Methods and compositions for cellular immunotherapy

Abstract

The present disclosure relates to genetically modified T cells comprising a transgene encoding an engineered antigen specific receptor, wherein expression of an endogenous gene selected from MNK1, MNK2, or both are inhibited in the genetically modified T cell in order to enhance central memory T cell subsets in cellular immunotherapy compositions.

1 - 32 . (canceled) 
     
     
         33 . A method of generating a modified T cell, the method comprising introducing a transgene encoding an engineered antigen specific receptor into a T cell collected from a subject treated with a MNK-specific inhibitor, thereby generating the modified T cell, wherein the subject treated with the MNK-specific inhibitor was treated 1, 2, 3, 4, 5, 6, 7, 14, 21, or 28 days prior to collection of the T cell from the subject, and wherein the MNK-specific inhibitor is a compound according to the following formula: 
       
         
           
           
               
               
           
         
       
       or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein:
 W 1  and W 2  are independently O, S or N—OR′, where R′ is lower alkyl; 
 Y is —N(R 5 )—, —O—, —S—, —C(O)—, —S═O, —S(O) 2 —, or —CHR 9 —; 
 R 1  is hydrogen, lower alkyl, cycloalkyl or heterocyclyl wherein any lower alkyl, cycloalkyl or heterocyclyl is optionally substituted with 1, 2 or 3 J groups; 
 n is 1, 2 or 3; 
 R 2  and R 3  are each independently hydrogen, alkyl, alkenyl, alkynyl, aryl, araalkylene, heteroaryl, heteroarylalkylene, cycloalkyl, cycloalkylalkylene, heterocyclyl, or heterocyclylalkylene, wherein any alkyl, aryl, araalkylene, heteroaryl, heteroarylalkylene, cycloalkyl, cycloalkylalkylene, heterocyclyl, or heterocyclylalkylene, is optionally substituted with 1, 2 or 3 J groups; 
 or R 2  and R 3  taken together with the carbon atom to which they are attached form a cycloalkyl or heterocyclyl, wherein any cycloalkyl or heterocyclyl is optionally substituted with 1, 2 or 3 J groups; 
 R 4a  and R 4b  are each independently hydrogen, halogen, hydroxyl, thiol, hydroxyalkylene, cyano, alkyl, alkoxy, acyl, thioalkyl, alkenyl, alkynyl, cycloalkyl, aryl, or heterocyclyl; 
 R 5  is hydrogen, cyano, or lower alkyl; 
 or R 5  and R 8  taken together with the atoms to which they are attached form a fused heterocyclyl optionally substituted with 1, 2 or 3 J groups; 
 R 6 , R 7  and R 8  are each independently hydrogen, hydroxy, halogen, cyano, amino, alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl, cycloalkylalkylene, cycloalkylalkenylene, alkylaminyl, alkylcarbonylaminyl, cycloalkylcarbonylaminyl, cycloalkylaminyl, heterocyclylaminyl, heteroaryl, or heterocyclyl, and wherein any amino, alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl, cycloalkylalkylene, cycloalkylalkenylene, amino, alkylaminyl, alkylcarbonylaminyl, cycloalkylcarbonylaminyl, cycloalkylaminyl, heterocyclylaminyl, heteroaryl, or heterocyclyl is optionally substituted with 1, 2 or 3 J groups; 
 or R 7  and R 8  taken together with the atoms to which they are attached form a fused heterocyclyl or heteroaryl optionally substituted with 1, 2 or 3 J groups; 
 J is —SH, —SR 9 , —S(O)R 9 , —S(O) 2 R 9 , —S(O)NH 2 , —S(O)NR 9 R 9 , —NH 2 , —NR 9 R 9 , —COOH, —C(O)OR 9 , —C(O)R 9 , —C(O)—NH 2 , —C(O)—NR 9 R 9 , hydroxy, cyano, halogen, acetyl, alkyl, lower alkyl, alkenyl, alkynyl, alkoxy, haloalkyl, thioalkyl, cyanoalkylene, alkylaminyl, NH 2 —C(O)-alkylene, NR 9 R 9 —C(O)-alkylene, —CHR 9 —C(O)-lower alkyl, —C(O)-lower alkyl, alkylcarbonylaminyl, cycloalkyl, cycloalkylalkylene, cycloalkylalkenylene, cycloalkylcarbonylaminyl, cycloalkylaminyl, —CHR 9 —C(O)-cycloalkyl, —C(O)-cycloalkyl, —CHR 9 —C(O)-aryl, —CHR 9 -aryl, —C(O)-aryl, —CHR 9 —C(O)-heterocycloalkyl, —C(O)-heterocycloalkyl, heterocyclylaminyl, or heterocyclyl; or any two J groups bound to the same carbon or hetero atom may be taken together to form oxo; and 
 R 9  is hydrogen, lower alkyl or —OH. 
 
