Bystander effect caused by suicide gene expression indicates the feasibility of gene therapy for hepatocellular carcinoma

doi:10.1016/0270-9139(95)90213-9

Hepatology

Volume 22, Issue 6, December 1995, Pages 1838-1846

https://doi.org/10.1016/0270-9139(95)90213-9 Get rights and content

Abstract

In the field of gene therapy using retroviral vectors, it appears impossible to introduce a foreign gene into all target cells. Therefore adjacent cell killing, the so-called bystander effect, caused by genetically modified cells provides therapeutic advantages for gene therapy against cancers. We retrovirally transduced the herpes simplex virus thymidine kinase (HSV-tk) gene into murine and rat hepatocellular carcinoma (HCC) cells. These HSV-tk gene-transduced HCC cells were cocultured with the corresponding parental cells in the presence of ganciclovir, at a concentration not at all cytotoxic to the parental cells. When parental HCC cells were cocultured with their HSV-tk gene-transduced counterparts at a high density at which most cells were in contact with one another, they were markedly eliminated. Conversely, when cocultured at a low density at which none of the cells were in contact, a weak but statistically significant bystander effect was observed. Addition of lysates of HSV-tk gene-transduced cells in the presence of ganciclovir did not cause any killing of parental cells. Furthermore, media conditioned by transduced cells with ganciclovir exhibited weak cytotoxic effects on parental cells. These results indicate that cell-cell contact plays a major causative role in the bystander effect and that minor contributors to this phenomenon are some cytotoxic substance released from transduced cells. Importantly, the bystander effect was induced in vivo as well as in vitro. When mixtures of transduced and untransduced HCC cells were implanted into the flank region of mice, intraperitoneal ganciclovir administration considerably inhibited tumor development, indicating the feasibility of gene therapy with HSV-tk gene and ganciclovir against HCC.

