Pharmacodynamic Studies of Fluorescent Diamond Carriers of Doxorubicin in Liver Cancer Cells and Colorectal Cancer Organoids

Ron Firestein; Cezary Marcinkiewicz; Linyan Nie; Hui Kheng Chua; Ines Velazquez Quesada; Marco Torelli; Mark Sternberg; Bojana Gligorijevic; Olga Shenderova; Romana Schirhagl; Giora Z Feuerstein

doi:10.2147/NSA.S321725

Pharmacodynamic Studies of Fluorescent Diamond Carriers of Doxorubicin in Liver Cancer Cells and Colorectal Cancer Organoids

Nanotechnol Sci Appl. 2021 Sep 7:14:139-159. doi: 10.2147/NSA.S321725. eCollection 2021.

Authors

Ron Firestein^#^{1

2}, Cezary Marcinkiewicz^#^{3

4}, Linyan Nie⁵, Hui Kheng Chua^{1

2}, Ines Velazquez Quesada⁴, Marco Torelli⁶, Mark Sternberg³, Bojana Gligorijevic⁴, Olga Shenderova⁶, Romana Schirhagl⁵, Giora Z Feuerstein³

Affiliations

¹ Centre for Cancer Research, Hudson Institute of Medical Research, Clayton, Victoria, 3168, Australia.
² Department of Molecular and Translational Sciences, Monash University, Clayton, Victoria, 3800, Australia.
³ Debina Diagnostics Inc., Newtown Square, PA, USA.
⁴ College of Engineering, Department of Bioengineering, Temple University, Philadelphia, PA, USA.
⁵ Groningen University, Groningen, 9727, the Netherlands.
⁶ Adámas Nanotechnologies, Inc., Raleigh, NC, 27617, USA.

^# Contributed equally.

Abstract

Background: We recently reported on preferential deposition of bare fluorescent diamond particles FDP-NV-700/800nm (FDP-NV) in the liver following intravenous administration to rats. The pharmacokinetics of FDP-NV in that species indicated short residency in the circulation by rapid clearance by the liver. Retention of FDP-NV in the liver was not associated with any pathology. These observations suggested that cancer therapeutics, such as doxorubicin, linked to FDP-NV, could potentially serve for anti-cancer treatment while sparing toxicities of peripheral organs.

Purpose: To generate proof-of-concept (POC) and detail mechanisms of action of doxorubicin-coated FDP-NV-700/800nm (FDP-DOX) as a prospective chemotherapeutic for metastatic liver cancer.

Methods: FDP-DOX was generated by adsorption chemistry. Experimental design included concentration and time-dependent efficacy studies as compared with naïve (baren) FDP-NV in in vitro liver cancer cells models. Uptake of FDP-NV and FDP-DOX by HepG-2, Hep-3B and hCRC organoids were demonstrated by flow-cytometry and fluorescent microscopy. FDP-DOX pharmacodynamic effects included metabolic as well as cell death biomarkers Annexin V, TUNEL and LDH leakage. DOX desorpted from FDP-DOX was assessed by confocal microscopy and chemical assay of cells fractions.

Results: FDP-DOX efficacy was dose- and time-dependent and manifested in both liver cancer cell lines and human CRC organoids. FDP-DOX was rapidly internalized into cancer cells/organoids leading to cancer growth inhibition and apoptosis. FDP-DOX disrupted cell membrane integrity as evident by LDH release and suppressing mitochondrial metabolic pathways (AlamarBlue assay). Access of free DOX to the nuclei was confirmed by direct UV-Visible fluorescent assay and confocal microscopy of DOX fluorescence.

Conclusion: The rapid uptake and profound cancer inhibition observed using FDP-DOX in clinically relevant cancer models, highlight FDP-DOX promise for cancer chemotherapeutics. We also conclude that the in vitro data justify further investment in in vivo POC studies.

Keywords: apoptosis; doxorubicin; fluorescent diamond particles-NV-700/800nm; human colorectal cancer organoids; liver cancer cell-lines.

Grants and funding

R01 CA230777/CA/NCI NIH HHS/United States