Humanized anti-ACTH antibodies and use thereof
The present invention is directed to antibodies and fragments thereof having binding specificity for ACTH. Embodiments of this invention relate to the binding fragments of antibodies described herein, comprising the sequences of the V H , V L and/or CDR polypeptides described herein, and the polynucleotides encoding them. The invention also contemplates anti-ACTH antibodies and binding fragments thereof conjugated to one or more functional or detectable moieties. The invention further contemplates methods of making said anti-ACTH antibodies and binding fragments thereof. Embodiments of the invention also pertain to the use of anti-ACTH antibodies and binding fragments thereof for the diagnosis, assessment, prevention and treatment of diseases and disorders associated with ACTH, such as CAH, FGD, Cushing's Disease, Cushing's Syndrome, Parkinson's disease, obesity, diabetes, sleep disorders, depression, anxiety disorders, cancer, muscle atrophy, hypertension, hyperinsulinemia, cognitive dysfunction, Alzheimer's disease, galactorrhea, stress related conditions, cardiac conditions, metabolic syndrome, hyperaldosteronism, Conn's syndrome and familial hyperaldosteronism.
1. A nucleic acid construct comprising a nucleic acid encoding a variable heavy chain (VH) or heavy chain polypeptide of an anti-human adrenocorticotrophic hormone (ACTH) antibody, wherein the encoded VH or heavy chain polypeptide comprises heavy chain CDR1, CDR2, and CDR3 polypeptides comprising the amino acid sequences of SEQ ID NO: 564; SEQ ID NO: 566; and SEQ ID NO: 568, respectively, and comprises at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 562.
2. The nucleic acid construct according to claim 1 , wherein the encoded VH polypeptide comprises the amino acid sequence of SEQ ID NO: 562.
3. The nucleic acid construct according to claim 1 , wherein the encoded heavy chain polypeptide comprises the amino acid sequence of SEQ ID NO: 561.
4. The nucleic acid construct according to claim 1 , which comprises the nucleic acid sequence of SEQ ID NO: 572 or 571.
5. A vector comprising the nucleic acid construct according to claim 1 .
6. A nucleic acid construct comprising a nucleic acid encoding a variable light chain (VL) or light chain polypeptide of an anti-human ACTH antibody, wherein the encoded VL or light chain polypeptide comprises light chain CDR1, CDR2, and CDR3 polypeptides comprising the amino acid sequences of SEQ ID NO: 584; SEQ ID NO: 586; and SEQ ID NO: 588, respectively, and comprises at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 582.
7. The nucleic acid construct according to claim 6 , wherein the encoded VL polypeptide comprises the amino acid sequence of SEQ ID NO: 582.
8. The nucleic acid construct according to claim 6 , wherein the encoded light chain polypeptide comprises the amino acid sequence of SEQ ID NO: 581.
9. The nucleic acid construct according to claim 6 , which comprises the nucleic acid sequence of SEQ ID NO: 592 or 591.
10. A vector comprising a nucleic acid construct according to claim 6 .
11. A nucleic acid construct comprising:
(a) a nucleic acid encoding a VII or heavy chain polypeptide of an anti-human ACTH antibody, wherein the encoded VH or heavy chain polypeptide comprises heavy chain CDR1, CDR2, and CDR3 polypeptides comprising the amino acid sequences of SEQ ID NO: 564; SEQ ID NO: 566; and SEQ ID NO: 568, respectively, and comprises at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 562; and
(b) a nucleic acid encoding a VL or light chain polypeptide of an anti-human ACTH antibody, wherein the encoded VL or light chain polypeptide comprises light chain CDR1, CDR2, and CDR3 polypeptides comprising the amino acid sequences of SEQ ID NO: 584; SEQ ID NO: 586; and SEQ ID NO: 588, respectively, and comprises at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 582.
12. The nucleic acid construct according to 11 , wherein:
(a) the encoded VH polypeptide comprises the amino acid sequence of SEQ ID NO: 562; and
(b) the encoded VL polypeptide comprises the amino acid sequence of SEQ ID NO: 582.
13. The nucleic acid construct according to 11 , wherein:
(a) the encoded heavy chain polypeptide comprises the amino acid sequence of SEQ ID NO: 561; and
(b) the encoded light chain polypeptide comprises the amino acid sequence of SEQ ID NO: 581.
14. A vector comprising the nucleic acid construct according to claim 13 .
15. The nucleic acid construct according to 11 , wherein:
(a) the nucleic acid encoding the VH or heavy chain polypeptide comprises the nucleic acid sequence of SEQ ID NO: 572 or 571, respectively: and
(b) the nucleic acid encoding the VL or light chain polypeptide comprises the nucleic acid sequence of SEQ ID NO: 592 or 591, respectively.
16. A vector comprising the nucleic acid construct according to claim 11 .
17. The nucleic acid construct according to claim 11 , which encodes a scFv comprising said VH and VL polypeptides.
18. The nucleic acid construct according to claim 11 , wherein the encoded constant domains are the constant domains of a human IgG1, IgG2, IgG3, or IgG4.
19. The nucleic acid construct according to claim 11 , wherein the encoded VH and VL polypeptides are humanized.
20. A host cell comprising:
(a) a nucleic acid which encodes a heavy chain polypeptide which comprises heavy chain CDR1, CDR2, and CDR3 polypeptides comprising the amino acid sequences of SEQ ID NO: 564; SEQ ID NO: 566; and SEQ ID NO: 568, respectively; and
(b) a nucleic acid which encodes a light chain polypeptide which comprises light chain CDR1, CDR2, and CDR3 polypeptides comprising the amino acid sequences of SEQ ID NO: 584; SEQ ID NO: 586; and SEQ ID NO: 588, respectively,
wherein both nucleic acid (i) and nucleic acid (ii) are operably linked to a promoter and further wherein the expression of nucleic acid (i) and nucleic acid (ii) in said host cell results in an anti-human ACTH antibody.
21. The host cell according to claim 20 , which comprises a bacterial, yeast, insect or mammalian cell.
22. The host cell according to claim 20 , which comprises a CHO or Pichia pastoris cell.
23. A method of producing an anti-human ACTH antibody, wherein the method comprises culturing the host cell according to claim 20 under conditions that result in the expression of said anti-human ACTH antibody.
24. The method of claim 23 , wherein the expressed ACTH antibody comprises:
(a) a VH polypeptide which comprises the amino acid sequence of SEQ ID NO: 562; and
(b) a VL polypeptide which comprises the amino acid sequence of SEQ ID NO: 582.
25. The method of claim 23 , wherein the expressed ACTH antibody comprises:
(a) a heavy chain polypeptide which comprises the amino acid sequence of SEQ ID NO: 561; and
(b) a light chain polypeptide which comprises the amino acid sequence of SEQ ID NO: 581.
26. A method of producing an anti-human ACTH Fab fragment, wherein the method comprises:
(i) culturing the host cell according to claim 20 under conditions that result in the expression of said anti-human ACTH antibody; and
(ii) enzymatically digesting the expressed anti-human ACTH antibody, thereby producing the anti-human ACTH Fab fragment.