Transcriptional targeting in ovarian cancer cells using the human epididymis protein 4 promoter.
OBJECTIVE
Limitations of current ovarian cancer gene therapies include lack of specificity and transduction of normal tissues. One strategy toward overcoming these limitations is to direct gene therapy specifically to ovarian cancer cells by using tissue- and tumor-specific promoters. The whey-acidic protein human epididymis protein 4 (HE4) is frequently overexpressed in ovarian cancer, suggesting that the HE4 promoter is highly transcriptionally active in the disease. The objective of this study was to isolate the HE4 promoter and examine its ability to selectively activate reporter gene expression in an ovarian cancer-specific manner.


METHODS
To investigate transcriptional targeting in ovarian cancer gene therapy, we isolated a region of the HE4 promoter from -530 to +122 (pHE4-652; relative to the ATG start site of HE4) and placed it upstream of a luciferase reporter gene plasmid to generate pHE4-652-luc. The activity of the pHE4-652-luc reporter construct was characterized in transient transfection assays in a panel of epithelial ovarian cancer cell lines (SKOV-3, SKOV-3x, CP70, HeyC2, A2780, A2780CP, OVCAR-3), non-ovarian tumor cell lines, and primary cultures of normal cells. The activity of two other candidate gene therapy promoters, human telomerase reverse transcriptase (hTERT) and OSP1, was also characterized in these cell lines.


RESULTS
The HE4 promoter was active in 5/7 ovarian cancer cell lines with the range of activity spanning 0.06- to 3-fold that observed for a positive control, cotransfected reporter construct (SV-40-luc). Minimal pHE4-652 promoter activity, defined as < or =5% of the activity detected with the SV-40-luc construct, was observed in the non-ovarian tumor cell lines and normal cells. The hTERT and the OSP1 promoters were active in the ovarian cancer lines. hTERT activity was highest in the CP70 cell line, and OSP1 activity was highest in the SKOV-3x cell line. Modest OSP1 and hTERT promoter activity was observed in normal cell lines and in selected non-ovarian cancer cell lines.


CONCLUSIONS
This is the first report using the pHE4-652 promoter to drive specific reporter gene expression in epithelial ovarian cancer cell lines, and we are continuing to develop this promoter for use in transcriptional targeting in ovarian cancer gene therapy.
Medical Sciences, Indiana University School of Medicine, Bloomington, IN 47405, USA.
Nicholas B
NB
Berry
Yong Mee
YM
Cho
Maureen A
MA
Harrington
Stephen D
SD
Williams
John
J
Foley
Kenneth P
KP
Nephew
D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins; D030501:Epididymal Secretory Proteins / Q000235:genetics*; D005260:Female; D017930:Genes, Reporter / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D010051:Ovarian Neoplasms / Q000235:genetics* / Q000628:therapy*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D019098:Telomerase / Q000235:genetics; D023083:beta-Defensins
C426184:DEFB126 protein, human; D004268:DNA-Binding Proteins; D030501:Epididymal Secretory Proteins; D023083:beta-Defensins; D008156:Luciferases; D019098:Telomerase