Glycopolycation-DNA Polyplex Formulation N/P Ratio Affects Stability, Hemocompatibility, and in Vivo Biodistribution

Haley R. Phillips, Zachary P. Tolstyka, Bryan C. Hall, Joseph K. Hexum, Perry B Hackett, Theresa M Reineke

Research output: Contribution to journalArticle

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Abstract

Genome editing therapies hold great promise for the cure of monogenic and other diseases; however, the application of nonviral gene delivery methods is limited by both a lack of fundamental knowledge of interactions of the gene-carrier in complex animals and biocompatibility. Herein, we characterize nonviral gene delivery vehicle formulations that are based on diblock polycations containing a hydrophilic and neutral glucose block chain extended with cationic secondary amines of three lengths, poly(methacrylamido glucopyranose-block-2-methylaminoethyl methacrylate) [P(MAG-b-MAEMt)-1, -2, -3]. These polymers were formulated with plasmid DNA to prepare polyelectrolyte complexes (polyplexes). In addition, two controls, P(EG-b-MAEMt) and P(MAEMt), were synthesized, formulated into polyplexes and the ex vivo hemocompatibility, or blood compatibility, and in vivo biodistribution of the formulations were compared to the glycopolymers. While both polymer structure and N/P (amine to phosphate) ratio were important factors affecting hemocompatibility, N/P ratio played a stronger role in determining polyplex biodistribution. P(EG-b-MAEMt) and P(MAEMt) lysed red blood cells at both high and low N/P formulations while P(MAG-b-MAEMt) did not significantly lyse cells at either formulation at short and medium polymer lengths. Conversely, P(MAG-b-MAEMt) did not affect coagulation at N/P = 5, but significantly delayed coagulation at N/P = 15. P(EG-b-MAEMt) and P(MAEMt) did not affect coagulation at either formulation. After polymer and pDNA cargo distribution was observed in vivo, P(EG-b-MAEMt) N/P = 5 and P(MAG-b-MAEMt) N/P = 5 both dissociated and deposited polymer in the liver, while pDNA cargo from P(MAG-b-MAEMt) N/P = 15 was found in the liver, lungs, and spleen. The contrast between P(MAG-b-MAEMt) at N/P = 5 and 15 demonstrates that polyplex stability in the blood can be improved with N/P ratio and potentially aid polyplex biodistribution through simply varying the formulation ratios.

Original languageEnglish (US)
Pages (from-to)1530-1544
Number of pages15
JournalBiomacromolecules
Volume20
Issue number4
DOIs
StatePublished - Apr 8 2019

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Methacrylates
DNA
Polymers
Genes
Coagulation
Blood
Polyelectrolytes
Liver
Amines
Biocompatibility
Glucose
Animals
Phosphates
Cells
Plasmids

PubMed: MeSH publication types

  • Journal Article

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Glycopolycation-DNA Polyplex Formulation N/P Ratio Affects Stability, Hemocompatibility, and in Vivo Biodistribution. / Phillips, Haley R.; Tolstyka, Zachary P.; Hall, Bryan C.; Hexum, Joseph K.; Hackett, Perry B; Reineke, Theresa M.

In: Biomacromolecules, Vol. 20, No. 4, 08.04.2019, p. 1530-1544.

Research output: Contribution to journalArticle

Phillips, Haley R. ; Tolstyka, Zachary P. ; Hall, Bryan C. ; Hexum, Joseph K. ; Hackett, Perry B ; Reineke, Theresa M. / Glycopolycation-DNA Polyplex Formulation N/P Ratio Affects Stability, Hemocompatibility, and in Vivo Biodistribution. In: Biomacromolecules. 2019 ; Vol. 20, No. 4. pp. 1530-1544.
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AU - Hexum, Joseph K.

