A quantitative reverse transcription and polymerase chain reaction assay for human IGF-II allows direct comparison of IGF-II mRNA levels in cancerous breast, bladder, and prostate tissues.

Abstract

Previously, we showed by in situ hybridization that insulin-like growth factor (IGF)-II is upregulated in approximately 50% of prostate, breast, and bladder tumours. In this study, a quantitative competitive reverse transcription and polymerase chain reaction (QC RT-PCR) assay was established and used to quantify human IGF-II mRNA levels in cells and tissues. In this QC RT-PCR assay, a competitor IGF-II RNA, prepared from a newly constructed plasmid encoding the human IGF-II sequence with a 110-bp fragment inserted, was added to RNA samples prior to RT-PCR. The human IGF-II specific QC RT-PCR assay has allowed us to readily compare the levels of IGF-II mRNA in human tissues and cultured cells. Consistent with our previous observations by in situ hybridization, IGF-II mRNA was up-regulated in 50% of cancerous breast tissues examined as compared to the matching benign tissues, and IGF-II mRNA levels were higher in bladder tumours than breast and prostate tumours. In summary, we present here quantitative data confirming that a subclass of breast cancer samples has elevated levels of IGF-II transcripts by the new competitive RT-PCR assay.