Monitoring of changes in lipid profiles during PLK1 knockdown in cancer cells using DESI MS.
The importance of the polo-like kinase 1 (PLK1) gene is increasing substantially both as a biomarker and as a target for highly specific cancer therapy. This is due to its involvement in multiple points of cell progression and carcinogenesis. PLK1 inhibitors' efficacy in treating human cancers has been limited due to the lack of a specific targeting strategy. Here, we describe a method of targeted downregulation of PLK1 in cancer cells and the concomitant rapid detection of surface lipidomic perturbations using desorption electrospray ionization mass spectrometry (DESI MS). The efficient delivery of siRNA targeting PLK1 gene selectively to the cancer cells is achieved by targeting overexpressed cell surface epithelial cell adhesion molecule (EpCAM) by the EpDT3 aptamer. The chimeric aptamer (EpDT3-siPLK1) showed the knockdown of PLK1 gene expression and PLK1 protein levels by quantitative PCR and western blotting, respectively. The abundant surface lipids, phosphatidylcholines (PCs), such as PC(32:1) (m/z 754.6), PC(34:1) (m/z 782.6), and PC(36:2) (m/z 808.6), were highly expressed in MCF-7 and WERI-RB1 cancer cells compared to normal MIO-M1 cells and they were observed using DESI MS. These overexpressed cell surface lipids in the cancer cells were downregulated upon the treatment of EpDT3-siPLK1 chimera indicating a novel role of PLK1 to regulate surface lipid expression in addition to the efficient selective cancer targeting ability. Our results indicate that DESI MS has a potential ability to rapidly monitor aptamer-mediated cancer therapy and accelerate the drug discovery process. Graphical abstract Binding of aptamer chimera to the cells and changes in lipid profile.
Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, 600006, India.|Centre for Biotechnology, Anna University, Chennai, Tamil Nadu, 600025, India.
Balasubramanyam
B
Jayashree
DST Unit of Nanoscience (DST UNS) and Thematic Unit of Excellence (TUE), Department of Chemistry, Indian Institute of Technology Madras, Chennai, Tamil Nadu, 600036, India.
Amitava
A
Srimany
Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, 600006, India.
Srinidhi
S
Jayaraman
Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, 600006, India.
Anjali
A
Bhutra
Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, 600006, India.
Narayanan
N
Janakiraman
Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, 600006, India.
Srujana
S
Chitipothu
Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, 600006, India.
Subramanian
S
Krishnakumar
Centre for Biotechnology, Anna University, Chennai, Tamil Nadu, 600025, India.
Lakshmi Subhadra
LS
Baddireddi
Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, 600006, India. sailaja.elchuri@gmail.com.
Sailaja
S
Elchuri
DST Unit of Nanoscience (DST UNS) and Thematic Unit of Excellence (TUE), Department of Chemistry, Indian Institute of Technology Madras, Chennai, Tamil Nadu, 600036, India. pradeep@iitm.ac.in.
Thalappil
T
Pradeep
D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006801:Humans; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry*; D061986:MCF-7 Cells; D009374:Neoplasms, Experimental / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D000097869:Polo-Like Kinase 1
D018797:Cell Cycle Proteins; D008055:Lipids; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D000097869:Polo-Like Kinase 1
Aptamer; Cancer; DESI MS; Lipid; siPLK1