Synergistic treatment of cancer using immunomers in conjunction with chemotherapeutic agents

Abstract

The invention relates to the therapeutic use of immunostimulatory oligonucleotides and/or immunomers in combination with chemotherapeutic agents to provide a synergistic therapeutic effect.

1 . A method for treating cancer in a cancer patient comprising administering to the patient an immunomer in combination with a chemotherapeutic agent,
 wherein the immunomer comprises at least one immunostimulatory oligonucleotide having the structure
   5′-Nn-N1-Y—Z—N1-Nn-3′  (III)
 
   wherein:   YZ is an immunostimulatory dinucleotide and   Y is cytidine, 2′ deoxycytidine, arabinocytidine, 2′-deoxythymidine, 2′-deoxy-2′-substitutedarabinocytidine, 2′-O-substitutedarabinocytidine, 2′-deoxy-5-hydroxycytidine, 2′-deoxy-N4-alkyl-cytidine, 2′-deoxy-4-thiouridine, other non-natural pyrimidine nucleosides, or 1-(2′-deoxy-β-D-ribofuranosyl)-2-oxo-7-deaza-8-methyl-purine;   Z is guanosine or 2′-deoxyguanosine, 2′ deoxy-7-deazaguanosine, 2′-deoxy-6-thioguanosine, arabinoguanosine, 2′-deoxy-2′ substituted-arabinoguanosine, 2′-O-substituted-arabinoguanosine, 2′ deoxyinosine or other non-natural purine nucleoside,   provided however, that when Y is cytidine or 2′ deoxycytidine then Z is not guanosine or 2′ deoxyguanosine, and when Z is guanosine then Y is not cytidine or 2′ deoxycytidine,   N1, at each occurrence, is a naturally occurring or a synthetic nucleoside or an immunostimulatory moiety selected from the group consisting of abasic nucleosides, arabinonucleosides, 2′-deoxyuridine, α-deoxyribonucleosides,   β-L-deoxyribonucleosides, and nucleosides linked by a phosphodiester or modified internucleoside linkage to the adjacent nucleoside on the 3′ side, the modified internucleotide linkage being selected from, without limitation, a linker having a length of from about 2 angstroms to about 200 angstroms, C2-C18 alkyl linker, poly(ethylene glycol) linker, 2-aminobutyl-1,3-propanediol linker, glyceryl linker, 2′-5′ internucleoside linkage, and phosphorothioate, phosphorodithioate, or methylphosphonate internucleoside linkage;   Nn, at each occurrence, is a naturally occurring nucleoside or synthetic nucleoside or an immunostimulatory moiety selected from the group consisting of abasic nucleosides, arabinonucleosides, 2′-deoxyuridine, α-deoxyribonucleosides, 2′-O-substituted ribonucleosides, and nucleosides linked by a modified internucleoside linkage to the adjacent nucleoside on the 3′ side, the modified internucleoside linkage preferably being selected from the group consisting of amino linker, C2-C18 alkyl linker, poly(ethylene glycol) linker, 2-aminobutyl-1,3-propanediol linker, glyceryl linker, 2′-5′ internucleoside linkage, and methylphosphonate internucleoside linkage;   wherein each n is independently a number from 0 to 30; and   wherein the 3′ end is linked directly or via a non-nucleotidic linker to another oligonucleotide, which may or may not be immunostimulatory.   wherein the immunomer comprises at least two linked oligonucleotides and has more than one 5′ end, wherein at least one of the oligonucleotides is an immunostimulatory oligonucleotide having an accessible 5′ end and comprises an immunostimulatory dinucleotide selected from the group consisting of C*pG, CpG* and C*pG*;   wherein C is cytidine or 2′ deoxycytidine, wherein C* is selected from arabinocytidine, 2′-deoxythymidine, 2′-deoxy-2′-substitutedarabinocytidine, 2′-O-substitutedarabinocytidine, 2′-deoxy-5-hydroxycytidine, 2′-deoxy-N4-alkyl-cytidine, 2′-deoxy-4-thiouridine, other non-natural pyrimidine nucleosides, or 1-(2′-deoxy-β-D-ribofuranosyl)-2-oxo-7-deaza-8-methyl-purine;   wherein G is guanosine or 2′ deoxyguanosine, wherein G* is selected from 2′ deoxy-7-deazaguanosine, 2′-deoxy-6-thioguanosine, arabinoguanosine, 2′-deoxy-2′ substituted-arabinoguanosine, 2′-O-substituted-arabinoguanosine, 2′ deoxyinosine or other non-natural purine nucleoside;   wherein the chemotherapeutic agent is selected from the group consisting of gemcitabine, taxotere, doxorubicin, docetaxel, herceptin and rituxan;   wherein the combination of the immunomer and chemotherapeutic agent create an enhanced anti-cancer effect relative to the immunomer or the chemotherapeutic agent alone.   
     
