HUMANIZED ANTIBODIES
First Claim
Patent Images
1. A humanized antibody or binding fragment thereof comprising an ultralong CDR3.
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Accused Products
Abstract
The present disclosure provides humanized antibodies, including antibodies comprising an ultralong CDR3 (e.g. bovine) and uses thereof.
9 Citations
179 Claims
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1. A humanized antibody or binding fragment thereof comprising an ultralong CDR3.
- View Dependent Claims (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, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 68, 69)
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2. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 is 35 amino acids in length or longer, 40 amino acids in length or longer, 45 amino acids in length or longer, 50 amino acids in length or longer, 55 amino acids in length or longer, or 60 amino acids in length or longer.
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3. The humanized antibody or binding fragment thereof of claim 2, wherein the ultralong CDR3 is 35 amino acids in length or longer.
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4. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises 3 or more cysteine residues, 4 or more cysteine residues, 5 or more cysteine residues, 6 or more cysteine residues, 7 or more cysteine residues, 8 or more cysteine residues, 9 or more cysteine residues, 10 or more cysteine residues, 11 or more cysteine residues, or 12 or more cysteine residues.
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5. The humanized antibody or binding fragment thereof of claim 4, wherein the ultralong CDR3 comprises 3 or more cysteine residues.
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6. The humanized antibody or binding fragment thereof of claim 1, wherein the antibodies or binding fragments thereof comprise a cysteine motif.
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7. The humanized antibody or binding fragment thereof of claim 6, wherein the cysteine motif is selected from the group consisting of:
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8. The humanized antibody or binding fragment thereof of claim 6, wherein the cysteine motif is selected from the group consisting of:
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9. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises 2 to 6 disulfide bonds.
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10. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises SEQ ID NO:
- 40 or a derivative thereof.
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11. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises amino acid residues 3-6 of any of one SEQ ID NO:
- 1-4.
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12. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a non-human DH or a derivative thereof.
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13. The humanized antibody or binding fragment thereof of claim 12, wherein the non-human DH is SEQ ID NO:
- 5, SEQ ID NO;
6, SEQ ID NO;
7, SEQ ID NO;
8, SEQ ID NO;
9, SEQ ID NO;
10, SEQ ID NO;
11, or SEQ ID NO;
12.
- 5, SEQ ID NO;
-
14. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a JH sequence or a derivative thereof.
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15. The humanized antibody or binding fragment thereof of claim 14, wherein the JH sequence is SEQ ID NO:
- 13, SEQ ID NO;
14, SEQ ID NO;
15, SEQ ID NO;
16, or SEQ ID NO;
17.
- 13, SEQ ID NO;
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16. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises:
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a non-human VH sequence or a derivative thereof; a non-human DH sequence or a derivative thereof; and
/ora JH sequence or derivative thereof.
-
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17. The humanized antibody or binding fragment thereof of claim 16, wherein the ultralong CDR3 comprises an additional amino acid sequence comprising two to six amino acid residues or more positioned between the VH sequence and the DH sequence.
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18. The humanized antibody or binding fragment thereof of claim 17, wherein the additional amino acid sequence is selected from the group consisting of:
- IR, IF, SEQ ID NO;
18, SEQ ID NO;
19, SEQ ID NO;
20 or SEQ ID NO;
21.
- IR, IF, SEQ ID NO;
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19. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a sequence derived from or based on SEQ ID NO:
- 22, SEQ ID NO;
23, SEQ ID NO;
24, SEQ ID NO;
25, SEQ ID NO;
26, SEQ ID NO;
27, or SEQ ID NO;
28.
- 22, SEQ ID NO;
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20. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a non-bovine sequence or a non-antibody sequence.
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21. The humanized antibody or binding fragment thereof of claim 20, wherein the non-antibody sequence is a synthetic sequence.
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22. The humanized antibody or binding fragment thereof of claim 20, wherein the non-antibody sequence is a cytokine sequence, a lymphokine sequence, a chemokine sequence, a growth factor sequence, a hormone sequence, or a toxin sequence.
