Bispecific OR-gate chimeric antigen receptor responsive to CD19 and CD20
A CD19-OR-CD20 chimeric antigen receptor (CAR) protein construct is provided. Also provided are nucleic acids encoding the CD19-OR-CD20 CAR; and methods of use, e.g. in the treatment of B cell malignancies. The CD19-OR-CD20 CAR of the invention is a bispecific CAR that can trigger T-cell activation upon detection of either CD19 or CD20 (or both). It is a single molecule that confers two-input recognition capability upon human T cells engineered to stably express this CAR.
1. A polypeptide comprising a CD19-OR-CD20 chimeric antigen receptor (CAR), wherein the CAR comprises from N- to C-terminus:
a) an anti-CD20 scFv with a variable heavy domain and a variable light domain obtained from ofatumumab;
b) a (G 4 S) n linker, wherein n is 1 (SEQ ID NO: 15), 3 (SEQ ID NO: 16), or 4 (SEQ ID NO: 19);
c) an anti-CD19 scFv comprising a variable heavy domain from SEQ ID NO:4 and a variable light domain from SEQ ID NO:4;
d) a spacer of SEQ ID NO:5;
e) a transmembrane domain of SEQ ID NO:6;
f) a co-stimulatory domain of SEQ ID NO:8; and
g) a CD3-zeta cytoplasmic signaling domain of SEQ ID NO:9.
2. The polypeptide of claim 1 , wherein the (G 4 S) n linker is (G 4 S) 1 (SEQ ID NO: 15).
3. The polypeptide of claim 1 , wherein the (G 4 S) n linker is (G 4 S) 3 (SEQ ID NO: 16).
4. The polypeptide of claim 1 , wherein the (G 4 S) n linker is (G 4 S) 4 (SEQ ID NO: 19).
5. A nucleic acid encoding the polypeptide of claim 1 .
6. A viral vector comprising the nucleic acid of claim 5 .
7. The viral vector of claim 6 , wherein the vector is a lentiviral vector.
8. A cell comprising the polypeptide of claim 1 .
9. The cell of claim 8 , wherein the cell is a T cell, a NK cell, or a NKT cell.