Effect Of Adeno-associated Virus Mediated ENO1 Human-Mouse Chimeric Antibody On Cervical Cancer HeLa Cells

Objective We constructed a recombinant adeno-associated virus(AAV)encoding the light and heavy chain variable region gene of mouse derived monoclonal antibody ENO1 and the human Ig G1 light and heavy chain constant region gene,and investigated the effect of recombinant adeno-associated virus mediated enolase 1(ENO1)human mouse chimeric antibody on the proliferation of cervical cancer He La cells.Methods 1.The recombinant plasmid r AAV-ENO1-ximab was constructed by PCR,p AAV2-MCS vector was selected,and the variable region gene of the light and weight chain of murine monoclonal antibody ENO1 was linked to the constant region gene of the light and weight chain of human Ig G1 by FMDV 2A sequence.2.On the basis of p Helper and p AAV-RC,recombinant plasmid r AAV-ENO1-ximab was transfected into AAV293 cells using an AAV vector system to prepare recombinant r AAV-ENO1-ximab virus.At the same time,the control group r AAV-GFP was prepared to express green fluorescent protein.3.The adeno-associated virus was isolated,concentrated and purified by PEG method,and the titer of the recombinant virus was determined according to its fluorescence expression.4.Cultured human cervical cancer He La cells,MTT method was used to detect the effect of r AAV-ENO1-ximab on cervical cancer cells in vitro.Result 1.The plasmid p AAV-ENO1-ximab was successfully constructed,and the target gene band was correct for PCR detection,enzyme digestion identification and sequencing.2.Recombinant plasmids r AAV-ENO1-ximab and r AAV-GFP were transfected into AAV293cells,and the transfection efficiency was 90%.3.The titers of recombinant viruses r AAV-ENO1-ximab and r AAV-GFP were 4.0×10~7PFU/ml after purification and concentration.4.MTT assay showed that recombinant virus r AAV-ENO1-ximab could significantly inhibit the proliferation of cervical cancer He La cells,and the difference was statistically significant compared with the negative control group(r AAV-GFP)and the blank control group(control)(P<0.001).Conclusion 1.Recombinant adeno-associated virus r AAV-ENO1-ximab was successfully constructed to mediate ENO1 human and mouse chimeric antibody,which could efficiently and targeted mediate ENO1 human and mouse chimeric antibody gene into the cytoplasm;2.Recombinant adeno-associated virus r AAV-ENO1-ximab can significantly inhibit the proliferation of He La cells of cervical cancer,providing new ideas for the treatment of cervical cancer.