Silencing of Astrocyte Elevated Gene-1 (AEG-1) inhibits the proliferative and invasive potential through interaction with Exostosin-1 (EXT-1) in primary and metastatic colon cancer cells

Colon cancer is the third major cause of cancer deaths,accounting for about 8%in terms of mortality globally.The present study aims to explore the effect of silencing Astrocyte Elevated Gene-1(AEG-1),a metastasis mediating factor,and how it interacts with Exostosin-1(EXT-1)protein to inhibit the proliferative and invasive potential in colon cancer cells.Forward siRNA transfection was performed using AEG-1 siRNA in SW480 and SW620 colon cancer cell lines,and the expression levels of mRNA and protein were analyzed by Real-time PCR and Immunofluorescence.A simple bioinformatics approach was carried out to identify the possible interactions between AEG-1 and EXT-1 using Easy Networks and Pathway Commons Database.Cell survival and clonal efficiency were determined using Cell Counting Kit-8 assay and clonogenic assay,apoptosis using flow cytometry analysis,migration and invasion using scratch and Transwell assays,respectively.Forward siRNA transfection significantly suppressed the expression of AEG-1 in mRNA and protein levels on SW480 and SW620 colon cancer cells.From our results,we found that EXT-1 mRNA and protein level was significantly upregulated in AEG-1 siRNA transfected cells.Moreover,treatment with AEG-1 siRNA inhibited the proliferation,clonogenic ability,migration,and invasion and also induced apoptosis.Through the bioinformatic approach,our data analyses pointed towards the crosstalk between AEG-1 and EXT-1 mediated through Patched-1(PTCH-1)protein.Our current results demonstrated that silencing AEG-1 can restrain cell proliferation,migration,and invasion,ultimately leading to apoptosis.In AEG-1 siRNA transfected cells,PTCH-1 activity might be modulated by several genes and,in turn,affects the EXT-1 activity.Collectively,these observations not only provide insight into the interplay between AEG-1 and EXT-1 but also suggest that AEG-1 may represent a possible candidate therapeutic target through interaction with EXT-1 in colon cancer.