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Introduction and Objectives: IGFBP2 and IGFBP5 are members of IGF-I axis that are known to be critical in the regulation of neoplastic tumor progression and differentiation. IGFBP2 is a major binding protein in the prostate cancer that is up regulated during progression. IGFBP5 is highly expressed in bone and may play a role in the site-specific metastasis seen with prostate cancer. Since both binding proteins are involved in prostate cancer development and progression, they represent potential targets for therapeutic inhibition. Methods: A prostate cancer tissue microarray spotted with 382 untreated and post hormonetherapy treated cancers was used to evaluate changes in IGFBP-2 and -5 after androgen ablation and in osseous metastases. Sequence similarity between the genes coding for IGFBP2 and IGFBP5 permits the design of bi-specific antisense oligonucleotide (bs-ASO) to target both IGFBP2 and IGBP5 mRNA. Dose-dependent sequence-specific effects of bs-ASO on mRNA and protein levels of IGFBP2 in LNCaP and C42 prostate cancer cell lines and IGFBP-5 in the MSF osteosarcoma cell line were evaluated using Northern and Western Blotting, while flow cytometry and MTT assay were performed to evaluate effects of bs-ASO treatment on cell cycle, cell growth, and apoptosis. Results: IGFBP2 increased after androgen ablation and during progression to the androgen independent (AI) state in the tissue microarray and LNCaP xenograft model. High level of IGFBP-5 was found in prostate cancer osseous metastasis. Northern blot showed dose-dependent sequence-specific down-regulation (up to 90%) of mRNA in cells expressing BP2 and BP5 respectively after bs-ASO treatment, which was confirmed on the protein level with Western Blot. bs-ASO treatment also resulted in dose-dependent sequence-specific cell growth inhibition (from 50% to 90% depending on cell type), a 2-fold increase in subG0 apoptotic fraction, and a 3-fold G2/M arrest in LNCaP and C4-2 cells. In order to identify the way by which bs-ASO may affect cell biological behavior, LNCaP and C4-2 cells were pretreated with the PI3K inhibitor LY294002. IGF-I overcomes LY toxicity, which is measured by AKT phosphorylation. bs-ASO completely blocked the ability of IGF-I to overcome LY toxicity compare to scrambled oligonucleotide control. Conclusion: Suppression of IGFBP-2 and -5 expression decreases the IGF-I responsiveness of targeted cells and so sensitizes cells to cytotoxic stress. IGFBP2 and IGFBP5 bs-ASO is a potential therapeutic in prostate cancer patients with combined local and antimetastatic activity.

[Proc Amer Assoc Cancer Res, Volume 45, 2004]

American Association for Cancer Research
2004