Antigen binding polypeptides
The present disclosure provides compositions comprising antigen binding polypeptides. More particularly, the disclosure relates to polypeptides comprising antibodies or antigen binding fragments thereof, nucleic acids encoding the polypeptides, and vectors for expressing the same.
1. An antibody or antigen binding fragment thereof comprising a heavy chain variable region (VH) comprising a CDRH1, a CDRH2, and a CDRH3 comprising the amino acid sequences set forth in SEQ ID NOs: 82, 83, and 84; a polypeptide linker; and a light chain variable region (VL) comprising a CDRL1, a CDRL2, and a CDRL3 comprising the amino acid sequences set forth in SEQ ID NOs: 86, 87, and 88.
2. The antibody or antigen binding fragment thereof of claim 1 , wherein the polypeptide linker is selected from the group consisting of: TGEKP (SEQ ID NO: 2); (GGGGS) n wherein n=1, 2, 3, 4 or 5 (SEQ ID NOs: 3, 976-979); EGKSSGSGSESKVD (SEQ ID NO: 4); KESGSVSSEQLAQFRSLD (SEQ ID NO: 5); LRQRDGERP (SEQ ID NO: 6); LRQKDGGGSERP (SEQ ID NO: 7); LRQKD(GGGS) 2 ERP (SEQ ID NO: 8), GEGTSTGSGGSGGSGGAD (SEQ ID NO: 9), and GSTSGSGKPGSGEGSTKG (SEQ ID NO: 10) and variants thereof comprising an amino acid sequence 95% identical thereto.
3. The antibody or antigen binding fragment thereof of claim 1 , wherein the VH comprises the amino acid sequence set forth in SEQ ID NO: 81 and the VL comprises the amino acid sequence set forth in SEQ ID NO: 85.
4. The antibody or antigen binding fragment thereof of claim 1 , wherein the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO: 89 or 90.
5. A bispecific antibody comprising the antibody or antigen binding fragment thereof of claim 1 .
6. The bispecific antibody of claim 5 , further comprising an anti-CD3 antibody that binds CD3δ, CD3ε, CD3γ, or CD3ζ.
7. An antibody conjugate comprising the antibody or antigen binding fragment thereof of claim 1 conjugated to a cytotoxic agent.
8. The antibody conjugate of claim 7 , wherein:
(a) the cytotoxic agent is a toxin selected from the group consisting of: saporin, diphtheria toxin, pseudomonas exotoxin A, Ricin A chain derivatives, a small molecule toxin, and combinations thereof;
(b) the cytotoxic agent is a radioisotope selected from the group consisting of: 131I, 90Y, 177Lu, 188Re, 67Cu, 213Bi, 211At, and 227Ac;
(c) the cytotoxic agent is an RNA polymerase II inhibitor and/or RNA polymerase III inhibitor selected from the group consisting of: an amatoxin, α-amanitin, β-amanitin, γ-amanitin, ¿-amanitin, amanin, amaninamide, amanullin, amanullinic acid and any functional fragments, derivatives or analogs thereof, or
(d) the cytotoxic agent is a DNA-damaging agent selected from the group consisting of: an antitubulin agent, a DNA crosslinking agent, a DNA alkylating agent and a mitotic disrupting agent.
9. A chimeric antigen receptor (CAR) comprising the antibody or antigen binding fragment thereof of claim 1 , a spacer domain, a transmembrane domain, and one or more intracellular signaling domains.
10. The CAR of claim 9 , wherein:
(a) the spacer domain comprises a hinge domain or fragment thereof selected from the group consisting of: a CD4 hinge, a CD8β hinge, a CD8α hinge, a CD28 hinge, a CD134 hinge, a CD137 hinge, a CD152 hinge, a CD278 hinge, an IgG1 hinge, an IgG2 hinge, an IgG3 hinge, and an IgG4 hinge;
(b) the transmembrane domain is isolated or derived from a polypeptide selected from the group consisting of an alpha, beta, gamma, or delta chain of the T-cell receptor, CD3δ, CD3ε, CD3γ, CD3ζ, CD4, CD5, CD8α, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1);
(c) the one or more intracellular signaling domains comprises a primary signaling domain isolated or derived from a polypeptide selected from the group consisting of FcRγ, FcRβ, CD3γ, CD3δ, CD3ε, CD3ζ, CD22, CD79a, CD79b, and CD66d; and
(d) the one or more intracellular signaling domains comprises a costimulatory signaling domain isolated or derived from a polypeptide selected from the group consisting of TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD28, CD30, CD40, ICAM, CD83, CD94, CD134 (OX40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, SLP76, TRAT1, TNFR2, TNFRS14, TNFRS18, TNFRS25, and ZAP70.
