Basic Research Of TRAIL Recombinant Live E.coli On Tumor-targeting Therapy

Traditional therapies of malignant tumors include surgical operation, chemotherapy and radiotherapy and there are many shortcomings about these therapies such as less specifically targeting to tumor cells, insufficient gene therapy products to kill tumors and more cytotoxicity on normal cells. Currently, a new concept named tumor-targeting therapy is produced. The main aim of tumor-targeting therapy are targeting drugs to tumor cells, sparing normal cells, increasing the drugs concentration in tumor sites and increasing the anti-tumor effects of drugs. So it is the most important that how to choose a tumor-targeting vector and an anti-tumor agent.Many bacteria such as Clostridum novyi, Bifidobacterium longum and Salmonella have a nature to targeting to tumor site and some of them per se could kill tumor cells. Escherichia coli is a gram-negative bacteria that naturally resides in the digestive tracts of humans and other animals. Yu et al. showed that Escherichia coli DH5αcould selectively target to tumors, but there have been no reports to date describing the use of E. coli as a vector to repress tumor growth in vivo. In this paper, we first have determined the safe dose of E. coli DH5αin mice and validated its tumor-targeting nature in different tumor models. At the same time, we found tumor-targeting of E. coli DH5αwas not limited by tumor size and they could colonise large tumors and metastatic tumor nodules as small as 1mm in diameter. High tolerance in mice and more extensive tumor-targeting make E. coli DH5αbe an ideal vector used in tumor theapy.Tumor necrosis factor-related apoptosis inducing ligand (TRAIL) is a new found member of TNF family and is made up of 281 amino acids. The soluble extracellular domains of TRAIL (amino acids 95-281, 104-281 or 114-281) induce apoptosis by activating the cell surface death receptors 4 and 5 (DR4/DR5) in a wide variety of tumor cell lines, but are not cytotoxic to normal cells in vitro. Repeated i.v. administration of recombinant, biologically-active TRAIL (rTRAIL) in preclinical experiments can induce tumor cell apoptosis, suppress tumor progression, and improve survival in tumor-bearing mice, suggesting TRAIL may be exploited as a powerful anti-tumor agent. Unfortunately, the use of TRAIL as a therapeutic agent is limited by severe cytotoxicity in normal hepatocytes, esophageal epithelial cells, prostate cells in vitro and keratinocytes, and its short half-life after systemic administration.E. coli DH5αis an extracellular bacterium and not enter into cells. They are suitable for expressing a therapeutic protein such as an extracellular inducer of apoptosis. An ideal candidate protein is sTRAIL, which induces apoptosis by binding with the cell surface death receptors. In our study, we first constructed a prokaryotic expression vector of sTRAIL and found E. coli DH5αexpressing sTRAIL had cytotoxic effects on tumor cells and induced tumor cells apoptosis in dose or time-dependent fashion in vitro when coincubated with tumor cells. These in vitro results encouraged us to evaluate whether tumor apoptosis from E. coli DH5αexpressing sTRAIL could be demonstrated in vivo. Our results showed that intratumoral and intravenous injection of E. coli DH5αexpressing sTRAIL could specically target to tumors and slowly release biologically active sTRAIL protein, which lead to a significant reduction in the rate of tumor growth and the prolonged survival of tumor-bearing mice. sTRAIL delivered by E. coli DH5αdid not cause detectable toxicity to any of the examined organs of the mice in vivo, suggesting that E. coli DH5α-mediated sTRAIL therapy may be further developed into a feasible and effective form of treatment for solid tumors. TRAIL protein delivery by E. coli DH5αused in tumor therapy combine the high tumor-targeting nature of E. coli DH5αand more efficient tumor killing character of TRAIL protein. Meanwhile, this method overcome little anti-tumor effect of E. coli DH5αand short half-life, less tumor-targeting and potential toxicity of TRAIL protein. TRAIL protein delivered by E. coli DH5αmay be providing a feasible and effective form of treatment for solid tumors.