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Polyamines and nucleic acid interaction : a structure property relationship study

summary We have synthesized polycationic compounds comprising of one, two and four spermines, having linear, cyclic, star-shaped, and quatrefoil architecture to investigate the effect of molecular weight and architecture on its interaction with nucleic acids. Our results demonstrate that increase in molecular weight increases the binding of linear as well as cyclic oligospermines. Further, molecular architecture of oligospermines affects their binding to nucleic acids with star-shaped tetraspermine exhibiting highest binding affinity compared to its linear and quatrefoil counterparts. Molecular architecture also affects cytotoxicity of oligospemines in a panel of cancer cell lines. Tetraspermines are more suited as carriers for oligonucleotides or siRNA than for plasmid DNA. Introduction Nucleic acid therapeutics such as plasmid DNA, small interfering RNA’s (siRNA) and antisense oligonucleotides are under investigation as therapeutic modalities. However, delivering nucleic acid is challenging due to their lower cellular uptake, susceptibility to nucleases and lower half-life. Delivery vehicles such as liposomes, polycationic polymers, and cationic lipids are often used to overcome these barriers. Although many cationic polymers are under investigation, there is a limited understanding of effect of architecture of polycations on interaction with nucleic acids. The long-term goal of this project is to find the best molecular architecture for polycationic compounds that exhibits lower intrinsic toxicity, higher transfection and optimum in-vivo characteristics. Here we selected spermine as a building block to synthesize oligospermines with different architecture. Spermine is an important component of a variety of delivery vehicles because of its high affinity towards nucleic acids that enables complete complexation and formation of stable complexes within nanometer size range. We report the synthesis of molecules having one, two, or four spermines arranged in a linear, starshaped, cyclic, and quatrefoil architecture and evaluate their intrinsic toxicity and affinity towards siRNA and plasmid DNA. Experimental methods Synthesis of oligospermines: Seven oligospermines were synthesized as depicted in Fig 1 using standard protection and deprotection chemistry. Figure1. Schematic representation for the synthesis of oligospermines. Cytotoxicity: Intrinsic toxicity of oligospermines after 48 h and 5 day treatment was evaluated in prostate cancer cell line DU 145, breast cancer cell line Hs578T and epithelial cell line HEK 293 using WST reagent. Binding of oligospermines with nucleic acids: Interaction of spermine-analogs with siRNA and plasmid DNA was investigated using SYBR gold assay. Results and discussion Table 1 describes the characteristics of the seven compounds synthesized. Fig 2 shows cell viability after incubation with oligospermines for 48h. Cytotoxicity of oligospermines was dependent on the cell lines used. Spermine was the most cytotoxic compound in all the cell lines studied, whereas no toxicity was observed for cyclic spermine as well as for amino-cyclic spermine. In general decease in cytotoxicity was observed with the increase in the number spermines conjugated together. Among tetraspermines dendritic architecture was less toxic than its linear counterpart. + NH2 N N NH2 Boc Boc