ABSTRACT
Objective: To establish an animal model of human lung cancer in NOD/SCID mice and its cell line with highly metastatic potential and observe their correlated biological features in metastasis so as to provide an important useful tool for studying the metastatic mechanism, diagnosis, prevention and therapy of lung cancer. Methods: Human lung cancer cells SPC-A-1 were inoculated subcutaneously into NOD/SCID mice, metastatic tumor masses in lung were harvested from curative resection of subcutaneous tumor, and re-implanted into NOD/SCID mice for the second round of in vivo selection. Besides resection of subcutaneous tumor, the same manipulation was triplicately repeated. Eventually, we established a highly metastatic animal model of human lung cancer in NOD/SCID mice and its cell line with highly metastatic behavior. The cell growth behavior, metastasis, morphological characteristics of the implanted tumors were observed by light microscopy. The biological features of the highly metastatic cell line were investigated. Results: The lung metastasis rate of SPC-A-1 cells was 66. 7% after curative resection of subcutaneous tumor in the first round in vivo screening. An animal model of human lung cancer with 100% lung metastasis and its cell line with unique metastatic characteristics were established by in vivo 4-round repeated screening. The cell growth behavior and karyotype analysis showed that SPC-A-1 cell line maintained the biological characteristics of primary human lung adenocarcinoma. Conclusion: A highly metastatic animal model of subcutaneous implanted human lung cancer and its cell line are successfully established by in vivo screening. Our study provides an ideal animal model for research of prevention of lung cancer and experimental therapy against metastasis.
ABSTRACT
Objective:To obtain phage-displayed ScFv library targeting tumor tissues and to screen for antibodies specifically binding to tumor vessels using in vivo phage display,so as to lay a foundation for diagnosis and treatment of cancer.Methods:The membrane proteins were extracted from the specimens of esophageal carcinoma,stomach carcinoma,brain cancer,lung cancer,and spinal cord tumor.The recombinant phage-antibody system was used to construct a single-chain Fv fragment(ScFv)cDNA library from the total RNA of the BALB/c mice immunized with purified membrane protein.The specific primers of VH and VL were used to amplify the cDNA of VH and VL,respectively,which were then assembled into ScFv gene with a specially constructed linker DNA.The ScFv gene was ligated into the phagemid vector pCANTAB 5E and the ligated samples were transformed into competent E.coli TG1.The transformed cells were infected with M13KO7 helper phage to yield recombinant phage.Using the animal model of human cervical carcinoma(HeLa cells),sepecific phage-ScFvs were selected by phage displaying and panning in vivo.After four rounds,24 phage-ScFvs,which were identified by PCR,were analyzed immunohistochemically.The ScFvs expressed in the tumor tissue slices and negative in control kidney tissue slices were sequenced.Results:Tomors-bearing animal models were established with 7 different kinds of carcinoma cell lines in BALB/c nude mice.It was found that inoculation with HeLa cells resulted in most satisfactory tumorigenesis in nude mice.A ScFv library of 1.6?106 was obtained and a tumor vessel specific phage-ScFv named ScFvH1(VH-linker-VL)was selected from the library.Conclusion:A tumor targeting ScFv library has been successfully constructed and a tumor vessel-specifrc antibody has been identified from the library,which provides a new way for the early diagnosis and therapy of cancer.