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Synthesis and structure-activity relationships of a series of increasingly hydrophobic cationic steroid lipofection reagents

Authors

  • Jeffrey A. Gruneich,

    1. Department of Bioengineering, Institute for Medicine and Engineering, 1024 Vagelos Research Laboratory, University of Pennsylvania, Philadelphia, PA 19104, USA
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  • Scott L. Diamond

    Corresponding author
    1. Department of Bioengineering, Institute for Medicine and Engineering, 1024 Vagelos Research Laboratory, University of Pennsylvania, Philadelphia, PA 19104, USA
    2. Department of Chemical and Biomolecular Engineering, Institute for Medicine and Engineering, 1024 Vagelos Research Laboratory, University of Pennsylvania, Philadelphia, PA 19104, USA
    • Department of Bioengineering, Institute for Medicine and Engineering, 1024 Vagelos Research Laboratory, University of Pennsylvania, Philadelphia, PA 19104, USA.
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Abstract

The use of cholesterol-based cationic lipids and the ability of glucocorticoids to reduce local inflammatory response to lipoplexes motivated an investigation of structure-activity relationships for cationic steroids. A one-step synthetic scheme using iminothiolane was developed to link spermine to the 21-OH position of steroids via an amidine linkage. Five steroids (cortisol, dexamethasone, corticosterone, 11-deoxycortisol, and 11-deoxycorticosterone) with increasing hydrophobicity of the parent steroid (Log Pster from 1.51 to 3.01) were conjugated with spermine, formulated with dioleoylphosphatidylethanolamine (DOPE) at DOPE : steroid mole ratios (R) of R = 0.5 to 2, and then complexed with 1 µg enhanced green fluorescent protein (EGFP) plasmid DNA at charge ratios (CR) = 2 to 24 amines per phosphate (0.5 to 6 steroids per phosphate). The resulting 105 different formulations of the cationic steroid series were used to lipofect bovine aortic endothelial cells. Transgene expression data at either 24 or 48 h post-lipofection for all formulations was collapsed onto master curves when plotted against a single empirical dimensionless parameter, the lipofection index (LI) = CR (Log Pliposome)(Log Pster/|ΔLog P|) [R/(R + 1)] where ΔLog P = Log PDOPE− Log Pster and Log Pliposome is a mole-weighted average of the DOPE/cationic steroid liposome hydrophobicity. For 7 < LI < 29, the EGFP expression at 24 or 48 h post-lipofection increased linearly with LI (EGFP ∼0 for LI < 7), but did not increase further for LI > 29, thus providing a predictive design rule based on Log P of the hydrophobic moiety of the cationic steroid lipid. Copyright © 2007 John Wiley & Sons, Ltd.

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