Regulatable chimeric antigen receptor

Abstract

Compositions and methods relating to regulatable chimeric antigen receptors (RCARs), where the intracellular signaling or proliferation of the RCAR can be controlled to optimize the use of an RCAR-expressing cell to provide an immune response, are provided. For example, a RCAR can comprise a dimerization switch that, upon the presence of a dimerization molecule, can couple an intracellular signaling domain to an extracellular recognition element, e.g., an antigen binding domain, an inhibitory counter ligand binding domain, or costimulatory ECD domain. An RCAR can be engineered to include an appropriate antigen binding domain that is specific to a desired antigen target and used in the treatment of a disease.

1 . A regulatable chimeric antigen receptor (RCAR), wherein the RCAR comprises:
 a) an intracellular signaling member comprising:
 an intracellular signaling domain, and 
 a first switch domain; 
   b) an antigen binding member comprising:
 an antigen binding domain, and 
 a second switch domain; and 
   c) a transmembrane domain, wherein,   (i) the antigen binding member comprises more than one intracellular signaling domains;   (ii) the first and second switch domains comprise a FKBP-FRB based switch, which comprises a switch domain comprising a FRB binding fragment or analog of FKBP and a switch domain comprising an FKBP binding fragment or analog of FRB, and the FKBP binding fragment or analog of FRB comprises one or more mutations;   (iii) the RCAR comprises four intracellular signaling domain wherein, (A) one of the first, second, third and fourth intracellular signaling domain is a primary intracellular signaling domain selected from the list in Table 1 and the other three are costimulatory signaling domains selected from the list in Table 2, or (B) two of the first, second, third, and fourth intracellular signaling domains are primary intracellular signaling domains selected from the list in Table 1, and the other two are costimulatory domain selected from the list in Table 2;   (iv) the antigen binding domain does not comprise a transmembrane domain or membrane anchoring domain;   (v) the RCAR further comprises c) an auxiliary antigen binding member comprising:
 an antigen binding domain that binds a second antigen; and 
 a transmembrane domain or membrane anchoring domain; 
   (vi) the RCAR further comprises
 an inhibitory counter ligand binding member comprising, 
 an inhibitory counter ligand binding domain from Table 4, and 
 a transmembrane domain or membrane anchoring domain; 
   (vii) the RCAR further comprises an inhibitory CAR (iCAR) member, wherein the iCAR member comprises:
 an antigen binding domain (or other extracellular domain) that recognizes an antigen on 
 a non-target cell; 
 a transmembrane domain; and, 
 a domain from an inhibitory molecule listed in Table 12; 
   (viii) the RCAR further comprising a second RCAR, or second antigen binding member, wherein the antigen binding domain of one of the RCARs or antigen binding members does not comprise a variable light domain and a variable heavy domain and the other is not an scFv;   (ix) the antigen binding member comprises: an antigen binding domain; a transmembrane domain; a first intracellular switch domain; and a primary signaling domain, or a costimulatory signaling domain;   (x) the first and second switch domains can form an extracellular dimerization switch; or   (xi) the RCAR comprises an inhibitor of an inhibitory molecule chosen from Table 3.   
     
     
         2 . The RCAR of  claim 1 , wherein the antigen binding member comprises: a plurality of costimulatory signaling domains chosen from Table 2, and no primary intracellular signaling domain. 
     
     
         3 .- 6 . (canceled) 
     
     
         7 . The RCAR of  claim 1 , wherein the intracellular signaling domain on the intracellular signaling member is a primary intracellular signaling domain from Table 1. 
     
     
         8 . (canceled) 
     
     
         9 . The RCAR of  claim 1 , wherein the intracellular signaling domain on the intracellular signaling member is a costimulatory signaling domain selected from Table 2. 
     
     
         10 .- 18 . (canceled) 
     
     
         19 . The RCAR of  claim 1 , wherein the RCAR comprises:
 a) an inhibitory extracellular domain member comprising,   an inhibitory extracellular domain selected from Table 4,   a transmembrane region, and   a switch domain;   b) an intracellular signaling member comprising,   an intracellular signaling domain, and   a switch domain.   
     
     
         20 . (canceled) 
     
     
         21 . The RCAR of  claim 1 , wherein the RCAR comprises:
 a) a costimulatory extracellular domain (ECD) member comprising   a costimulatory ECD domain selected from Table 5, other than that of CD27;   a transmembrane region, and   a switch domain;   b) an intracellular signaling member comprising   an intracellular signaling domain, and   a switch domain.   
     
