Heat-activated transgene expression from adenovirus vectors infected into human prostate cancer cells

Replication-deficient adenovirus expression vectors were used to introduce a recombinant DNA construct containing enhanced green fluorescent protein ( EGFP) under control of a truncated, human heat shock promoter into human pr ostate cancer cells growing either exponentially or in plateau phase. This was done to measure controlled, heat shock-induced EGFP expression under co nditions relevant to treating human cancers with heat-activated gene therap y, Both the temporal duration and magnitude of EGFP expression increased pr oportionately with stronger heat shocks (time at temperature) up to maximum values that were induced by 4 h at 41.0 degreesC or 2 b at 42.0 degreesC, Longer heat shocks at either temperature yielded no additional EGFP express ion and ultimately reduced it. Maximal EGFP expression was induced in expon ential cultures by heat shocks delivered 12-24 h after virus infection. Ind uction at progressively later postinfection times induced increasingly lowe r, peak EGFP expression. Maximal EGFP expression could not be induced until 48 h after infection of plateau phase cultures but could still be induced 180 h after virus infection. However, peak EGFP levels in plateau cultures were approximately 25-50% of those observed in identically induced exponent ial cultures, Ostensibly, the differences in expression from the heat shock promoter observed in exponential and plateau cultures were attributable to cell division diluting the vector within exponential cultures and the lowe r metabolic activity in serum-starved plateau cultures. For all experimenta l conditions, EGFP expression induced from the heat shock promoter was comp arable with or higher than that from the constitutively active cytomegalovi rus promoter over any 24-h period. The experimental results demonstrated that EGFP expression from the heat sh ock promoter was controllable in both exponential and plateau phase culture s and support the plausibility of using controlled heat shock activation of this promoter as a means of regulating both the spatial and temporal expre ssion of therapeutic DNA constructs within human tissues. The ability to lo calize and regulate expression from the heat shock promoter may prove parti cularly advantageous for many cancer applications, especially if the therap eutic products are highly toxic, e.g., proteotoxins or cytokines, However, the results of this study suggest that differential growth conditions withi n tumors could markedly affect the expression of recombinant DNA under cont rol of both inducible and constitutive promoters. Consequently, inducing sc hemes may need to be spatially adjusted to obtain the desired therapeutic r esults in all tumor domains using heat-activated gene therapy.