Chimeric antigen receptor
The present invention provides a chimeric antigen receptor (CAR) which binds a target antigen having a bulky extracellular domain, wherein the CAR comprises a Fab antigen binding domain. The present invention also provides nucleic acid sequences and constructs encoding such a CAR, cells expressing such a CAR and their therapeutic uses.
1. An anti-CD22 antigen-binding domain which comprises:
a) a heavy chain variable region (VH) having complementarity determining regions (CDRs) with the following sequences:
(SEQ ID NO: 93)
CDR1 - NFAMA,
(SEQ ID NO: 94)
CDR2 - SISTGGGNTYYRDSVKG,
and
(SEQ ID NO: 95)
CDR3 - QRNYYDGSYDYEGYTMDA,
and
b) a light chain variable region (VL) having complementarity determining regions (CDRs) with the following sequences:
(SEQ ID NO: 96)
CDR1 - RSSQDIGNYLT,
(SEQ ID NO: 97)
CDR2 - GAIKLED,
and
(SEQ ID NO: 98)
CDR3 - LQSIQYP.
2. An antigen-binding domain according to claim 1 , which comprises a VH domain having the sequence shown as SEQ ID NO: 65; and a VL domain having the sequence shown as SEQ ID NO: 66.
3. An anti-CD22 chimeric antigen receptor (CAR) which comprises an antigen-binding domain according to claim 1 .
4. A nucleic acid sequence which encodes a CAR according to claim 3 .
5. A nucleic acid construct which comprises a first nucleic acid sequence according to claim 4 and a second nucleic acid sequence encoding an anti-CD19 CAR.
6. A nucleic acid construct according to claim 5 , wherein the antigen binding domain of the anti-CD19 CAR comprises:
a) a heavy chain variable region (VH) having complementarity determining regions (CDRs) with the following sequences:
(SEQ ID NO: 69)
CDR1 - GYAFSSS,
(SEQ ID NO: 70)
CDR2 - YPGDED,
and
(SEQ ID NO: 71)
CDR3 - SLLYGDYLDY,
and
b) a light chain variable region (VL) having CDRs with the following sequences:
(SEQ ID NO: 72)
CDR1 - SASSSVSYMH,
(SEQ ID NO: 73)
CDR2 - DTSKLAS,
and
(SEQ ID NO: 74)
CDR3 - QQWNINPLT.
7. A nucleic acid construct according to claim 5 , wherein the antigen-binding domain of the anti-CD19 CAR comprises a VH domain as shown in SEQ ID NO: 75 and a VL domain as shown as SEQ ID NO: 76.
8. A nucleic acid construct according to claim 5 , wherein the anti-CD22 CAR is in an ScFv format, the nucleic acid construct having the general structure:
AgBD1-spacer1-TM1-endo1-coexpr-AgBD2-spacer2-TM2-endo2 or
AgBD2-spacer2-TM2-endo2-coexpr-AgBD1-spacer1-TM1-endo1,
wherein:
AgBD1 is a nucleic acid sequence encoding an antigen binding domain of the first CAR,
Spacer1 is a nucleic acid sequence encoding a spacer of the first CAR,
TM1 is a a nucleic acid sequence encoding a transmembrane domain of the first CAR,
Endo1 is a nucleic acid sequence encoding an endodomain of the first CAR,
Coexpr is a nucleic acid sequence enabling co-expression of the first and second CARs,
AgBD2 is a nucleic acid sequence encoding an antigen binding domain of the second CAR,
Spacer2 is a nucleic acid sequence encoding a spacer of the second CAR,
TM2 is a a nucleic acid sequence encoding a transmembrane domain of the second CAR, and
Endo2 is a nucleic acid sequence encoding an endodomain of the second CAR.
9. A vector which comprises a nucleic acid sequence according to claim 4 .
10. A method for making a cell, which comprises the step of introducing a nucleic acid sequence according to claim 4 into a cell ex vivo.
11. A cell which expresses a CAR according to claim 3 .
12. A pharmaceutical composition which comprises a plurality of cells according to claim 11 , together with a pharmaceutically acceptable carrier, diluent or excipient.
13. A cell which co-expresses a first CAR according to claim 3 and a second CAR which is an anti-CD19 CAR.
14. A cell according to claim 13 , wherein the antigen binding domain of the anti-CD19 CAR comprises
a) a heavy chain variable region (VH) having complementarity determining regions (CDRs) with the following sequences:
(SEQ ID NO: 69)
CDR1 - GYAFSSS,
(SEQ ID NO: 70)
CDR2 - YPGDED,
and
(SEQ ID NO: 71)
CDR3 - SLLYGDYLDY,
and
b) a light chain variable region (VL) having CDRs with the following sequences:
(SEQ ID NO: 72)
CDR1 - SASSSVSYMH,
(SEQ ID NO: 73)
CDR2 - DTSKLAS,
and
(SEQ ID NO: 74)
CDR3 - QQWNINPLT.
15. A cell according to claim 14 , wherein the antigen-binding domain of the anti-CD19 CAR comprises a VH domain as shown in SEQ ID NO: 75 and a VL domain as shown as SEQ ID NO: 76.
16. A method for treating a B cell leukemia or B cell lymphoma which comprises the step of administering a pharmaceutical composition according to claim 12 to a subject.