     
     
         34 . The method of  claim 33 , further comprising contacting the collected T cell with the MNK-specific inhibitor sequentially, concurrently, or simultaneously with the introduction of the transgene encoding an engineered antigen specific receptor. 
     
     
         35 . The method of  claim 33 , wherein the method further comprises introducing an internal MNK-specific inhibitor into the T cell sequentially, concurrently, or simultaneously with the introduction of the transgene encoding an engineered antigen specific receptor, wherein expression of endogenous MNK1, MNK2, or both is inhibited in the T cell. 
     
     
         36 . (canceled) 
     
     
         37 . A method of generating a modified T cell, the method comprising contacting a T cell with a MNK-specific inhibitor; and introducing a transgene encoding an engineered antigen specific receptor into the T cell, wherein the contacting step occurs simultaneously, concurrently, sequentially with the introduction of the transgene encoding the engineered antigen specific receptor into the T cell, thereby generating the modified T cell, wherein the MNK-specific inhibitor is a compound according to the following formula: 
       
         
           
           
               
               
           
         
       
       or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein:
 W 1  and W 2  are independently 0, S or N—OR′, where R′ is lower alkyl; 
 Y is —N(R 5 )—, —O—, —S—, —C(O)—, —S═O, —S(O) 2 —, or —CHR 9 —; 
 R 1  is hydrogen, lower alkyl, cycloalkyl or heterocyclyl wherein any lower alkyl, cycloalkyl or heterocyclyl is optionally substituted with 1, 2 or 3 J groups; 
 n is 1, 2 or 3; 
 R 2  and R 3  are each independently hydrogen, alkyl, alkenyl, alkynyl, aryl, araalkylene, heteroaryl, heteroarylalkylene, cycloalkyl, cycloalkylalkylene, heterocyclyl, or heterocyclylalkylene, wherein any alkyl, aryl, araalkylene, heteroaryl, heteroarylalkylene, cycloalkyl, cycloalkylalkylene, heterocyclyl, or heterocyclylalkylene, is optionally substituted with 1, 2 or 3 J groups; 
 or R 2  and R 3  taken together with the carbon atom to which they are attached form a cycloalkyl or heterocyclyl, wherein any cycloalkyl or heterocyclyl is optionally substituted with 1, 2 or 3 J groups; 
 R 4a  and R 4b  are each independently hydrogen, halogen, hydroxyl, thiol, hydroxyalkylene, cyano, alkyl, alkoxy, acyl, thioalkyl, alkenyl, alkynyl, cycloalkyl, aryl, or heterocyclyl; 
 R 5  is hydrogen, cyano, or lower alkyl; 
 or R 5  and R 8  taken together with the atoms to which they are attached form a fused heterocyclyl optionally substituted with 1, 2 or 3 J groups; 
 R 6 , R 7  and R 8  are each independently hydrogen, hydroxy, halogen, cyano, amino, alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl, cycloalkylalkylene, cycloalkylalkenylene, alkylaminyl, alkylcarbonylaminyl, cycloalkylcarbonylaminyl, cycloalkylaminyl, heterocyclylaminyl, heteroaryl, or heterocyclyl, and wherein any amino, alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl, cycloalkylalkylene, cycloalkylalkenylene, amino, alkylaminyl, alkylcarbonylaminyl, cycloalkylcarbonylaminyl, cycloalkylaminyl, heterocyclylaminyl, heteroaryl, or heterocyclyl is optionally substituted with 1, 2 or 3 J groups; 
 or R 7  and R 8  taken together with the atoms to which they are attached form a fused heterocyclyl or heteroaryl optionally substituted with 1, 2 or 3 J groups; 
 J is —SH, —SR 9 , —S(O)R 9 , —S(O) 2 R 9 , —S(O)NH 2 , —S(O)NR 9 R 9 , —NH 2 , —NR 9 R 9 , —COOH, —C(O)OR 9 , —C(O)R 9 , —C(O)—NH 2 , —C(O)—NR 9 R 9 , hydroxy, cyano, halogen, acetyl, alkyl, lower alkyl, alkenyl, alkynyl, alkoxy, haloalkyl, thioalkyl, cyanoalkylene, alkylaminyl, NH 2 —C(O)-alkylene, NR 9 R 9 —C(O)-alkylene, —CHR 9 —C(O)-lower alkyl, —C(O)-lower alkyl, alkylcarbonylaminyl, cycloalkyl, cycloalkylalkylene, cycloalkylalkenylene, cycloalkylcarbonylaminyl, cycloalkylaminyl, —CHR 9 —C(O)-cycloalkyl, —C(O)-cycloalkyl, —CHR 9 —C(O)-aryl, —CHR 9 -aryl, —C(O)-aryl, —CHR 9 —C(O)-heterocycloalkyl, —C(O)-heterocycloalkyl, heterocyclylaminyl, or heterocyclyl; or any two J groups bound to the same carbon or hetero atom may be taken together to form oxo; and 
 R 9  is hydrogen, lower alkyl or —OH. 
 
