Abstract

Background

The goal of this study was to synthesise and characterise oligonucleotide (ON)-functionalised gold nanoparticle (AuNP)-based radiopharmaceuticals for the selective targeting of telomerase-positive tumours.

Method

A 2’O-methyl ON sequence complementary to the RNA template of telomerase (5’-CAGUUAGGGUUAG-3') was used for telomerase targeting. A scramble ON sequence (5’-GCAGUGUGAUGAU-3’) was used as control. ONs were conjugated to ¹¹¹In-DTPA for internalisation and clonogenic survival studies or the fluorophore Cy3 for confocal microscopy. AuNP were synthesised by citrate reduction and functionalised with ON-conjugates (ON-AuNP) or ON-conjugates plus the cell penetrating peptide, TAT (ON-AuNP-TAT). All cell studies were performed in the telomerase-positive melanoma cell line MDA-MB-435. The effect of both constructs on telomerase activity was tested using a telomeric repeat amplification protocol (TRAP). Subcellular distribution was analysed by internalisation assays, confocal microscopy and transmission electron microscopy (TEM). Cell survival was determined in clonogenic survival assays.

Results

AuNP constructs were successfully synthesised. ONs inhibited telomerase activity in a dose-dependent manner (logIC50 -6.93±0.072). ON-AuNP (logIC50 -6.43±0.04) and ON-AuNP-TAT (logIC50 -6.50±0.06) constructs showed similar dose-response effects. Internalisation of ¹¹¹In-DTPA-ON-AuNP-TAT was significantly higher than ¹¹¹In-DTPA-ON-AuNP (5.609% ± 0.969% vs 3.618% ± 0.302%, p=0.0005). Confocal microscopy and TEM showed that ON-AuNP were predominantly located in endosomal vesicles. However, endosomal escape of ON-AuNP-TAT, was demonstrated as AuNP were observed in the nucleus. ¹¹¹In-DTPA-ON-AuNP-TAT significantly reduced the clonogenic survival of MDA-MB-435 cells (Survival fraction 0.224±0.099 [1 μg/mL ON, 3 MBq/μg]).

Conclusion

Telomerase-targeting AuNP-based radiopharmaceuticals specifically inhibit telomerase activity and are cytotoxic in telomerase-positive cancer cells.