Hepatitis B immunotherapy agent targeting ccl19/ccr7 and its use

A-CCL19, hepatitis B technology, applied in the field of biomedicine, can solve problems such as the role has not yet been announced

Active Publication Date: 2021-11-16
WUXI NO 5 PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Cytokines are widely used as immunomodulators in antiviral therapy. For example, IFN-α is recommended in the "Treatment Guidelines" in anti-HBV therapy and anti-new coronavirus therapy (16) , Chemokine CCL19 is a special cytokine that normally exists in the human body. The conventionally known function is that it can promote the directional migration of lymphocytes containing its receptor CCR7 molecule and enhance the immune response. It can also be used as a molecular adjuvant or Immunopotentiator to enhance antiviral immunity against HIV-1 and HSV-2 (14,15) , but its role in HBV immune clearance has not yet been revealed

Method used

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  • Hepatitis B immunotherapy agent targeting ccl19/ccr7 and its use
  • Hepatitis B immunotherapy agent targeting ccl19/ccr7 and its use
  • Hepatitis B immunotherapy agent targeting ccl19/ccr7 and its use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1 Immunomodulators CCL19 and shCCR7 are used for peripheral blood lymphocytes of patients with HBV chronic infection

[0049] The reason why the virus in the liver of patients with chronic hepatitis B cannot be cleared is mainly because the liver is an immune tolerance organ, and at the same time, there is exhaustion of CD8+ T lymphocytes in patients with hepatitis B. Functional validation was performed on peripheral blood lymphocytes (PBMC). The specific steps are:

[0050] (1) Extract peripheral blood lymphocytes and add them to 24-well plate, 1×10 per well 6 Lymphocytes per 500ml, CCL19 group was stimulated with human CCL19 protein (10ng / well) for 72 hours, HBc group was stimulated with HBV specific antigen HBc polypeptide (10ng / well) for 72h; HBc+CCL19 group was stimulated with human CCL19 protein (10ng / well) and HBV-specific antigen HBc polypeptide stimulation for 72h; HBc+CCL19+L294002 group was stimulated with human CCL19 protein (10ng / well) and HBV-spe...

Embodiment 2

[0051] The establishment of embodiment 2 mouse model

[0052] C57 / BJ mice chronically infected with HBV were used to construct the HBV-infected mouse model, and the HBV1.2 plasmid was injected through the high-pressure tail vein, 10ng / mouse. LinYJ, Huang LR, Yang HC, Tzeng HT, Hsu PN, Wu HL, et al. Hepatitis B virus coreantigen determines viral persistence in a C57BL / 6mouse model. Proceedings of the National Academy of Sciences of the United States of America (2010) 107 (20):9340-5. doi:10.1073 / pnas.1004762107.

[0053] The HBV-infected mouse model constructed according to the above method was used to construct a model with high expression of CCL19 and decreased expression of its receptor CCR7.

[0054] (1) Mouse model of chronic HBV infection with high expression of CCL19 molecules: the recombinant expression mouse CCL19 plasmid on pcDNA3.1(+) constructed in our laboratory (plasmid map as shown in Figure 7所示,编码CCL19的核苷酸序列为:ATGGCCCCCCGTGTGACCCCACTCCTGGCCTTCAGCCTGCTGGTTCTCTG...

Embodiment 3

[0057] Example 3 Immunomodulators CCL19 and shCCR7 are used for HBV chronically infected mice

[0058] The immunomodulator CCL19 and the immunomodulator shCCR7 were functionally verified on HBV chronically infected mice, and the control mouse model constructed in Example 2, the CCR7 knockdown HBV chronically infected mouse model and the high expression of CCL19 molecules were detected respectively The proportion of liver non-parenchymal cells, immune modulation and the effect on cells in chronic HBV infection mouse model.

[0059] (1) Changes in the proportion of liver non-parenchymal cells 2 weeks after the action of immunomodulators: the proportion of KCs cells (F4 / 80+, CD11c-) in mice with high expression of CCL19 was significantly increased (about 70%), and the molecular knockdown of CCR7 Cause the proportion of KCs cells to decline (about 60%), and the proportion difference in each group of CHB mice is not obvious (about 50%) ( figure 2 A-2B). However, compared with th...

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Abstract

The invention discloses a hepatitis B immunotherapy agent targeting CCL19 / CCR7 and its use, and belongs to the technical field of biomedicine. The present invention provides a CCL19 protein with immune enhancing effect and a regulator AAV-shRNA-CCR7 (shCCR7) that knocks down the expression of its receptor CCR7 in promoting and increasing the ratio of CD8+ T cells and the activation and apoptosis of HBV antigen-specific T cells , applications in accelerating HBV clearance. It can be used to accelerate the clearance of HBV in mammals chronically infected with HBV, and provides a theoretical basis for the preparation of HBV preventive drugs or therapeutic vaccines.

Description

technical field [0001] The invention relates to an immunotherapeutic agent for hepatitis B targeting CCL19 / CCR7 and its application, belongs to the technical field of biomedicine, and specifically relates to an immunomodulator and its application. Background technique [0002] Approximately 250 million people worldwide are chronically infected with hepatitis B virus (HBV) and are at high risk of developing hepatic steatosis, cirrhosis, end-stage liver failure or hepatocellular carcinoma (1,2) . Currently approved therapeutic strategies, including interferon-alpha (IFN-α), nucleoside and nucleotide analogs, offer only limited efficacy in maintaining long-term HBV 'eradication' (1,3,4) , the occurrence of adverse events leading to treatment discontinuation or poor compliance (5) . In primary HBV infection, the virus is completely eradicated in approximately 90% of adults, with the remaining individuals developing viral persistence and disease progression. Immune defense ag...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K45/00A61K9/00A61K48/00A61K31/713A61P1/16A61P31/20
CPCA61K9/0019A61K31/713A61K45/00A61K48/005A61P1/16A61P31/20
Inventor 阎岩
Owner WUXI NO 5 PEOPLES HOSPITAL
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