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. Author manuscript; available in PMC: 2008 Mar 4.
Published in final edited form as: Cancer Res. 2007 Jul 15;67(14):6872–6881. doi: 10.1158/0008-5472.CAN-06-3244

Figure 1.

Figure 1

Hypoxia-dependent E1A and mIL4 protein expression by HYPR-Ad-mIL4–infected cells. A, schematic of HYPR-Ad-mIL4. HYPR-Ad-mIL4 is a novel hypoxia/HIF-dependent oncolytic Ad that is armed with the antitumorigenic mouse interleukin-4 cytokine gene. This virus contains an exogenous bidirectional hypoxia-responsive promoter composed of six tandem copies of the HRE of the human VEGF gene (6× VEGF-HRE) bordered by two independent mini-CMV promoter elements (12). The right arm of this promoter drives the expression of the Ad E1A replication gene, whereas the left arm regulates expression of the mouse IL-4 gene. The E1B/IX gene region was also reconstituted. This modified Ad type 5 E1 gene region was cloned into a replication-deficient Ad5 genome lacking the E1 and E3 genomic regions. B, LN229 human glioblastoma cells were infected at MOI 1 with HYPR-Ad-mIL4, dl309-Ad, HYPR-Ad#1 viruses or mock infected and then maintained under normoxia (N) or hypoxia (H). Total cell lysates were examined by Western blot analysis for Ad E1A and cellular actin (loading control) at days 1 and 3 postinfection. C, LN229 cells were infected at MOI 1 with AdLacZ, dl309-Ad, HYPR-Ad#1, HYPR-Ad-mIL4, or mock infected and then maintained under normoxia (N, white) or hypoxia (Hyp, black). mIL4 in the conditioned media was measured by ELISA at days 3 and 6 postinfection. Columns, mean mIL4 concentration; bars, SD (pg/mL). *, statistically significant (Student’s t test) increase in mIL4 levels under hypoxia.