Yonsei Med J.  2019 Jun;60(6):509-516. 10.3349/ymj.2019.60.6.509.

Induction of Melanoma Cell-Selective Apoptosis Using Anti-HER2 Antibody-Conjugated Gold Nanoparticles

Affiliations
  • 1Department of Oral & Maxillofacial Surgery, School of Dentistry, Pusan National University, Yangsan, Korea. kuksjs@pusan.ac.kr
  • 2Department of Oral Anatomy, School of Dentistry, Pusan National University, Yangsan, Korea. ki91000m@pusan.ac.kr
  • 3Feagle Co., Ltd., Yangsan, Korea.

Abstract

PURPOSE
This study was conducted to verify the induction and mechanism of selective apoptosis in G361 melanoma cells using anti-HER2 antibody-conjugated gold nanoparticles (GNP-HER2).
MATERIALS AND METHODS
Following GNP-HER2 treatment of G361 cells, cell cycle arrest and apoptosis were measured by WST-1 assay, Hemacolor staining, Hoechst staining, immunofluorescence staining, fluorescence-activated cell sorting analysis, and Western blotting.
RESULTS
G361 cells treated with GNP-HER2 showed condensation of nuclei, which is an apoptotic phenomenon, and translocation of apoptosis-inducing factor and cytochrome c from mitochondria into the nucleus and cytoplasm, respectively. Increases in BAX in cells undergoing apoptosis, activation of caspase-3 and -9, and fragmentation of poly (ADP-ribose) polymerase and DNA fragmentation factor 45 (inhibitor of caspase-activated DNase) were observed upon GNP-HER2 treatment. Following GNP-HER2 treatment, an increase of cells in sub-G1 phase, which is a signal of cell apoptosis, was observed. This resulted in the down-regulation of cyclin A, cyclin D1, cyclin E, cdk2, cdk4, and cdc2 and the up-regulation of p21. Thus, GNP-HER2 treatment was confirmed to induce the cessation of cell cycle progression. Also, decreases in phospho-focal adhesion kinase and phospho-human epidermal growth factor receptor, which activate cellular focal adhesion, and decreases in phospho-paxillin, which stimulates the disassembly of filamentous actin, were observed. Reduced cell adhesion and disassembly of the intracellular structure indicated cell deactivation.
CONCLUSION
GNP-HER2 can selectively kill G361 melanoma cells without affecting normal cells. The mechanism of G361 cell death upon treatment with GNP-HER2 was apoptosis accompanied by activation of caspases.

Keyword

HER2; melanoma cells; cell cycle arrest; apoptosis; conjugation

MeSH Terms

Actins
Apoptosis Inducing Factor
Apoptosis*
Blotting, Western
Caspase 3
Caspases
Cell Adhesion
Cell Cycle
Cell Cycle Checkpoints
Cell Death
Cyclin A
Cyclin D1
Cyclin E
Cyclins
Cytochromes c
Cytoplasm
DNA Fragmentation
Down-Regulation
Flow Cytometry
Fluorescent Antibody Technique
Focal Adhesions
Melanoma*
Mitochondria
Nanoparticles*
Phosphotransferases
Receptor, Epidermal Growth Factor
Up-Regulation
Actins
Apoptosis Inducing Factor
Caspase 3
Caspases
Cyclin A
Cyclin D1
Cyclin E
Cyclins
Cytochromes c
Phosphotransferases
Receptor, Epidermal Growth Factor
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