     
         2 . The method according to  claim 1  wherein the purine is 6-thioguanine or 7-deazaguanine. 
     
     
         3 . The method according to  claim 1  wherein the non-naturally occurring pyrimidine base is selected from the group consisting of 5-hydroxycytosine, 5-hydroxymethylcytosine, N4-alkylcytosine, and 4-thiouracil. 
     
     
         4 . The method according to  claim 1 , wherein the non-nucleotidic linker selected from the group consisting of a linker from about 2 angstroms to about 200 angstroms in length, a metal, a soluble or insoluble biodegradable polymer bead, an organic moiety having functional groups that permit attachment to the 3′-terminal nucleoside of the oligonucleotide, a biomolecule, a cyclic or acyclic small molecule, an aliphatic or aromatic hydrocarbon, either of which optionally can include, either in the linear chain connecting the oligonucleotides or appended to it, one or more functional groups selected from the group consisting of hydroxy, amino, thiol, thioether, ether, amide, thioamide, ester, urea, and thiourea; amino acids, carbohydrates, cyclodextrins, adamantane, cholesterol, haptens, antibiotics, glycerol and a glycerol homolog of the formula HO—(CH 2 ) o —CH(OH)—(CH 2 ) p —OH, wherein o and p independently are integers from 1 to about 6, and a derivative of 1,3-diamino-2-hydroxypropane. 
     
     
         5 . The method of  claim 1  further comprising administering a vaccine. 
     
     
         6 . The method of  claim 5  wherein the immunomer or the vaccine, or both, are linked to an immunogenic protein. 
     
     
         7 . The method of  claim 5  further comprising administering an adjuvant. 
     
     
         8 . The method according to  claim 1 , wherein at least one N1 or Nn is an immunostimulatory moiety. 
     
     
         9 . A pharmaceutical formulation comprising an immunomer, a chemotherapeutic agent and a physiologically acceptable carrier; wherein the immunomer comprises at least two oligonucleotides linked by a non-nucleotidic linker and having more than one 5′ end, wherein at least one of the oligonucleotides is an immunostimulatory oligonucleotide having an accessible 5′ end
 wherein at least one oligonucleotide of the immunomer has the structure
   5′-Nn-N1-Y—Z—N1-Nn-3′  (III)
 
