Use of antibody against o-acetylated gd2 ganglioside to improve the therapeutic potential of drugs

Abstract

Disclosed is a method for delivery of an anti-cancer agent into a cell expressing the OAcGD2 ganglioside by using an antibody recognizing the OAcGD2 ganglioside.

1 . An in vitro/ex vivo method of identifying synergistic combination of (i) at least one anti-cancer agent having a molecular mass ranging from 100 Daltons to 200,000 Daltons and, (ii) at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside, comprising the steps of:
 a) incubating primary tumor cells or cancer cell lines cells expressing the O-acetylated form of GD2 ganglioside in vitro with a composition comprising (i) at least one anti-cancer agent having a molecular mass ranging from 100 Daltons to 200,000 Daltons and, (ii) at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside, and   b) measuring the CI50 index according to the method as disclosed by Chou and Talalay, wherein a CI50 below 1 is indicative of synergism.   
     
     
         2 . A kit suitable for treating cancer expressing OAcGD2 ganglioside comprising (i) at least one anti-cancer agent having a molecular mass ranging from 100 Daltons to 200,000 Daltons, (ii) at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside, and optionally (iii) a pharmaceutically acceptable carrier. 
     
     
         3 . The method of  claim 1 , wherein the at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside comprises
 a) a light chain variable region (VL) polypeptide having the amino acid sequence SEQ ID NO: 1 and   b) a heavy chain variable region (VH) having the amino acid sequence SEQ ID NO: 2.   
     
     
         4 . The method of  claim 1 , wherein the at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside comprises
 the light chain variable region (VL) polypeptide having the amino acid sequence selected in the group comprising or consisting of SEQ ID NO: 39 and SEQ ID NO: 40; and   the heavy chain variable region (VH) having the amino acid sequence selected in the group comprising or consisting of SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50 and SEQ ID NO: 51.   
     
     
         5 . The method of  claim 1 , wherein the multimeric fragment of the antibody recognizing the OAcGD2 ganglioside is in the form of single chain fragments of heavy and light chain variable regions (scFv). 
     
     
         6 . The method of  claim 1 , wherein the multimeric fragment of the antibody recognizing the OAcGD2 ganglioside is a single-chain part of a chimeric antigen receptor (CAR). 
     
     
         7 . The method of  claim 1 , wherein the at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside comprises
 the light chain variable region (VL) polypeptide having the amino acid sequence selected in the group comprising or consisting of SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO:39 and SEQ ID NO: 40, and   the heavy chain variable region (VH) having the amino acid sequence selected in the group comprising or consisting of SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO:44, SEQ ID NO:45, SEQ ID NO:46, SEQ ID NO: 47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50 and SEQ ID NO: 51.   
     
     
         8 . The method of  claim 1 , wherein the at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside comprises:
 a) a light chain variable region (VL) polypeptide having the amino acid sequence SEQ ID NO:14; and   b) a heavy chain variable region (VH) having the amino acid sequence SEQ ID NO:15.   
     
     
         9 . The method of  claim 1 , wherein the at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside comprises:
 a) a heavy chain comprising three heavy chain complementary regions (CDRs) having the amino acid sequences SEQ ID NO:16, SEQ ID NO:17 and SEQ ID NO:18, and a heavy chain framework sequence from an immunoglobulin heavy chain, and   b) a light chain comprising three light chain complementary regions (CDRs) having the amino acid sequences SEQ ID NO:19, SEQ ID NO:20 and SEQ ID NO:21, and a light chain framework sequence from an immunoglobulin light chain.   
     
     
         10 . The method of  claim 1 , wherein the at least one multimeric antibody or multimeric fragment thereof recognizing the OAcGD2 ganglioside is an immunoconjugate. 
     
     
         11 . The method of  claim 1 , wherein the at least one anti-cancer agent having a molecular mass ranging from 100 Daltons to 200,000 Daltons is selected from the group comprising or consisting of anti-cancer agents such as alkylating agents, anti-metabolites, anti-tumor antibodies, anti-tumor antibiotics, topoisomerase inhibitors, mitotic inhibitors, tyrosine kinase inhibitors, corticosteroids, hormones or hormone-like drugs, cytokines, nucleoside analogs, nucleic acids, such double-stranded synthetic short RNA molecules (miRNAs) or synthetic DNA/RNA-like oligonucleotides (ASOs). 
     
     
         12 . The method of  claim 1 , wherein the at least one anti-cancer agent having a molecular mass ranging from 100 Daltons to 200,000 Daltons is unable to cross the cell membrane of cancer cells by itself. 
     
     
         13 . The method of  claim 1 , wherein the tumor or cancer expressing the OAcGD2 ganglioside is selected from the group comprising or consisting of neuroblastoma, glioma, retinoblastoma, Ewing's family of tumors, sarcoma small cell lung cancer, breast cancer, melanoma, metastatic renal carcinoma, head and neck cancer and hematological cancers. 
     
     
         14 . The method of  claim 1 , wherein the at least one anti-cancer agent is selected from the group comprising or consisting of cyclophosphamide, doxorubicin, topotecan, irinotecan, temozolomide (TMZ), retinoic acid (RA), 5-Fluorouracil (5-FU), fludarabine, carboplatin and cisplatin. 
     
     
         15 . The method of  claim 1 , wherein the at least one anti-cancer agent is temozolomide, topotecan, irinotecan, fludarabine, cyclophosphamide or a mixture thereof and the cancer expressing the OAcGD2 ganglioside is neuroblastoma. 
     
     
         16 . The kit of  claim 2 , wherein the kit is suitable for delivering the at least one anti-cancer agent into a cell expressing the OAcGD2 ganglioside, and wherein the multimeric antibody or multimeric fragment thereof causes permeability defects within the cell membrane.