CHIMERIC ANTIGEN RECEPTOR (CAR) COMPRISING A CD19-BINDING DOMAIN
There is provided a chimeric antigen receptor (CAR) comprising a CD19-binding domain which comprises a) a heavy chain variable region (VH) having complementarity determining regions (CDRs) with the following sequences: (SEQ ID No. 1) CDR1 - GYAFSSS; (SEQ ID No. 2) CDR2 - YPGDED (SEQ ID No. 3) CDR3 - SLLYGDYLDY; and b) a light chain variable region (VL) having CDRs with the following sequences: (SEQ ID No. 4) CDR1 - SASSSVSYMH; (SEQ ID No. 5) CDR2 - DTSKLAS (SEQ ID No. 6) CDR3 - QQWNINPLT. There is also provided a cell comprising such a CAR, and the use of such a cell in the treatment of cancer, in particular a B cell malignancy.
1 . A chimeric antigen receptor (CAR) comprising a CD19-binding domain which comprises
a) a heavy chain variable region (VH) having complementarity determining regions (CDRs) with the following sequences:
(SEQ ID NO: 1)
CDR1 - GYAFSSS;
(SEQ ID NO: 2)
CDR2 - YPGDED
(SEQ ID NO: 3)
CDR3 - SLLYGDYLDY;
and
b) a light chain variable region (VL) having CDRs with the following sequences:
(SEQ ID NO: 4)
CDR1 - SASSSVSYMH;
(SEQ ID NO: 5)
CDR2 - DTSKLAS
(SEQ ID NO: 6)
CDR3 - QQWNINPLT.
2 . A CAR according to claim 1 , wherein the CD19 binding domain comprises a VH domain having the sequence shown as SEQ ID No. 7 and/or or a VL domain having the sequence shown as SEQ ID No 8 or a variant thereof having at least 95% sequence identity.
3 . A CAR according to claim 1 , wherein the CD19 binding domain comprises an scFv in the orientation VH-VL.
4 . A CAR according to claim 3 , wherein the CD19 binding domain comprises the sequence shown as SEQ ID No 9 or a variant thereof having at least 90% sequence identity.
5 . A CAR according to claim 1 , wherein the CD19 binding domain comprises the 6 CDRs defined in claim 1 grafted on to a human antibody framework.
6 . A CAR according to any preceding claim , wherein CD19-binding domain and the transmembrane domain are connected by a spacer.
7 . A CAR according to claim 6 , wherein the spacer comprises one of the following: a human an IgG1 Fc domain; an IgG1 hinge; or a CD8 stalk.
8 . A CAR according to claim 7 , wherein the spacer comprises an IgG1 hinge or a CD8 stalk.
9 . A CAR according to any preceding claim which also comprises an intracellular T cell signalling domain.
10 . A CAR according to claim 9 wherein the intracellular T cell signalling domain comprises one or more of the following endodomains: CD28 endodomain; 41BB endodomain, OX40 endodomain and the CD3-Zeta endodomain.
11 . A CAR according to claim 10 wherein the intracellular T-cell signalling domain comprises the 41BB endodomain and the CD3-Zeta endodomain.
12 . A CAR according to claim 10 wherein the intracellular T-cell signalling domain comprises the OX40 endodomain and the CD3-Zeta endodomain.
13 . A CAR according to claim 10 wherein the intracellular T cell signalling domain comprises all of the following endodomains: CD28 endodomain; OX40 and CD3-Zeta endodomain.
14 . A CAR according to any preceding claim , which comprises the sequence shown as any of SEQ ID No. 10 to 15 or a variant thereof which has at least 80% sequence identity but retains the capacity to i) bind CD19 and ii) induce T cell signalling.
15 . A CAR according to any preceding claim which, when expressed by a T-cell and used to target a CD19 expressing cell, causes lower IFNγ release by the CD19-expressing target cell than that caused by a T-cell expressing a CAR comprising a CD19-binding domain which comprises
a) a heavy chain variable region (VH) having complementarity determining regions (CDRs) with the following sequences:
(SEQ ID NO. 16)
CDR1 - GVSLPDY;
(SEQ ID NO. 17)
CDR2 - WGSET;
(SEQ ID NO. 18)
CDR3 - HYYYGGSYAMDY;
and
b) a light chain variable region (VL) having CDRs with the following sequences:
(SEQ ID NO. 19)
CDR1 - RASQDISKYLN;
(SEQ ID NO. 20)
CDR2 - HTSRLHS
(SEQ ID NO. 21)
CDR3 - QQGNTLPYT.
16 . A CAR according to any preceding claim wherein the CDRs are grafted on to a human or humanised framework.
17 . A nucleic acid sequence which encodes a CAR according to any preceding claim .
18 . A vector which comprises a nucleic acid sequence according to claim 17 .
19 . A cell which comprises a CAR according to any of claims 1 to 16 .
20 . A cell according to claim 19 , which is a T cell or a natural killer (NK) cell.
21 . A cell composition which comprises a plurality of cells according to claim 19 or 20 .
22 . A method for making a cell according to claim 19 or 20 , which comprises the step of transducing or transfecting a cell with a vector according to claim 18 .
23 . A method for making a cell composition according to claim 21 which comprises the step of transducing or transfecting a sample of cells from a subject ex vivo with a vector according to claim 18 .
24 . A pharmaceutical composition which comprises a cell according to claim 19 or 20 , or a cell composition according to claim 21 , together with a pharmaceutically acceptable carrier, diluent or excipient.
25 . A method for treating cancer which comprises the step of administering a cell according to claim 19 or 20 , a cell composition according to claim 21 or a pharmaceutical composition according to claim 24 to a subject.
26 . A method according to claim 25 which comprises the step of transducing or transfecting cells from the subject ex vivo with a vector according to claim 18 , then administering transfected cells back to the subject.
27 . A method according to claim 25 or 26 wherein the cancer is a B cell malignancy.
28 . A pharmaceutical composition according to claim 24 for use in treating cancer.
29 . The use of a cell according to claim 20 or 21 in the manufacture of a pharmaceutical composition for treating cancer.