Effect of serum on transfection by polyethylenimine/virus-like particle hybrid gene delivery vectors

David M. Drake; Rahul K. Keswani; Daniel W. Pack

doi:10.1007/s11095-010-0238-z

Effect of serum on transfection by polyethylenimine/virus-like particle hybrid gene delivery vectors

David M. Drake, Rahul K. Keswani, Daniel W. Pack

Research output: Contribution to journal › Article › peer-review

13 Scopus citations

Abstract

Purpose: Murine leukemia virus-like particles (M-VLP) complexed with polymers to promote cellular uptake and endosomal escape represent a new class of effective gene delivery vectors. Building upon recent studies of viral-synthetic hybrid vectors, we report the effects of serum on the formation, activity and stability of PEI/M-VLP complexes. Methods: M-VLP were produced by cells grown in serum-supplemented media (M-VLP-S), serum-free media (M-VLP-SF) or serum-free Opti-MEM® I (M-VLP-OM). PEI/M-VLP stoichiometry was varied to investigate complex formation and optimal transfection conditions. The effects of prolonged storage, freeze-thaw cycles, and ultracentrifugation of M-VLP on the stability of vector transduction efficiency were also observed. Results: M-VLP-S required more PEI to form infective complexes than M-VLP-SF and M-VLP-OM. The stoichiometry of PEI/M-VLP-S was dependent on total PEI concentration (7-8 μg/100 μL M-VLP supernatant), while optimal infectivity of PEI/M-VLP-SF and PEI/M-VLP-OM depended on PEI/M-VLP ratios (12-17 μg and 10-14 μg PEI/10⁹ M-VLP, respectively). PEI/M-VLP-SF and PEI/M-VLP-OM complexes were significantly more efficient than PEI/M-VLP-S. Stability of the hybrid vectors was not significantly affected by serum. Conclusions: PEI/M-VLP complexes exhibiting increased efficiency were constructed by producing M-VLP in serum-free media. M-VLP could be stored by freezing or refrigeration and concentrated by ultracentrifugation without unacceptable loss of infectivity.

Original language	English
Pages (from-to)	2457-2465
Number of pages	9
Journal	Pharmaceutical Research
Volume	27
Issue number	11
DOIs	https://doi.org/10.1007/s11095-010-0238-z
State	Published - Nov 2010

Bibliographical note

Funding Information:
This work was supported by the National Science Foundation (BES 06-02636) and the American Cancer Society (RSG-05-019-01-CDD). Flow cytometry was performed at the Flow Cytometry Facility of the Roy J. Carver Biotechnology Center at the University of Illinois, and confocal microscopy was performed at the Center for Microscopic Imaging at the University of Illinois College of Veterinary Medicine.

Keywords

gene delivery
hybrid vectors
murine leukemia virus
polyethylenimine
virus-like particles

ASJC Scopus subject areas

Biotechnology
Molecular Medicine
Pharmacology
Pharmaceutical Science
Organic Chemistry
Pharmacology (medical)

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10.1007/s11095-010-0238-z

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title = "Effect of serum on transfection by polyethylenimine/virus-like particle hybrid gene delivery vectors",

abstract = "Purpose: Murine leukemia virus-like particles (M-VLP) complexed with polymers to promote cellular uptake and endosomal escape represent a new class of effective gene delivery vectors. Building upon recent studies of viral-synthetic hybrid vectors, we report the effects of serum on the formation, activity and stability of PEI/M-VLP complexes. Methods: M-VLP were produced by cells grown in serum-supplemented media (M-VLP-S), serum-free media (M-VLP-SF) or serum-free Opti-MEM{\textregistered} I (M-VLP-OM). PEI/M-VLP stoichiometry was varied to investigate complex formation and optimal transfection conditions. The effects of prolonged storage, freeze-thaw cycles, and ultracentrifugation of M-VLP on the stability of vector transduction efficiency were also observed. Results: M-VLP-S required more PEI to form infective complexes than M-VLP-SF and M-VLP-OM. The stoichiometry of PEI/M-VLP-S was dependent on total PEI concentration (7-8 μg/100 μL M-VLP supernatant), while optimal infectivity of PEI/M-VLP-SF and PEI/M-VLP-OM depended on PEI/M-VLP ratios (12-17 μg and 10-14 μg PEI/109 M-VLP, respectively). PEI/M-VLP-SF and PEI/M-VLP-OM complexes were significantly more efficient than PEI/M-VLP-S. Stability of the hybrid vectors was not significantly affected by serum. Conclusions: PEI/M-VLP complexes exhibiting increased efficiency were constructed by producing M-VLP in serum-free media. M-VLP could be stored by freezing or refrigeration and concentrated by ultracentrifugation without unacceptable loss of infectivity.",

keywords = "gene delivery, hybrid vectors, murine leukemia virus, polyethylenimine, virus-like particles",

