Domain 5 of kininogen inhibits proliferation of human colon cancer cell line (HCT-116) by interfering with G1/S in the cell cycle.
BACKGROUND
Domain 5 (D5) of kininogen inhibits endothelial cell adhesion, migration, proliferation and angiogenesis by inducing apoptosis and disrupting a signaling pathway initiated by binding to the urokinase receptor (uPAR).


OBJECTIVE
Because tumor cells frequently overexpress uPAR, we hypothesized that D5 can directly inhibit proliferation of colon carcinoma cells.


RESULTS
A recombinant fusion protein of D5 and glutathione S-transferase (GST-D5) but not GST at 280 nm inhibited proliferation of human colon carcinoma cells (HCT-116) in vitro by 75-86%. We found that treatment with GST-D5 did not affect the survival pathway, phosphatidylinositol 3-kinase or the apoptotic pathway. In contrast, the G1/S phase transition of the cell cycle was downregulated as evidenced by an increase of cells in G0/G1 and a decrease in cells in S by flow cytometry. We found a decrease in serine phosphorylation of the retinoblastoma protein Rb (p107) after incubation with GST-D5. Less E2F-1 transcription factor and p107 were released and fewer cells overcame the G1/S growth restriction point. Expression levels of cyclins D1, A and E were reduced as measured by densiometric analysis of western blots. Cyclin-dependent protein kinase activities were downregulated and p27, the cyclin-dependent kinase inhibitor, was activated by GST-D5.


CONCLUSIONS
These findings indicate that D5 of high molecular weight kininogen interferes with the G1 to S phase transition, reducing the proliferation of human colon carcinoma cells.
Bior
A D
AD
The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA 19140, USA.
Pixley
R A
RA
Colman
R W
RW
D002453:Cell Cycle / Q000187:drug effects; D018797:Cell Cycle Proteins / Q000032:analysis; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000473:pathology*; D016193:G1 Phase; D006801:Humans; D007399:Interphase / Q000187:drug effects*; D007704:Kininogens / Q000737:chemistry / Q000502:physiology*; D010446:Peptide Fragments / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D016160:Retinoblastoma Protein / Q000378:metabolism; D016196:S Phase
D018797:Cell Cycle Proteins; D007704:Kininogens; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D016160:Retinoblastoma Protein