Induction of the human heat shock promoter HSP70B by nutritional stress: implications for cancer gene therapy.
BACKGROUND
We designed and tested, in vitro, an adenoviral construct containing the feline interleukin-12 (IL-12) gene under control of the heat-inducible promoter HSP70B. This construct, AdhspfIL12, was used in a phase I trial in feline soft tissue sarcomas. During the course of our experiments, we noted that IL-12 was being produced in the transfected Crandell Feline Kidney (CrFK) cells under certain conditions even in the absence of hyperthermia. This observation was further explored to identify the cause of this unintended HSP70B induction.


METHODS
We used real-time PCR as a sensitive method to quantitatively detect the presence of even small amounts of IL-12 mRNA. This served as a surrogate indicator of HSP70B induction. Various conditions were tested to induce the heat shock promoter, including nutritional deprivation, radiation and changes in pH.


RESULTS
Nutritional stresses, specifically the absence of glucose and glutamine, could induce the heat shock promoter, thus, resulting in production of the downstream gene product. Other factors known to trigger the heat shock response, pH change, and reactive oxygen species production were also studied but were not found to contribute to heat shock promoter induction in our setting.


CONCLUSIONS
The human heat shock promoter (HSP70B) is reported to be an efficient and tightly regulated promoter. We discovered, using sensitive real-time PCR techniques, that it can also be induced in response to cellular nutrient stresses. The pros and cons of this phenomenon and its implications for cancer gene therapy are discussed.
Siddiqui
Farzan
F
Department of Environmental, Colorado State University, Fort Collins, Colorado, USA. fsiddiq1@hfhs.org
Avery
Paul R
PR
Li
Chuan-Yuan
CY
Zhang
Xiuwu
X
LaRue
Susan M
SM
Dewhirst
Mark W
MW
Ullrich
Robert L
RL
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D050256:Antimitotic Agents / Q000494:pharmacology; D016590:Aphidicolin / Q000494:pharmacology; D002415:Cats; D002460:Cell Line; D049109:Cell Proliferation; D003470:Culture Media / Q000378:metabolism; D015316:Genetic Therapy / Q000009:adverse effects / Q000379:methods*; D005822:Genetic Vectors; D005947:Glucose / Q000172:deficiency; D005973:Glutamine / Q000172:deficiency; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006979:Hyperthermia, Induced; D018664:Interleukin-12 / Q000096:biosynthesis* / Q000235:genetics; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis; D017382:Reactive Oxygen Species / Q000378:metabolism; D013312:Stress, Physiological / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D015533:Transcriptional Activation / Q000187:drug effects; D014162:Transfection
D050256:Antimitotic Agents; D003470:Culture Media; D018840:HSP70 Heat-Shock Proteins; C491129:HSPA7 protein, human; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D005973:Glutamine; D018664:Interleukin-12; D016590:Aphidicolin; D005947:Glucose