Carbonic anhydrase IX-specific antibodies and uses thereof

1. An isolated or purified antibody or antigen-binding fragment thereof, comprising

a) a light chain comprising a complementarity determining region (CDR) L1 sequence selected from the group consisting of:

(SEQ ID NO: 1)

RASGNIHNYLA;

(SEQ ID NO: 7)

RSSQSLVHSNGNTYLH;

and

(SEQ ID NO: 13)

KSSQSLLDSDGKTYLN,

a CDR L2 sequence selected from the group consisting of:

(SEQ ID NO: 2)

NTITLAD;

(SEQ ID NO: 8)

KVSNRFS;

and

(SEQ ID NO: 14)

LVSKLDS,

and

a CDR L3 sequence selected from the group consisting of:

(SEQ ID NO: 3)

QHFWNIPFT;

(SEQ ID NO: 9)

SQNTHVPPT;

and

(SEQ ID NO: 15)

CQGTHFPW,

and

b) a heavy chain comprising a complementarity determining region (CDR) H1 sequence selected from the group consisting of:

(SEQ ID NO: 4)

GFTFTSCYIH;

(SEQ ID NO: 10)

GFTFNTYAMY;

and

(SEQ ID NO: 16)

GYTFTNYGMN,

a CDR H2 sequence selected from the group consisting of:

(SEQ ID NO: 5)

WIYPGNGNTKYNEIFKG;

(SEQ ID NO: 11)

RIRSKSNNYAIYYADSVKD;

and

(SEQ ID NO: 17)

WINTYTGEPTYADDFKG,

and

a CDR H3 sequence selected from the group consisting of:

(SEQ ID NO: 6)

GDTTANTMDY;

(SEQ ID NO: 12)

GWDWFAY;

and

(SEQ ID NO: 18)

GGIATPTSY,

wherein the antibody or antigen-binding fragment thereof specifically binds the extracellular domain of Carbohydrate Anhydrase IX.

2. The isolated or purified antibody or antigen-binding fragment thereof of claim 1 , wherein the antibody or antigen-binding fragment thereof is selected from the group consisting of:

a) a light chain comprising CDR L1 of sequence RASGNIHNYLA (SEQ ID NO:1), CDR L2 of sequence NTITLAD (SEQ ID NO:2), and CDR L3 of sequence QHFWNIPFT (SEQ ID NO:3); and a heavy chain comprising CDR H1 of sequence GFTFTSCYIH (SEQ ID NO:4), CDR H2 of sequence WIYPGNGNTKYNEIFKG (SEQ ID NO:5), and CDR H3 of sequence GDTTANTMDY (SEQ ID NO:6); and wherein the antibody or antigen-binding fragment thereof binds the catalytic domain of CA-IX;

b) a light chain comprising CDR L1 of sequence RSSQSLVHSNGNTYLH (SEQ ID NO:7), CDR L2 of sequence KVSNRFS (SEQ ID NO:8), CDRL3 of sequence SQNTHVPPT (SEQ ID NO:9); and a heavy chain comprising CDR H1 of sequence GFTFNTYAMY (SEQ ID NO:10), CDR H2 of sequence RIRSKSNNYAIYYADSVKD (SEQ ID NO:11), and CDR H3 of sequence GWDWFAY(SEQ ID NO:12); and wherein the antibody or antigen-binding fragment thereof binds the PG-like domain of CA-IX; and

c) a light chain comprising CDR L1 of sequence KSSQSLLDSDGKTYLN (SEQ ID NO:13), CDR L2 of sequence LVSKLDS (SEQ ID NO:14), CDRL3 of sequence CQGTHFPW (SEQ ID NO:15); and a heavy chain comprising CDR H1 of sequence GYTFTNYGMN (SEQ ID NO:16), CDR H2 of sequence WINTYTGEPTYADDFKG (SEQ ID NO:17), and CDR H3 of sequence GGIATPTSY (SEQ ID NO:18); and wherein the antibody or antigen-binding fragment thereof binds the PG-like domain of CA-IX.

3. An isolated or purified antibody or antigen-binding fragment thereof, comprising:

a) a variable light (VL) domain of sequence selected from the group consisting of:

(SEQ ID NO: 19)

DIQMTQSPASLSASVGETVTITCRASGNIHNYLAWYQQKQGKSPQLLVYN

TITLADGVPSRFSGSGSGTQYSLKINSLQPEDFGSYYCQHFWNIPFTFGA

GTKLELK,

(SEQ ID NO: 21)

DVVMTQTPLSLPVSLGDQASISCRSSQSLVHSNGNTYLHWYLQKPGQSPK

WYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQNTHVPPT

FGGGTKLEIK,

and

(SEQ ID NO: 23)

DVVMTQTPLTLSVTIGQPASISCKSSQSLLDSDGKTYLNWLLQRPGQSPK

RLIYLVSKLDSGVPDRFTGSGSGTDFTLKISRVEAEDLGVYYCCQGTHFP

WTFGGGTKLEIK;

and

b) a variable heavy (V H ) domain of sequence selected from the group consisting of:

