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Study of the selective effect of FITC-CRD13 dendrimer locked in liposomes on the viability of adhesive breast and colon cancer cell lines in comparison with the normal human fibroblasts


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Study of the selective effect of FITC-CRD13 dendrimer locked in liposomes on the viability of adhesive breast and colon cancer cell lines in comparison with the normal human fibroblasts

Šimon Šutý 1 Jakub Magiera 2 Veronika Šubjaková 1 Zuzana Garaiová 1 Sylwia Michlewska 2 Marcin Holota 2 Maksim Ionov 2 Natalia Sanz-del Olmo 3 Dzmitry Shcharbin 4 Iveta Waczulíková 1 Francisco Javier de la Mata 3 Maria Bryszewska 2 Tibor Hianik 1

1Faculty of Mathematics, Physics and Informatics Comenius University, Department of Nuclear Physics and Biophysics, Bratislava, Slovakia
2Faculty of Biology and Environmental protection, Department of General Biophysics, University of Lodz, Laboratory of Microscopic Imaging and Specialized Biological Techniques, Lodz, Poland
3Department of Organic and Inorganic Chemistry, and Research Institute in Chemistry ”Andrés M. Del Río” (IQAR), University of Alcalá, Madrid, Spain
4The State Scientific Institution "Institute of Biophysics and Cell Engineering of the National Academy of Sciences of Belarus, Minsk, Belarus
simurda.suter@gmail.com

Since malignancies have long been the second most common cause of death, it is important to explore new approaches to the treatment of cancer patients. In this context, we focused on a novel dual nanoparticle-based drug delivery platform. Specifically, we combined a fluorescently labelled carbosilane-ruthenium dendrimer FITC-CRD13 possessing anticancer properties [1] with liposomes and investigated its cytotoxic effect on adhesive breast (MCF-7) and colon (HT-29) cancer cells with expected minimal effect on the viability of healthy human fibroblasts (BJ). Liposomes were prepared by a hydration method using zwitterionic 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC; 15mM). FITC-CRD13 dendimer (0.15mM) was added during the hydration step of the lipid film. In order to reduce the size of liposomes and remove unencapsulated dendrimers, prepared samples were extruded through a 400 nm polycarbonate membrane and centrifuged thereafter.

Subsequently, the effect of DMPC+FITC-CRD13 liposomes (0.5; 1; 2.5; 5; 10 µM; presumed dendrimer concentration) on the viability of selected cell lines was investigated using MTT assay [2]. Non-treated cells as 100% viability reference (negative control) and methanol (10µM, positive control) were used.

Treatment of the cells with DMPC+FITC-CRD13 liposomes along the whole tested concentration range caused a viability drop to or below 80% for HT-29 and MCF-7 cancer cell lines. On the other hand, in BJ cell line, the viability was maintained at 80% or more. This suggests a selective effect of DMPC+FITC-CRD13 liposomes on the cell viability, with a desirable toxicity towards cancer cells. For all three tested cell lines, a larger decrease in cell viability upon the 3h incubation time in comparison to 24h, was observed. Concentration dependent relation between the applied DMPC+FITC-CRD13 and viability of cells was not significant, probably as a result of narrow concentration range chosen.

Obtained preliminary results will be used for further protocol optimization. Stability of DMPC+FITC-CRD13 liposomes as well as the effect of lipid composition should be tested.

This work was supported by the Slovak Research and Development Agency, APVV (Projects No. SK-PL-18-0080, APVV-14-0267, SK-BY-RD-19-0019), VEGA 1/0756/20, KEGA 041UK-4/2020 and by PL-SK 2019–2020 bilateral project of NAWA PPN/BIL/2018/1/00150; Polish National Agency for Academic Exchange, NAWA, PPI/APM/2018/1/00007/U/001.
[1] Michlewska et al. 2019: Synthesis and Characterization of FITC Labelled Ruthenium Dendrimer as a Prospective Anticancer Drug; In Biomolecules 2019, 9, 411, doi:10.3390/biom9090411.
[2] Chonco L. et al.: Carbosilane dendrimer nanotechnology outlines of the broad HIV blocker profile; In Journal of Controlled Release, 2012, 161, 949–958, doi:10.1016/j.jconrel.2012.04.050.
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Discussion

Prospects for further research 16.05.2020 22:04
Hello Simon,
Thank you for the interesting contribution - there is no doubt that development of efficient and safe drug delivery systems for tumor targeting is of utmost importance. The combination of ...
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Šimon Šutý
Passive
Re: Prospects for further research 24.05.2020 11:54
Hello Iveta,
Thank you for your comments and questions.
1. We where thinking about estimating the final amount of entrapped liposomes using STED-microscopy and measuring the fluorescence of FITC...
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Interesting contribution 22.05.2020 13:40
Hello Simon,
it is the interesting contribution,
why did you add dendrimers during the hydration step of the lipid film?
Šimon Šutý
Passive
Re: Interesting contribution 24.05.2020 11:11
Hello Lucia
Thank you for your question. From previous experiments we estimated that liposome-CRD-13 complex is more stable when CRD-13 is added during the hydration step, in comparison when CRD-13 was...
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nice work 22.05.2020 13:48
Hi Šimon,
can you explain why size of DMPC is larger than size of DMPC+FITC-CRD13?
Šimon Šutý
Passive
Re: nice work 24.05.2020 11:32
Hello Daniela,
Thank you for your question. Previous work of Michlewska S. et al. (2019) indicates, that CRD-13 interacts with both hydrophobic and hydrophilic part of the membrane of liposomes. Theref...
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