Cyclooxygenase-2 is a target of KRASD12, which facilitates the outgrowth of murine C26 colorectal liver metastases.
OBJECTIVE
Mutational activation of the KRAS oncogene and overexpression of cyclooxygenase-2 (COX-2) contribute to colorectal carcinoma (CRC) development, but the relationship between these two events is unclear. This study was designed to clarify that relationship and to assess the contribution of KRAS-dependent COX-2 to the seeding of CRC cells in the liver and to their outgrowth as liver metastases in an experimental mouse model.


METHODS
The effect of RNA interference-mediated KRAS knockdown on COX-2 expression and activity was tested in murine C26 CRC cells. The contribution of KRAS-dependent COX-2 to early metastatic tumor cell seeding (by intravital microscopy) and outgrowth of metastases in the liver (by bioluminescence imaging) was studied by using parecoxib, a novel and highly selective liver-activated COX-2 inhibitor. Intratumoral cell proliferation, apoptosis, and tumor-associated angiogenesis were assessed by immunohistochemistry on liver tissue sections.


RESULTS
Stable knockdown of mutant KRAS(D12) in murine C26 CRC cells by RNA interference lead to a dramatic reduction of COX-2 synthesis and prostaglandin E2 production. Inhibition of host or tumor cell COX-2 activity had no effect on early metastatic cell seeding in the liver but greatly reduced intrahepatic tumor cell proliferation and the rate of liver metastasis outgrowth. COX-2 inhibition had no effect on early tumor vascularization or on tumor cell apoptosis.


CONCLUSIONS
The high levels of COX-2 enzyme and prostaglandin production in C26 CRC cells are primarily caused by the presence of endogenous mutant KRAS(D12). Furthermore, COX-2 inhibition affects the tumoral rather than the vascular compartment during the early stages of C26 liver metastasis outgrowth.
Smakman
Niels
N
Department of Surgery, University Medical Center Utrecht, Utrecht, The Netherlands.
Kranenburg
Onno
O
Vogten
Jan M
JM
Bloemendaal
Alexander L A
AL
van Diest
Paul
P
Borel Rinkes
Inne H M
IH
D000818:Animals; D017209:Apoptosis; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D015179:Colorectal Neoplasms / Q000473:pathology*; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D015232:Dinoprostone / Q000378:metabolism; D004195:Disease Models, Animal; D011905:Genes, ras / Q000235:genetics*; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D007555:Isoxazoles / Q000494:pharmacology; D019394:Ki-67 Antigen / Q000096:biosynthesis; D008099:Liver / Q000378:metabolism; D008113:Liver Neoplasms / Q000556:secondary*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008833:Microcirculation; D009154:Mutation; D009362:Neoplasm Metastasis; D009368:Neoplasm Transplantation; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D034622:RNA Interference; D013997:Time Factors
D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D007555:Isoxazoles; D019394:Ki-67 Antigen; C409945:parecoxib; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; D016283:Proto-Oncogene Proteins p21(ras); D015232:Dinoprostone