(SEQ ID NO: 20)

QVQLQQSGPELVKPGASVRISCKASGFTFTSCYIHWMKQRPGQGLEWIGW

IYPGNGNTKYNEIFKGRATLTTDKSSSTAYMQLSSLTSEDSAVYFCARGD

TTANTMDYWGQGTSVTVSS;

(SEQ ID NO: 22)

EVQLVESGGRLVQPKGSLKLSCAASGFTFNTYAMYWIRQAPGKGLEWVAR

IRSKSNNYAIYYADSVKDRFTISRDDSQSMLYLQMNNLKTEDTAMYYCVR

GWDWFAYWGQGTPVTVSA;

and

(SEQ ID NO: 24)

QIQLVQSGPELKKPGETVKISCKASGYTFTNYGMNWVQQAPGKGLKWMGW

INTYTGEPTYADDFKGRFAFSLETSASTAYLQINNLKNEDMATYFCARGG

IATPTSYWGQGTTLTVSS;

wherein the antibody or antigen-binding fragment thereof specifically binds to the extracellular domain of CA-IX.

4. The isolated or purified antibody or antigen-binding fragment thereof of claim 1 , wherein the isolated or purified antibody or antigen-binding fragment thereof comprises

a) a variable light (VL) domain of sequence

(SEQ ID NO: 19)

DIQMTQSPASLSASVGETVTITCRASGNIHNYLAWYQQKQGKSPQLLVYN

TITLADGVPSRFSGSGSGTQYSLKINSLQPEDFGSYYCQHFWNIPFTFGA

GTKLELK

and a variable heavy (V H ) domain of sequence

(SEQ ID NO: 20)

QVQLQQSGPELVKPGASVRISCKASGFTFTSCYIHWMKQRPGQGLEWIGW

IYPGNGNTKYNEIFKGRATLTTDKSSSTAYMQLSSLTSEDSAVYFCARGD

TTANTMDYWGQGTSVTVSS;

b) a variable light (V L ) domain of sequence

(SEQ ID NO: 21)

DVVMTQTPLSLPVSLGDQASISCRSSQSLVHSNGNTYLHWYLQKPGQSPK

LLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQNTHVP

PTFGGGTKLEIK

and a variable heavy (V H ) domain of sequence

(SEQ ID NO: 22)

EVQLVESGGRLVQPKGSLKLSCAASGFTFNTYAMYWIRQAPGKGLEWVAR

IRSKSNNYAIYYADSVKDRFTISRDDSQSMLYLQMNNLKTEDTAMYYCVR

GWDWFAYWGQGTPVTVSA;

or

c) a variable light (V L ) domain of sequence

(SEQ ID NO: 23)

DVVMTQTPLTLSVTIGQPASISCKSSQSLLDSDGKTYLNWLLQRPGQSPK

RLIYLVSKLDSGVPDRFTGSGSGTDFTLKISRVEAEDLGVYYCCQGTHFP

WTFGGGTKLEIK

and a variable heavy (V H ) domain of sequence

(SEQ ID NO: 24)

QIQLVQSGPELKKPGETVKISCKASGYTFTNYGMNWVQQAPGKGLKWMGW

INTYTGEPTYADDFKGRFAFSLETSASTAYLQINNLKNEDMATYFCARGG

IATPTSYWGQGTTLTVSS.

5. The isolated or purified antibody or antigen-binding fragment thereof claim 1 , wherein the antibody or antigen-binding fragment thereof is a full-length IgG, Fv, scFv, Fab, or F(ab′) 2 , or wherein the antibody or antigen-binding fragment thereof comprises framework regions from IgA, IgD, IgE, IgG, or IgM.

6. The isolated or purified antibody or antigen-binding fragment thereof of claim 1 , wherein the antibody or antigen-binding fragment thereof is chimeric.

7. The isolated or purified antibody or antigen-binding fragment thereof of claim 6 , wherein the chimeric antibody or antigen-binding fragment thereof comprises constant regions from human IgG1, or wherein the chimeric antibody or antigen-binding fragment thereof comprises constant regions from human kappa 1 light chain and human IgG1 heavy chain.