 
 wherein: 
 YZ is an immunostimulatory dinucleotide and 
 Y is cytidine, 2′ deoxycytidine, arabinocytidine, 2′-deoxythymidine, 2′-deoxy-2′-substitutedarabinocytidine, 2′-O-substitutedarabinocytidine, 2′-deoxy-5-hydroxycytidine, 2′-deoxy-N4-alkyl-cytidine, 2′-deoxy-4-thiouridine, other non-natural pyrimidine nucleosides, or 1-(2′-deoxy-β-D-ribofuranosyl)-2-oxo-7-deaza-8-methyl-purine; 
 Z is guanosine or 2′-deoxyguanosine, 2′ deoxy-7-deazaguanosine, 2′-deoxy-6-thioguanosine, arabinoguanosine, 2′-deoxy-2′ substituted-arabinoguanosine, 2′-O-substituted-arabinoguanosine, 2′ deoxyinosine or other non-natural purine nucleoside, 
 provided, however that when Y is cytidine or 2′ deoxycytidine, then Z is not guanosine or 2′ deoxyguanosine, 
 N1, at each occurrence, is a naturally occurring or a synthetic nucleoside or an immunostimulatory moiety selected from the group consisting of abasic nucleosides, arabinonucleosides, 2′-deoxyuridine, α-deoxyribonucleosides, 
 β-L-deoxyribonucleosides, and nucleosides linked by a phosphodiester or modified internucleoside linkage to the adjacent nucleoside on the 3′ side, the modified internucleotide linkage being selected from, without limitation, a linker having a length of from about 2 angstroms to about 200 angstroms, C2-C18 alkyl linker, poly(ethylene glycol) linker, 2-aminobutyl-1,3-propanediol linker, glyceryl linker, 2′-5′ internucleoside linkage, and phosphorothioate, phosphorodithioate, or methylphosphonate internucleoside linkage; 
 Nn, at each occurrence, is a naturally occurring nucleoside or synthetic nucleoside or an immunostimulatory moiety selected from the group consisting of abasic nucleosides, arabinonucleosides, 2′-deoxyuridine, α-deoxyribonucleosides, 2′-O-substituted ribonucleosides, and nucleosides linked by a modified internucleoside linkage to the adjacent nucleoside on the 3′ side, the modified internucleoside linkage preferably being selected from the group consisting of amino linker, C2-C18 alkyl linker, poly(ethylene glycol) linker, 2-aminobutyl-1,3-propanediol linker, glyceryl linker, 2′-5′ internucleoside linkage, and methylphosphonate internucleoside linkage; 
 wherein each n is independently a number from 0 to 30; and 
 wherein, the 3′ end is linked directly or via a non-nucleotidic linker to another oligonucleotide, which may or may not be immunostimulatory; and 
 wherein the chemotherapeutic agent is selected from the group consisting of gemcitabine, taxotere, doxorubicin, docetaxel, herceptin and rituxan. 
 
     
     
         10 . The pharmaceutical formulation according to  claim 9  wherein the purine is 6-thioguanine or 7-deazaguanine. 
     
     
         11 . The pharmaceutical formulation according to  claim 9  wherein the non-naturally occurring pyrimidine base is selected from the group consisting of 5-hydroxycytosine, 5-hydroxymethylcytosine, N4-alkylcytosine, and 4-thiouracil. 
     
     
         12 . The pharmaceutical formulation according to  claim 9 , wherein the non-nucleotidic linker selected from the group consisting of a linker from about 2 angstroms to about 200 angstroms in length, a metal, a soluble or insoluble biodegradable polymer bead, an organic moiety having functional groups that permit attachment to the 3′-terminal nucleoside of the oligonucleotide, a biomolecule, a cyclic or acyclic small molecule, an aliphatic or aromatic hydrocarbon, either of which optionally can include, either in the linear chain connecting the oligonucleotides or appended to it, one or more functional groups selected from the group consisting of hydroxy, amino, thiol, thioether, ether, amide, thioamide, ester, urea, and thiourea; amino acids, carbohydrates, cyclodextrins, adamantane, cholesterol, haptens, antibiotics, glycerol and a glycerol homolog of the formula HO—(CH 2 ) o —CH(OH)—(CH 2 ) p —OH, wherein o and p independently are integers from 1 to about 6, and a derivative of 1,3-diamino-2-hydroxypropane. 
     
     
         13 . The pharmaceutical formulation of  claim 9  further comprising a vaccine. 
     
     
         14 . The pharmaceutical formulation of  claim 13  wherein the immunomer or the vaccine, or both, are linked to an immunogenic protein. 
     
     
         15 . The pharmaceutical formulation of  claim 13  further comprising an adjuvant. 
     
     
         16 . The pharmaceutical formulation according to  claim 9 , wherein at least one N1 or Nn is an immunostimulatory moiety.