EXPERIMENTAL

SUMMARY: The selective over expression of ferritin receptor (Fr) in breast cancer cells provide a unique opportunity to deliver therapeutic agents to cancer cells via receptor mediated endocytosis mechanism. The present study is aimed at designing and development of doxorubicin bearing solid lipid nanoparticles coupled with ferritin for their selective and preferential presentation at tumour cells and thus assessing their targetability.

Akash Chaurasiya1, Mayank Singhal and Anil K. Chaurasiya2

1Faculty of Pharmacy, Jamia Hamdard, New Delhi, India2Pharmacy Department, S.N. Medical College, Agra, India

e-mail: akashchaurasiya@rediffmail.com

Solid lipid nanoparticles have been widely investigated clinically for various purposes such as selective drug delivery and diagnostic agents [1-2]. Solid lipid nanoparticles encapsulated anticancer drugs reveal their potential for increased therapeutic efficacy and decreased nonspecific toxicities due to their ability to enhance the delivery of chemotherapeutic agents to tumors. The ferritin molecule is a hollow protein shell (outside diameter 12–13 nm, inside 7–8 nm), composed of 24 polypeptide chains and capable of storing up to 4500 Fe (III) atoms as an inorganic complex [3] Iron is vital element for the metabolism, viability and proliferation of normal and of cancer cells. In the work done by various researchers, ferritin has been identified as one of the successful targets commonly found on tumour cells of breast, liver, colon, etc, while cell types with normal phenotype express low or frequently undetectable levels of ferritin [4]. Researchers observed that ferritin level is increased in tumour cells or malignant cells which could be due to an increase in number of receptors responsible for uptake of ferritin (i.e. iron storage protein). The present study is aimed at designing and development of doxorubicin bearing solid lipid nanoparticles coupled with ferritin for their selective and preferential presentation at tumour cells and thus assessing their targetability.

[1] C. Schwarz, W. Mehnert, J.S. Lucks, R.H. Muller, J Control Rel., 1994, 30, pp. 83–96.[2] R.H. Muller, W. Mehnert, J.S. Lucks, C. Schwarz, A.Z. Muhlen, H. Weyhers, C. Freitas , D. Ruhl, Eur J Pharm Biopharm, 1995, 41, pp. 62–69.[3] G.C. Ford, P.M. Harrison, D.W. Rice, J.M.A. Smith, A. Treffry, J.L. White, J. and Yariv, Phil. Trans. R. Soc. Lond. B, 304, 1984, pp. 551-565.[4] S.B. Mark, J.N. Thomson, The EMBO J, 1983, 2, pp. 1599–603.

TEM Photomicrogra

ph of Optimized SLN-PEG-Fr

SEM Photomicrogra

ph of optimized

SLN-PEG-Fr

B

Size, PDI and Entrapment Efficiency of Optimized SLN

Formulations

INTRODUCTION

Surface Morphology

FLUORESCENCE MICROSCOPY

PBS (pH 7.4) containing 1%

tween-80

Cellular uptake studies Cellular uptake studies were conducted with SLNs formulation encapsulating calcein dye diluted in 1 ml of RPMI-1640 and added to monolayers of MDA-MB-468 breast cancer cells grown in 25 mm culture flask and incubating 2x105 cells for 0.5, 1, 2, 3 and 4 hours at 37º C.

Fr-conjugated SLNs

Solvent dilution method

Characterization of SLNs•Shape and morphology by transmission electron microscopy and Scanning electron microscopy

•Vesicle size and size distribution using Zetasizer 3000 (Malvern Inst. Ltd., UK).

•Entrapment of doxorubicin in SLNs formulation using minicolunm centrifugation technique.

•In vitro drug release from the formulations using dialysis membrane.

Tristearin, HSPC, cholesterol and Fr-PEG-

DSPE conjugate (40:40:20:1) dissolved in

ethanol at 40ºC

SLN Preparation

Formulation

code

Size (nm)

PDI EntrapmentEfficiency

(%)SLN-PEG 154.1±3.

540.08

247.34±2.45

SLN-PEG-Fr

189.2±2.46

0.094

59.25±1.86

RESULTS AND DISCUSSION

0

10

20

30

40

50

60

70

0 2 4 6 8 10 12 14 16 18 20 22 24

Time (hr)

Cu

mu

lati

ve

(%

) D

rug

Re

lea

se

Percent drug release of uncoupled SLN

Percent drug release of ferritin coupled SLN

In vitro Release Study

Cell Uptake Study

Fluorescence microscopic images of

(A) Fr coupled SLN and (B) uncoupled SLNs

Doxorubicin loaded ferritin coupled

SLNs were successfully developed and

characterized. In-vitro release study clearly shows

that incorporation of doxorubicin in

SLNs decreases the rate of release

therefore the drug remains for longer

duration of time in the body. Conjugation of ferritin to the

pegylated SLNs enhance in vitro cell

uptake upto 10.4 times as compared to

uncoupled pegylated SLNs. The Fluorescence microscopy further

confirms the higher uptake of ferritin

coupled SLNs in MDA-MB-468 breast

cancer cell lines. Therefore, the developed system

appear promising in the cancer

treatment by selectively targeting the

tumour cell lines. The results warrant

further evaluation of this delivery

system.

CONCLUSIONS

REFERENCES

National Conference on Impact of Nanotechnology on Drug Discovery & Development, Shimla 2011