Pathways of Formation of Polymer Coated Lipid-DNA Nanoparticles: Competing Electrostatic Attractions and Polymer Repulsions Cyrus R. Safinya, University.

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Pathways of Formation of Polymer Coated Lipid-DNA Nanoparticles: Competing Electrostatic Attractions and Polymer Repulsions Cyrus R. Safinya, University of California-Santa Barbara, DMR Figure 1 Formation of polymer coated lipid-DNA nanoparticles (LD-NPs) along different pathways. Left Panel: LD-NPs formed in brine have to overcome a strong repulsive barrier due to the steric repulsion of the polymer coat while their attractive electrostatic interactions (between DNA and cationic membrane) are reduced due to the ionic screening. The resulting LD- NPs contain only a fewer layers and give rise to broad small-angle-x-ray- scattering profiles (Middle panel, red curve). Right Panel: LD-NPs formed in de-ionized water, where electrostatic attractions are strongest, result in onion-like LD-NPs with many layers. In de-ionized water the attractive interactions (between DNA and membrane) overwhelm the repulsive steric barrier due to the polymer coat. PUBLICATION: Silva, B. F. B.; Majzoub, R. N.; Chan, C.-L.; Li, Y.; Olsson, U.; Safinya C. R.: PEGylated Cationic Liposome–DNA Complexation in Brine is Pathway-Dependent. Biochim. Biophys. Acta – Biomembranes 2014, 1838, DOI: /j.bbamem The use of lipid nanoparticles as gene or drug carriers in targeted cell therapeutics is currently unprecedented. Here we report on the discovery that the order of mixing cationic lipids, DNA, and preparation media, has a profound effect on the formation of polymer coated lipid-DNA nanoparticles (LD-NPs). LD-NPs prepared in water where electrostatic forces dominate, and transferred to saline media (mimicking in-vivo ionic strengths), exhibit onion-like structures with a large number of layers (Figure, right panel). Conversely, LD-NPs prepared in salt with weakened electrostatic forces, exhibit drastically reduced number of layers due to the polymer- induced repulsion between layers (Figure, left panel). Significance: The results are of wide interest to the worldwide community of researchers studying pathways of formation of lipid-DNA nanoparticles employed in targeted drug delivery. The discovery allows for controlled formation of multilayered onion-like LD-NPs with dramatically increased loading capacity (i.e. due to the large number of DNA molecules per particle) and thus, therapeutic efficacy, of particles entering targeted cells.

Broader Impacts: Education and Outreach Research Training Cyrus R. Safinya, University of California-Santa Barbara, DMR Education: Undergraduate and graduate students, and postdoctoral scholars with backgrounds in materials science, physics, chemistry, and biology, are educated in methods to discover nature’s rules for assembling molecular building blocks in distinct shapes and sizes for particular functions. The learned concepts enable development of advanced materials for applications Outreach/Participation of undergraduate/ underrepresented students: gene therapy in a multidisciplinary environment (bottom photo, shown from right is Bryan Palmer, student in nursing; Zita Espinoza, student in nursing; and Glen Robinson, Chemical Engineering student). Emily Wonder also participated in the Jack Kent Cooke Bridges for Engineering and Science Transfers Program, which involved mentoring Community College students from across California. This program’s goal is to encourage Community College students to pursue an education in science and engineering at a four year University. (For more information see Materials PhD graduate student Emily Wonder (bottom photo, left) participated in the Condor Techs outreach program, a joint internship program between UCSB’s Center for Science and Engineering Partnership and Oxnard Community College. This program is partially funded by a Federal Department of Education Hispanic Serving Institution Grant Program with a goal of increasing Hispanic student representation in STEM majors. Emily mentored three Oxnard Community College students exposing them to research in Christine Tchounwou (top photo) is a Masters graduate student at Jackson State University working towards her Biology degree. Christine participated in UCSB’s RISE program (Research in Science and Engineering). Her project was centered around optical microscopy and small-angle-x-ray-scattering studies of lipid-DNA complexes used in gene delivery applications. She was trained in these modern techniques by her mentor Ramsey Majzoub (physics PhD graduate student, top photo).