Role of oncogenic K-Ras in cancer stem cell activation by aberrant Wnt/β-catenin signaling.
BACKGROUND
Adenomatous polyposis coli (APC) loss-of-function mutations and K-Ras gain-of-function mutations are common abnormalities that occur during the initiation and intermediate adenoma stages of colorectal tumorigenesis, respectively. However, little is known about the role these mutations play in cancer stem cells (CSCs) associated with colorectal cancer (CRC) tumorigenesis.


METHODS
We analyzed tissue from CRC patients (n = 49) to determine whether K-Ras mutations contributed to CSC activation during colorectal tumorigenesis. DLD-1-K-Ras-WT and DLD-1-K-Ras-MT cells were cultured and evaluated for their ability to differentiate, form spheroids in vitro, and form tumors in vivo. Interaction between APC and K-Ras mutations in colorectal tumorigenesis was evaluated using APC (Min/+)/K-Ras (LA2) mice and DLD-1-K-Ras-WT and DLD-1-K-Ras-MT cell xenografts. (n = 4) Group differences were determined by Student t test. All statistical tests were two-sided.


RESULTS
The sphere-forming capability of DLD-1-K-Ras-MT cells was statistically significantly higher than that of DLD-1-K-Ras-WT cells (DLD-1-K-Ras-MT mean = 86.661 pixel, 95% confidence interval [CI] = 81.701 to 91.621 pixel; DLD-1-K-Ras-WT mean = 42.367 pixel, 95% CI = 36.467 to 48.267 pixel; P = .003). Moreover, both the size and weight of tumors from DLD-1-K-Ras-MT xenografts were markedly increased compared with tumors from DLD-1-K-Ras-WT cells. Expression of the CSC markers CD44, CD133, and CD166 was induced in intestinal tumors from APC (Min/+)/K-Ras (LA2)mice, but not K-Ras (LA2) mice, indicating that APC mutation is required for CSC activation by oncogenic K-Ras mutation.


CONCLUSIONS
K-Ras mutation activates CSCs, contributing to colorectal tumorigenesis and metastasis in CRC cells harboring APC mutations. Initial activation of β-catenin by APC loss and further enhancement through K-Ras mutation induces CD44, CD133, and CD166 expression.
Affiliations of authors: Department of Biotechnology (B-SM, W-JJ, JP, K-YC), Translational Research Center for Protein Function Control (B-SM, W-JJ, JP, DSM, K-YC), and Department of Internal Medicine and Institute of Gastroenterology, College of Medicine (TIK), Yonsei University, Seoul, Korea; Department of Molecular Biology, College of Natural Science, Pusan National University, Pusan, Korea (DSM).
Byoung-San
BS
Moon
Woo-Jeong
WJ
Jeong
Jieun
J
Park
Tae Il
TI
Kim
Do Sik
do S
Min
Kang-Yell
KY
Choi
D000071916:AC133 Antigen; D011125:Adenomatous Polyposis Coli / Q000378:metabolism; D025601:Adenomatous Polyposis Coli Protein; D000328:Adult; D000368:Aged; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D015816:Cell Adhesion Molecules, Neuronal / Q000378:metabolism; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism*; D015179:Colorectal Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D005260:Female; D005326:Fetal Proteins / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D011905:Genes, ras; D006023:Glycoproteins / Q000378:metabolism; D064593:Heterografts; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D008297:Male; D051379:Mice; D008875:Middle Aged; D009154:Mutation; D014411:Neoplastic Stem Cells / Q000378:metabolism* / Q000473:pathology; D010455:Peptides / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D056910:Republic of Korea; D015398:Signal Transduction; D051153:Wnt Proteins / Q000378:metabolism*; D051176:beta Catenin / Q000378:metabolism*; D018631:ras Proteins / Q000235:genetics*; D020410:Activated-Leukocyte Cell Adhesion Molecule
D000071916:AC133 Antigen; D025601:Adenomatous Polyposis Coli Protein; D015703:Antigens, CD; D015816:Cell Adhesion Molecules, Neuronal; D005326:Fetal Proteins; D006023:Glycoproteins; D018960:Hyaluronan Receptors; D010455:Peptides; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D051153:Wnt Proteins; D051176:beta Catenin; D018631:ras Proteins; C105613:ALCAM protein, human; D020410:Activated-Leukocyte Cell Adhesion Molecule; C555337:APC protein, human; C497483:CD44 protein, human; C495270:CTNNB1 protein, human; C117307:KRAS protein, human; C000606023:PROM1 protein, human; C000606024:Prom1 protein, mouse