| Colon cancer is one of the most common gastrointestinal cancers,there are about 102 million new cases and 530 thousand deaths each year across the world.In China,the incidence of colon cancer ranked fifth in that of mainly cancers,and has been increasing with a growth rate of 4.2% per year,which was significant higher than the international standard of 2%.Colon cancer has seriously affected the health of our citizens.At present,the surgery supplemented with radiotherapy and chemotherapy is the predominant way to cure colon cancer so far,easily lead to tumor recurrence and make great harm to patents’ body and mind.In recent years,there was a great progress in molecular targeting treatment which opened up a new way for colon cancer treatment.The combination of molecular targeted drugs can prolong the survival time of patients.Molecular targeting treatment is a way to inhibite tumor cells growth by selecting a specific blocker that worked on the molecular which over-expressed on tumor cells.Recent researches found that the quantity of cholecystokinin-2-receptor(CCK2R)on tumors cells especially on some colon cancer cells is significantly higher than that on normal cells,so it can be used as a tumor target.In this sdudy,we contrasted a novel recombinant toxin which fused cholecystokinin(CCK)that could target CCK2 R and Pseudomonas exotoxin(PE)that has strong cell toxicity.The vitro experiment on cells and vivo experiment on model of colon cancer in nude mice indicate that the recombinant toxin has anti-colon cancer ability,which lay a foundation for precilinical experiment,and provide guidance for non-surgery treatment of colon cancer.Researches found that cholecytokinin played an important role in the invasion and metastasis of tumor cells,which was realized by the high affinity to CCK2 R.In this study,the 96th-104 th amino acids of CCK that has the function of targeting CCK2 R were reversed to r CCK96-104 and conjuct to the anmino terminal of PE38,which constructed a novel recombinant toxin,r CCK96-104PE38.A prokaryotic expression vector BL21(DE3)(p ET28a-r CCK96-104PE38)was constructed for the expression of r CCK96-104PE38.The recombinant protein was expressed in soluble form by inducing the E.coil with 0.1 m M IPTG and incubated it for 18 h at 16℃.The recombinant protein was identified by wetern-blotting.The Indirect immunofluorescent assay and flow cytometry analysis indicated that r CCK96-104PE38 could specifically snap to conlon cancer which over-expressed CCK2 R and induced cell apoptosis by internalization;on the contrary,r CCK96-104PE38 could not have a significant impact on cells which hardly expressed CCK2 R.A CCK-8 Kit was used to map the tumor inhibitory curve of r CCK96-104PE38 on colon cancer cell line,HCT 116.The IC50 value of variety of cells was measured,which preliminary comfirmed that the colon cancer targeting of r CCK96-104PE38.A colon cancer,HCT116 model of nude mice was established.By the way of tail vein administration,it was found that the recombinant toxin had high metabolic rate and low toxicity in animals and could inhibit the growth and spread of tumor cells.These results suggested that recombinant toxin,r CCK96-104PE38 was a promising drug for colon cancer and might provide a new approach for the treatment of colon cancer. |
PDF Full Text Request