Polypeptide capable of inducing cytotoxic T lymphocytes and application thereof
A technology of lymphocytes and induced cells, which is applied to polypeptides with the ability to induce cytotoxic T lymphocytes and its application fields, which can solve the problems of gene variation in the ligand binding domain, etc., to reduce the recurrence rate, improve the effect of immune response, and improve the cure rate effect
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Embodiment 1
[0069] A screening method for a polypeptide having the ability to induce cytotoxic T lymphocytes, comprising the steps of:
[0070] (a). A conventional genetic engineering peptide production method was used to design and synthesize 39 polypeptides, including polypeptides numbered A1-A37, negative control group polypeptides, and positive control group polypeptides.
[0071] (b). Spread T&B hybrid cells (Shanghai Heyuan Biological Co., Ltd.) on 0.5×10 6 100 μmol / L of the above polypeptides were respectively added to the 24-well plate of the cells, and cultured in an incubator at 37° C. and 5% carbon dioxide for 4 hours.
[0072] (c). The cultured cells were collected, washed with physiological saline, and stained with an anti-human histocompatibility (MHC) class I antibody for 30 minutes. No polypeptide was added as a negative control, and a known positive reaction polypeptide (YLLPAIVHI, cited in Cytometry 41:271-278, 2000) was set as a positive control. After staining, wash ...
Embodiment 2
[0076] Anticancer activity experiment of eight polypeptide-specific CD8 T killer cells whose amino acid sequences are SEQ ID NO.1~SEQ ID NO.8
[0077] (1). Dendritic cells expressing CD14 and CD11c were enriched from peripheral blood cells of healthy people
[0078] Human peripheral blood cells were placed in an incubator at 37°C for 1-2 hours. Adherent cells were used as dendritic cell precursors, and suspension cells were used as T cell aggregates. Interleukin-7 (IL7) and interleukin-2 ( IL2) and interleukin-15 (IL15) stimulation for 5d.
[0079] (2). Polypeptide combinations stimulate the acquisition of effector cells
[0080] After five days, the above cells were mixed and respectively treated with single polypeptide SEQ ID No.1, single polypeptide SEQ ID No.3, single polypeptide SEQ ID No.5, double peptide combination (ie SEQ ID No.1 and SEQ ID No.3, or SEQ ID No.1 and SEQ ID No.5), polypeptide combination 1 (ie the combination of SEQ ID No.1, SEQ ID No.3 and SEQ ID No....
Embodiment 3
[0090] Experiment of Anticancer Effect of External Tumor Animal Model
[0091] (1). SCID female mice (provided by Shanghai Heyuan Biological Co., Ltd.) with double deletion of immune cells T&B cells at the age of 8 weeks, and the mammary glands were treated with 10 6 The cells were inoculated with estrogen receptor-positive breast cancer cells MCF7, which were pre-labeled with biofluorescein. After culturing for about 1 week to 10 days, there is a palpable tumor in the mammary gland, and the initial tumor size is recorded with a bioluminescent instrument.
[0092] (2). Polypeptide combination 1 (ie the combination of SEQ ID No.1, SEQ ID No.3 and SEQ ID No.5), polypeptide combination 2 (ie SEQ ID No.2, SEQ ID No. .4 and the combination of SEQ ID No.7), polypeptide combination 3 (that is, the combination of SEQ ID No.6 and SEQ ID No.8) is administered by intraperitoneal injection (IP) twice a week at a concentration of 100 μmol / L, and the duration of administration is For four...
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