US 12,735,697 B2Granted

Method for selecting antibody fragments, recombinant antibodies produced therefrom, and uses thereof

AssigneeAcademia Sinica

InventorsAn-Suei Yang, Chung-Ming Yu, Ing-Chien Chen, Chao-Ping Tung, Hung-Pin Peng

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Abstract

Disclosed herein are methods for selecting an antibody fragment specific to an influenza virus. According to certain embodiments of the present disclosure, the influenza virus may be influenza virus type A (IAV) or influenza virus type B (IBV). Also disclosed herein are the selected antibodies, recombinant antibody produced from the selected antibodies, and the uses thereof in the diagnosis of influenza virus infection.

Claims10

Dependent claims

A method for selecting an antibody fragment specific to an influenza virus, comprising,

(a) providing a phage-displayed single-chain variable fragment (scFv) library that comprises a plurality of phage-displayed scFvs, wherein the heavy chain variable (VH) domain of each phage-displayed scFvs has a binding affinity to protein A, and the light chain variable (VL) domain of each phage-displayed scFvs has a binding affinity to protein L;
(b) exposing the phage-displayed scFv library of the step (a) to a target nucleoprotein comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:
1-6;
(c) selecting, from the phage-displayed scFv library of the step (b), a first plurality of phages that respectively express scFvs exhibiting binding affinity to the target nucleoprotein;
(d) exposing the first plurality of phages selected in the step (c) to the target nucleoprotein in the presence of at least one scrambled nucleoprotein, wherein the scrambled nucleoprotein comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:
1-6, and the amino acid sequence of the scrambled nucleoprotein is different from the amino acid sequence of the target nucleoprotein;
(e) selecting, from the first plurality of phages of the step (d), a second plurality of phages that respectively express scFvs exhibiting binding affinity to the target nucleoprotein in the presence of the scrambled nucleoprotein;
(f) respectively enabling the second plurality of phages selected in the step (e) to express a plurality of soluble scFvs;
(g) exposing the plurality of soluble scFvs of the step (f) to the target nucleoprotein;
(h) determining the respective binding affinity of the plurality of soluble scFvs in the step (g);
and
(i) based on the results determined in the step (h), selecting one soluble scFv that exhibits superior affinity over the other soluble scFvs of the plurality of soluble scFvs as the antibody fragment.

9 more elements

A recombinant antibody or an antigen binding fragment thereof, comprising a VL domain and a VH domain, wherein the VL domain comprises a first light chain complementarity determining region (CDR-L1), a second light chain CDR (CDR-L2) and a third light chain CDR (CDR-L3), and the VH domain comprises a first heavy chain CDR (CDR-H1), a second heavy chain CDR (CDR-H2) and a third heavy chain CDR (CDR-H3), wherein

the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
7-12;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
13-18;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
19-24;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
25-30;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
31-36;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
37-42;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
43-48;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
49-54;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
55-60;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
61-66;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
67-72;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
73-78;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
79-84;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
85-90;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
91-96;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
97-102;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
103-108;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
109-114;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
115-120;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H-1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
121-126;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
127-132;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
133-138;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
139-144;
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
145-150;
or
the CDR-L1, the CDR-L2, the CDR-L3, the CDR-H1, the CDR-H2 and the CDR-H3 respectively comprise the amino acid sequences of SEQ ID NOs:
151-156.

48 more elements

Dependencies are inferred from the claim text.

Facts

Application: 18/008,753Filed 7 Jun 2021 · us provisional application
Priority: 8 Jun 2020
Published: US 2023/0212554 A16 Jul 2023
Granted: 15 Sep 2026
Term ends: Not estimatedThe source records don’t support an estimate.

Classification10 codes

C12N 15/1037
Isolating an individual clone by screening libraries: screening libraries presented on the surface of microorganisms, e.g. phage display, E. coli…
C07K 16/108
RNA viruses: orthomyxoviridae , e.g. influenza virus
G01N 33/56983
For microorganisms, e.g. protozoa, bacteria, viruses: viruses
C07K 2317/622
Comprising only variable region components: single chain antibody

Family

Application63/036,143Us provisional application · filed 8 Jun 2020
US 2023/0212554 A1Pre-grant publication · published 6 Jul 2023
US 12,735,697 B2This patent

From the US filing history. Foreign family members aren’t listed.

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Abstract

A method for selecting an antibody fragment specific to an influenza virus, comprising,

(a) providing a phage-displayed single-chain variable fragment (scFv) library that comprises a plurality of phage-displayed scFvs, wherein the heavy chain variable (VH) domain of each phage-displayed scFvs has a binding affinity to protein A, and the light chain variable (VL) domain of each phage-displayed scFvs has a binding affinity to protein L;
(b) exposing the phage-displayed scFv library of the step (a) to a target nucleoprotein comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:
1-6;
(c) selecting, from the phage-displayed scFv library of the step (b), a first plurality of phages that respectively express scFvs exhibiting binding affinity to the target nucleoprotein;
(d) exposing the first plurality of phages selected in the step (c) to the target nucleoprotein in the presence of at least one scrambled nucleoprotein, wherein the scrambled nucleoprotein comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:
1-6, and the amino acid sequence of the scrambled nucleoprotein is different from the amino acid sequence of the target nucleoprotein;
(e) selecting, from the first plurality of phages of the step (d), a second plurality of phages that respectively express scFvs exhibiting binding affinity to the target nucleoprotein in the presence of the scrambled nucleoprotein;
(f) respectively enabling the second plurality of phages selected in the step (e) to express a plurality of soluble scFvs;
(g) exposing the plurality of soluble scFvs of the step (f) to the target nucleoprotein;
(h) determining the respective binding affinity of the plurality of soluble scFvs in the step (g);
and
(i) based on the results determined in the step (h), selecting one soluble scFv that exhibits superior affinity over the other soluble scFvs of the plurality of soluble scFvs as the antibody fragment.

9 more elements

Facts

Application

18/008,753Filed 7 Jun 2021 · us provisional application

Priority

8 Jun 2020

Published

US 2023/0212554 A16 Jul 2023

Granted

15 Sep 2026

Term ends

Not estimatedThe source records don’t support an estimate.

Classification10 codes

C12N 15/1037
Isolating an individual clone by screening libraries: screening libraries presented on the surface of microorganisms, e.g. phage display, E. coli…
C07K 16/108
RNA viruses: orthomyxoviridae , e.g. influenza virus
G01N 33/56983
For microorganisms, e.g. protozoa, bacteria, viruses: viruses
C07K 2317/622
Comprising only variable region components: single chain antibody