TY - JOUR
T1 - Kinetic Control in Assembly of Plasmid DNA/Polycation Complex Nanoparticles
AU - Hu, Yizong
AU - He, Zhiyu
AU - Hao, Yue
AU - Gong, Like
AU - Pang, Marion
AU - Howard, Gregory P.
AU - Ahn, Hye Hyun
AU - Brummet, Mary
AU - Chen, Kuntao
AU - Liu, Heng Wen
AU - Ke, Xiyu
AU - Zhu, Jinchang
AU - Anderson, Caleb F.
AU - Cui, Honggang
AU - Ullman, Christopher G.
AU - Carrington, Christine A.
AU - Pomper, Martin G.
AU - Seo, Jung Hee
AU - Mittal, Rajat
AU - Minn, Il
AU - Mao, Hai Quan
N1 - Funding Information:
This work was supported by Cancer Targeting Systems Inc., the National Institutes of Health (R01 EB018358, P41 EB024495, and P50 CA058236), and Maryland Advanced Research Computing Center. The authors thank Dr. Will West from Cancer Targeting Systems, Ms. Beihang Yu from University of California Santa Barbara (SLS analysis), Dr. Michael Bevan and Ms. Elena Alexandra Garcia from Johns Hopkins University (SLS instrumentation), and Dr. Michael McCaffery from Johns Hopkins Integrated Imaging Center (TEM analysis) for their helpful discussions and technical assistance.
Publisher Copyright:
© 2019 American Chemical Society.
PY - 2019/4/30
Y1 - 2019/4/30
N2 - Polyelectrolyte complex (PEC) nanoparticles assembled from plasmid DNA (pDNA) and polycations such as linear polyethylenimine (lPEI) represent a major nonviral delivery vehicle for gene therapy tested thus far. Efforts to control the size, shape, and surface properties of pDNA/polycation nanoparticles have been primarily focused on fine-tuning the molecular structures of the polycationic carriers and on assembly conditions such as medium polarity, pH, and temperature. However, reproducible production of these nanoparticles hinges on the ability to control the assembly kinetics, given the nonequilibrium nature of the assembly process and nanoparticle composition. Here we adopt a kinetically controlled mixing process, termed flash nanocomplexation (FNC), that accelerates the mixing of pDNA solution with polycation lPEI solution to match the PEC assembly kinetics through turbulent mixing in a microchamber. This achieves explicit control of the kinetic conditions for pDNA/lPEI nanoparticle assembly, as demonstrated by the tunability of nanoparticle size, composition, and pDNA payload. Through a combined experimental and simulation approach, we prepared pDNA/lPEI nanoparticles having an average of 1.3 to 21.8 copies of pDNA per nanoparticle and average size of 35 to 130 nm in a more uniform and scalable manner than bulk mixing methods. Using these nanoparticles with defined compositions and sizes, we showed the correlation of pDNA payload and nanoparticle formulation composition with the transfection efficiencies and toxicity in vivo. These nanoparticles exhibited long-term stability at -20 °C for at least 9 months in a lyophilized formulation, validating scalable manufacture of an off-the-shelf nanoparticle product with well-defined characteristics as a gene medicine.
AB - Polyelectrolyte complex (PEC) nanoparticles assembled from plasmid DNA (pDNA) and polycations such as linear polyethylenimine (lPEI) represent a major nonviral delivery vehicle for gene therapy tested thus far. Efforts to control the size, shape, and surface properties of pDNA/polycation nanoparticles have been primarily focused on fine-tuning the molecular structures of the polycationic carriers and on assembly conditions such as medium polarity, pH, and temperature. However, reproducible production of these nanoparticles hinges on the ability to control the assembly kinetics, given the nonequilibrium nature of the assembly process and nanoparticle composition. Here we adopt a kinetically controlled mixing process, termed flash nanocomplexation (FNC), that accelerates the mixing of pDNA solution with polycation lPEI solution to match the PEC assembly kinetics through turbulent mixing in a microchamber. This achieves explicit control of the kinetic conditions for pDNA/lPEI nanoparticle assembly, as demonstrated by the tunability of nanoparticle size, composition, and pDNA payload. Through a combined experimental and simulation approach, we prepared pDNA/lPEI nanoparticles having an average of 1.3 to 21.8 copies of pDNA per nanoparticle and average size of 35 to 130 nm in a more uniform and scalable manner than bulk mixing methods. Using these nanoparticles with defined compositions and sizes, we showed the correlation of pDNA payload and nanoparticle formulation composition with the transfection efficiencies and toxicity in vivo. These nanoparticles exhibited long-term stability at -20 °C for at least 9 months in a lyophilized formulation, validating scalable manufacture of an off-the-shelf nanoparticle product with well-defined characteristics as a gene medicine.
KW - DNA/polycation nanoparticle
KW - gene delivery
KW - kinetic control
KW - linear polyethylenimine
KW - polyelectrolyte complex
KW - transfection
KW - turbulent mixing
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U2 - 10.1021/acsnano.9b03334
DO - 10.1021/acsnano.9b03334
M3 - Article
C2 - 31503450
AN - SCOPUS:85073303709
VL - 13
SP - 10161
EP - 10178
JO - ACS Nano
JF - ACS Nano
SN - 1936-0851
IS - 9
ER -