MULTI-SPECIFIC MOLECULES AND MENTOD OF USE AND MAKING THEREOF
Provided are multi-specific molecules that can simultaneously engage T cells and target cancer or tumor cells. Also provided are methods of treating a cell proliferative disorder such as a cancer or tumor using the multi-specific molecules. In certain embodiments, a multi-specific molecule, comprises a first targeting domain that specifically binds to human CD3; a second targeting domain that specifically binds to human CD28; a third targeting domain that specifically binds to human Muc17, or human DLL3, or human CLDN18.2; and an Fc fragment that lacks antibody-dependent cellular cytotoxicity.
1 . A multi-specific molecule, comprising:
a) a first targeting domain that specifically binds to human CD3 with a binding affinity that equals to, or is greater than, 1 nmol/L;
b) a second targeting domain that specifically binds to human CD28 with a binding affinity that equals to, or is greater than, 100 nmol/L;
c) a third targeting domain that specifically binds to human Muc17, or human DLL3, or human CLDN18.2; and
d) an Fc fragment that lacks antibody-dependent cellular cytotoxicity.
2 . The multi-specific molecule of claim 1 , wherein the third targeting domain specifically binds to human Muc17.
3 . The multi-specific molecule of claim 2 , wherein the third targeting domain comprises
(1) an HCDR1 of SEQ ID NO:1, an HCDR2 of SEQ ID NO:2, an HCDR3 of SEQ ID NO: 3, an LCDR1 of SEQ ID NO:4, an LCDR2 of SEQ ID NO:5, and an LCDR3 of SEQ ID NO: 6, or
(2) an HCDR1 of SEQ ID NO:27, an HCDR2 of SEQ ID NO:28, an HCDR3 of SEQ ID NO: 29, an LCDR1 of SEQ ID NO:30, an LCDR2 of SEQ ID NO:31, and an LCDR3 of SEQ ID NO:32,
4 . The multi-specific molecule of claim 2 , wherein the second targeting domain comprises:
(1) an HCDR1 of SEQ ID NO: 107, an HCDR2 of SEQ ID NO: 108, an HCDR3 of SEQ ID NO: 124, an LCDR1 of SEQ ID NO: 110, an LCDR2 of SEQ ID NO: 111, and an LCDR3 of SEQ ID NO:112; or
(2) an HCDR1 of SEQ ID NO: 107, an HCDR2 of SEQ ID NO: 120, an HCDR3 of SEQ ID NO:115, an LCDR1 of SEQ ID NO:110, an LCDR2 of SEQ ID NO: 111, and an LCDR3 of SEQ ID NO:112.
5 . The multi-specific molecule of claim 2 , wherein the first targeting domain comprises:
(1) an HCDR1 of SEQ ID NO:93, an HCDR2 of SEQ ID NO:94, an HCDR3 of SEQ ID NO:95, an LCDR1 of SEQ ID NO:96, an LCDR2 of SEQ ID NO:97, and an LCDR3 of SEQ ID NO:98; or
(2) an HCDR1 of SEQ ID NO:93, an HCDR2 of SEQ ID NO:101, an HCDR3 of SEQ ID NO:95, an LCDR1 of SEQ ID NO:96, an LCDR2 of SEQ ID NO:97, and an LCDR3 of SEQ ID NO:98; or
(3) an HCDR1 of SEQ ID NO:93, an HCDR2 of SEQ ID NO:94, an HCDR3 of SEQ ID NO: 105, an LCDR1 of SEQ ID NO:96, an LCDR2 of SEQ ID NO:97, and an LCDR3 of SEQ ID NO:98.
6 . The multi-specific molecule of claim 1 , wherein the third targeting domain specifically binds to human DLL3
7 . The multi-specific molecule of claim 6 , wherein the third targeting domain comprises:
(1) an HCDR1 of SEQ ID NO:35, an HCDR2 of SEQ ID NO:36, an HCDR3 of SEQ ID NO:37, an LCDR1 of SEQ ID NO:38, an LCDR2 of SEQ ID NO:39, and an LCDR3 of SEQ ID NO:40; or
(2) an HCDR1 of SEQ ID NO:43, an HCDR2 of SEQ ID NO:44, an HCDR3 of SEQ ID NO:45, an LCDR1 of SEQ ID NO:46, an LCDR2 of SEQ ID NO:47, and an LCDR3 of SEQ ID NO:48.
