CD133 (+) human colon carcinoma)), not to CD133-negative cells (parental 293 and LS174T (CD133 (-) human colon carcinoma)). In addition, the anti-CD133 antibody inhibited the binding of AdML-TYML to CD133. To assess the in vitro oncolysis of the CD133-targeted OAd, human colon cancer cell lines were infected and the cytocidal effect was assessed by crystal violet staining. AdML-TYML showed strong and selective oncolysis in LoVo cells (CD133 (+)), whereas there was no effect on LS174T cells (CD133 (-)).In order to assess the in vivo function of AdML-TYML in CSCs, tumor formation assay was performed. While 100% of the mice inoculated with LoVo cells without OAd treatment had developed tumors, none of the mice with AdML-TYML-treated LoVo cells generated a tumor. Contrarily, there was no significant difference between non-infected cells (100%) and infected cells (75%) in CD133-negative LS174T cells. These results indicate that CD133-targeted OAd leads to a reduction in tumor formation of CD133-positive cell lines in vivo. When the anti-tumor effect of the CD133-targeted OAd was analyzed in established tumors of LoVo colon cancer subcutaneous xenografts, intra-tumor (i. t.) administration of AdML-TYML exhibited significantly stronger antitumor effect compared to its equivalent AdML-5WT (wild type Ad5 fiber).In this study, we focused on the CD133 as a target molecule of colon cancer CSCs and successfully identified potent CD133-targeting OAd using high-throughput screening system. The CD133-targeted OAd exhibited selective infectivity and oncolysis to CD133-positive cells. Inhibition of tumorigenicity and therapeutic effect in established tumor was observed in colon cancer in vivo models. The same approach can be applied to other cancers whose CSC marker is CD133. The ability of CSCs-targeted Ad such as CD133-targeted OAd to infect and kill CSCs has important implications for the prevention of metastases and relapses in a variety of cancers.