Membrane-anchored  -glucuronidase Steve Roffler Institute of Biomedical Sciences Academia Sinica Taipei, Taiwan

MEMzyme Transgene Transgene Expression ß-glucuronidase glucuronide Glucuronide probe or drug B7 TM glucuronide Potential advantage: Signal amplification due to enzymatic hydrolysis of glucuronide probes or drugs Memzyme activation of imaging or therapeutic agents

ßG in lysosomes – inaccessible to glucuronides Low serum ßG – little systemic activation Low toxicity Fast renal clearance – rapid imaging Endogenous - low immunogenicity Many possible glucuronide probes and drugs Potential advantages of a ßG reporter

Fluorescent probe can be activated and accumulate at cells that express enzyme In vitro imaging of gene expression red = enzyme expression green = activated probe accumulation

Sites of Ad transgene expression could be imaged Imaging of adenovirus memzyme expression Kodak IS2000MM

In vitro activation of HAMG HAMG can be activated at EJ/mßG cells O HO OH COOH O N Cl HAMG Glucuronide prodrug N Cl HO pHAM Active drug

HAMG prodrug toxicity to mice HAMG is well tolerated by mice

Prodrug therapy of EJ tumors HAMG displayed selective activity against EJ/mßG tumors EJ PBS EJ HAMG EJ/mßG PBS EJ/mßG HAMG Days Tumor volume (mm 3 ) (150 mg/kg HAMG on days 15 and 19)

Summary Surface enzyme allow both imaging and therapy Immunogenicity is low but selectivity is high May possess advantages for cancer gene therapy

Dr. Kuang-Wen Liao Shey-Cherng Tzo Tang-Bi Liu Surface scFv T cell activation Surface expression Shih-en Chang Wen-Jun Chung Bing-Mae Chen Dr. Yu-Ling Leu Chia-Nan College of Pharmacy and Science Dr. Ji-Wang Chern National Taiwan University Dr. Tian-Lu Cheng Kaohsiung Medical University Chin-Chuan Chen Dr. Hsin-Ell Wang National Yang Ming University Yi-Hsuan Chiang Pei-Yu Wu Chien-I Su