Heat-directed cancer gene therapy

The primary goal of cancer gene therapy is to transfer and express genes in tumour cells or related tissues in order to facilitate disease eradication. This is challenging because therapeutic genes are often highly toxic to normal cells and the vectors that deliver them lack specificity for tumour cells. Furthermore, current vectors are unable to transduce a large percentage of cells within a given tumour. To address this, researchers have striven to harness toxic genes capable of killing not only the cells that they are expressed in, but also surrounding, untransduced “bystander” cells. The broad toxicity of genes with large “bystander effects” necessitates precise control over gene expression. Combining gene therapy with existing conventional treatments is one way to address this problem. Clinical hyperthermia has shown promise as an adjuvant form of cancer therapy when combined with ionizing radiation in clinical trials. Adoption of hyperthermia into clinical practice however, has been hampered by difficulties in maintaining uniformly elevated temperatures in human tumours. The human genome contains promoters that respond to mild heat exposure by dramatically up-regulating the genes that they control. This presents an attractive route for spatio-temporal control of gene expression in locally heated tumours. This thesis explores the hypothesis that this “heat shock response” could be harnessed to control toxic gene expression with heat-inducible promoters. This would not only augment the therapeutic effects of mild hyperthermia (exploiting the bystander effect) but would also limit normal tissue toxicity by constraining gene expression to the heated volume. Heat-targeted gene therapy is explored using the hsp70b promoter to control expression of a marker gene (β-galactosidase); an enzyme (cytosine deaminase/thymidine kinase) that converts non-toxic prodrugs into toxic metabolites; or a highly toxic gene (the active form of the Gibbon-Ape Leukemia Virus envelop protein) which causes widespread cell:cell fusion. Cell culture and animal studies using human tumour cells demonstrate that: (1) heat-responsiveness of the hsp70b promoter can be augmented through DNA sequence manipulation; (2) hyperthermia can effectively focus gene expression to heated tumours; and (3) heat-directed toxic gene expression can significantly enhance heat-induced cytotoxicity.