T-cell modulatory multimeric polypeptides and methods of use thereof

Abstract

The present disclosure provides variant immunomodulatory polypeptides, and fusion polypeptides comprising the variant immunomodulatory peptides. The present disclosure provides T-cell modulatory multimeric polypeptides, and compositions comprising same, where the T-cell modulatory multimeric polypeptides comprise a variant immunomodulatory polypeptide of the present disclosure. The present disclosure provides nucleic acids comprising nucleotide sequences encoding the T-cell modulatory multimeric polypeptides, and host cells comprising the nucleic acids. The present disclosure provides methods of modulating the activity of a T cell; the methods comprise contacting the T cell with a T-cell modulatory multimeric polypeptide of the present disclosure.

1 .- 132 . (canceled) 
     
     
         133 . A molecule comprising:
 a) a first polypeptide comprising:
 i) an HPV16 cancer-associated peptide epitope; and 
 ii) first class I major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class I MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         134 . A composition comprising one or more nucleic acids encoding the first and second polypeptides of  claim 133 . 
     
     
         135 . A method of producing the molecule of  claim 133 , the method comprising culturing a host cell comprising a composition according to  claim 134  in vitro in a culture medium under conditions such that the host cell produces the molecule. 
     
     
         136 . A method of activating an epitope-specific T cell, the method comprising contacting in vivo the T cell with the molecule of  claim 133 , wherein said contacting activates the epitope-specific T cell. 
     
     
         137 . A method of activating an epitope-specific T cell, the method comprising contacting ex vivo the T cell with the molecule of  claim 133 , wherein said contacting activates the epitope-specific T cell. 
     
     
         138 . A method of treating cancer in an individual, the method comprising administering to the individual an effective amount of the molecule of  claim 133 . 
     
     
         139 . A method of detecting, in a mixed population of T cells obtained from an individual, the presence of a target T cell that binds an epitope of interest, the method comprising:
 a) contacting in vitro the mixed population of T cells with the molecule of  claim 133 , wherein the molecule comprises the epitope of interest; and   b) detecting activation and/or proliferation of target T cells in response to said contacting.   
     
     
         140 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a cancer-associated peptide epitope other than an HPV16 cancer-associated epitope; and 
 ii) first class I major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class I MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL2 polypeptide comprising the IL2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         141 . A composition comprising one or more nucleic acids encoding the first and second polypeptides of  claim 140 . 
     
     
         142 . A method of producing the molecule of  claim 140 , the method comprising culturing a host cell comprising a composition according to  claim 141  in vitro in a culture medium under conditions such that the host cell produces the molecule. 
     
     
         143 . A method of activating an epitope-specific T cell, the method comprising contacting in vivo the T cell with the molecule of  claim 140 , wherein said contacting activates the epitope-specific T cell. 
     
     
         144 . A method of activating an epitope-specific T cell, the method comprising contacting ex vivo the T cell with the molecule of  claim 140 , wherein said contacting activates the epitope-specific T cell. 
     
     
         145 . A method of treating cancer in an individual, the method comprising administering to the individual an effective amount of the molecule of  claim 140 . 
     
     
         146 . A method of detecting, in a mixed population of T cells obtained from an individual, the presence of a target T cell that binds an epitope of interest, the method comprising:
 a) contacting in vitro the mixed population of T cells with the molecule of  claim 140 , wherein the molecule comprises the epitope of interest; and   b) detecting activation and/or proliferation of target T cells in response to said contacting.   
     
     
         147 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a peptide epitope present in a self antigen; and 
 ii) first class I major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class I MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         148 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a pathogen-associated peptide epitope; and 
 ii) first class I major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class I MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         149 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a peptide epitope other than an epitope of a cancer-associated antigen, a self-antigen, or a pathogen-associated epitope; and 
 ii) first class I major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class I MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         150 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a cancer-associated peptide epitope; and 
 ii) first class II major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class II MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         151 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a peptide epitope present in a self antigen; and 
 ii) first class II major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class II MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         152 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a pathogen-associated peptide epitope; and 
 ii) first class II major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class II MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.   
     
     
         153 . A molecule comprising:
 a) a first polypeptide comprising:
 i) a peptide epitope other than an epitope of a cancer-associated antigen, a self-antigen, or a pathogen-associated epitope; and 
 ii) first class II major histocompatibility complex (MHC) polypeptide; 
   b) a second polypeptide comprising a second class II MHC polypeptide; and   c) one or more immunomodulatory polypeptides,   wherein the first and second polypeptide form a heterodimer,   wherein the first and/or the second polypeptide comprises the one or more immunomodulatory polypeptides,   wherein the one or more immunomodulatory polypeptides comprises one or more variant interleukin-2 (IL-2) polypeptides comprising an amino acid sequence having at least 85% amino acid sequence identity to set forth in SEQ ID NO:1, and having one or more amino acid substitutions relative to set forth in SEQ ID NO:1, and   wherein at least one of the one or more immunomodulatory polypeptides that is a variant IL-2 polypeptide exhibits reduced binding affinity to an IL-2 receptor (IL2R) comprising alpha, beta, and gamma polypeptides having amino acid sequences set forth in SEQ ID NOs:54-56, compared to the binding affinity of a control IL-2 polypeptide comprising the IL-2 amino acid sequence set forth in SEQ ID NO:1 for the IL2R polypeptide.