Tumor-specific intravenous gene delivery using oncolytic adenoviruses

Abstract

In this report, we describe a vector system that specifically delivers transgene products to tumors following intravenous (i.v.) administration. The Escherichia coli cytosine deaminase (CD) gene was placed in the E3B region of the tumor-selective, replicationcompetent adenovirus ONYX-411, under the control of endogenous viral late gene regulatory elements. Thus, CD expression was directly coupled to the tumor-selective replication of the viral vector. In vitro, CD was expressed efficiently in various human cancer cell lines tested but not in cultured normal human cells, including human hepatocytes. Following i.v. administration into nude mice carrying human tumor xenografts, robust CD activity was detected only in tumors but not in liver or other normal tissues. Levels of CD activity in the tumors increased progressively following i.v. virus administration, correlating closely with virus replication in vivo. Subsequent administration of 5-fluorocytosine (5-FC) demonstrated a trend to improve the antitumor efficacy of these viruses in a mouse xenograft model, presumably due to the intratumoral conversion of 5-FC to the chemotherapeutic drug 5-fluorouracil. We show that the combination of a highly selective oncolytic virus, ONYX-411, with the strategic use of the viral E3B region for transgene insertion provides a powerful platform that allows for tumor-specific, persistent and robust transgene expression after i.v. administration. This technology provides an opportunity to enhance greatly both safety and efficacy of cancer gene therapy.

Comparison of the E3 and L3 regions for arming oncolytic adenoviruses to achieve a high level of tumor-specific transgene expression

“…Transgene expression is restricted from Ad-14.7-GM and Ad-23K-GM viruses after intravenous administration in nude mice Previous studies have shown that expression of the transgenes can be targeted to the tumor but prevented in non-target organs such as the liver by linking transgene expression to viral replication, 15 thereby improving the safety profile of the vectors. To determine if inserting the transgene into the E3 or L3 regions results in a similarly improved safety profile as observed for the Ad-14.7K-GM or Ad-23K-GM vectors, an in vivo study was performed in which virus was injected into mice i.v.…”

“…15,18 This tumor-specific transgene expression improves the safety of the vector. Several insertion sites, such as the E3A region 11,18,22 and E3B region, 15,18 were identified on the basis of detailed knowledge of transcription in adenoviruses. More recently, a novel transposonbased mechanism has been described that identified other potential insertion sites, even though the exact transcriptional regions had not been previously identified.…”

“…9 Potential anticancer transgenes include tumor suppressor genes, for example, p53; 10 immunomodulatory genes, for example, granulocyte macrophage colony-stimulating factor (GM-CSF) 11,12 and interleukin-24; 13 pro-drug-converting enzymes, for example, thymidine kinase 14 and cytosine deaminase 15 and proapoptotic genes, for example, tumor necrosis factor-a 16 and TRAIL. 17 Several strategies for arming the vectors and for controlling transgene expression have been used, with the most recent attempts utilizing adenoviral promoters and transcriptional control elements to regulate the expression of inserted transgenes.…”

“…17 Several strategies for arming the vectors and for controlling transgene expression have been used, with the most recent attempts utilizing adenoviral promoters and transcriptional control elements to regulate the expression of inserted transgenes. 15,[18][19][20][21] These studies showed that linking transgene expression to adenoviral genes that are expressed late in infection, that is, after DNA replication, restricted transgene expression to tumor cells. 15,18 This tumor-specific transgene expression improves the safety of the vector.…”

“…15,[18][19][20][21] These studies showed that linking transgene expression to adenoviral genes that are expressed late in infection, that is, after DNA replication, restricted transgene expression to tumor cells. 15,18 This tumor-specific transgene expression improves the safety of the vector. Several insertion sites, such as the E3A region 11,18,22 and E3B region, 15,18 were identified on the basis of detailed knowledge of transcription in adenoviruses.…”