Evaluation of cytotoxicity of a purified venom protein from Naja kaouthia (NKCT1) using gold nanoparticles for targeted delivery to cancer cell.
In our earlier report, gold nanoparticle (GNP) and snake venom protein toxin NKCT1 were conjugated and primary characteristics were done. In this communication, further characteristics of GNP-NKCT1 were done with TGA, BET, Zeta potential, ICP-MS, FTIR, XPS, and in vitro release kinetics for its physicochemical, molecular nature and bonding. TGA and ICP-MS showed that the number of conjugation was 40 ± 5 to 90 ± 8 NKCT1 per gold nanoparticles. FTIR and XPS corresponding to (CO), (NH), (SS) reformulated the conjugation of GNP with NKCT1. The efficacy of GNP-NKCT1 on cancer cells were analyzed by MTT assay which demonstrated superior cytotoxic effects as compared to native NKCT1. IC50dose of GNP-NKCT1 was less than 4 μg/ml in cancer cell lines, whereas in case of NKCT1 it was average 8 μg/ml. Twice dose of IC50of GNP-NKCT1 even showed less toxicity compared to unconjugated NKCT1, towards normal epithelial or fibroblast cell and also in peripheral blood mononuclear lymphocytes. Flow cytometry analysis revealed that percentage of apoptotic C6 cells was much higher in GNP-NKCT1 treatment (54.58%) than that of NKCT1 treatment (26.79%). Flow cytometric analysis of cell cycle using GNP-NKCT1 on C6 cancer cells revealed that it arrested the cell cycle at Go/G1phases. In diethylnitrosamine (DEN) induced in vivo hepatocarcinoma mice, the activities of hepatic enzymes- aspartate transaminase (AST) and alanine transaminase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and activities of antioxidant enzymes- superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) and glutathione peroxidase (GPx) were restored by GNP-NKCT1. This study indicated the capability of gold nanoparticles in enhancing the cancer cell uptake of NKCT1 and also suggested that GNP-NKCT1 might be a good source of anti-carcinoma or anti-sarcoma targeted agent.
Laboratory of Toxinology & Experimental Pharmacodynamics, Department of Physiology, University of Calcutta, 92, APC Road, Kolkata, 700 009, India.
Tanmoy
T
Bhowmik
Laboratory of Toxinology & Experimental Pharmacodynamics, Department of Physiology, University of Calcutta, 92, APC Road, Kolkata, 700 009, India.
Partha Pratim
PP
Saha
Laboratory of Toxinology & Experimental Pharmacodynamics, Department of Physiology, University of Calcutta, 92, APC Road, Kolkata, 700 009, India.
Amrita
A
Sarkar
Laboratory of Toxinology & Experimental Pharmacodynamics, Department of Physiology, University of Calcutta, 92, APC Road, Kolkata, 700 009, India. Electronic address: agomescu@gmail.com.
Antony
A
Gomes
D000595:Amino Acid Sequence; D000818:Animals; D017304:Annexin A5 / Q000378:metabolism; D000975:Antioxidants / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000378:metabolism; D015153:Blotting, Western; D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D016503:Drug Delivery Systems; D004546:Elapid Venoms / Q000737:chemistry / Q000494:pharmacology*; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D006046:Gold / Q000737:chemistry*; D057809:HEK293 Cells; D006801:Humans; D020128:Inhibitory Concentration 50; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D053768:Metal Nanoparticles / Q000737:chemistry*; D051379:Mice; D041681:NIH 3T3 Cells; D056951:Photoelectron Spectroscopy; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D051785:Smad Proteins / Q000378:metabolism; D017550:Spectroscopy, Fourier Transform Infrared; D013818:Thermogravimetry
D017304:Annexin A5; D000975:Antioxidants; D014408:Biomarkers, Tumor; D004546:Elapid Venoms; C000591546:NKCT1 protein, Naja kaouthia; D019253:Proto-Oncogene Proteins c-bcl-2; D051785:Smad Proteins; D006046:Gold; D045304:Cytochromes c; D053148:Caspase 3; D016650:Fluorescein-5-isothiocyanate
Apoptosis; Cancer cell; Gold nanoparticle; NKCT1; Snake venom; Toxicity