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23. The humanized antibody or binding fragment thereof of claim 20, wherein the non-antibody sequence is an IL-8 sequence, an IL-21 sequence, an SDF-1 (alpha) sequence, a somatostatin sequence, a chlorotoxin sequence, a Pro-TxII sequence, or a ziconotide sequence.
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24. The humanized antibody or binding fragment thereof of claim 20, wherein the non-antibody sequence is any one of SEQ ID NOS:
- 475-481.
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25. The humanized antibody or binding fragment thereof of claim 20, wherein the non-antibody sequence replaces at least a portion of the ultralong CDR3.
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26. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a X1X2X3X4X5 motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q). -
27. The humanized antibody or binding fragment thereof of claim 26, wherein the X1X2X3X4X5 motif is TTVHQ (SEQ ID NO:
- 153), TSVHQ (SEQ ID NO;
154), SSVTQ (SEQ ID NO;
155), STVHQ (SEQ ID NO;
156), ATVRQ (SEQ ID NO;
157), TTVYQ (SEQ ID NO;
158), SPVHQ (SEQ ID NO;
159), ATVYQ (SEQ ID NO;
160), TAVYQ (SEQ ID NO;
161), TNVHQ (SEQ ID NO;
162), ATVHQ (SEQ ID NO;
163), STVYQ (SEQ ID NO;
164), TIVHQ (SEQ ID NO;
165), AIVYQ (SEQ ID NO;
166), TTVFQ (SEQ ID NO;
167), AAVFQ (SEQ ID NO;
168), GTVHQ (SEQ ID NO;
169), ASVHQ (SEQ ID NO;
170), TAVFQ (SEQ ID NO;
171), ATVFQ (SEQ ID NO;
172), AAAHQ (SEQ ID NO;
173), VVVYQ (SEQ ID NO;
174), GTVFQ (SEQ ID NO;
175), TAVHQ (SEQ ID NO;
176), ITVHQ (SEQ ID NO;
177), ITAHQ (SEQ ID NO;
178), VTVHQ (SEQ ID NO;
179);
AAVHQ (SEQ ID NO;
180), GTVYQ (SEQ ID NO;
181), TTVLQ (SEQ ID NO;
182), TTTHQ (SEQ ID NO;
183), or TTDYQ (SEQ ID NO;
184).
- 153), TSVHQ (SEQ ID NO;
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28. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a CX1X2X3X4X5 motif.
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29. The humanized antibody or binding fragment thereof of claim 28, wherein the CX1X2X3X4X5 motif is CTTVHQ (SEQ ID NO:
- 185), CTSVHQ (SEQ ID NO;
186), CSSVTQ (SEQ ID NO;
187), CSTVHQ (SEQ ID NO;
188), CATVRQ (SEQ ID NO;
189), CTTVYQ (SEQ ID NO;
190), CSPVHQ (SEQ ID NO;
191), CATVYQ (SEQ ID NO;
192), CTAVYQ (SEQ ID NO;
193), CTNVHQ (SEQ ID NO;
194), CATVHQ (SEQ ID NO;
195), CSTVYQ (SEQ ID NO;
196), CTIVHQ (SEQ ID NO;
197), CAIVYQ (SEQ ID NO;
198), CTTVFQ (SEQ ID NO;
199), CAAVFQ (SEQ ID NO;
200), CGTVHQ (SEQ ID NO;
201), CASVHQ (SEQ ID NO;
202), CTAVFQ (SEQ ID NO;
203), CATVFQ (SEQ ID NO;
204), CAAAHQ (SEQ ID NO;
205), CVVVYQ (SEQ ID NO;
206), CGTVFQ (SEQ ID NO;
207), CTAVHQ (SEQ ID NO;
208), CITVHQ (SEQ ID NO;
209), CITAHQ (SEQ ID NO;
210), CVTVHQ (SEQ ID NO;
211);
CAAVHQ (SEQ ID NO;
212), CGTVYQ (SEQ ID NO;
213), CTTVLQ (SEQ ID NO;
214), CTTTHQ (SEQ ID NO;
215), or CTTDYQ (SEQ ID NO;
216).