8. The isolated or purified antibody or antigen-binding fragment thereof of claim 7 , wherein the isolated or purified antibody or antigen-binding fragment thereof comprises

a) a variable light (V L ) domain comprising the sequence

(SEQ ID NO: 25)

DIQMTQSPASLSASVGETVTITCRASGNIHNYLAWYQQKQGKSPQLLVYN

TITLADGVPSRFSGSGSGTQYSLKINSLQPEDFGSYYCQHFWNIPFTFGA

GTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKV

DNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQG

LSSPVTKSFNRGEC

and a variable heavy (V H ) domain comprising the sequence

(SEQ ID NO: 26)

QVQLQQSGPELVKPGASVRISCKASGFTFTSCYIHWMKQRPGQGLEWIGW

IYPGNGNTKYNEIFKGRATLTTDKSSSTAYMQLSSLTSEDSAVYFCARGD

TTANTMDYWGQGTSVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKD

YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTY

ICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPK

DTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNS

TYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQV

YTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVL

DSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG;

b) a variable light (V L ) domain comprising the sequence

(SEQ ID NO: 27)

DVVMTQTPLSLPVSLGDQASISCRSSQSLVHSNGNTYLHWYLQKPGQSPK

LLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQNTHVP

PTFGGGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAK

VQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACE

VTHQGLSSPVTKSFNRGEC

and a variable heavy (V H ) domain comprising the sequence

(SEQ ID NO: 28)

EVQLVESGGRLVQPKGSLKLSCAASGFTFNTYAMYWIRQAPGKGLEWVAR

IRSKSNNYAIYYADSVKDRFTISRDDSQSMLYLQMNNLKTEDTAMYYCVR

GWDWFAYWGQGTPVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY

FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYI

CNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKD

TLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNST

YRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY

TLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLD

SDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG;

or

c) a variable light (V L ) domain comprising the sequence

(SEQ ID NO: 29)

DVVMTQTPLTLSVTIGQPASISCKSSQSLLDSDGKTYLNWLLQRPGQSPK

RLIYLVSKLDSGVPDRFTGSGSGTDFTLKISRVEAEDLGVYYCCQGTHFP

WTFGGGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAK

VQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACE

VTHQGLSSPVTKSFNRGEC

and a variable heavy (V H ) domain comprising the sequence

(SEQ ID NO: 30)

QIQLVQSGPELKKPGETVKISCKASGYTFTNYGMNWVQQAPGKGLKWMGW

INTYTGEPTYADDFKGRFAFSLETSASTAYLQINNLKNEDMATYFCARGG

IATPTSYWGQGTTLTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY

FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYI

CNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKD

TLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNST

YRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY

TLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLD

SDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG.

9. The isolated or purified antibody or antigen-binding fragment thereof of claim 1 , wherein the antibody or antigen-binding fragment thereof is immobilized onto a surface, or wherein the antibody or antigen-binding fragment thereof is linked to a cargo molecule.

10. The isolated or purified antibody or antigen-binding fragment thereof of claim 9 , wherein the cargo molecule is a detectable agent, a therapeutic agent, a drug, a peptide, an enzyme, a growth factor, a cytokine, a receptor trap, an antibody or antigen-binding fragment thereof, a chemical compound, a carbohydrate moiety, DNA-based molecules, a cytotoxic agent, viral vector, one or more liposomes or nanocarriers loaded with any of the previously recited types of cargo molecules, or one or more nanoparticle, nanowire, nanotube, or quantum dots.

11. A composition comprising one or more than one isolated or purified antibody or antigen-binding fragment thereof of claim 1 and a pharmaceutically-acceptable carrier, diluent, or excipient.

12. An in vitro method of detecting CA-IX, comprising

a) contacting a tissue sample with one or more than one isolated or purified antibody or antigen-binding fragment thereof of claim 1 linked to a detectable agent; and

b) detecting the detectable agent linked to the antibody or antigen-binding fragment thereof bound to CA-IX in the tissue sample.

13. The method of claim 12 , wherein method detects CA-IX in circulating cells and the sample is a serum sample.

14. The method of claim 12 , wherein the step of detecting (step b)) is performed using optical imaging, immunohistochemistry, molecular diagnostic imaging, enzyme-linked immunosorbent assay (ELISA).

15. An in vivo method of detecting CA-IX expression in a subject, comprising:

a) administering one or more than one isolated or purified antibody or antigen-binding fragment thereof of claim 1 linked to a detectable agent to the subject; and

b) detecting the detectable agent linked to the antibody or antigen-binding fragment thereof bound to CA-IX.

16. A method of transporting a molecule of interest into cells expressing CA-IX, comprising administering one or more than one isolated or purified antibody or antigen-binding fragment thereof of claim 1 linked to the molecule of interest to a subject, wherein the one or more than one isolated or purified antibody or fragment thereof delivers the molecule of interest to the subject's cells expressing CA-IX.

17. The method of claim 16 , wherein the step of detecting (step b)) is performed using positron emission tomography (PET), single photon emission computed tomography (SPECT), fluorescence imaging.

18. The method of claim 16 , wherein the molecule of interest is selected from the group consisting of a detectable agent, a therapeutic agent, a drug, a peptide, an enzyme, a growth factor, a cytokine, a receptor trap, an antibody or antigen-binding fragment thereof, a chemical compound, a carbohydrate moiety, DNA-based molecules, a cytotoxic agent, viral vector, one or more liposomes or nanocarriers loaded with any of the previously recited types of cargo molecules, or one or more nanoparticle, nanowire, nanotube, or quantum dots.