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Tumor cell vaccine targeting CAFs, and preparation method and application thereof

A tumor cell vaccine, tumor cell technology, applied in the direction of tumor/cancer cells, botanical equipment and methods, biochemical equipment and methods, etc., can solve the problem that tumor vaccines are not suitable for mass production and transformation applications, and tumor cells express antigens that are not stable enough , unfavorable tumor immune activation and other problems, to achieve the effect of enhancing radiosensitivity, improving fibrosis, and good effect

Active Publication Date: 2021-04-06
SICHUAN CANCER HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the vaccine prepared by this method can effectively slow down tumor growth in the early stage of treatment, the tumor growth is accelerated in the later stage. The possible reason is that the antigen expressed by tumor cells after transient transfection is not stable enough, which is not conducive to the activation of anti-tumor immunity. The obtained tumor vaccine is also not suitable for mass production and transformation application
In addition, the effect of vaccine immunotherapy alone is limited, so it is necessary to further improve this

Method used

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  • Tumor cell vaccine targeting CAFs, and preparation method and application thereof
  • Tumor cell vaccine targeting CAFs, and preparation method and application thereof
  • Tumor cell vaccine targeting CAFs, and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] A method for preparing a tumor cell vaccine targeting CAFS, comprising the following steps:

[0055] The NCBI gene number of FAP (fibroblast activation protein) is NM_007986.3, and the length of its coding region is 2285bp; PCR amplification is carried out with FAP cDNA as a template: the primer sequences used are shown in Table 1 below:

[0056] Table 1 Primer sequences used in PCR:

[0057]

[0058] Mix the following solutions in a PCR tube:

[0059]

[0060] The negative control is corresponding to no template; PCR reaction conditions: 98°C, 10sec, 55°C, 5sec, 72°C, 30sec, a total of 35 cycles, then 72°C for 2min, 16°C for 2min;

[0061] Through the above PCR reaction, amplified fragments with consistent band sizes were successfully amplified, such as figure 1 As shown in band 2 in the middle, this PCR amplified fragment was seamlessly cloned and connected to the lentiviral vector pCDH digested with EcoRI and BamHI to construct the lentiviral vector pCDH-FA c...

Embodiment 2

[0065] The inventor's previous research found that a tumor cell vaccine expressing human FAPα can inhibit tumor growth and improve tumor interstitial fibrosis, but the efficacy of the vaccine alone is limited. Studies have shown that SABR combined with tumor-specific vaccines will produce an in situ vaccine effect and further improve the body's anti-tumor immune response. The therapeutic effect is affected by factors such as radiotherapy regimens and the timing of combined treatment.

[0066] This example established a mouse LLC lung cancer model, compared the anti-tumor effects of different SABR regimens (single dose of 16.4Gy×1 time or single dose of 8Gy×3 times) and different treatment sequences, and observed tumor-bearing mice Tolerance to treatment.

[0067] Experimental animal grouping: 6 days after tumor inoculation, 48 mice were randomly divided into 8 groups, 6 in each group, as follows: Figure 6 as shown,

[0068] The specific method is as follows: female C57 / bl m...

Embodiment 3

[0071] In this example, mouse 4T1 breast cancer and CT26 colon cancer subcutaneous tumor models were established, and it was further clarified that the therapeutic effect of tumor cell vaccine expressing human FAP combined with SABR was superior to radiotherapy or vaccine immunotherapy alone.

[0072] Female Balb / c mice aged 6-8 weeks were selected and subcutaneously inoculated with 4×10 5 / 100ml 4T1 breast cancer or CT26 colon cancer cells. On the 8th day, 30 mice were randomly divided into 5 groups, 6 in each group, specifically:

[0073] Untreated group: control group

[0074] SABR group: start ablative radiotherapy on the 10th day 8Gy×3 times

[0075] Vaccine group: Immunotherapy with subcutaneous injection of tumor cell vaccine expressing human FAP on days 7, 14, and 21

[0076] SABR+pCDH group: On the 10th day, ablation radiotherapy 8Gy×3 times was started, and on the 7th, 14th, and 21st days, the tumor cells infected with the inactivated pCDH empty vector were used f...

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Abstract

The invention discloses a preparation method of a tumor cell vaccine targeting CAFs, and belongs to the technical field of biological medicines, the method is as follows: lentiviral packaging plasmids are adopted to infect tumor cells to obtain a stable expression strain, and then the stable expression strain is inactivated to obtain the tumor cell vaccine targeting CAFs; the invention also discloses application of the vaccine prepared by the method. On the basis of a brand-new preparation method, the prepared vaccine can more stably express an antigen, the immunotherapy response rate is higher, the effect is better and more stable, and the obtained vaccine is a broad-spectrum vaccine and can treat more than 90% of tumors from epithelium, part of soft tissue tumors and melanoma; besides, the vaccine can also enhance the radiosensitivity of tumors, can achieve a better effect when used in combination with radiotherapy, especially ablation radiotherapy, and can improve fibrosis caused by radiotherapy and reduce toxic and side effects of radiotherapy.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a tumor cell vaccine targeting CAFs, its preparation method and its application. Background technique [0002] For a long time, the focus of cancer research has been mainly on the tumor cells themselves. With the deepening of research, it has been discovered that stromal cells in the tumor microenvironment (TME) not only provide "fertile soil" for tumor occurrence, development, invasion, and metastasis, but also help tumor cells escape immune surveillance. More and more studies have shown that, like tumor cells, stromal cells in the TME have also become important targets for tumor therapy. [0003] Cancer associated fibroblasts (cancer associated fibroblasts; CAFs or CAFs) are key components of the TME. In different tumors, CAFs constitute 5-90% of the TME, and in breast and pancreatic cancers, 80% of the tumor volume is composed of CAFs and the connective tissue they drive...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/00A61K39/39A61K41/00C12N5/10C12N15/867A61P35/00
CPCA61K39/0011A61K39/39A61P35/00A61K41/0038C07K14/705C12N5/0693C12N15/86A61K2039/53C12N2740/10043
Inventor 陈梅华郎锦义谢继锐
Owner SICHUAN CANCER HOSPITAL
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