Engineered DC cells and methods for promoting TNF-alpha cytokine production by helper T cells

An engineering, alpha cell technology, applied in the fields of immunology and medicine, can solve the problems of improving the effect of immunotherapy, failing to obtain clinical effect, and prolonging the survival period of patients.

Active Publication Date: 2020-08-21
宜兴辰功新药开发有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Recent studies have found that DC vaccines can significantly prolong the survival of patients, but the use of DC vaccines alone usually does not lead to the expected improvement in the effect of immunotherapy, and cannot obtain satisfactory clinical results
Current clinical trials show that the response rate of DC therapeutic vaccines rarely exceeds 15%, and the overall response rate is low

Method used

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  • Engineered DC cells and methods for promoting TNF-alpha cytokine production by helper T cells
  • Engineered DC cells and methods for promoting TNF-alpha cytokine production by helper T cells
  • Engineered DC cells and methods for promoting TNF-alpha cytokine production by helper T cells

Examples

Experimental program
Comparison scheme
Effect test

preparation example 1

[0049] This preparation is used to prepare DNA and mRNA encoding antigens and precursors of different interleukin complexes

[0050] 1. Preparation of DNA and mRNA Constructs

[0051] DNA sequences for producing IL12p70 and IL15 / IL15Rα mRNA were constructed respectively, and used for subsequent in vitro transcription reactions. Following the coding sequence is a polyadenosine segment. The DNA sequence information is shown in Table 1 below.

[0052] In addition, the coding sequences of human tumor antigens AKR1B10 and GPC3 for in vitro sensitization were constructed. The sequences of AKR1B10 and GPC3 are available through the Genebank database. In this example, the antigen disclosed in CN107583042A was used.

[0053] Table-1 DNA sequence list

[0054] name serial number IL-15\IL-15Rα SEQ ID No.2 IL-12 SEQ ID No.5

[0055] 2. In vitro transcription

[0056] Firstly, the prepared corresponding DNA plasmid was linearized using restriction endon...

Embodiment 1

[0058] This example is used to study the effect of the composition of the present invention on T cell response.

[0059] 1. Induction culture of DC cells in vitro

[0060] Aseptically extract 50ml of venous blood from patients with hepatocellular carcinoma, separate peripheral blood mononuclear cells with lymphocyte separation medium in an ultra-clean workbench, add mononuclear cells to AIM-V medium, and place them in 37°C, 5% CO 2 Incubate in an incubator to allow monocytes to adhere to the wall. After 2h, the non-adherent cells were removed, and the adherent cells were added to iDC medium (GM-CSF with a final concentration of 800U / mL and IL-4 at 500U / mL were added to the AIM-V medium), and placed at 37°C for 5 %CO 2 Cultured in the incubator for 6 days. Transfer half of the cell culture medium to a centrifuge tube, collect the cells by centrifugation at 500g, remove the supernatant, and add an equal volume of fresh mDC medium (configuration of fresh medium for mDC: add AIM-...

Embodiment 2

[0080] In this implementation, experiments similar to those in the examples were carried out using the GPC3 antigen. In this embodiment, the mRNA combinations used are as follows:

[0081] 1) Control without any mRNA (mDC control group)

[0082] 2) Add only mRNA encoding GPC3 antigen (GPC3 control group)

[0083] 3) mRNA encoding GPC3 antigen and IL-12p70 mRNA (IL12 group)

[0084] 4) mRNA encoding GPC3 antigen and IL15 / IL15Rα mRNA (IL15 group)

[0085] 5) mRNA encoding GPC3 antigen and mRNA of IL-12p70 and IL15 / IL15Rα (experimental group)

[0086] The results are shown in Table 2 and image 3 and Figure 4 .

[0087] Compared with using the composition provided by the invention and using IL-12 and IL15 / IL15Rα alone, the composition of the invention has a synergistic immune enhancing effect. Among the CD8 T cell subsets, the proportion of IFN-γ positive cells in the experimental group was 3.82%, which was 56.18 times that of the GPC3 control, and the proportion of IFN-γ...

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Abstract

The invention discloses a composition for enhancing T cell sensitization capacity of antigen presenting cells (APCs) and application. The composition comprises a first interleukin complex or a precursor capable of forming the first interleukin complex, and a second interleukin complex or a precursor capable of forming the second interleukin complex, wherein the first interleukin complex is an IL-15 / IL-15 R[alpha] complex, and the second interleukin complex is an IL-12 complex. The composition provided by the invention can synergistically enhance the T cell sensitization capability of APCs.

Description

technical field [0001] The invention relates to the fields of immunology and medicine, in particular to a composition and application for enhancing the ability of antigen-presenting cells to sensitize T cells. Background technique [0002] In the process of the body's immune response, antigen-presenting cells (APC) play an important role. Studies have shown that such cells can assist and regulate T cells and B cells to recognize antigens and respond to antigens. According to the expression of APC cell surface membrane molecules There are two types of APCs: professional and non-professional APCs. Among them, dendritic cells (DC) are the professional APCs with the strongest function in the body, and their biggest feature is that they can stimulate the proliferation of initial T cells. Therefore, APCs Occupying a unique position in the immune system, it is the initiator of the immune response. [0003] CN101360827A discloses an IL-15Rα sushi domain as a selective and potent en...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0783C12P21/00
CPCC12N5/0636C12N2501/2312C12N2501/2315C12P21/00
Inventor 孙圣楠林鑫谢桦函
Owner 宜兴辰功新药开发有限公司
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