Eradication of Breast Cancer Xenografts by Hyperthermic Suicide Gene Therapy under the Control of the Heat Shock Protein Promoter

Abstract

To investigate the usefulness of heat shock protein (HSP) promoter for breast cancer gene therapy, hyperthermia and HSV thymidine kinase (tk) suicide gene combination therapy was examined with mouse mammary cancer cell line FM3A. HSP promoter activity was markedly increased after heat shock (41-45 degrees C), with maximum activation (about 400-fold) at 3 hr. An in vitro cytotoxic assay showed that HSP-tk-transduced FM3A cells became more sensitive (more than 50,000 times) to ganciclovir (GCV) with heat shock, but untreated cells showed no increased cytotoxic sensitivity to GCV compared with control FM3A cells. In addition to promoter-oriented selective cell killing, a "chemosensitization effect" as a bystander effect was demonstrated by hyperthermia and suicide gene combination therapy, using a non-heat-inducible promoter. Immunohistochemical analysis revealed that this synergistic killing effect was dependent on apoptotic cell death with upregulation of both Fas and FasL (Fas ligand) expression. We also examined the efficacy of HSP-tk gene therapy in vivo by implanting breast cancer in subcutaneous and intraperitoneal models of BALB/c nude mice targeted by the HVJ-anionic liposome method. Significant tumor regression was observed in HSP-tk-transduced tumors followed by hyperthermia therapy, but no such inhibition was noted in either the mock vector transfection or hyperthermia group compared with control tumor-bearing mice. Our results demonstrate that this combination system is synergistically effective in mediating Fas-dependent apoptosis for a specific gene therapy targeting HSP-expressing mammary carcinomas, even in advanced and heat-resistant breast cancer.

Heat-directed suicide gene therapy for breast cancer

“…This is consistent with the decrease in clonogenic survival of PC-3 prostate cancer cells following heat-targeted gene expression of CDTK, 7,35 and the beneficial effects of heat-targeted TK expression observed in a syngeneic mouse breast carcinoma model. 8 Together these results support the notion that, as an adjunct to radiotherapy for recurrent breast cancer, heat-directed CDTK suicide gene therapy may significantly improve treatment outcome.…”

“…Previous studies in our laboratory 6 and in others [7][8][9][10][11][12] demonstrated that the human hsp70b promoter exhibits low levels of basal activity at 371C, but is induced as much as 800-fold in response to heat treatment of in vitro and in vivo tumor models. To examine whether this heat-directed gene targeting strategy can complement hyperthermia in breast cancer treatment, an E. coli cytosine deaminase/ herpes simplex virus 1 thymidine kinase (CDTK) fusion gene was cloned downstream of the hsp70b promoter in an adenoviral vector to generate Ad.70b.CDTK (Fig 1a).…”

“…In this study, we have constructed a recombinant adenoviral vector (Ad.70b.CDTK) in which the CDTK fusion gene is under the control of the hsp70b promoter. Previous studies in our laboratory 6 and in others [7][8][9][10][11][12] have shown that adenoviral vectors incorporating the hsp70b promoter can provide high levels of therapeutic gene expression in tumor cells treated with mild hyperthermia. Here we examine the ability of heat-directed CDTK suicide gene therapy to complement hyperthermia in heat-resistant breast cancer cells.…”

“…We 6 and others [7][8][9][10][11][12] have been working to harness hyperthermia as a modality to target therapeutic gene expression to tumor cells. Studies to date have demonstrated that hyperthermia shows significant promise as a way to achieve treatment-targeted, spatiotemporal control of gene expression in vitro 6,[8][9][10][11][12][13] and in vivo.…”

“…Studies to date have demonstrated that hyperthermia shows significant promise as a way to achieve treatment-targeted, spatiotemporal control of gene expression in vitro 6,[8][9][10][11][12][13] and in vivo. 6,[8][9][10][11][12][13] LRBC is an excellent choice for the clinical application of heat-directed cancer gene therapy in that it is prevalent, responds to hyperthermia and is relatively uncomplicated in terms of treatment planning for heat dosimetry. It is also amenable to gene therapy because its superficial and localized nature renders it easily accessible for vector administration.…”