- 185), CTSVHQ (SEQ ID NO;
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30. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a (XaXb)z motif,
wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
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31. The humanized antibody or binding fragment thereof of claim 30, wherein the (XaXb)z motif is CYTYNYEF (SEQ ID NO:
- 217), HYTYTYDF (SEQ ID NO;
218), HYTYTYEW (SEQ ID NO;
219), KHRYTYEW (SEQ ID NO;
220), NYIYKYSF (SEQ ID NO;
221), PYIYTYQF (SEQ ID NO;
222), SFTYTYEW (SEQ ID NO;
223), SYIYIYQW (SEQ ID NO;
224), SYNYTYSW (SEQ ID NO;
225), SYSYSYEY (SEQ ID NO;
226), SYTYNYDF (SEQ ID NO;
227), SYTYNYEW (SEQ ID NO;
228), SYTYNYQF (SEQ ID NO;
229), SYVWTHNF (SEQ ID NO;
230), TYKYVYEW (SEQ ID NO;
231), TYTYTYEF (SEQ ID NO;
232), TYTYTYEW (SEQ ID NO;
233), VFTYTYEF (SEQ ID NO;
234), AYTYEW (SEQ ID NO;
235), DYIYTY (SEQ ID NO;
236), IHSYEF (SEQ ID NO;
237), SFTYEF (SEQ ID NO;
238), SHSYEF (SEQ ID NO;
239), THTYEF (SEQ ID NO;
240), TWTYEF (SEQ ID NO;
241), TYNYEW (SEQ ID NO;
242), TYSYEF (SEQ ID NO;
243), TYSYEH (SEQ ID NO;
244), TYTYDF (SEQ ID NO;
245), TYTYEF (SEQ ID NO;
246), TYTYEW (SEQ ID NO;
247), AYEF (SEQ ID NO;
248), AYSF (SEQ ID NO;
249), AYSY (SEQ ID NO;
250), CYSF (SEQ ID NO;
251), DYTY (SEQ ID NO;
252), KYEH (SEQ ID NO;
253), KYEW (SEQ ID NO;
254), MYEF (SEQ ID NO;
255), NWIY (SEQ ID NO;
256), NYDY (SEQ ID NO;
257), NYQW (SEQ ID NO;
258), NYSF (SEQ ID NO;
259), PYEW (SEQ ID NO;
260), RYNW (SEQ ID NO;
261), RYTY (SEQ ID NO;
262), SYEF (SEQ ID NO;
263), SYEH (SEQ ID NO;
264), SYEW (SEQ ID NO;
265), SYKW (SEQ ID NO;
266), SYTY (SEQ ID NO;
267), TYDF (SEQ ID NO;
268), TYEF (SEQ ID NO;
269), TYEW (SEQ ID NO;
270), TYQW (SEQ ID NO;
271), TYTY (SEQ ID NO;
272), or VYEW (SEQ ID NO;
273).
- 217), HYTYTYDF (SEQ ID NO;
-
32. The humanized antibody or binding fragment thereof of claim 30, wherein the (XaXb)z motif is YXYXYX.
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33. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a X1X2X3X4X5Xn motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), wherein X5 is glutamine (Q), and wherein n is 27-54. -
34. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises Xn(XaXb)z motif,
wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), wherein n is 27-54, and wherein z is 1-4.
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35. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises a X1X2X3X4X5Xn(XaXb)z motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q), wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), wherein n is 27-54, and wherein z is 1-4.
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36. The humanized antibody or binding fragment thereof of claim 35, wherein the X1X2X3X4X5 motif is TTVHQ (SEQ ID NO:
- 153) or TSVHQ (SEQ ID NO;
154), and wherein the (XaXb)z motif is YXYXYX.
