title: The potential of the human osteopontin promoter and single-nucleotide polymorphisms for targeted cancer gene therapy.
abstract: Regulatory elements of the osteopontin (opn) gene are attractive candidates for expressiontargeted gene therapy because numerous malignant cancers are marked by opn overexpression. The maximum opn promoter ((P)opn)-driven reporter intensity obtained for tested cancer cell lines was as strong (102.69%) as positive-control transfections. At the same time, (P)opn-driven reporter expression was reduced by ~90% in non-cancer cell lineages. Deletion analysis of the -922 bp region opn promoter did not confirm published reports of a repressor area within 922 bases upstream of the transcriptional start site. Further enhancements to targeting and expression were obtained through incorporation of single-nucleotide polymorphisms (SNPs) into the promoter sequence. It was found that the SNPs -443C, -155GG, -66T led to increased (P)opn-driven transfection in cancer cells (fold increase of 1.23 ~ 3.48), with a concomitant decrease in reporter expression in normal controls (fold change of 0.69). Further investigations to confirm a correlation between endogenous opn mRNA levels and (P)opn-driven reporter expression produced a surprising lack of correlation (R(2)=0.24). However, taking into account opn mRNA splicing variants showed a strong negative correlation between mRNA levels of the variant opn-a and (P) opn-driven transgene activity (R(2)=0.95). These data have implications on how future searches for expression-targeting promoters should be conducted.
authors.forename: X G
authors.initials: XG
authors.lastname: Chen
authors.affiliation: Laboratory for Gene Therapy and Cellular Engineering, Department of Chemical and Biomolecular Engineering, Tulane University, New Orleans, LA 70118, USA. godbey@tulane.edu.
authors.forename: W T
authors.initials: WT
authors.lastname: Godbey
mesh_terms: D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D045744:Cell Line, Tumor; D018390:Gene Targeting; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000628:therapy*; D053495:Osteopontin / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D012333:RNA, Messenger; D053495:Osteopontin