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Citation Excerpt :
Several HSV vectors take advantage of underlying defects in molecular pathways of tumor cells. HSV-1 vectors express the tk gene, which encodes thymidine kinase, which results in apoptosis of host cells (Kuriyama et al., 1995; Vile & Hart, 1993) Modification of envelope proteins can increase infectivity of tumor cells, which bear the corresponding receptors (Grandi, Spear, Breakefield, & Wang, 2004; Grandi, Wang, et al., 2004)
Gene transfer is an increasingly utilized approach for research and clinical applications involving the central nervous system (CNS). Vectors for gene transfer can be as simple as an unmodified plasmid, but more commonly involve complex modifications to viruses to make them suitable gene delivery vehicles. This chapter will explain how tools for CNS gene transfer have been derived from naturally occurring viruses. The current capabilities of plasmid, retroviral, adeno-associated virus, adenovirus, and herpes simplex virus vectors for CNS gene delivery will be described. These include both focal and global CNS gene transfer strategies, with short- or long-term gene expression. As is described in this chapter, an important aspect of any vector is the cis-acting regulatory elements incorporated into the vector genome that control when, where, and how the transgene is expressed.
Encapsulation of ganciclovir in albumin nanoparticles enhances the thymidine kinase suicide gene therapy
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The therapeutic combination of the herpesvirus thymidine kinase (tk) and ganciclovir has shown great clinical promise as gene therapy for the treatment of hepatocarcinoma. The aim of this study was to evaluate the efficacy of the loading of ganciclovir in albumin nanoparticles to induce hepatoxicity in animals previously treated with adenoviral vectors expressing the tk gene (AdCMVtk). The use of these carriers dramatically increased the drug toxic effects, including high transaminase levels (AST and ALT levels). In addition, the histologic analysis of the liver also revealed a great cellular damage with necrotizing area and perivascular inflammatory reactions. These nanoparticulate carriers offered the additional advantage of increasing the plasmatic half-life of the drug and its tissue distribution, as it was determined by the pharmacokinetic analysis. Thereby, they constitute a very effective system for the prolonged delivery of ganciclovir in vivo, improving the disposition of ganciclovir in the liver.
An optimal therapeutic expression level is crucial for suicide gene therapy for hepatic metastatic cancer in mice
2003, Hepatology
The most serious problem in current gene therapy is discrepancies between experimental data and actual clinical outcomes, which may be due to insufficient analyses and/or inappropriate animal models. We have explored suicide gene therapy by using various clinically relevant animal models and doubt the clinical use of maximal suicide gene expression, which has been generally recommended. To explore this subject further, we studied what expression level of suicide gene and what promoter led to the maximal clinical benefit in the case of hepatic metastatic cancer in mice. Therapeutic and adverse side effects of 4 adenoviral vectors that express herpes simplex virus thymidine kinase (HSV-tk) under different promoters were scrupulously investigated in 2 mouse models of hepatic metastasis of gastric cancer that possess clinical characteristics. Surprisingly, increases in HSV-tk expression beyond a certain point, achieved by the Rous sarcoma virus long terminal repeat promoter, not only enhanced the adverse side effects of lethal hepatotoxicity and ganciclovir-independent cytotoxicity but also failed to further increase therapeutic potential. Moreover, the carcinoembryonic antigen (CEA) tumor-specific promoter, the therapeutic potential of which had been underestimated, was much more useful—even in the case of low CEA-producing cancer—than had been previously reported. In conclusion, the optimal therapeutic expression level of a suicide gene is a novel concept and a crucial factor for successful cancer gene therapy. The present results, which contradict those of previous studies, alert researchers about possible problems with ongoing and future clinical trials that lack this concept. (HEPATOLOGY 2003;37:155-163.)
Polyethylenimine-mediated suicide gene transfer induces a therapeutic effect for hepatocellular carcinoma in Vivo by using an Epstein-Barr virus-based plasmid vector
2002, Biochemical and Biophysical Research Communications
The present study aimed to establish a novel efficient nonviral strategy for suicide gene transfer in hepatocellular carcinoma (HCC) in vivo. We employed branched polyethylenimine (PEI) and combined it with Epstein–Barr virus (EBV)-based plasmid vectors. The HCC cells transfected with an EBV-based plasmid carrying the herpes simplex virus-1 thymidine kinase (HSV-1 Tk) gene (pSES.Tk) showed up to 30-fold higher susceptibilities to ganciclovir (GCV) than those transfected with a conventional plasmid vector carrying the HSV-1 Tk gene (pS.Tk). The therapeutic effect in vivo was tested by intratumoral injection of the plasmids into HuH-7 hepatomas transplanted into C.B-17 scid/scid mutant (SCID) mice and subsequent GCV administrations. Treatment with pSES.Tk, but not pS.Tk, markedly suppressed growth of hepatomas in vivo, resulting in a significantly prolonged survival period of the mice. These findings suggest that PEI-mediated gene transfer system can confer efficient expression of the suicide gene in HCC cells in vivo by using EBV-based plasmid vectors.
Hydroxyapatite ceramics as a carrier of gene-transduced bone marrow cells
2002, Journal of Orthopaedic Science
The aim of this study was to develop an efficient exogenous gene delivery system using cultured marrow cells and porous hydroxyapatite ceramics. Bone marrow cells were obtained from the femoral shaft of a Fischer 344 rat and cultured in a medium containing 15% fetal bovine serum until confluent. After trypsinization, cells were subcultured at a cell density of 1 ×104 cells/cm2 in the presence of fetal bovine serum. The subcultured bone marrow cells were infected with recombinant retroviruses carrying the lacZ gene. The retrovirus infection was performed seven times from day 1 to day 7 during the culturing procedure. Cells expressing the lacZ gene were stained blue with the X-gal staining and represented approximately 80%. Composites of virus-infected bone marrow cells and hydroxyapatite ceramics were implanted at the subcutaneous site of recipient Fischer 344 rats. Four weeks after the implantation the ceramics were harvested. The histological sections of the ceramics showed abundant bone formation in the pores of the ceramics and obviously blue-stained osteoblasts and osteocytes. Other cell types that were stained blue were some fibroblastic cells and endothelial cells in the newly formed capillaries. These findings indicate that osteoblasts and osteocytes in the newly formed bone were derived from the cultured bone marrow cells, and therefore gene transfection by retroviruses did not disturb the bone formation process. Because of the durability of the newly formed bone tissue, creating composites of cultured bone marrow cells and hydroxyapatite ceramics might be an ideal method for exogenous gene transfection.
Suicide gene therapy of human hepatoma and its peritonitis carcinomatosis by a vector of replicative-deficient herpes simplex virus
2002, Biochemical and Biophysical Research Communications
Herpes simplex virus type 1 (HSV-1) deleted for the immediate-early gene was applied for treatment of hepatoma cells of SKHep 1 and Huh-7. Hepatoma cells were cultured in medium containing HSV1 expressing GFP gene (QOZ/HG) to determine its transfection rate, and both cell lines infected by MOI 1 of QOZ/HG were found to have high expression of GFP without cytotoxicity. Subcutaneous growth of SKHep 1 cell tumor in nude mice was significantly reduced by injection of replicative-deficient herpes virus (TOZ.1) containing Tk-gene with administration of GCV, in comparison with that of noninjected tumor. SCID mice of peritonitis carcinomatosis due to Huh-7 hepatoma cells infected with TOZ.1 could survive longer under administration of GCV than those without TOZ.1. Therefore replicative-deficient HSV1 is a useful vector for treatment of human hepatoma cells, and TOZ.1 with GCV may be applied to suicide gene therapy for hepatoma and peritonitis carcinomatosis of hepatoma cells.

