Chimeric antigen receptors for phagocytosis

Abstract

The present disclosure generally relate to novel chimeric antigen receptors (CARs) that bind an engulfment receptor expressed on the surface of a phagocytic cell and activate the endogenous phagocytic signaling pathway. Also disclosed are compositions and methods useful for producing such CARs, nucleic acids encoding same, phagocytic cells that have been modified to include a targeted effector activity directed towards a cell of interest such as, e.g., a cancer cell, as well as for modifying a cell and/or for the treatment of various health disorders such as cancer, including solid tumor and hematologic malignancy.

What is claimed is: 
     
         1 . A recombinant polynucleotide comprising a sequence encoding a chimeric antigen receptor (CAR), wherein the CAR comprises:
 (a) an extracellular domain comprising an antigen-binding region specific for a cell surface antigen,   (b) a transmembrane domain, and   (c) an intracellular domain comprising a p85-recruitment domain that binds a p85 regulatory subunit of phosphoinositide 3-kinase (PI3K).   
     
     
         2 . The recombinant polynucleotide of  claim 1 , wherein the p85-recruitment domain is from CD19, Gab2, IREM-1, PDGF receptor, CSFR-1, c-Kit, ErbB3 or CD7. 
     
     
         3 . The recombinant polynucleotide of  claim 1 , wherein the antigen-binding region comprises an amino acid sequence encoding an antibody or a binding fragment thereof selected from the group consisting of an antigen-binding fragment (Fab), a single-chain variable fragment (scFv), a nanobody, a VH domain, a VL domain, a single domain antibody (sdAb), a VNAR domain, and a VHH domain, a bispecific antibody and a diabody. 
     
     
         4 . The recombinant polynucleotide of  claim 1 , wherein the cell surface antigen is selected from the group consisting of CD19, CD22, HER2 (ERBB2/neu), Mesothelin, PSCA, CD123, CD30, CD171, CD138, CS-1, CLECL1, CD33, CD79b, EGFRvIII, GD2, GD3, BCMA, PSMA, ROR1, FLT3, TAG72, CD38, CD44v6, CEA, EPCAM, B7H3 (CD276), KIT (CD117), CD213A2, IL-1 IRa, PRSS21, VEGFR2, FSHR, TROP2, CD24, MUC-16, PDGFR-beta, SSEA-4, CD20, MUC1, EGFR, NCAM, Prostase, PAP, ELF2M, Ephrin B2, FAP, EphA2, GM3, TEM1/CD248, TEM7R, CLDN6, TSHR, GPRC5D, CD97, CD179a, ALK and IGLL1. 
     
     
         5 . The recombinant polynucleotide of  claim 1 , wherein the transmembrane domain comprises a stalk and/or transmembrane domain from CD8, Megf10, FcRγ, Bai1, MerTK, TIM4, Stabilin-1, Stabilin-2, RAGE, CD300f, Integrin subunit αv, Integrin subunit β5, CD36, LRP1, SCARF1, C1Qa, Axl, CD45 or CD86. 
     
     
         6 . The recombinant polynucleotide of  claim 1 , wherein the intracellular domain further comprises an intracellular signaling domain from an engulfment receptor. 
     
     
         7 . The recombinant polynucleotide of  claim 6 , wherein the p85-recruitment domain is operably linked downstream to the intracellular signaling domain from an engulfment receptor. 
     
     
         8 . The recombinant polynucleotide of  claim 6 , wherein the intracellular signaling domain from an engulfment receptor is an intracellular signaling domain from Megf10, FcRγ, Bai1, MerTK, TIM4, Stabilin-1, Stabilin-2, RAGE, CD300f, Integrin subunit av, Integrin subunit β5, CD36, LRP1, SCARF1, C1Qa, or Axl. 
     
     
         9 . The recombinant polynucleotide of  claim 1 , wherein the intracellular domain comprises at least one ITAM motif. 
     
     
         10 . The recombinant polynucleotide of  claim 1 , wherein the intracellular domain further comprises an intracellular signaling domain from CD3zeta. 
     
     
         11 . The recombinant polynucleotide of  claim 1 , wherein the recombinant polynucleotide further comprises a promoter sequence or a sequence encoding a signal peptide. 
     
