Identification and optimization of small molecules that restore E-cadherin expression and reduce invasion in colorectal carcinoma cells.
E-cadherin is a transmembrane protein that maintains intercellular contacts and cell polarity in epithelial tissue. The down-regulation of E-cadherin contributes to the induction of the epithelial-to-mesenchymal transition (EMT), resulting in an increased potential for cellular invasion of surrounding tissues and entry into the bloodstream. Loss of E-cadherin has been observed in a variety of human tumors as a result of somatic mutations, chromosomal deletions, silencing of the CDH1 gene promoter, and proteolytic cleavage. To date, no compounds directly targeting E-cadherin restoration have been developed. Here, we report the development and use of a novel high-throughput immunofluorescent screen to discover lead compounds that restore E-cadherin expression in the SW620 colon adenocarcinoma cell line. We confirmed restoration of E-cadherin using immunofluorescent microscopy and were able to determine the EC(50) for selected compounds using an optimized In-Cell Western assay. The profiled compounds were also shown to have a minimal effect on cell proliferation but did decrease cellular invasion. We have also conducted preliminary investigations to elucidate a discrete molecular target to account for the phenotypic behavior of these small molecules and have noted a modest increase in E-cadherin mRNA transcripts, and RNA-Seq analysis demonstrated that potent analogues elicited a 10-fold increase in CDH1 (E-cadherin) gene expression.
Department of Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37215, USA.
Sydney L
SL
Stoops
A Scott
AS
Pearson
Connie
C
Weaver
Alex G
AG
Waterson
Emily
E
Days
Chris
C
Farmer
Suzanne
S
Brady
C David
CD
Weaver
R Daniel
RD
Beauchamp
Craig W
CW
Lindsley
D015820:Cadherins / Q000096:biosynthesis* / Q000235:genetics; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000378:metabolism* / Q000473:pathology*; D004353:Drug Evaluation, Preclinical / Q000379:methods; D058750:Epithelial-Mesenchymal Transition; D006801:Humans; D009361:Neoplasm Invasiveness / Q000517:prevention & control*; D012333:RNA, Messenger / Q000378:metabolism
D015820:Cadherins; D012333:RNA, Messenger
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