Transitions between distinct compaction regimes in complexes of multivalent cationic lipids and DNA

¹ Ben Gurion University
² University of California, Santa Barbara
³ University of California, Davis

^* To whom correspondence should be addressed. E-mail: safinya{at}mrl.ucsb.edu.

Submitted on October 31, 2007
Revised on December 13, 2007
Accepted on 29 February 2008

	Abstract

Cationic lipids (CLs) have found widespread use as nonviral gene carriers (vectors), including applications in clinical trials of gene therapy. However, their observed transfection efficiencies (TEs) are inferior to those of viral vectors, providing a strong incentive for a detailed understanding of CL-DNA complex behavior. In recent systematic studies employing monovalent as well as newly synthesized multivalent lipids (MVLs), the membrane cationic charge density has been identified as a key parameter governing the TE of lamellar CL-DNA complexes. In this work, we use X-ray scattering and molecular simulations to investigate the structural properties of complexes containing MVLs. At low mole fraction of neutral lipids (NLs), _NL, the complexes show dramatic DNA compaction, down to essentially close packed DNA arrays with a DNA interaxial spacing d_DNA = 25 Å. A gradual increase in _NL does not lead to a continuous increase in d_DNA as observed for DNA complexes of monovalent CLs. Instead, distinct spacing regimes exist, with sharp transitions between them. Three packing states have been identified: (i) close packed, (ii) condensed, but not close packed, with d_DNA = 27-28 Å, and (iii) an expanded state, where d_DNA increases gradually with _NL. Based on our experimental and computational results, we conclude that the DNA condensation is mediated by the multivalent cationic lipids, which assemble between the negatively charged DNA rods. Quite remarkably, the computational results show that the less tightly packed structure in regime (ii) is thermodynamically more stable than the close packed structure in regime (i). Accordingly, the constant DNA spacing observed in regime (ii) is attributed to lateral phase coexistence between this stable CL-DNA complex and neutral membranes. This finding may explain the reduced TE measured for such complexes: Transfection involves endosomal escape and disassembly of the complex, and these processes are inhibited by the high thermodynamic stability. Our results, which demonstrate the existence of an inverse correlation between the stability and transfection activity of lamellar CL-DNA complexes are, therefore, consistent with a recently proposed model of cellular entry.

Key Words: X-ray scattering, coarse-grained simulations, electrostatic interactions, gene therapy, lipid-DNA complexes, structure-function relationship