Small Molecule Therapeutics PET Imaging of b-Glucuronidase Activity by an Activity-Based I-Trapping Probe for the Personalized Glucuronide Prodrug Targeted Therapy

Beta-glucuronidase (bG) is a potential biomarker for cancer diagnosis and prodrug therapy. The ability to image bG activity in patients would assist in personalized glucuronide prodrug cancer therapy. However, whole-body imaging of bG activity for medical usage is not yet available. Here, we developed a radioactive bG activity–based trapping probe for positron emission tomography (PET). We generated a I-tyramine–conjugated difluoromethylphenol beta-glucuronide probe (TrapG) to form I-TrapG that could be selectively activated by bG for subsequent attachment of I-tyramine to nucleophilic moieties near bG-expressing sites. We estimated the specificity of a fluorescent FITC-TrapG, the cytotoxicity of tyramine-TrapG, and the serum half-life of I-TrapG. bG targeting of I-TrapG in vivo was examined by micro-PET. The biodistribution of I-TrapG was investigated in different organs. Finally, we imaged the endogenous bG activity and assessed its correlation with therapeutic efficacy of 9-aminocamptothecin glucuronide (9ACG) prodrug in native tumors. FITC-TrapG showed specific trapping at bG-expressing CT26 (CT26/mbG) cells but not in CT26 cells. The native TrapG probe possessed low cytotoxicity. ITrapG preferentially accumulated in CT26/mbG but not CT26 cells. Meanwhile, micro-PET and wholebody autoradiography results demonstrated that I-TrapG signals in CT26/mbG tumors were 141.4-fold greater than in CT26 tumors. Importantly, Colo205 xenografts in nude mice that express elevated endogenous bG can be monitored by using infrared glucuronide trapping probes (NIR-TrapG) and suppressed by 9ACG prodrug treatment. I-TrapG exhibited low cytotoxicity allowing long-term monitoring of bG activity in vivo to aid in the optimization of prodrug targeted therapy. Mol Cancer Ther; 13(12); 2852–63. 2014 AACR. Introduction b-Glucuronidase (bG) can catalyze the hydrolysis of b-D-glucuronic acid residues for the breakdown of mucopolysaccharides (e.g., heparan sulfate) in lysosomes (1). It has been widely used as an attractive enzyme for reporter imaging (2–4) and cancer prodrug therapies (5–8). Several glucuronide prodrugs have been used in antibody-directed enzyme prodrug therapy (ADEPT; refs. 9–11) and gene-directed enzyme prodrug therapy (GDEPT; refs. 6, 12, 13) as well as directly in prodrug monotherapy, which relies on the elevated levels of bG found in the tumor microenvironment (14–16) to selectively convert prodrug into active drug. For instance, Albin and colleagues reported that the level of bG human breast tumors was up to 6-times greater than that in normal tissues (17). Sperker and colleagues also reported that pancreatic adenocarcinoma exhibited higher bG levels than did normal pancreatic tissue (14). Furthermore, human tumor xenografts also showed elevated levels of bG, including human breast (MCF-7, BT20, and HS578T), colon (HT29 and SW480), and small-cell lung cancer (OH3 and SW2) cell Institute ofMicrobiologyand Immunology,NationalYang-MingUniversity, Taipei, Taiwan. Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan. Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan. Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan. Department of Pharmacy, Chia Nan University of Pharmacy andScience, Tainan, Taiwan. Department ofSurgery, Faculty of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan. Institute of Basic Medical Sciences, National Cheng Kung University, Tainan, Taiwan. Department of Biomedical Imaging andRadiological Sciences, National Yang-Ming University, Taipei, Taiwan. Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan. Department of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung, Taiwan. Center for Biomarkers and Biotech Drugs, Kaohsiung Medical University, Kaohsiung, Taiwan. Note: Supplementary data for this article are available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/). Y.-C. Su and T.-C. Cheng contributed equally to this article. Corresponding Authors: Tian-Lu Cheng, Department of Biomedical and Environmental Biology, KaohsiungMedical University, 100Shih-Chuan 1st Road, Kaohsiung, Taiwan. Phone: 886-7-3121101, ext. 2360; Fax: 886-73227508; E-mail: [email protected]; and Hsin-Ell Wang, Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, No. 155, Sec. 2, Li-Nong Street, Taipei, Taiwan. Phone: 886-228267215; Fax: 886-2-28201095; E-mail: [email protected] doi: 10.1158/1535-7163.MCT-14-0212 2014 American Association for Cancer Research. Molecular Cancer Therapeutics Mol Cancer Ther; 13(12) December 2014 2852 on June 22, 2017. © 2014 American Association for Cancer Research. mct.aacrjournals.org Downloaded from Published OnlineFirst October 2, 2014; DOI: 10.1158/1535-7163.MCT-14-0212