Anti-KC4 humanized monoclonal antibody and cells that express the antibody

Abstract

An anti-KC-4 humanized monoclonal antibody that comprises the variable regions of the light and heavy chains of the anti-KC-4 murine antibody, wherein the light chain has 7 amino acids and the heavy chain has 12 amino acids of the framework regions substituted with amino acid present in equivalent positions in antibodies of a species other than munne, and the constant regions of a human antibody. The antibody may be labeled and/or glycosylated, and is presented as a composition with a carrier. The anti-KC-4 monoclonal antibody is used in diagnostic kits for cancer and in in vivo methods of imaging and treating a primary or metastasized cancer, and in vitro diagnosis and ex vivo purging neoplastic cells from a biological fluid. RNAs and DNAs encode the monoclonal antibody, and a hybrid vector carrying the nucleotides and transfected cells express the peptides.

1. A modified chimeric  humanized anti-KC- 4  antibody which selectively binds to the human KC-4 antigen, comprising (1) the humanized variable regions of the light and heavy chains of an anti-KC-4 murine antibody and (2) light and heavy chain constant regions of a human antibody, wherein the amino acid sequence of the variable regions comprises SEQ ID NOS: 50 and SEQ ID NO: 51. 
     
     
       2. The antibody of  claim 1 , wherein 12 amino acids in the light chain framework regions (FRs) are substituted and 7 amino acids in the heavy chain FRs are substituted. 
     
     
       3. The antibody of  claim 2 , wherein said amino acids in said framework regions are amino acids present in equivalent positions in antibodies of species other than murine. 
     
     
       4. An antibody according to  claim 1 , further comprising at least one glycosyl residue attached thereto. 
     
     
       5. A composition of matter, comprising the antibody of  claim 1  and a carrier. 
     
     
       6. A cancer diagnostic kit, comprising the composition of  claim 5 , wherein said carrier is a pharmaceutically acceptable carrier for in vivo administration, and wherein said antibody is in radiolabeled form. 
     
     
       7. A cancer therapy kit, comprising the composition of  claim 4  in a pharmaceutically-acceptable form, wherein said antibody is in radiolabeled form. 
     
     
       8. An in vitro cancer diagnostic kit, comprising the antibody of  claim 1 ; and a solid support. 
     
     
       9. The in vitro kit of  claim 8 , further comprising one or more heterologous immunoglobulins that selectively bind to the constant region of the anti-KC-4 antibody. 
     
     
       10. The in vitro kit of  claim 9 , further comprising a label selected from the group consisting of a radioisotope, an enzyme, a phosphorescent molecule, and a fluorescent molecule, wherein said label is conjugated to said antibody of said immunoglobulin. 
     
     
       11. A hybridoma  cell expressing the antibody of  claim 1 . 
     
     
       12. The hybridoma  cell of  claim 11 , having the ATCC Accession No. HB 11455 (HuKC4V2). 
     
     
       13. A composition, comprising the hybridoma  cell of  claim 11 , and a diluent or a carrier. 
     
     
       14. A method of determining the presence of cancer cells in a tissue or a blood sample thereof, comprising contacting a tissue or a blood sample with the anti-KC- 4  antibody of  claim 1  or an antigen binding fragment thereof, and detecting the presence of an immune complex formed between said anti-KC- 4  antibody and the KC- 4  antigen of said tissue or said blood sample thereof. 
     
     
       15. A method according to  claim 14 , wherein the tissue is excised from a subject prior to being contacted with the anti-KC- 4  antibody, and wherein said contacting and detecting steps are carried out in vitro. 
     
     
       16. A method according to  claim 15 , further comprising comparing results obtained by analyzing said formed complexes with a standard cut-off value determined by comparing results obtained for normal and cancer patients' samples wherein a result above a cut-off value is indicative of cancer. 
     
     
       17. A method of imaging a cancer, comprising administering a pharmaceutically-acceptable composition comprising an effective imaging amount of the anti-KC- 4  antibody of  claim 1  or an antigen binding fragment thereof, in radiolabeled form, to a subject suspected of having a primary or metastasized cancer; and detecting any binding of the labeled antibody to cancer cells in the subject by external imaging. 
     
     
       18. A method of treating cancer in a subject, comprising administering to a subject in need of treatment an effective therapeutic amount of the antibody of  claim 1 , wherein the antibody is conjugated to a therapeutic agent. 
     
     
       19. A method of purging cancer cells from a biological sample, comprising contacting a biological sample obtained from a subject suspected of having cancer with the antibody of  claim 1  and allowing said antibody to bind the KC- 4  antigen associated with cancer cells in the sample to form an immune complex between said antibody and the antigen of said biological sample; and removing said immune complex from the biological sample. 
     
     
       20. A method according to  claim 14 , wherein said tissue sample is prepared for immunohistochemical analysis. 
     
     
       21. A method of purging cancer cells from a biological sample and replenishing the purged sample to a subject, comprising the method of  claim 19 , wherein said method further comprises reintroducing the purged biological sample to the subject.