     
     
         38 . The method of  claim 37 , wherein the contacting step and introduction step occur ex vivo. 
     
     
         39 . The method of  claim 37 , further comprising introducing an internal MNK-specific inhibitor into the T cell, wherein expression of endogenous MNK1, MNK2, or both is inhibited in the T cell. 
     
     
         40 . The method according to  claim 39 , wherein the internal MNK-specific inhibitor is introduced into the T cell simultaneously, concurrently, sequentially with the transgene encoding the engineered antigen specific receptor. 
     
     
         41 . A method of generating a modified T cell, the method comprising introducing a transgene encoding an engineered antigen specific receptor into a T cell, and introducing an internal MNK-specific inhibitor, wherein expression of endogenous MNK1, MNK2, or both is inhibited in the T cell, thereby generating the modified T cell. 
     
     
         42 . The method of  claim 37 , wherein the engineered antigen specific receptor is a chimeric antigen receptor (CAR), an engineered T cell receptor (TCR), a TCR-CAR, or any combination thereof. 
     
     
         43 - 47 . (canceled) 
     
     
         48 . The method of  claim 37 , wherein the MNK-specific inhibitor is a compound according to the following formula: 
       
         
           
           
               
               
           
         
       
     
     
         49 - 51 . (canceled) 
     
     
         52 . The method of  claim 41 , wherein the internal INK-specific inhibitor is an inhibitory nucleic acid. 
     
     
         53 . (canceled) 
     
     
         54 . The method of  claim 41 , wherein the internal MNK-specific inhibitor is a chromosomal MNK gene knock out by chromosomal editing via an endonuclease selected from a CRISPR/Cas nuclease system, a zinc finger nuclease (ZFN), a Transcription Activator Like Effector nuclease (TALEN), and a meganuclease. 
     
     
         55 - 59 . (canceled) 
     
     
         60 . The method of  claim 37 , further comprising inhibiting expression of an endogenous gene selected from a TCR gene, an HLA gene, an immunosuppression component gene, or any combination thereof. 
     
     
         61 - 63 . (canceled) 
     
     
         64 . The method of  claim 37 , wherein the antigen is a tumor antigen, a pathogenic microorganism antigen, a neurodegenerative disease antigen, or an autoimmune disease antigen. 
     
     
         65 . (canceled) 
     
     
         66 . The method of  claim 37 , wherein the T cell is a CD4+ T cell or a CD8+ T cell. 
     
     
         67 - 88 . (canceled) 
     
     
         89 . The method of  claim 41 , wherein the engineered antigen specific receptor is a chimeric antigen receptor (CAR), an engineered T cell receptor (TCR), a TCR-CAR, or any combination thereof. 
     
     
         90 . The method of  claim 33 , wherein the MNK-specific inhibitor is a compound according to the following formula: 
       
         
           
           
               
               
           
         
       
     
     
         91 . The method of  claim 41 , further comprising inhibiting expression of an endogenous gene selected from a TCR gene, an HLA gene, an immunosuppression component gene, or any combination thereof. 
     
     
         92 . The method of  claim 41 , wherein the antigen is a tumor antigen, a pathogenic microorganism antigen, a neurodegenerative disease antigen, or an autoimmune disease antigen. 
     
     
         93 . The method of  claim 41 , wherein the T cell is a CD4+ T cell or a CD8+ T cell.