8 . The multi-specific molecule of claim 6 , wherein the second targeting domain comprises:
(1) an HCDR1 of SEQ ID NO:107, an HCDR2 of SEQ ID NO:108, an HCDR3 of SEQ ID NO: 124, an LCDR1 of SEQ ID NO:110, an LCDR2 of SEQ ID NO: 111, and an LCDR3 of SEQ ID NO:112; or
(2) an HCDR1 of SEQ ID NO:107, an HCDR2 of SEQ ID NO: 120, an HCDR3 of SEQ ID NO: 115, an LCDR1 of SEQ ID NO:110, an LCDR2 of SEQ ID NO: 111, and an LCDR3 of SEQ ID NO:112.
9 . The multi-specific molecule of claim 6 , wherein the first targeting domain comprises:
(1) an HCDR1 of SEQ ID NO:93, an HCDR2 of SEQ ID NO:94, an HCDR3 of SEQ ID NO:95, an LCDR1 of SEQ ID NO:96, an LCDR2 of SEQ ID NO:97, and an LCDR3 of SEQ ID NO:98; or
(2) an HCDR1 of SEQ ID NO:93, an HCDR2 of SEQ ID NO: 101, an HCDR3 of SEQ ID NO:95, an LCDR1 of SEQ ID NO:96, an LCDR2 of SEQ ID NO:97, and an LCDR3 of SEQ ID NO:98.
10 . The multi-specific molecule of claim 1 , wherein the third targeting domain specifically binds to human CLDN18.2.
11 . The multi-specific molecule of claim 10 , wherein the third targeting domain comprises an HCDR1 of SEQ ID NO:65, an HCDR2 of SEQ ID NO:66, an HCDR3 of SEQ ID NO:67, an LCDR1 of SEQ ID NO:68, an LCDR2 of SEQ ID NO:69, and an LCDR3 of SEQ ID NO:70.
12 . The multi-specific molecule of claim 10 , wherein the second targeting domain comprises:
(1) an HCDR1 of SEQ ID NO:107, an HCDR2 of SEQ ID NO:108, an HCDR3 of SEQ ID NO: 124, an LCDR1 of SEQ ID NO: 110, an LCDR2 of SEQ ID NO: 111, and an LCDR3 of SEQ ID NO:112; or
(2) an HCDR1 of SEQ ID NO: 107, an HCDR2 of SEQ ID NO: 120, an HCDR3 of SEQ ID NO: 115, an LCDR1 of SEQ ID NO: 110, an LCDR2 of SEQ ID NO: 111, and an LCDR3 of SEQ ID NO:112.
13 . The multi-specific molecule of claim 11 , wherein the first targeting domain comprises an HCDR1 of SEQ ID NO:93, an HCDR2 of SEQ ID NO: 101 and 103, an HCDR3 of SEQ ID NO:95, an LCDR1 of SEQ ID NO:96, an LCDR2 of SEQ ID NO:97, and an LCDR3 of SEQ ID NO:98.
14 . The multi-specific molecule of claim 1 , wherein the first targeting domain specifically binds to human CD3 with a binding affinity in the range of 1-2000 nmol/L.
15 . The multi-specific molecule of claim 1 , wherein the second targeting domain specifically binds to human CD28 with a binding affinity in the range of 100-10000 nmol/L.
16 . The multi-specific molecule of claim 1 , wherein the first targeting domain specifically binds to human CD3 with a binding affinity in the range of 1-2000 nmol/L and wherein the second targeting domain specifically binds to human CD28 with a binding affinity in the range of 100-10000 nmol/L.
17 . The multi-specific molecule of claim 1 , wherein the first targeting domain specifically binds to human CD3 with a binding affinity in the range of 1-500 nmol/L and wherein the second targeting domain specifically binds to human CD28 with a binding affinity in the range of 100-2000 nmol/L.
18 . The multi-specific molecule of claim 1 , wherein each targeting domain comprises at least one antibody fragment selected from the group consisting of single domain antibody (sdAb), a fragment variable (Fv) heterodimer, a single chain Fv (scFv), Fab fragment and combinations thereof.
19 . A method of stimulating T-cell activity in a subject, comprising the step of administering to the subject, an effective amount of the multi-specific molecule of claim 1 .
20 . A method to treating cancer in a subject, comprising the step of administering to the subject, an effective amount of the multi-specific molecule of claim 1 .