| Background and objective:Interfeon is a natural potent pleiotropic cytokine with multiple biological functions such as anti-virus,anti-tumor and immune regulation.Interferon combines with the interferon receptor on the surface of the cells,and then activates the JAK-STAT signal pathway.With the regulation of the interferon related genes,the cells express the various life activities related proteins for the variable functions.Pancreatic cancer is the fourth leading cause of cancer-associated death in both men and women.Currently,surgery is the main therapy for the early stage patients.For the most pancreatic cancer patients who are diagnosed at the advanced stage,no effective therapeutic regimens are able to significantly ameliorate the progress of the disease.Accumulating evidence shows that IFN?,has antitumor effect and restitutes the chemosensitivity in pancreatic cancer and other solid tumors.However,the potency of IFN? therapy is significantly limited by its systemic toxicity.Long-term parental administration of IFN? is required to maintain therapeutic efficacy,and this often induces high-grade toxicity and significantside effects in many patients.Methods:To potentiate the effect of interferon in cell proliferation,we developed a“c DNA in-frame fragment” library screening technology.We constructed the SIF? plasmids,and then studied the effect in the pancreatic cancer cell proliferation.At last,we explored the underlying mechanism of the SIF?.(1)We screened the IEP by “c DNA in-frame fragment library” and conducted the SIF? recombinant plasmids.Then,the pancreatic cancer cells were transfected with SIF? plasmids by lentivirus vector.We checked the IFN?secretion by the ELISA and the Western blot analysis.(2)We used the SRB to test the cells viability,made the cell cycle analysis,treated the cancer cells combined SIF? with chemotherapeutic drug gemcitabin.(3)Treated pancreatic cancer cells with the secreted SIF? and IFN?.(4)Checked the interferon related genes expression by RT-PCR and q PCR,and detected the cell binding assay.Results:(1)By screening,we identified short c DNA fragments that enhance the activity of IFN?.Interestingly,three peptides contain a short stretch of positively charged amino acids derived from placental growth factor-2(PLGF-2).The IEP had 23 amino acids,the sequence was RRRPKGRGKPPPEKQRPTDCHLC.We combined this novel IEP peptide to the 3' terminal of the IFN?.We detected the transfected cells secreted the SIF? protein,and the IEP did not affect the protein secretion.(2)Compared with IFN?,the synthetic SIF? plasmid inhibited tumor cell growth and blocked cells at the S phase more significantly by the lenti virus vector.At the same time,we found IEP could modulate the effect of the chemotherapeutic drug gemcitabin(GEM)in human pancreatic cancer cells.(3)The secreted SIF? inhibited the pancreatic cancer cells proliferation more obviously.(4)By RT-PCR and qPCR,we found the SIF? group had higher expression of interferon related genes by activating the STAT pathway.And the SIF? enhanced the cell binding to the pancreatic cancer cells.Conclusion:(1)We have identified a short positively charged peptide(IEP)using the“c DNA in-frame” fragment.When fused the IEP to the 3' terminal of the IFN?,we conducted the SIF? plasid.(2)IEP significantly potentiated the antitumor activity of interferon alpha in the proliferation of pancreatic cancer cells.And the synthetic SIF? blocked more cells at the S phase.(3)At the same time,we found IEP could modulate the effect of the chemotherapeutic drug gemcitabin(GEM)in human pancreatic cell lines,by activating the STAT pathway and enhancing the IFN? binding to the cell membrane. |
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