Daralnakhla Haneen, Saher Osama, Zamolo Susanna, Bazaz Safa, P Bost Jeremy, Heitz Marc, Lundin Karin E, El Andaloussi Samir, Darbre Tamis, Reymond Jean-Louis, Zain Rula, Smith C I Edvard
Clinical Research Center, Department of Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, 141 86 Huddinge, Sweden.
Department of Pharmaceutics and Industrial Pharmacy, Faculty of Pharmacy, Cairo University, Cairo 11562, Egypt.
Pharmaceutics. 2021 Jan 18;13(1):116. doi: 10.3390/pharmaceutics13010116.
Non-viral transfection reagents are continuously being developed in attempt to replace viral vectors. Among those non-viral vectors, dendrimers have gained increasing interest due to their unique molecular structure and multivalency. However, more improvements are still needed to achieve higher efficacy and lower toxicity. In this study, we have examined 18 peptide dendrimers conjugated to lipophilic moieties, such as fatty acids or hydrophobic amino acids, that were previously explored for siRNA. Reporter cells were employed to investigate the transfection of single strand splice-switching oligonucleotides (ONs) using these peptide dendrimers. Luciferase level changes reflecting efficiency varied with amino acid composition, stereochemistry, and complexation media used. 3rd generation peptide dendrimers with D-amino acid configuration were superior to L-form. Lead formulations with 3rd generation, D-amino acid peptide dendrimers increased the correction level of the delivered ON up to 93-fold over untreated HeLa Luc/705 cells with minimal toxicity. To stabilize the formed complexes, Polyvinyl alcohol 18 (PVA18) polymer was added. Although PVA18 addition increased activity, toxicity when using our best candidates G 2,3KL-(Leu) (D) and G 2,3KL-diPalmitamide (D) was observed. Our findings demonstrate the potential of lipid-conjugated, D-amino acid-containing peptide dendrimers to be utilized as an effective and safe delivery vector for splice-switching ONs.
为了取代病毒载体，非病毒转染试剂正在不断研发。在这些非病毒载体中，树枝状大分子因其独特的分子结构和多价性而越来越受到关注。然而，仍需要更多改进以实现更高的疗效和更低的毒性。在本研究中，我们检测了18种与亲脂性基团（如脂肪酸或疏水氨基酸）偶联的肽树枝状大分子，这些亲脂性基团先前已用于小干扰RNA（siRNA）的研究。使用报告细胞来研究这些肽树枝状大分子对单链剪接转换寡核苷酸（ONs）的转染情况。反映转染效率的荧光素酶水平变化因氨基酸组成、立体化学和所用的复合介质而异。具有D-氨基酸构型的第三代肽树枝状大分子优于L-型。第三代D-氨基酸肽树枝状大分子的先导制剂可使递送的ON的校正水平比未处理的HeLa Luc/705细胞提高93倍，且毒性最小。为了稳定形成的复合物，添加了聚乙烯醇18（PVA18）聚合物。虽然添加PVA18提高了活性，但在使用我们的最佳候选物G 2,3KL-(Leu) (D)和G 2,3KL-二棕榈酰胺（D）时观察到了毒性。我们的研究结果表明，脂质偶联的含D-氨基酸的肽树枝状大分子有潜力作为一种有效且安全的剪接转换ONs递送载体。
