Humanized anti-Tn-MUC1 antibodies and their conjugates

1. An isolated humanized antibody that binds to Tn-MUC1, wherein the isolated humanized antibody comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 29, a light chain variable region having the amino acid sequence of SEQ ID NO: 31, 32, 33, or 34, and optionally comprises a constant region derived from one or more human antibodies.

2. The isolated humanized antibody according to claim 1 , wherein the isolated humanized antibody comprises:

(i) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 1 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(ii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(iii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 3 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(iv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 4 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(v) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 5 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(vi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 6 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(vii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 7 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(viii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(ix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 9 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(x) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 10 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 11 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 12 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xiii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 13 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xiv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 14 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 15 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 16 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 17 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 18 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 19 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xx) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 20 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 21 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 22 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxiii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 23 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxiv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 24 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 25 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 26 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 27 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 28 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 29 and a light chain variable region having the amino acid sequence of SEQ ID NO: 31;

(xxx) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 1 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 3 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxiii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 4 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxiv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 5 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 6 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 7 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 9 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xxxix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 10 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xl) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 11 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xli) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 12 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xlii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 13 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xliii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 14 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xliv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 15 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xlv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 16 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xlvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 17 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xlvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 18 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xlviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 19 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(xlix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 20 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(l) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 21 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(li) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 22 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(lii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 23 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(liii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 24 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(liv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 25 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(lv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 26 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(lvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 27 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(lvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 28 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(lviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 29 and a light chain variable region having the amino acid sequence of SEQ ID NO: 32;

(lix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 1 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lx) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 3 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 4 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxiii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 5 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxiv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 6 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 7 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 9 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 10 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 11 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxx) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 12 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 13 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 14 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxiii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 15 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxiv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 16 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 17 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 18 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 19 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 20 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 21 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxx) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 22 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 23 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 24 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxiii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 25 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxiv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 26 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 27 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 28 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 29 and a light chain variable region having the amino acid sequence of SEQ ID NO: 33;

(lxxxviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 1 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(lxxxix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xc) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 3 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xci) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 4 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xcii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 5 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xciii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 6 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xciv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 7 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xcv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xcvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 9 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xcvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 10 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xcviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 11 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(xcix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 12 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(c) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 13 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(ci) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 14 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 15 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(ciii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 16 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(civ) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 17 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 18 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 19 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cvii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 20 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cviii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 21 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cix) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 22 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cx) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 23 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cxi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 24 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cxii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 25 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cxiii) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 26 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cxiv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 27 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34;

(cxv) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 28 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34; or

(cxvi) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 29 and a light chain variable region having the amino acid sequence of SEQ ID NO: 34.

3. The humanized antibody according to claim 1 , wherein said antibody binds Tn-MUC1 with an affinity (Kd) of at least 10 −6 M.

4. The humanized antibody according to claim 3 , wherein said antibody binds Tn-MUC1 with an affinity (Kd) of at least 10 −9 M.

5. The humanized antibody according to claim 1 , wherein said antibody competitively inhibits the binding to Tn-MUC1 of an antibody comprising a heavy chain variable region having the amino acid sequence of SEQ ID NO: 1 and a light chain variable region having the amino acid sequence of SEQ ID NO: 30.

6. The humanized antibody according to claim 1 , wherein said antibody competitively inhibits the binding to Tn-MUC1 of the mouse 5E5 antibody.

7. The humanized antibody according to claim 1 , wherein said antibody or antibody fragment substantially neutralizes at least one activity of at least one Tn-MUC1.

8. The humanized antibody according to claim 1 , wherein said antibody or antibody fragment expresses at a level of at least 10 micrograms/ml in a static HEK293T transient expression system.

9. The humanized antibody according to claim 1 , wherein said antibody or antibody fragment expresses at a level of at least 20 micrograms/ml in a shaken HEK293T transient expression system.

10. The humanized antibody according to claim 1 , wherein said antibody has a melting temperature (T m ) of at least 62° C.

11. The humanized antibody according to claim 1 , wherein said antibody or antibody fragment has a constant region of either isotype IgG1, IgG2, IgG3 or IgG4, or a mutated IgG constant region, and optionally a light chain constant region of isotype kappa or lambda.

12. The humanized antibody according to claim 1 , wherein the humanized antibody fragment is a scFv, Fab or F(ab′) 2 .

13. A polynucleotide encoding a humanized antibody according to claim 1 .

14. A conjugate comprising the humanized antibody according to claim 1 coupled to a functional moiety.

15. The conjugate according to claim 14 , wherein the functional moiety is selected from a drug, a reporter, a toxin, an organic moiety, and a binding member.

16. The conjugate according to claim 15 wherein the reporter is a fluorescent compound, a radionuclide, or an enzyme.

17. The conjugate according to claim 15 wherein the binding member is an antibody or antibody fragment.

18. The conjugate according to claim 14 , wherein the humanized antibody is covalently bonded to the functional moiety.

19. A pharmaceutical composition comprising the humanized antibody according to claim 1 , and a pharmaceutically acceptable diluent, carrier or excipient.

20. The pharmaceutical composition of claim 19 further comprising a therapeutically effective amount of a chemotherapeutic agent.

21. A method of treating cancer comprising administering to a patient the pharmaceutical composition according to claim 19 .

22. The method of claim 21 wherein the patient is administered a chemotherapeutic agent, in combination with the humanized antibody.

23. A method of treating a proliferative disease in a subject, wherein the proliferative disease is associated with raised levels of Tn-MUC1, the method comprising administering to the subject the conjugate according to claim 14 .