- 153) or TSVHQ (SEQ ID NO;
-
37. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises:
-
a CX1X2X3X4X5 motif, wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q), a cysteine motif selected from the group consisting of;
CX10CX5CX5CXCX7C (SEQ ID NO;
41), CX10CX6CX5CXCX15C (SEQ ID NO;
42), CX11CXCX5C (SEQ ID NO;
43), CX11CX5CX5CXCX7C (SEQ ID NO;
44), CX10CX6CX5CXCX13C (SEQ ID NO;
45), CX10CX5CXCX4CX8C (SEQ ID NO;
46), CX10CX6CX6CXCX7C (SEQ ID NO;
47), CX10CX4CX7CXCX8C (SEQ ID NO;
48), CX10CX4CX7CXCX7C (SEQ ID NO;
49), CX13CX8CX8C (SEQ ID NO;
50), CX10CX6CX5CXCX7C (SEQ ID NO;
51), CX10CX5CX5C (SEQ ID NO;
52), CX10CX5CX6CXCX7C (SEQ ID NO;
53), CX10CX6CX5CX7CX9C (SEQ ID NO;
54), CX9CX7CX5CXCX7C (SEQ ID NO;
55), CX10CX6CX5CXCX9C (SEQ ID NO;
56), CX10CXCX4CX5CX11C (SEQ ID NO;
57), CX7CX3CX6CX5CXCX5CX10C (SEQ ID NO;
58), CX10CXCX4CX5CXCX2CX3C (SEQ ID NO;
59), CX16CX5CXC (SEQ ID NO;
60), CX6CX4CXCX4CX5C (SEQ ID NO;
61), CX11CX4CX5CX6CX3C (SEQ ID NO;
62), CX8CX2CX6CX5C (SEQ ID NO;
63), CX10CX5CX5CXCX10C (SEQ ID NO;
64), CX10CXCX6CX4CXC (SEQ ID NO;
65), CX10CX5CX5CXCX2C (SEQ ID NO;
66), CX14CX2CX3CXCXC (SEQ ID NO;
67), CX15CX5CXC (SEQ ID NO;
68), CX4CX6CX9CX2CX11C (SEQ ID NO;
69), CX6CX4CX5CX5CX12C (SEQ ID NO;
70), CX7CX3CXCXCX4CX5CX9C (SEQ ID NO;
71), CX10CX6CX5C (SEQ ID NO;
72), CX7CX3CX5CX5CX9C (SEQ ID NO;
73), CX7CX5CXCX2C (SEQ ID NO;
74), CX10CXCX6C (SEQ ID NO;
75), CX10CX3CX3CX5CX7CXCX6C (SEQ ID NO;
76), CX10CX4CX5CX12CX2C (SEQ ID NO;
77), CX12CX4CX5CXCXCX9CX3C (SEQ ID NO;
78), CX12CX4CX5CX12CX2C (SEQ ID NO;
79), CX10CX6CX5CXCX11C (SEQ ID NO;
80), CX16CX5CXCXCX14C (SEQ ID NO;
81), CX10CX5CXCX8CX6C (SEQ ID NO;
82), CX12CX4CX5CX8CX2C (SEQ ID NO;
83), CX12CX5CX5CXCX8C (SEQ ID NO;
84), CX10CX6CX5CXCX4CXCX9C (SEQ ID NO;
85), CX11CX4CX5CX8CX2C (SEQ ID NO;
86), CX10CX6CX5CX8CX2C (SEQ ID NO;
87), CX10CX6CX5CXCX8C (SEQ ID NO;
88), CX10CX6CX5CXCX3CX8CX2C (SEQ ID NO;
89), CX10CX6CX5CX3CX8C (SEQ ID NO;
90), CX10CX6CX5CXCX2CX6CX5C (SEQ ID NO;
91), CX7CX6CX3CX3CX9C (SEQ ID NO;
92), CX9CX8CX5CX6CX5C (SEQ ID NO;
93), CX10CX2CX2CX7CXCX11CX5C (SEQ ID NO;
94), and CX10CX6CX5CXCX2CX8CX4C (SEQ ID NO;
95),a (XaXb)z motif, wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of;
tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
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38. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 comprises:
-
a CX1X2X3X4X5 motif, wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q); a cysteine motif selected from the group consisting of;
