JPWO2019175885A5

JPWO2019175885A5 -

Info

Publication number: JPWO2019175885A5
Application number: JP2020549021A
Authority: JP
Publication date: 2022-03-23

Claims

A drug that binds to the membrane CD28 (mCD28) and inhibits the proteolytic cleavage of the mCD28.

The drug
a. Neither a CD28 agonist nor an antagonist ,
b. It does not degrade the mCD28 or inhibit mCD28-mediated immune cell activation.
c . Does not induce antibody-dependent cellular cytotoxicity (ADCC) or complement-dependent cellular cytotoxicity (CDC) ,
d. Binding within the stalk region of CD28, the stalk region optionally comprises the amino acid sequence GKHLPCPSPLFPGPSKP (SEQ ID NO: 9) or KGKHLPCPSPLFPGPS (SEQ ID NO: 36); and optionally consists of the amino acid sequence HVKGKHLPCPSPLFPGPSKP (SEQ ID NO: 10) or both.
e. Inhibits proteolytic cleavage by at least one protease and, optionally, said at least one protease is a metalloprotease, or
f. A combination of them,
The agent according to claim 1 .

The drug
a. It is an antibody;
b. Contains IgG2 or IgG4;
c. Includes Fc domains designed to reduce CDC, ADCC, or both;
d. Deficient in Fc domain;
e. Fab fragment;
f. It is a single chain antibody;
g. Single domain antibody;
h. Small molecule;
i. A peptide that specifically binds to mCD28; or j. A combination of them,
The agent according to claim 1 or 2 .

The agent is an antibody or antigen binding fragment thereof and comprises three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L).
CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 30 (GFTFSSYYMS), CDR-H2 comprises the amino acid sequence set forth in SEQ ID NO: 31 (TISDGGDNTYYAGTVTG), and CDR-H3 comprises the amino acid sequence set forth in SEQ ID NO: 32 (IHWPYYFDS). Containing the amino acid sequence shown, CDR-L1 comprises the amino acid sequence set forth in SEQ ID NO: 33 (RASSSVSYMN), CDR-L2 comprises the amino acid sequence set forth in SEQ ID NO: 34 (ATSDLAS), and CDR-L3 comprises. The agent according to any one of claims 1 to 3 , which comprises the amino acid sequence shown in SEQ ID NO: 35 (QQWSSHPPT).

a. Heavy chain containing the amino acid sequence of SEQ ID NO: 53; and b. The agent of claim 4 , comprising at least one of the light chains comprising the amino acid sequence of SEQ ID NO: 55.

A drug that binds to soluble CD28 (sCD28) and is neither a CD28 agonist nor an antagonist.

The agent according to claim 6 , which does not bind to the membrane CD28 (mCD28).

The drug
a. Specific binding to an antibody or its antigen-binding fragment, Fab fragment, Fv, Fab, F (ab') ₂ , scFV, scFV ₂ fragment, humanized antibody, single chain antibody, single domain antibody, small molecule, and sCD28. Selected from peptides with
b. Does not inhibit the binding of sCD28 to the ligand ,
c. Binds to dimer sCD28, monomeric sCD28 or both ,
d. Binds to the outside of the IgV domain of sCD28 ,
e. Binding of the drug to sCD28 in an organism results in at least one of the degradation of the bound sCD28, the removal of sCD28 from the blood, and the transport of the bound sCD28 to lysosomes, endosomes, proteasomes or combinations thereof.
f. Does not induce antibody-dependent cellular cytotoxicity (ADCC) or complement-dependent cellular cytotoxicity (CDC),
g. A combination of them,
The agent according to claim 6 or 7 .

The agent is an antibody or antigen binding fragment thereof and comprises three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L).
CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 12 (GYTLTNY), CDR-H2 comprises the amino acid sequence set forth in SEQ ID NO: 13 (NTYTGK), and CDR-H3 comprises the amino acid sequence set forth in SEQ ID NO: 14 (GDANQQFAY). Containing the amino acid sequence shown, CDR-L1 comprises the amino acid sequence set forth in SEQ ID NO: 15 (KASQDINSYS), CDR-L2 comprises the amino acid sequence set forth in SEQ ID NO: 16 (RANRLVD), and CDR-L3 comprises. Includes the amino acid sequence set forth in SEQ ID NO: 17 (LQYDEFPT);
CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 18 (GYTFTSY), CDR-H2 comprises the amino acid sequence set forth in SEQ ID NO: 19 (YPGDGD), and CDR-H3 comprises the amino acid sequence set forth in SEQ ID NO: 20 (NYRYSSFGY). Containing the amino acid sequence shown, CDR-L1 comprises the amino acid sequence set forth in SEQ ID NO: 21 (KSSQSLLNSGNQKNYLT), CDR-L2 comprises the amino acid sequence set forth in SEQ ID NO: 22 (WASTRES), and CDR-L3 comprises. Contains the amino acid sequence set forth in SEQ ID NO: 23 (QSDYSYSTEM); or CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 24 (GYTFTDY), and CDR-H2 comprises the amino acid sequence set forth in SEQ ID NO: 25 (NPNYDS). CDR-H3 comprises the amino acid sequence set forth in SEQ ID NO: 26 (SSPYYDSNHFDY), CDR-L1 comprises the amino acid sequence set forth in SEQ ID NO: 27 (SARSSINYMH), and CDR-L2 comprises the amino acid sequence set forth in SEQ ID NO: 28 (SSPYYDSNHFDY). DTSKLAS) The agent according to any one of claims 6 to 8 , wherein CDR-L3 comprises the amino acid sequence shown in SEQ ID NO: 29 (HQRNSYPFT).

a. A heavy chain comprising an amino acid sequence selected from SEQ ID NO: 41, 45 or 49; and b. 9. The agent of claim 9 , comprising at least one of the light chains comprising an amino acid sequence selected from SEQ ID NO: 43, 47 or 51.

