阅读说明：本技术 一种抗SARS-CoV-2的单克隆抗体1F2 (Monoclonal antibody 1F2 for resisting SARS-CoV-2 ) 是由 杨晓明 潘勇兵 王炯 詹珊珊 吴小丽 宋刚 桂芳 邓小杰 张囡 敬兆飞 杨溢民 于 2020-06-19 设计创作，主要内容包括：本发明涉及一种抗SARS-CoV-2的单克隆抗体1F2,该抗体的六个CDR区为：(1)重链CDR1(VHCDR1)包含如SEQ ID NO.1所示的氨基酸序列；(2)重链CDR2(VHCDR2)包含如SEQ ID NO.2所示的氨基酸序列；(3)重链CDR3(VHCDR3)包含如SEQ ID NO.3所示的氨基酸序列；(4)轻链CDR1(VLCDR1)包含如SEQ ID NO.4所示的氨基酸序列；(5)轻链CDR2(VLCDR2)包含如SEQ ID NO.5所示的氨基酸序列；(6)轻链CDR3(VLCDR3)包含如SEQ ID NO.6所示的氨基酸序列。(The invention relates to a monoclonal antibody 1F2 for resisting SARS-CoV-2, six CDR areas of the antibody are: (1) heavy chain CDR1(VHCDR1) comprises the amino acid sequence shown in SEQ ID No. 1; (2) heavy chain CDR2(VHCDR2) comprises the amino acid sequence shown in SEQ ID No. 2; (3) heavy chain CDR3(VHCDR3) comprises the amino acid sequence shown in SEQ ID No. 3; (4) the light chain CDR1(VLCDR1) comprises the amino acid sequence shown in SEQ ID No. 4; (5) the light chain CDR2(VLCDR2) comprises the amino acid sequence shown in SEQ ID NO. 5; (6) the light chain CDR3(VLCDR3) comprises the amino acid sequence shown in SEQ ID NO. 6.)

1. Monoclonal antibody 1F2 directed against SARS-CoV-2 virus, comprising the six CDR regions:

(1) the heavy chain CDR1(VHCDR1) comprises the amino acid sequence shown in SEQ ID No. 1: GFTVSSNYMN, respectively;

(2) the heavy chain CDR2(VHCDR2) comprises the amino acid sequence shown in SEQ ID NO. 2: VIYSGGSTFYADSVKG, respectively;

(3) the heavy chain CDR3(VHCDR3) comprises the amino acid sequence shown in SEQ ID NO. 3: DLDILGGMDV, respectively;

(4) the light chain CDR1(VLCDR1) comprises the amino acid sequence shown in SEQ ID No. 4: RASQSVSSSYLA, respectively;

(5) the light chain CDR2(VLCDR2) comprises the amino acid sequence shown in SEQ ID NO. 5: a GASSRAT;

(6) the light chain CDR3(VLCDR3) comprises the amino acid sequence shown in SEQ ID NO. 6: QQYGSSPPT are provided.

2. The antibody of claim 1,

the full length of the heavy chain variable region of the monoclonal antibody 1F2 comprises an amino acid sequence shown as SEQ ID NO. 7:

the full length of the variable region of the light chain of the monoclonal antibody 1F2 comprises an amino acid sequence shown in SEQ ID NO. 8.

3. The antibody of claim 1 or 2, wherein monoclonal antibody 1F2 is a human IgG-type antibody.

4. The antibody of claim 1 or 2, wherein the antigen bound by monoclonal antibody 1F2 is spike protein S1 of SARS-CoV-2 virus, and specifically, the antigenic domain bound by monoclonal antibody 1F2 is RBD domain of spike protein S1 of SARS-CoV-2 virus.

5. A nucleic acid fragment encoding the antibody of any one of claims 1-4.

6. Use of the monoclonal antibody 1F2 of any one of claims 1-4 in the preparation of a reagent for detecting SARS-CoV-2 virus.

7. Use of monoclonal antibody 1F2 according to any one of claims 1 to 4 in the preparation of a reagent for inhibiting SARS-CoV-2 virus.

8. Use of the monoclonal antibody 1F2 of any one of claims 1-4 in the manufacture of a medicament for the prevention and/or treatment of a disease caused by SARS-CoV-2 virus infection.

Technical Field

The invention belongs to the technical field of biological medicines, and particularly relates to a monoclonal antibody 1F2 for resisting SARS-CoV-2.

Background

Convalescent plasma containing high concentrations of anti-SARS-CoV-2 antibody has shown positive effects in therapy, suggesting that anti-SARS-CoV-2 specific antibodies are effective in blocking virus binding to cells. In addition, a series of monoclonal antibodies having neutralizing activity have been developed during outbreaks of severe infectious diseases such as SARS and MERS, and have been proven to be safe and effective in the prevention and treatment of diseases. These suggest that in response to SARS-CoV-2 virus, it is possible to consider the preparation of monoclonal antibodies against SARS-CoV-2, particularly fully human monoclonal antibodies, which not only protect against virus invasion by blocking the binding of SARS-CoV-2 to the recipient cell, but also have the advantage of less side effects than humanized or murine chimeric antibodies, and will provide a novel means for the specific prevention and treatment of COVID-19.

