Steinhauser Isabel M, Langer Klaus, Strebhardt Klaus M, Spänkuch Birgit
Institute of Pharmaceutical Technology, Biocenter of Goethe-University, Frankfurt, Germany.
Biomaterials. 2008 Oct;29(29):4022-8. doi: 10.1016/j.biomaterials.2008.07.001. Epub 2008 Jul 23.
Nanoparticles represent a promising tool for targeted drug delivery to tumour cells and are able to protect drugs against degradation. In our present study we developed targeted nanoparticles loaded with antisense oligonucleotides (ASOs) against Plk1 (polo-like kinase 1) prepared by heat denaturation instead of using glutaraldehyde. Glutaraldehyde can lead to an inactivation of ASOs through chemical crosslinking and is a toxic entity. We examined the ideal preparation conditions and characterised the resulting particles in terms of physico-chemical properties, ASO recovery after enzymatic degradation and stability. Stable monodisperse nanoparticles with an ASO recovery of more than 80% could be prepared at a temperature of 105 degrees C for 10 min. Furthermore we performed quantitative real-time PCR and Western blot to detect an ASO-mediated effect on Plk1 in BT-474 cells. We observed a significant reduction of Plk1 mRNA and protein expression. Thus, this is the first report of ASO-loaded HSA nanoparticles prepared by heat denaturation, where an impact on gene expression could be observed. The data provide the basis for the further development of carrier systems for ASOs to reduce off-target effects evoked by systemically administered ASOs and to achieve a better penetration into primary and metastatic target cells.
纳米颗粒是一种将药物靶向递送至肿瘤细胞的有前景的工具，并且能够保护药物不被降解。在我们目前的研究中，我们开发了负载有针对Plk1（polo样激酶1）的反义寡核苷酸（ASO）的靶向纳米颗粒，其通过热变性而非使用戊二醛来制备。戊二醛可通过化学交联导致ASO失活，并且是一种有毒物质。我们研究了理想的制备条件，并从物理化学性质、酶促降解后的ASO回收率和稳定性方面对所得颗粒进行了表征。在105℃的温度下加热10分钟可制备出稳定的单分散纳米颗粒，其ASO回收率超过80%。此外，我们进行了定量实时PCR和蛋白质免疫印迹，以检测ASO对BT-474细胞中Plk1的影响。我们观察到Plk1 mRNA和蛋白质表达显著降低。因此，这是关于通过热变性制备的负载ASO的人血清白蛋白纳米颗粒的首次报道，在此观察到了对基因表达的影响。这些数据为进一步开发用于ASO的载体系统提供了基础，以减少全身给药的ASO引起的脱靶效应，并实现更好地渗透到原发性和转移性靶细胞中。