11. A CAR comprising:
(a) an antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO: 89 or 90;
(b) a spacer domain comprising a hinge domain or fragment thereof selected from the group consisting of: a CD8α hinge, a CD28 hinge, and an IgG4 hinge;
(c) a transmembrane domain isolated from a polypeptide selected from the group consisting of: CD8α and CD28;
(d) one or more intracellular signaling domains comprising a costimulatory signaling domain isolated from a polypeptide selected from the group consisting of: CD28, CD137 (4-1BB), and CD278 (ICOS); and
(e) a primary signaling domain isolated from CD3ζ.
12. The CAR of claim 11 , wherein the CAR comprises the amino acid sequence set forth in any one of SEQ ID NOs: 501, 502, 504, 505, 510, 511, 525, 526, 528, 529, 534, and 535.
13. A polynucleotide encoding the antibody or antigen binding fragment thereof of claim 1 .
14. A polynucleotide encoding the CAR of claim 11 .
15. A polynucleotide encoding the CAR of claim 12 .
16. A polynucleotide encoding or comprising a promoter operably linked to a polynucleotide encoding the CAR of claim 11 .
17. The polynucleotide of claim 16 , wherein the promoter is a CAG promoter, an EF1α promoter, an MNDU3 promoter, a PGK1 promoter, an SFFV promoter, or a UBC promoter.
18. A DNA comprising a polynucleotide encoding the CAR of claim 11 .
19. An RNA encoded by the DNA of claim 18 .
20. A vector comprising a polynucleotide encoding the CAR of claim 11 .
21. A vector encoding or comprising an MNDU3 or EF1α promoter operably linked to a polynucleotide encoding the CAR of claim 11 .
22. The vector of claim 21 , wherein the vector is a plasmid, transfer plasmid or viral vector.
23. The vector of claim 21 , wherein the vector is a viral vector selected from the group consisting of an adenoviral (Ad) vector, an adeno-associated virus (AAV) vector, a herpes simplex virus (HSV) vector, a parvovirus vector, a rhabdovirus vector, a vesiculovirus vector, a paramyxovirus vector, a morbillovirus vector, a henipavirus vector, an alphavirus vector, a flavivirus vector, a retroviral vector, and a lentiviral vector (LVV).
24. The vector of claim 23 , wherein the lentiviral vector is engineered or derived from the genome of a lentivirus selected from the group consisting of: HIV (HIV type 1 or HIV type 2); visna-maedi virus (VMV); caprine arthritis-encephalitis virus (CAEV); equine infectious anemia virus (EIAV); feline immunodeficiency virus (FIV); bovine immune deficiency virus (BIV); and simian immunodeficiency virus (SIV).
25. A lentiviral vector comprising: a 5′ long terminal repeat (LTR) comprising R and U5 regions; a Psi (Y) packaging signal; a cPPT/FLAP; an export element; a polynucleotide encoding or comprising a promoter operably linked to the polynucleotide encoding a signal peptide and the CAR of claim 11 ; a 3′ LTR comprising U3 and R regions; a polyadenylation signal, and a poly(A) tail.
26. A recombinant lentivirus comprising one or more copies of the lentiviral vector of claim 25 .
27. A composition comprising the recombinant lentivirus of claim 26 .
28. An RNA comprising: a 5′ long terminal repeat (LTR) comprising R and U5 regions; a Psi (Ψ) packaging signal; a cPPT/FLAP; an export element; a polynucleotide encoding a promoter operably linked to a polynucleotide encoding a signal peptide and the CAR of claim 11 ; a 3′ LTR comprising U3 and R regions; a polyadenylation signal, and a poly(A) tail.
29. A recombinant lentivirus comprising one or more copies of the RNA of claim 28 .
30. A composition comprising the recombinant lentivirus of claim 29 .