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^☆: This work was supported in part by a Grant-in-Aid for Scientific Research (07457141) from the Ministry of Education, Science and Culture Japan.

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Bystander effect caused by suicide gene expression indicates the feasibility of gene therapy for hepatocellular carcinoma☆

Abstract

Cancer Lett

J Biol Chem

Int Hepatol Commun

Cell

Cell

TIBTECH

Electron Microsc Rev

Semin Cell Biol

Retroviruses

Science

Gene transfer by retrovirus vectors occurs only in cells that are actively replicating at the time of infection

Mol Cell Biol

Selective killing of glioma cells in culture and in vivo by retrovirus transfer of the herpes simplex virus thymidine kinase gene

New Biol

Selective elimination of recombinant genes in vivo with a suicide retroviral vector

New Biol

Selective killing of CD4+ cells harboring a human immunodeficiency virus-inducible suicide gene prevents viral spread in an infected cell population

Retroviral-mediated gene therapy for the treatment of hepatocellular carcinoma: an innovative approach for cancer therapy

Transfer of the bacterial gene for cytosine deaminase to mammalian cells confers lethal sensitivity to 5-fluorocytosine: a negative selection system

A first step in the development of gene therapy for colorectal carcinoma: cloning, sequencing and expression of Escherichia coli cytosine deaminase

Mol Pharmacol

A new nucleoside analog, 9-[[2- hydroxy-1-(hydroxymethyl)ethoxy] methyl] guanine, highly active in vitro against herpes simplex virus type 1 and 2

Antimicrob Agents Chemother

Anti-herpesvirus activity of the acyclic nucleoside 9-(1,3-dihydroxy-2-propoxymethyl) guanine

Antimicrob Agents Chemother

Unique spectrum of activity of 9-[(1,3-dihydroxy-2-propoxy) methyl]-guanine against herpesviruses in vitro and its mode of action against herpes simplex virus type 1

9-{[2 hydroxy-1-(hydroxymethyl) ethoxy] methyl} guanine: a selective inhibitor of herpes group virus replication

Lymphoma regression induced by ganciclovir in mice bearing a herpes thymidine kinase transgene

Hum Gene Ther

Herpes simplex-1 virus thymidine kinase gene is unable to completely eliminate live, nonimmunogenic tumor cell vaccines

J Immunother

Gene transfer of herpes simplex virus type 1 thymidine kinase gene as a drug sensitivity gene into human lung cancer cell lines using retroviral vectors

Am J Respir Cell Mol Biol

An experimental model of retrovirus gene therapy for malignant brain tumors

J Neurosurg

Use of tissue-specific expression of the herpes simplex virus thymidine kinase gene inhibits growth of established murine melanomas following direct intratumoral injection of DNA

Cancer Res

Thymidine kinase-mediated killing of rat brain tumors

J Neurosurg

In situ retroviral-mediated gene transfer for the treatment of brain tumors

Cancer Res

Toxicity studies of retroviral-mediated gene transfer for the treatment of brain tumors

J Neurosurg

Selective expression of foreign genes in glioma cells: use of the mouse myelin basic protein gene promoter to direct toxic gene expression

J Neurosci Res

Retrovirus-mediated gene therapy toward hepatoma using a liver-specific promoter

Hepatology

Tumor chemosensitivity conferred by inserted herpes thymidine kinase genes: paradigm for a prospective cancer control strategy

Cancer Res

Selective killing of CD4⁺ cells harboring a human immunodeficiency virus-inducible suicide gene prevents viral spread in an infected cell population