Employing tumor hypoxia to enhance oncolytic viral therapy in breast cancer.
BACKGROUND
Hypoxia is a common tumor condition correlated with therapeutic resistance. Ribonucleotide reductase (RR) is a rate-limiting enzyme for viral replication. We hypothesize that hypoxia-driven transcription of UL39, the gene encoding the large subunit of RR, would enhance herpes oncolytic viral therapy in breast cancer.


METHODS
Hypoxia-inducible factor 1alpha (HIF-1alpha) ELISA was performed on MCF7 human breast cancer cells in hypoxia (1% O2) or normoxia (21% O2). A multimerized hypoxia-responsive enhancer was constructed (10xHRE) and functionally tested in a luciferase assay. 10xHRE was cloned upstream of the UL39 gene (10xHRE-UL39). MCF7 cells were transfected with 10xHRE-UL39, incubated in hypoxia or normoxia, and infected with G207. Cytotoxicity assays and viral titers were performed.


RESULTS
HIF-1alpha levels increased 7-fold in hypoxic MCF7 cells (P < .001). 10xHRE increased luciferase gene expression 61-fold in hypoxia versus controls (P < .01). G207 cytotoxicity of 10xHRE-UL39-transfected, hypoxic MCF7 cells increased 74% versus mock-transfected, hypoxic MCF7 cells (P < .001). In normoxia, 10xHRE-UL39 transfection did not significantly improve G207 cytotoxicity. 10xHRE-UL39 transfection improved peak viral titers 69-fold in hypoxia (P < .005), with no significant difference in normoxia.


CONCLUSIONS
Hypoxia-driven RR production significantly enhances G207 cytotoxicity in hypoxic breast cancer cells, which would otherwise be resistant to herpes viral therapy alone.
Department of Surgery, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Richard H
RH
Pin
Maura
M
Reinblatt
Yuman
Y
Fong
D001943:Breast Neoplasms / Q000378:metabolism / Q000628:therapy*; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D005260:Female; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008156:Luciferases / Q000378:metabolism; D018139:Simplexvirus / Q000235:genetics*; D014157:Transcription Factors / Q000032:analysis; D014779:Virus Replication
C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D014157:Transcription Factors; D008156:Luciferases