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Relationships between Liposome Properties, Cell Membrane Binding, Intracellular Processing, and Intracellular Bioavailability

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Abstract

Positive surface charge enhances liposome uptake into cells. Pegylation, used to confer stealth properties to enable in vivo applications of cationic liposomes, compromises internalization. The goal of this study was to determine the quantitative relationships between these two liposome properties (separately and jointly), liposomes binding to cell membrane, and the subsequent internalization and residence in intracellular space (referred to as intracellular bioavailability). The results, obtained in pancreatic Hs-766T cancer cells, revealed nonlinear and inter-dependent relationships, as well as substantial qualitative and quantitative differences. The proportionality constant K of intracellular and membrane-bound liposomes at equilibrium (i.e., I eq and B eq) showed a positive triphasic relationship with surface charge and a negative biphasic relationship with pegylation. Near-neutral liposomes showed little internalization of the membrane-bound moiety, increasing to a constant K value for medium charge liposomes (+15 to +35 mV zeta potential), followed by a further increase for highly charged liposomes (greater than or equal to +46 mV). The decline of pegylation with K value showed a breakpoint at 2%. The negative consequences of pegylation (%PEG) were partially offset by increasing charge (ZP). The best-fitting regression equations are: B eq = −1.36 × %PEG + 0.33 × ZP; I eq = −1.52 × %PEG + 0.34 × ZP. It suggested that 1% pegylation increase can be offset with 4 mV ZP. The differences are such that it may be possible to balance these parameters to simultaneously maximize the stealth property and intracellular bioavailability of cationic liposomes.

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Abbreviations

AUC:

Area-under-concentration-time curve

B and B eq :

Concentration of liposomes bound to cell membrane at time t and at equilibrium

CX-Y :

Cationic liposomes with X mol.% DOTAP and Y% pegylation

DAPI:

4′,6-Diamidino-2-phenylindole, dihydrochloride

DOPE:

1,2-Dioleoyl-sn-glycero-3-phosphoethanolamine

DOTAP:

1,2-Dioleoyl-3-trimethylammonium-propane

DPPC:

1,2-Dipalmitoyl-sn-glycero-3-phosphocholine

EC:

Initial extracellular concentration

EPR:

Enhanced permeability and retention

I and I eq :

Intracellular concentrations of liposomes at time t and at equilibrium

mPEG-DSPE:

1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000]

PBS:

Phosphate-buffered saline

PEG:

Polyethylene glycol

PEG-DSPE:

1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-N-carbamoyl-methoxypoly-ethylene glycol-2000

Rhod-DOPE:

DOPE-N-(lissamine rhodamine B sulfonyl) ammonium salt

TC and TCeq :

Total concentrations of cell-associated liposomes at time t and at equilibrium

T eq :

Time to reach equilibrium

ZP:

Zeta potential

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ACKNOWLEDGEMENTS

This work was supported in part by research grants R43CA134047, R01EB015253 and R01CA158300 from the National Cancer Institute, DHHS. Yinghuan Li is supported by the China Scholarship Council Fellowship. Images used in this article were generated at The Campus Microscopy and Imaging Facility, The Ohio State University.

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Authors and Affiliations

  1. Division of Pharmaceutics, College of Pharmacy, China Pharmaceutical University, Nanjing, 210009, Jiangsu, People’s Republic of China

    Yinghuan Li & Jiabi Zhu

  2. Division of Pharmaceutics, College of Pharmacy, The Ohio State University, 500 West 12th Ave, Columbus, Ohio, 43210, USA

    Yinghuan Li, Yue Gao, M. Guillaume Wientjes & Jessie L.-S. Au

  3. Optimum Therapeutics LLC, OSU Science Tech Village, Columbus, Ohio, 43212, USA

    Jie Wang

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  1. Yinghuan Li
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Correspondence to Jessie L.-S. Au.

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Li, Y., Wang, J., Gao, Y. et al. Relationships between Liposome Properties, Cell Membrane Binding, Intracellular Processing, and Intracellular Bioavailability. AAPS J 13, 585–597 (2011). https://doi.org/10.1208/s12248-011-9298-1

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  • DOI: https://doi.org/10.1208/s12248-011-9298-1

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