Theranostic Properties of a Survivin-\ud Directed Molecular Beacon in Human\ud Melanoma Cells

摘要

Survivin is an inhibitor of apoptosis overexpressed in different types of tumors and\udundetectable in most terminally differentiated normal tissues. In the current study,\udwe sought to evaluate the in vitro theranostic properties of a molecular beaconoligodeoxynucleotide\ud(MB) that targets survivin mRNA. We used laser scanning\udconfocal microscopy to study MB delivery in living cells and real-time PCR and\udwestern blot to assess selective survivin-targeting in human malignant melanoma\udcells. We further assess the pro-apoptotic effect of MB by measuring\udinternucleosomal DNA fragmentation, dissipation of mitochondrial membrane\udpotential (MMP) and changes in nuclear morphology. Transfection of MB into A375\udand 501 Mel cells generated high signal intensity from the cytoplasm, while no\udsignal was detected in the extracellular environment and in survivin-negative cells\ud(i.e., human melanocytes and monocytes). MB time dependently decreased\udsurvivin mRNA and protein expression in melanoma cells with the maximum effect\udreached at 72 h. Treatment of melanoma cells with MB induced apoptosis by\udsignificant changes in MMP, accumulation of histone-complexed DNA fragments in\udthe cytoplasm and nuclear condensation. MB also enhanced the pro-apoptotic\udeffect of standard chemotherapeutic drugs tested at clinically relevant\udconcentrations. The MB tested in the current study conjugates the ability of imaging\udwith the pharmacological silencing activity against survivin mRNA in human\udmelanoma cells and may represent an innovative approach for cancer diagnosis\udand treatment.