A drug for use in treating or preventing cancer in a subject in need of treatment or prevention of the cancer, optionally, the subject's blood prior to the treatment or prevention is at least 5 ng / ml sCD28. The agent according to any one of claims 1 to 10, comprising the above.

A drug for use in improving PD-1 or PD-L1 based immunotherapy in subjects requiring PD-1 or PD-L1 based immunotherapy, the treatment or prevention of which is optional. 13. The drug according to any one item.

The agent according to any one of claims 11 or 12, wherein the cancer is selected from melanoma, head and neck cancer, non-small cell lung cancer, ovarian cancer, kidney cancer, gastric cancer and colorectal cancer.

The agent according to any one of claims 11 to 13, wherein the use does not degrade mCD28 or reduce mCD28-mediated immune cell activation.

A method for producing the agent according to any one of claims 1 to 10 .
a. Drugs that bind to the CD28 extracellular domain or fragment thereof are obtained and optionally the CD28 extracellular domain or fragment thereof comprises the CD28 stalk region and tested for the ability of the drug to block the cleavage of mCD28 by proteases. Selecting at least one agent that blocks cleavage of mCD28 by the protease and, optionally, assaying mCD28 downstream signaling in the presence of said obtained agent to substantially activate mCD28 signaling. Choosing at least one drug that does not substantially antagonize ; or
b. Culturing a host cell containing one or more vectors containing a nucleic acid sequence encoding a drug, said nucleic acid sequence.
i. Obtaining a drug that binds to the CD28 extracellular domain or fragment thereof, optionally said that the CD28 extracellular domain or fragment thereof comprises the CD28 stalk region ;
ii. To test the ability of the drug to block the cleavage of mCD28 by proteases ;
i ii. Select at least one agent that blocks the cleavage of mCD28 by the protease ; and optionally.
iv. Assaying mCD28 downstream signaling in the presence of the obtained drug to select at least one drug that does not substantially actuate or substantially antagonize mCD28 signaling.
Culturing , which is of the drug selected by
c. A drug that binds to the extracellular domain of CD28 or a fragment thereof is obtained and assayed for mCD28 downstream signaling in the presence of the obtained drug, which may substantially activate or substantially antagonize mCD28 signaling. Do not select at least one drug and optionally test the binding of the obtained drug to mCD28 and select at least one drug that does not bind to mCD28, or a cancer patient of the obtained drug. To test binding to sCD28 from a cancer patient and select at least one agent that binds to said sCD28 from a cancer patient ; or
d. Culturing a host cell containing one or more vectors containing a nucleic acid sequence encoding a drug, said nucleic acid sequence.
i. Obtaining a drug that binds to the CD28 extracellular domain or fragments thereof;
ii. Assaying mCD28 downstream signaling in the presence of the obtained agent ;
i ii. Selecting at least one agent that does not substantially actuate or substantially antagonize mCD28 signaling , and optionally.
iv. The binding of the obtained drug to mCD28 is tested and at least one drug that does not bind to mCD28 is selected, or the obtained drug is tested for binding to sCD28 from a cancer patient and from a cancer patient. Selecting at least one agent that binds to the sCD28.
The method of producing the agent according to any one of claims 1 to 10 , comprising culturing, which is of the agent selected by.

Obtaining the drug
a. Immunizing an organism with the CD28 extracellular domain or fragment thereof and collecting antibodies from the immunized organism , optionally the CD28 extracellular domain or fragment thereof in a dimer or monomer. There, the organism is selected from rabbits, mice, rats, sharks, camels, chickens, goats and phage, or the antibody can be collected to extract B cells from the spleen of the immunized organism. Immunize an organism with the CD28 extracellular domain or fragment thereof, comprising fusing the extracted B cells with myeloma cells to generate a hybridoma, and collecting antibodies from the hybridoma. Collecting antibodies from living organisms ;
b. Screening a library of agents for binding to the CD28 extracellular domain or fragments thereof and selecting the agent to be bound, optionally selecting the agent to be bound is said to the selected agent. Screening a library of drugs for binding to the CD28 extracellular domain or fragments thereof, including sequencing and generating recombinant forms of the drug from the sequence, and selecting the drug to bind.
15. The method of claim 15 , comprising at least one of the above.

A method of determining the suitability of a subject to be treated using the agent according to any one of claims 1-10 , wherein a sample is obtained from the subject and the sCD28 level in the sample is determined. Including determining, sCD28 levels above 5 ng / ml indicate that the subject is suitable for treatment with the agent according to any one of claims 1-10 , and optionally said sample. Is a blood sample, the method.

a. Anti-PD-1 and / or PD-L1 immunotherapy;
b. Labels indicating that the agents of the invention are intended for use with PD-1 and / or PD-L1-based immunotherapy; and c. A secondary detection molecule for detecting at least one of the agents according to any one of claims 1 to 10 .
A kit comprising at least one of the agents according to any one of claims 1 to 10 , further comprising at least one of the above-mentioned agents.