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Opening Up the Optical Imaging Window Using Nano-Luciferin

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Abstract

Purpose

The objective of this study was to formulate nanoparticles of D-luciferin (Nano-Luc), DiR (Nano-DiR) and dual functional nanoparticles with DiR and luciferin (Nano-LucDiR) for in-vivo imaging as well as tracking of the nanoparticles in tumors.

Methods

Nano-Luc and Nano-LucDiR were prepared using different lipids, and subsequently characterized for loading and entrapment efficiency, physical properties, release profile, toxicity and stability. We utilized Response Surface Methodology (RSM) to optimize the nanoparticles using design of experiment (DOE Vr.8.0). Nano-Luc was evaluated against free luciferin to establish its pharmacokinetic parameters in mice. In-vivo imaging of tumors and tracking of nanoparticles was carried out with an IVIS® Spectrum-CT (Caliper) using xenograft, orthotopic and metastatic tumor models in BALB/c nude mice with different cell lines and different routes of nanoparticle administration (subcutaneous, intraperitoneal and intravenous).

Results

Particle size of both Nano-Luc and Nano-LucDiR were found to be <200 nm. Nano-Luc formulation showed a slow and controlled release upto 72 h (90%) in vitro. The optimized Nano-Luc had loading efficiency of 5.0 mg/ml with 99% encapsulation efficiency. Nano-Luc and Nano-LucDiR formulations had good shelf stability. Nano-Luc and Nano-LucDiR enhanced plasma half-life of luciferin compared to free luciferin thus providing longer circulation of luciferin in plasma enabling imaging of tumors for more than 24 h. Nano-LucDiR allowed simultaneous bioluminescent and fluorescent imaging to be conducted, with three-dimensional reconstruct of tumors without losing either signal during the acquisition time.

Conclusion

Nano-Luc and Nano-LucDiR allowed prolonged reproducible in-vivo imaging of tumors, especially during multimodality 3D imaging.

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Abbreviations

CCD:

Central Composite Design

CT:

Computed Tomography

DOE:

Design Of Experiment

DSC:

Differential Scanning Calorimetry

FOV:

Field Of View

IP:

Intraperitoneal

IV:

Intravenous

Nano-DiR:

Nanoparticles of DiR

Nano-Luc:

Nanoparticles of D-Luciferin

Nano-LucDiR:

Nanoparticles with DiR and Luciferin

NCs:

Nano lipid carriers

QbB:

Quality by Design

ROI:

Region of Interest

RSM:

Response Surface Methodology

SD:

Standard Deviations

SQ/SC:

Subcutaneous

TPGS:

α-Tocopherol Polyethylene Glycol Succinate

Xenolight DiR:

DiIC18(7) or 1,1′-dioctadecyltetramethyl indotricarbocyanine iodide

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ACKNOWLEDGMENTS AND DISCLOSURES

This work was financially supported by National Institute of Health - MBRS-SC1 Program (Grant # SC1 GM092779-01). The authors report no financial interest that might pose a potential, perceived, or real conflict of interest. U.S. Patent filed “Modified Nanodelivery System and Method for Enhanced In vivo Medical and Preclinical Imaging.” (U.S. 13/851610).

Author information

Authors and Affiliations

  1. College of Pharmacy and Pharmaceutical Sciences, Florida A&M University, Tallahassee, Florida, 32307, USA

    Apurva R. Patel & Mandip Singh

  2. Caliper - A PerkinElmer Company, Alameda, California, 94501, USA

    Ed Lim & Kevin P. Francis

Authors
  1. Apurva R. Patel
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  2. Ed Lim
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Corresponding author

Correspondence to Mandip Singh.

Electronic supplementary material

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Figure S1

Affect of Lipid and Oil type on loading efficiency (A), entrapment efficiency (B), 24 h release rate (C) and mean particle size (D) in the RSM. (GIF 291 kb)

High resolution image (TIFF 34 kb)

Figure S2

Response surface plots (A) and countor plots (B) showing the effect of different concentration of lipid and oil on release rate, loading efficiency and entrapment efficiency of Nano-Luc. (GIF 529 kb)

High resolution image (TIFF 338 kb)

Figure S3

Differential Scanning Calorimetry of Nano-Luc formulations. A) Luciferin, B) Geleol, C) Nano-Luc, D) Precirol, E) Nano-LucDiR and F) Miglyol. (GIF 275 kb)

High resolution image (TIFF 35 kb)

Figure S4

A) A graph of Log (% luciferin recovery) vs time for the effect of temperature (30°C, 40°C and 50°C) on Nano-Luc stability. B) In-vitro Comparison of Nano-luciferin and luciferin at 150 μg/mL with background subtraction (Plot of total flux Vs cell number). (n = 3) (GIF 228 kb)

High resolution image (TIFF 61 kb)

Figure S5

Nano-luc Tumor Imaging in mice with A) subcutaneous 4 T1-luc2 tumors and B) orthotopic 4 T1-luc2 tumors using 150 mg/kg luciferin equivalent Nano-Luc by IP. (GIF 232 kb)

High resolution image (TIFF 238 kb)

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Patel, A.R., Lim, E., Francis, K.P. et al. Opening Up the Optical Imaging Window Using Nano-Luciferin. Pharm Res 31, 3073–3084 (2014). https://doi.org/10.1007/s11095-014-1400-9

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  • DOI: https://doi.org/10.1007/s11095-014-1400-9

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