Screening and preparation method and application for antibody medicament for malignant lymphoma and autoimmune diseases

Abstract

The invention relates to screening and a preparation method and application for an antibody medicament for malignant lymphoma and autoimmune diseases, and discloses a lymphoma specificity humanized antibody for resisting HLA-DR10, a person and whole person monoclonal antibodies and segments thereof, and a radioactive isotope labeling antibody thereof including a heavy chain and a light chain, wherein the heavy chain contains a constant region, a heavy chain variable region amino acid sequence is SEQID NO:1, the light chain contains a constant region, a light chain variable region amino acid sequence is SEQID NO:2, and the heavy chain is connected with the light chain through a disulfide bond. The invention obtains NSO and CHO engineering cell strains of stable high-level expression lymphoma specificity humanized, person and the whole person monoclonal antibodies (chlym-1 antibody) for resisting the HLA-DR10, adopts a serum-free high density culture technique so that the expression of the chlym-1 is more than 500 milligrams per liter, and discloses application of the monoclonal antibodies in treating the malignant lymphoma and the autoimmune diseases.

Description

technical field

[0001] The invention relates to the screening, preparation and application of monoclonal antibody drugs. Background technique

[0002] Malignant lymphoma (ML) is a malignant tumor originating from lymph nodes and extralymphatic lymphoid tissues. ML can be divided into two categories: Hodgkin's lymphoma (HD) and non-Hodgkin's lymphoma (NHL). In my country, NHL patients account for 85-90% of new cases of malignant lymphoma. In NHL, the incidence of low-grade lymphoma with good prognosis is low in my country, accounting for about 5% of NHL; while the incidence of high-grade lymphoma with poor prognosis is relatively high, accounting for about 30% of NHL. The survey shows that with the passage of time and the development of social economy, malignant lymphoma presents an increasing trend, in which HD is relatively stable, while the morbidity and mortality of NHL are increasing. The etiology and pathogenesis of non-Hodgkin's lymphoma have not yet been determined,...

Examples

Embodiment 1

[0027] Example 1 Preparation of lymphoma-specific humanized antibody against HLA-DR10

[0028] The NSO engineered cell line was prepared by the method in the example of the Chinese patent (patent number: 200410053872.0). It mainly includes the following steps:

[0029] 1. Preparation of mouse-derived anti-lymphoma monoclonal antibody:

[0030] 2. Preparation of specific chimeric monoclonal antibody against HLA-DR10, including preparation of heavy chain variable region and light chain variable region; fusion of variable region gene and constant region gene; establishment of expression cell seed cell line and its identification.

[0031] 3. After obtaining the NSO engineered cell line, use the following process conditions to prepare the lymphoma-specific humanized antibody against HLA-DR10

[0032] Expression process conditions: by determining the medium and other nutrients suitable for large-scale production needs, determine the way of supplementing nutrients, determine the ...

Embodiment 2

[0035] Example 2 131 Preparation of I-labeled lymphoma-specific humanized antibody against HLA-DR10

[0036] Using chloramine-T iodide labeling method, the 131 I was labeled on the lymphoma-specific humanized antibody against HLA-DR10 obtained in Example 1, and the radioactive specific activity of the product was 11 mCi / mg.

Embodiment 3

[0037] Example 3 131 Determination of Affinity Constant of I-labeled Anti-HLA-DR10 Lymphoma Specific Humanized Antibody

[0038] Adopt the method for the embodiment in the Chinese patent (patent number: 200410053872.0) to measure the obtained in embodiment 2 131 I-labeled lymphoma-specific humanized antibody against HLA-DR10, the affinity constant of the monoclonal antibody Ka=3.40×10 8 mol -1 .

technical field

[0001] The invention relates to the screening, preparation and application of monoclonal antibody drugs. Background technique

[0002] Malignant lymphoma (ML) is a malignant tumor originating from lymph nodes and extralymphatic lymphoid tissues. ML can be divided into two categories: Hodgkin's lymphoma (HD) and non-Hodgkin's lymphoma (NHL). In my country, NHL patients account for 85-90% of new cases of malignant lymphoma. In NHL, the incidence of low-grade lymphoma with good prognosis is low in my country, accounting for about 5% of NHL; while the incidence of high-grade lymphoma with poor prognosis is relatively high, accounting for about 30% of NHL. The survey shows that with the passage of time and the development of social economy, malignant lymphoma presents an increasing trend, in which HD is relatively stable, while the morbidity and mortality of NHL are increasing. The etiology and pathogenesis of non-Hodgkin's lymphoma have not yet been determined,...

Examples

Embodiment 1

[0027] Example 1 Preparation of lymphoma-specific humanized antibody against HLA-DR10

[0028] The NSO engineered cell line was prepared by the method in the example of the Chinese patent (patent number: 200410053872.0). It mainly includes the following steps:

[0029] 1. Preparation of mouse-derived anti-lymphoma monoclonal antibody:

[0030] 2. Preparation of specific chimeric monoclonal antibody against HLA-DR10, including preparation of heavy chain variable region and light chain variable region; fusion of variable region gene and constant region gene; establishment of expression cell seed cell line and its identification.

[0031] 3. After obtaining the NSO engineered cell line, use the following process conditions to prepare the lymphoma-specific humanized antibody against HLA-DR10

[0032] Expression process conditions: by determining the medium and other nutrients suitable for large-scale production needs, determine the way of supplementing nutrients, determine the ...

Embodiment 2

[0035] Example 2 131 Preparation of I-labeled lymphoma-specific humanized antibody against HLA-DR10

[0036] Using chloramine-T iodide labeling method, the 131 I was labeled on the lymphoma-specific humanized antibody against HLA-DR10 obtained in Example 1, and the radioactive specific activity of the product was 11 mCi / mg.

Embodiment 3

[0037] Example 3 131 Determination of Affinity Constant of I-labeled Anti-HLA-DR10 Lymphoma Specific Humanized Antibody

[0038] Adopt the method for the embodiment in the Chinese patent (patent number: 200410053872.0) to measure the obtained in embodiment 2 131 I-labeled lymphoma-specific humanized antibody against HLA-DR10, the affinity constant of the monoclonal antibody Ka=3.40×10 8 mol -1 .