     
         22 . (canceled) 
     
     
         23 . The RCAR of  claim 1 , wherein the RCAR comprises:
 a) an intracellular signaling member comprising:   a transmembrane domain or membrane anchoring domain;   a co-stimulatory signaling domain, selected from Table 2, and   a switch domain; and   b) an antigen binding member comprising:   an antigen binding domain,   a transmembrane domain, and   a primary intracellular signaling domain selected from Table 1, wherein the antigen binding member does not comprise a switch domain, or does not comprise a switch domain that dimerizes with a switch domain on the intracellular signaling member.   
     
     
         24 . A nucleic acid encoding a RCAR, or an element of an RCAR of  claim 1 . 
     
     
         25 .- 26 . (canceled) 
     
     
         27 . A nucleic acid encoding an RCAR comprising:
 a) an intracellular signaling member comprising:   an intracellular signaling domain,   a first switch domain, and   optionally, a transmembrane domain;   b) an antigen binding member comprising:   an antigen binding domain, and   a second switch domain, and   optionally, a transmembrane domain; and   wherein:   i) a sequence encoding the antigen binding member (a) and a sequence encoding the intracellular signaling member (b), are present on a single nucleic acid molecule, are transcribed as a single transcription product, and are configured as follows:   a promoter, is operably linked to (a), (b), and (c), wherein (c) encodes a cleavable peptide, and element (c) is disposed between (a) and (b); or   ii) a sequence encoding the antigen binding member (a) and a sequence encoding the intracellular signaling member (b), are present on a single nucleic acid molecule, are transcribed as a single transcription product, and are configured as follows:   a promoter is operably linked to (a), (b), and (c), wherein element (c) encodes an IRES, and element (c) is disposed between (a) and (b).   
     
     
         28 . A vector system comprising a nucleic acid of  claim 24 . 
     
     
         29 . (canceled) 
     
     
         30 . A cell comprising an RCAR of  claim 1 . 
     
     
         31 .- 33 . (canceled) 
     
     
         34 . A method of making a cell comprising an RCAR, comprising introducing a nucleic acid of  claim 24  into said cell. 
     
     
         35 . A method of treating a subject comprising administering to the subject an effective amount of a RCARX cell of  claim 30 , or providing a subject comprising the cell. 
     
     
         36 . The method of  claim 35 , wherein the RCARX cell is an autologous T cell, an allogeneic T cell, an autologous NK cell, or an allogeneic NK cell. 
     
     
         37 .- 38 . (canceled) 
     
     
         39 . The method of  claim 35 , comprising treating the subject for cancer, wherein the subject is a human. 
     
     
         40 . (canceled) 
     
     
         41 . The method of  claim 35 , comprising administering a dimerization molecule to the subject. 
     
     
         42 .- 43 . (canceled) 
     
     
         44 . A method of treating a subject comprising:
 (a) administering to the subject an effective amount of a RCARX cell, or providing a subject comprising the RCARX cell, wherein the cell comprises: an RCAR, a nucleic acid encoding the RCAR, or a nucleic acid encoding the RCAR; and   (b) administering   (i) a dimerization molecule which comprises RADOOI; or   (ii) a low, immune enhancing, dose of an allosteric mTOR inhibitor, wherein the RCAR comprises:   A) an intracellular signaling member comprising:   an intracellular signaling domain, and   a first switch domain;   B) an antigen binding member comprising:   an antigen binding domain, and   a second switch domain; and   C) a transmembrane domain,   wherein the first and second switch domains comprise a FKBP-FRB based switch.   
     
     
         45 . (canceled) 
     
     
         46 . The method of  claim 44 , wherein the RCARX cell is an autologous T cell, an allogeneic T cell, an autologous NK cell, or an allogeneic NK cell. 
     
     
         47 .- 48 . (canceled) 
     
     
         49 . The method of  claim 44 , comprising treating the subject for cancer, wherein the subject is a human. 
     
     
         50 .- 52 . (canceled) 
     
     
         53 . A method of providing an RCARX cell comprising:
 providing an immune effector cell to a recipient entity; and   receiving from said entity, an RCARX cell derived from said immune effector cell, or a daughter cell thereof, wherein the RCARX comprises an RCAR of  claim 1 , or a nucleic acid or vector encoding the RCAR.   
     
     
         54 .- 71 . (canceled)