Analysis of Protein-Protein Interactions Identifies NECTIN2 as a Target of N,N-Bis (5-Ethyl-2-hydroxybenzyl) Methylamine for Inhibition of Lung Cancer Metastasis

Sunisa Thongsom; Nithikoon Aksorn; Korrakod Petsri; Sittiruk Roytrakul; Nicharat Sriratanasak; Worawat Wattanathana; Pithi Chanvorachote

doi:10.21873/cgp.20347

Analysis of Protein-Protein Interactions Identifies NECTIN2 as a Target of N,N-Bis (5-Ethyl-2-hydroxybenzyl) Methylamine for Inhibition of Lung Cancer Metastasis

Cancer Genomics Proteomics. 2022 Sep-Oct;19(5):624-635. doi: 10.21873/cgp.20347.

Authors

Sunisa Thongsom^{1

2}, Nithikoon Aksorn³, Korrakod Petsri^{1

2}, Sittiruk Roytrakul⁴, Nicharat Sriratanasak^{1

2}, Worawat Wattanathana⁵, Pithi Chanvorachote^{6

2}

Affiliations

¹ Center of Excellence in Cancer Cell and Molecular Biology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, Thailand.
² Department of Pharmacology and Physiology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, Thailand.
³ Department of Clinical Pathology, Faculty of Medicine Vajira Hospital, Navamindradhiraj University, Bangkok, Thailand.
⁴ Functional Ingredients and Food Innovation Research Group, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Pathumthani, Thailand.
⁵ Department of Materials Engineering, Faculty of Engineering, Kasetsart University, Bangkok, Thailand.
⁶ Center of Excellence in Cancer Cell and Molecular Biology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, Thailand; pithi.c@chula.ac.th pithi_chan@yahoo.com.

Abstract

Background/aim: Metastasis negatively affects the survival of lung cancer patients, however, relatively few compounds have potential in metastasis suppression. This study investigated the molecular targets of N,N-bis (5-ethyl-2-hydroxybenzyl) methylamine (EMD) for metastatic inhibition.

Materials and methods: Proteins were analyzed by proteomic and bioinformatic analyses. Protein-protein interaction (PPI) networks were created with the Search Tool for the Retrieval of Interacting Genes. The Kyoto Encyclopedia of Genes and Genomes database and hub genes were used to determine dominant pathways. Immunofluorescence and western blot analyses validated the proteomic results and investigated signaling pathways in NCI-H23 lung cancer cells.

Results: A total of 1,751 proteins were common to the control, EMD and N,N-bis(5-methoxy-2-hydroxybenzyl) methylamine (MeMD) groups; 1,980 different proteins were categorized using metastatic capacity category and analyzed for unique proteins affected by EMD. Fifteen proteins were associated with cell adhesion and six with cell migration. Nectin cell adhesion molecule 2 (NECTIN2) was expressed in the control and MeMD-treated groups but not the EMD-treated group, suggesting NECTIN2 as an EMD target. PPI network showed association of NECTIN2 with proteins regulating cancer metastasis. Kyoto Encyclopedia of Genes and Genomes pathways revealed that NECTIN2 is an upstream target of cytoskeletal regulation via SRC signaling. Western blot and immunofluorescence analyses confirmed that EMD suppressed NECTIN2, and its downstream targets, including p-SRC (Y146 and Y527) and the epithelial-to-mesenchymal transition markers tight junction protein 1, vimentin, β-catenin, snail family transcriptional repressor 1 (SNAI1), and SNAI2, while increasing E-cadherin.

Conclusion: EMD suppressed NECTIN2-induced activation of EMT signaling. These data support the development of EMD to prevent metastasis of lung cancer.

Keywords: Lung cancer metastasis; NECTIN2; PPI; networks; protein–protein interaction; proteomics.

MeSH terms

Cell Line, Tumor
Cell Movement
Epithelial-Mesenchymal Transition
Humans
Lung Neoplasms* / drug therapy
Lung Neoplasms* / genetics
Methylamines / pharmacology
Nectins* / drug effects
Nectins* / metabolism
Proteomics

Substances

Methylamines
Nectins
N,N-bis(5-ethyl-2-hydroxybenzyl)methylamine