Pseudorabies virus for tumor treatment
A pseudorabies virus, tumor technology, applied in the field of virus and tumor treatment
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Embodiment 1
[0147] Example 1: Acquisition and preparation of PRV and its modified form
[0148] 1.1 Isolation of pseudorabies virus PRV from specimens
[0149] (1) The throat swabs and anal swabs of sick pigs are from China, Xiamen City Center for Disease Control and Prevention, pig embryo kidney cells (PK-15; ATCC NumberCCL-33) TM ) It is preserved by Xiamen University, China, National Infectious Disease Diagnostic Reagents and Vaccine Engineering Technology Research Center, and cultured in DMEM medium supplemented with 10% fetal bovine serum, glutamine, penicillin and streptomycin.
[0150] (2) Specimen processing: Fully stir the throat swab and anal swab in the specimen storage solution to wash off the virus and virus-containing cells on the swab, and then put the specimen storage solution at 4000 rpm at 4°C Centrifuge at high speed for 30 minutes;
[0151] (3) Vaccination and observation:
[0152] A. Spread PK-15 cells on a 24-well plate, the number of cells in each well is 1×10 5 Aspirate t...
Embodiment 2
[0165] Example 2: Evaluation of in vitro anti-tumor activity of wild-type PRV
[0166] 2.1 Virus and cell line used
[0167] (1) Virus: This example uses the PRV-WT (SEQ ID NO:1) provided in Example 1.
[0168] (2) Cell line: human rhabdomyosarcoma cell RD( Number:CCL-136 TM ); Human colorectal cancer cell line SW1116( Number:CCL-233 TM ), SW480( Number:CCL-228 TM ) And HT-29 ( Number:HTB-38 TM ); Human gastric cancer cell line AGS ( Number:CRL-1739 TM ), SGC7901 (CCTCC deposit number: GDC150), BGC823 (CCTCC deposit number: GDC151) and NCI-N87 ( Number:CRL-5822 TM ); human esophageal cancer cell line TE-1 (purchased from the Cell Resource Center, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, number: 3131C0001000700089); human small cell lung cancer cell line DMS114 ( Number:CRL-2066 TM ); Human non-small cell lung cancer cell line SPC-A-1 (CCTCC deposit number: GDC050), NCI-H1975 ( Number:CRL-5908 TM ), NCI-H1299( Number:CRL-5803 TM ), A5...
Embodiment 3
[0185] Example 3: In vivo anti-tumor experiment of wild-type PRV
[0186] 3.1 Viruses, cell lines and laboratory animals
[0187] (1) Virus: The PRV-WT provided in Example 1 is used in this example. Refer to Examples 2.2 and 2.3 for virus culture and virus titer determination methods, respectively.
[0188] (2) Cell line: human nasopharyngeal carcinoma cell line CNE1, human Burkitt's lymphoma cell Raji ( Number:CCL-86 TM ), human glioma cell line GBM (primary tumor cell line isolated from patient tumor tissue), mouse colon cancer cell CT26, mouse liver cancer cell Hep1-6, mouse kidney cancer cell Renca and mouse breast cancer cell 4T1 . The above cells except Raji use RPMI-1640 medium, all other cells are cultured in DMEM medium, and the above medium is supplemented with 10% fetal bovine serum, glutamine and penicillin-streptomycin double antibody. All the above cells are at 37℃, 5% CO 2 Culture under standard conditions.
[0189] (3) Experimental animals: Female C.B17SCID mice ...
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