wherein the cysteine motif is selected from the group consisting of;
CCX3CXCX3CX2CCXCX5CX9CX5CXC (SEQ ID NO;
96), CX6CX2CX5CX4CCXCX4CX6CXC (SEQ ID NO;
97), CX7CXCX5CX4CCCX4CX6CXC (SEQ ID NO;
98), CX9CX3CXCX2CXCCCX6CX4C (SEQ ID NO;
99), CX5CX3CXCX4CX4CCX10CX2CC (SEQ ID NO;
100), CX5CXCX1CXCX3CCX3CX4CX10C (SEQ ID NO;
101), CX9CCCX3CX4CCCX5CX6C (SEQ ID NO;
102), CCX8CX5CX4CX3CX4CCXCX1C (SEQ ID NO;
103), CCX6CCX5CCCX4CX4CX12C (SEQ ID NO;
104), CX6CX2CX3CCCX4CX5CX3CX3C (SEQ ID NO;
105), CX3CX5CX6CX4CCXCX5CX4CXC (SEQ ID NO;
106), CX4CX4CCX4CX4CXCX11CX2CXC (SEQ ID NO;
107), CX5CX2CCX5CX4CCX3CCX7C (SEQ ID NO;
108), CX5CX5CX3CX2CXCCX4CX7CXC (SEQ ID NO;
109), CX3CX7CX3CX4CCXCX2CX5CX2C (SEQ ID NO;
110), CX9CX3CXCX4CCX5CCCX6C (SEQ ID NO;
111), CX9CX3CXCX2CXCCX6CX3CX3C (SEQ ID NO;
112), CX8CCXCX3CCX3CXCX3CX4C (SEQ ID NO;
113), CX9CCX4CX2CXCCXCX4CX3C (SEQ ID NO;
114), CX10CXCX3CX2CXCCX4CX5CXC (SEQ ID NO;
115), CX9CXCX3CX2CXCCX4CX5CXC (SEQ ID NO;
116), CX6CCXCX5CX4CCXCX5CX2C (SEQ ID NO;
117), CX6CCXCX3CXCCX3CX4CC (SEQ ID NO;
118), CX6CCXCX3CXCX2CXCX4CX8C (SEQ ID NO;
119), CX4CX2CCX3CXCX4CCX2CX3C (SEQ ID NO;
120), CX3CX5CX3CCCX4CX9C (SEQ ID NO;
121), CCX9CX3CXCCX3CX5C (SEQ ID NO;
122), CX9CX2CX3CX4CCX4CX5C (SEQ ID NO;
123), CX9CX7CX4CCXCX7CX3C (SEQ ID NO;
124), CX9CX3CCCX10CX2CX3C (SEQ ID NO;
125), CX3CX5CX5CX4CCX10CX6C (SEQ ID NO;
126), CX9CX5CX4CCXCX5CX4C (SEQ ID NO;
127), CX7CXCX6CX4CCCX10C (SEQ ID NO;
128), CX8CX2CX4CCX4CX3CX3C (SEQ ID NO;
129), CX7CX5CXCX4CCX7CX4C (SEQ ID NO;
130), CX11CX3CX4CCCX8CX2C (SEQ ID NO;
131), CX2CX3CX4CCX4CX5CX15C (SEQ ID NO;
132), CX9CX5CX4CCX7C (SEQ ID NO;
133), CX9CX7CX3CX2CX6C (SEQ ID NO;
134), CX9CX5CX4CCX14C (SEQ ID NO;
135), CX9CX5CX4CCX8C (SEQ ID NO;
136), CX9CX6CX4CCXC (SEQ ID NO;
137), CX5CCX7CX4CX12 (SEQ ID NO;
138), CX10CX3CX4CCX4C (SEQ ID NO;
139), CX9CX4CCX5CX4C (SEQ ID NO;
140), CX10CX3CX4CX7CXC (SEQ ID NO;
141), CX7CX7CX2CX2CX3C (SEQ ID NO;
142), CX9CX4CX4CCX6C (SEQ ID NO;
143), CX7CXCX3CXCX6C (SEQ ID NO;
144), CX7CXCX4CXCX4C (SEQ ID NO;
145), CX9CX5CX4C (SEQ ID NO;
146), CX3CX6CX8C (SEQ ID NO;
147), CX10CXCX4C (SEQ ID NO;
148), CX10CCX4C (SEQ ID NO;
149), CX15C (SEQ ID NO;
150), CX10C (SEQ ID NO;
151), and CX9C (SEQ ID NO;
152); anda (XaXb)z motif, wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of;
tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
-
-
39. The humanized antibody or binding fragment thereof of claim 20, wherein the ultralong CDR3 comprises an additional sequence that is a linker.