author = "Drake, {David M.} and Keswani, {Rahul K.} and Pack, {Daniel W.}",

note = "Funding Information: This work was supported by the National Science Foundation (BES 06-02636) and the American Cancer Society (RSG-05-019-01-CDD). Flow cytometry was performed at the Flow Cytometry Facility of the Roy J. Carver Biotechnology Center at the University of Illinois, and confocal microscopy was performed at the Center for Microscopic Imaging at the University of Illinois College of Veterinary Medicine.",

year = "2010",

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doi = "10.1007/s11095-010-0238-z",

language = "English",

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T1 - Effect of serum on transfection by polyethylenimine/virus-like particle hybrid gene delivery vectors

AU - Drake, David M.

AU - Keswani, Rahul K.

AU - Pack, Daniel W.

N1 - Funding Information: This work was supported by the National Science Foundation (BES 06-02636) and the American Cancer Society (RSG-05-019-01-CDD). Flow cytometry was performed at the Flow Cytometry Facility of the Roy J. Carver Biotechnology Center at the University of Illinois, and confocal microscopy was performed at the Center for Microscopic Imaging at the University of Illinois College of Veterinary Medicine.

PY - 2010/11

Y1 - 2010/11

N2 - Purpose: Murine leukemia virus-like particles (M-VLP) complexed with polymers to promote cellular uptake and endosomal escape represent a new class of effective gene delivery vectors. Building upon recent studies of viral-synthetic hybrid vectors, we report the effects of serum on the formation, activity and stability of PEI/M-VLP complexes. Methods: M-VLP were produced by cells grown in serum-supplemented media (M-VLP-S), serum-free media (M-VLP-SF) or serum-free Opti-MEM® I (M-VLP-OM). PEI/M-VLP stoichiometry was varied to investigate complex formation and optimal transfection conditions. The effects of prolonged storage, freeze-thaw cycles, and ultracentrifugation of M-VLP on the stability of vector transduction efficiency were also observed. Results: M-VLP-S required more PEI to form infective complexes than M-VLP-SF and M-VLP-OM. The stoichiometry of PEI/M-VLP-S was dependent on total PEI concentration (7-8 μg/100 μL M-VLP supernatant), while optimal infectivity of PEI/M-VLP-SF and PEI/M-VLP-OM depended on PEI/M-VLP ratios (12-17 μg and 10-14 μg PEI/109 M-VLP, respectively). PEI/M-VLP-SF and PEI/M-VLP-OM complexes were significantly more efficient than PEI/M-VLP-S. Stability of the hybrid vectors was not significantly affected by serum. Conclusions: PEI/M-VLP complexes exhibiting increased efficiency were constructed by producing M-VLP in serum-free media. M-VLP could be stored by freezing or refrigeration and concentrated by ultracentrifugation without unacceptable loss of infectivity.

AB - Purpose: Murine leukemia virus-like particles (M-VLP) complexed with polymers to promote cellular uptake and endosomal escape represent a new class of effective gene delivery vectors. Building upon recent studies of viral-synthetic hybrid vectors, we report the effects of serum on the formation, activity and stability of PEI/M-VLP complexes. Methods: M-VLP were produced by cells grown in serum-supplemented media (M-VLP-S), serum-free media (M-VLP-SF) or serum-free Opti-MEM® I (M-VLP-OM). PEI/M-VLP stoichiometry was varied to investigate complex formation and optimal transfection conditions. The effects of prolonged storage, freeze-thaw cycles, and ultracentrifugation of M-VLP on the stability of vector transduction efficiency were also observed. Results: M-VLP-S required more PEI to form infective complexes than M-VLP-SF and M-VLP-OM. The stoichiometry of PEI/M-VLP-S was dependent on total PEI concentration (7-8 μg/100 μL M-VLP supernatant), while optimal infectivity of PEI/M-VLP-SF and PEI/M-VLP-OM depended on PEI/M-VLP ratios (12-17 μg and 10-14 μg PEI/109 M-VLP, respectively). PEI/M-VLP-SF and PEI/M-VLP-OM complexes were significantly more efficient than PEI/M-VLP-S. Stability of the hybrid vectors was not significantly affected by serum. Conclusions: PEI/M-VLP complexes exhibiting increased efficiency were constructed by producing M-VLP in serum-free media. M-VLP could be stored by freezing or refrigeration and concentrated by ultracentrifugation without unacceptable loss of infectivity.

KW - gene delivery

KW - hybrid vectors

KW - murine leukemia virus

KW - polyethylenimine

KW - virus-like particles

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JF - Pharmaceutical Research

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ER -

Effect of serum on transfection by polyethylenimine/virus-like particle hybrid gene delivery vectors

Abstract

Bibliographical note

Keywords

ASJC Scopus subject areas

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