     
         12 . The recombinant polynucleotide of  claim 1 , wherein the p85-recruitment domain comprises amino acids 345-379 of SEQ ID NO: 4. 
     
     
         13 . The recombinant polynucleotide of  claim 1 , wherein the CAR comprises SEQ ID NO: 5. 
     
     
         14 . The recombinant polynucleotide of  claim 1 , wherein the sequence encoding the CAR comprises SEQ ID NO: 14. 
     
     
         15 . The recombinant polynucleotide of  claim 1 , wherein the recombinant polynucleotide is selected from the group consisting of a viral vector, a DNA vector, an mRNA, a cDNA, a synthetic polynucleotide, and a plasmid. 
     
     
         16 . The recombinant polynucleotide of  claim 1 , wherein the recombinant polynucleotide is within or associated with a viral particle, a liposome, a lipid, a colloid, an exosome, a cell, a macromolecule complex, a nanocapsule, a microsphere, a bead, an oil-in-water emulsion and a micelle. 
     
     
         17 . The recombinant polynucleotide of  claim 1 , wherein the recombinant polynucleotide is isolated. 
     
     
         18 . A cell comprising the recombinant polynucleotide of  claim 1 . 
     
     
         19 . The cell of  claim 18 , wherein the cell is selected from the group consisting of a macrophage, a dendritic cell, a mast cell, a monocyte, a neutrophil, a microglia and an astrocyte. 
     
     
         20 . The cell of  claim 18 , wherein the cell is a human cell. 
     
     
         21 . A method of treating cancer in a subject in need thereof comprising administering the recombinant polynucleotide of  claim 1  to the subject, thereby treating the cancer in the subject. 
     
     
         22 . A method of treating cancer in a subject in need thereof comprising administering the cell of  claim 18  to the subject, thereby treating the cancer in the subject. 
     
     
         23 . A method of manufacturing a population of cells comprising:
 (a) providing a population of cells; and   (b) introducing a recombinant polynucleotide with a sequence encoding a chimeric antigen receptor (CAR) into the population of cells, wherein the CAR comprises:
 (i) an extracellular domain comprising an antigen-binding region specific for a cell surface antigen, 
 (ii) a transmembrane domain, and 
 (iii) an intracellular domain comprising a p85-recruitment domain that binds a p85 regulatory subunit of phosphoinositide 3-kinase (PI3K). 
   
     
     
         24 . The method of  claim 23 , wherein providing comprises obtaining a population of cells from a subject. 
     
     
         25 . The method of  claim 24 , wherein the subject has cancer. 
     
     
         26 . The method of  claim 23 , wherein introducing comprises transfection, lipofection, particle bombardment, microinjection, electroporation, iontophoresis, or infection with a viral vector. 
     
     
         27 . The method of  claim 23 , wherein the population if cells is selected from the group consisting of a population of macrophages, a population of dendritic cells, a population of mast cells, a population of monocytes, a population of neutrophils, a population of microglia and a population of astrocytes. 
     
     
         28 . The method of  claim 23 , wherein the p85-recruitment domain is from CD19, Gab2, IREM-1, PDGF receptor, CSFR-1, c-Kit, ErbB3 or CD7. 
     
     
         29 . The method of  claim 23 , wherein the intracellular domain further comprises an intracellular signaling domain from an engulfment receptor. 
     
     
         30 . The method of  claim 29 , wherein the p85-recruitment domain is operably linked downstream to the intracellular signaling domain from an engulfment receptor. 
     
     
         31 . The method of  claim 29 , wherein the intracellular signaling domain from an engulfment receptor is an intracellular signaling domain from Megf10, FcRγ, Bai1, MerTK, TIM4, Stabilin-1, Stabilin-2, RAGE, CD300f, Integrin subunit αv, Integrin subunit β5, CD36, LRP1, SCARF1, C1Qa, or Axl. 
     
     
         32 . The method of  claim 23 , wherein the intracellular domain comprises at least one ITAM motif. 
     
     
         33 . The method of  claim 23 , wherein the intracellular domain further comprises an intracellular signaling domain from CD3zeta