Disclosure of Invention

The invention firstly relates to a monoclonal antibody 1F2 aiming at SARS-CoV-2 virus, which is characterized in that the six CDR regions of the antibody are:

(1) the heavy chain CDR1(VHCDR1) comprises the amino acid sequence shown in SEQ ID No. 1: GFTVSSNYMN, respectively;

(2) the heavy chain CDR2(VHCDR2) comprises the amino acid sequence shown in SEQ ID NO. 2: VIYSGGSTFYADSVKG, respectively;

(3) the heavy chain CDR3(VHCDR3) comprises the amino acid sequence shown in SEQ ID NO. 3: DLDILGGMDV, respectively;

(4) the light chain CDR1(VLCDR1) comprises the amino acid sequence shown in SEQ ID No. 4: RASQSVSSSYLA, respectively;

(5) the light chain CDR2(VLCDR2) comprises the amino acid sequence shown in SEQ ID NO. 5: a GASSRAT;

(6) the light chain CDR3(VLCDR3) comprises the amino acid sequence shown in SEQ ID NO. 6: QQYGSSPPT are provided.

Further, in the above-mentioned case,

the full length of the heavy chain variable region of the monoclonal antibody 1F2 comprises an amino acid sequence shown as SEQ ID NO. 7: EVQLLESGGGLVQPGGSLRLSCAASGFTVSSNYMNWVRQAPGKGLEWVSVIYSGGSTFYADSVKGRFTISRDNSKNTLYLQ MNSLRAEDTAVYYCARDLDILGGMDVWGQGTTVTVSS；

The full length of the variable region of the light chain of the monoclonal antibody 1F2 comprises an amino acid sequence shown as SEQ ID NO. 8: EIVMTQSPGTLSLSPGERATLSCRASQSVSSSYLAWYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDF AVYYCQQYGSSPPTFGQGTKVEIK。

Further, the monoclonal antibody 1F2 is a human IgG type antibody.

Furthermore, the antigen bound by the monoclonal antibody 1F2 is spike protein S1 of SARS-CoV-2 virus, and specifically, the antigenic structural region bound by the monoclonal antibody 1F2 is RBD structural region in spike protein S1 of SARS-CoV-2 virus.

Most preferably, the light chain variable region and the heavy chain variable region of monoclonal antibody 1F2 are the amino acid sequences shown in SEQ ID NO.8 and SEQ ID NO.7, respectively.

The invention also relates to a nucleic acid fragment encoding the monoclonal antibody 1F 2.

The invention also relates to an antibody, wherein the light chain of the antibody is:

(1) the amino acid sequence shown in SEQ ID NO.8 is replaced, deleted or added with one or more amino acids to form a sequence with the same function;

or (2) an amino acid sequence with homology of more than 95 percent with the amino acid sequence shown in SEQ ID NO. 8;

the heavy chain of the antibody is:

(1) the amino acid sequence shown in SEQ ID NO.7 is replaced, deleted or added with one or more amino acids to form a sequence with the same function;

or (2) an amino acid sequence with homology of more than 95 percent with the amino acid sequence shown in SEQ ID NO. 7.

The invention also includes the application of the monoclonal antibody 1F2 in the preparation of a reagent for detecting SARS-CoV-2 virus.

The invention also includes the application of the monoclonal antibody 1F2 in the preparation of a reagent for inhibiting SARS-CoV-2 virus.

The invention also includes the application of the monoclonal antibody 1F2 in preparing medicaments for preventing and/or treating diseases caused by SARS-CoV-2 virus infection.

The invention has the beneficial effects that: because there are no vaccines and specific drugs on the market for the diseases caused by SARS-CoV-2 virus, although the convalescent plasma treatment has been proved to be a promising treatment method, the large-scale preparation is limited, and the method mainly aims at patients with severe and critical illness, while the monoclonal antibody has the advantages of high purity, strong targeting property, small side effect, large-scale preparation and the like. The experimental result of the 1F2 antibody shows that the neutralizing activity and the blocking activity of the monoclonal antibody 1F2 are both high. Therefore, the human monoclonal antibody 1F2 obtained by the invention provides a new candidate drug for specific prevention and treatment of COVID-19.

Drawings

FIG. 1, SDS-PAGE electrophoresis detection of monoclonal antibody 1F 2: lane 1 is a non-reducing SDS-PAGE electrophoresis, and lane 2 is a reducing SDS-PAGE electrophoresis.

FIG. 2 SEC-HPLC detection of monoclonal antibody 1F 2.

FIG. 3, results of the detection of the binding activity of monoclonal antibody 1F 2. A is the binding activity against SARS-CoV-2 spike protein S1; b is the binding activity against the RBD domain of the SARS-CoV-2 spike protein S1.

FIG. 4, results of the live virus neutralization assay of monoclonal antibody 1F 2.

FIG. 5 shows the results of affinity detection of monoclonal antibody 1F 2.

Detailed Description

Unless otherwise specified, the technical means used in the following examples are conventional means well known to those skilled in the art, and all reagent consumables are commercially available.