AU - Hackett, Perry B

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N2 - Genome editing therapies hold great promise for the cure of monogenic and other diseases; however, the application of nonviral gene delivery methods is limited by both a lack of fundamental knowledge of interactions of the gene-carrier in complex animals and biocompatibility. Herein, we characterize nonviral gene delivery vehicle formulations that are based on diblock polycations containing a hydrophilic and neutral glucose block chain extended with cationic secondary amines of three lengths, poly(methacrylamido glucopyranose-block-2-methylaminoethyl methacrylate) [P(MAG-b-MAEMt)-1, -2, -3]. These polymers were formulated with plasmid DNA to prepare polyelectrolyte complexes (polyplexes). In addition, two controls, P(EG-b-MAEMt) and P(MAEMt), were synthesized, formulated into polyplexes and the ex vivo hemocompatibility, or blood compatibility, and in vivo biodistribution of the formulations were compared to the glycopolymers. While both polymer structure and N/P (amine to phosphate) ratio were important factors affecting hemocompatibility, N/P ratio played a stronger role in determining polyplex biodistribution. P(EG-b-MAEMt) and P(MAEMt) lysed red blood cells at both high and low N/P formulations while P(MAG-b-MAEMt) did not significantly lyse cells at either formulation at short and medium polymer lengths. Conversely, P(MAG-b-MAEMt) did not affect coagulation at N/P = 5, but significantly delayed coagulation at N/P = 15. P(EG-b-MAEMt) and P(MAEMt) did not affect coagulation at either formulation. After polymer and pDNA cargo distribution was observed in vivo, P(EG-b-MAEMt) N/P = 5 and P(MAG-b-MAEMt) N/P = 5 both dissociated and deposited polymer in the liver, while pDNA cargo from P(MAG-b-MAEMt) N/P = 15 was found in the liver, lungs, and spleen. The contrast between P(MAG-b-MAEMt) at N/P = 5 and 15 demonstrates that polyplex stability in the blood can be improved with N/P ratio and potentially aid polyplex biodistribution through simply varying the formulation ratios.

AB - Genome editing therapies hold great promise for the cure of monogenic and other diseases; however, the application of nonviral gene delivery methods is limited by both a lack of fundamental knowledge of interactions of the gene-carrier in complex animals and biocompatibility. Herein, we characterize nonviral gene delivery vehicle formulations that are based on diblock polycations containing a hydrophilic and neutral glucose block chain extended with cationic secondary amines of three lengths, poly(methacrylamido glucopyranose-block-2-methylaminoethyl methacrylate) [P(MAG-b-MAEMt)-1, -2, -3]. These polymers were formulated with plasmid DNA to prepare polyelectrolyte complexes (polyplexes). In addition, two controls, P(EG-b-MAEMt) and P(MAEMt), were synthesized, formulated into polyplexes and the ex vivo hemocompatibility, or blood compatibility, and in vivo biodistribution of the formulations were compared to the glycopolymers. While both polymer structure and N/P (amine to phosphate) ratio were important factors affecting hemocompatibility, N/P ratio played a stronger role in determining polyplex biodistribution. P(EG-b-MAEMt) and P(MAEMt) lysed red blood cells at both high and low N/P formulations while P(MAG-b-MAEMt) did not significantly lyse cells at either formulation at short and medium polymer lengths. Conversely, P(MAG-b-MAEMt) did not affect coagulation at N/P = 5, but significantly delayed coagulation at N/P = 15. P(EG-b-MAEMt) and P(MAEMt) did not affect coagulation at either formulation. After polymer and pDNA cargo distribution was observed in vivo, P(EG-b-MAEMt) N/P = 5 and P(MAG-b-MAEMt) N/P = 5 both dissociated and deposited polymer in the liver, while pDNA cargo from P(MAG-b-MAEMt) N/P = 15 was found in the liver, lungs, and spleen. The contrast between P(MAG-b-MAEMt) at N/P = 5 and 15 demonstrates that polyplex stability in the blood can be improved with N/P ratio and potentially aid polyplex biodistribution through simply varying the formulation ratios.

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