-
40. The humanized antibody or binding fragment thereof of claim 39, wherein the linker is linked to a C-terminus, a N-terminus, or both C-terminus and N-terminus of the non-antibody sequence.
-
41. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 is a ruminant CDR3.
-
42. The humanized antibody or binding fragment thereof of claim 41, wherein the ruminant is a cow.
-
43. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprises a human heavy chain germline sequence that is a VH4 family member germline sequence.
-
44. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprises a VH4-34 germline sequence.
-
45. The humanized antibody or binding fragment thereof of claim 44, wherein the humanized antibody or binding fragment thereof comprises a human heavy chain germline sequence that is derived from the VH4-34 germline sequence.
-
46. The humanized antibody or binding fragment thereof of claim 45, wherein the VH4-34 germline sequence comprises a CDR1 comprising a Gly31Asp and Tyr32Lys substitution based on Kabat numbering.
-
47. The humanized antibody or binding fragment thereof of claim 45, wherein the VH4-34 germline sequence comprises a CDR2 comprising a Glu50Ser substitution based on Kabat numbering.
-
48. The humanized antibody or binding fragment thereof of claim 45, wherein the VH4-34 germline sequence comprises a CDR1 comprising Ala23Thr, Val24Ala, Tyr25Ser, Gly27Phe, Phe29Leu, Gly31Asp, Tyr32Lys, Tyr33Ala, Trp34Val, and Ser35Gly substitutions, and a CDR2 comprising Ile48Leu, Glu50Ser, Asn52Asp, His53Thr, Ser54Gly, Ser56Asn, and Asn58Gly substitutions, based on Kabat numbering.
-
49. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:
- 461, 462, 463, 464, 465, 466, 467, 468, or 469.
-
50. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:
- 470.
-
51. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:
- 461, 462, 463, 464, 465, 466, 467, 468, or 469; and
the amino acid sequence set forth in SEQ ID NO;
470.
- 461, 462, 463, 464, 465, 466, 467, 468, or 469; and
-
52. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:
- 461, 462, 463, 464, 465, 466, 467, 468, or 469;
the amino acid sequence set forth in SEQ ID NO;
470; and
a peptide sequence, wherein the peptide sequence is positioned between SEQ ID NO;
461, 462, 463, 464, 465, 466, 467, 468, or 469, and SEQ ID NO;
470.
- 461, 462, 463, 464, 465, 466, 467, 468, or 469;
-
53. The humanized antibody or binding fragment thereof of claim 52, wherein the peptide sequence is a bovine sequence, a non-bovine sequence, an antibody sequence, or a non-antibody sequence.
-
54. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprises a light chain variable region sequence that is a lambda light chain variable region sequence.
-
55. The humanized antibody or binding fragment thereof of claim 54, wherein the lambda light chain variable region sequence is a bovine lambda light chain variable region sequence.
-
56. The humanized antibody or binding fragment thereof of claim 54, wherein the lambda light chain variable region sequence is a human lambda light chain variable region sequence.
-
57. The humanized antibody or binding fragment thereof of claim 56, wherein the human lambda light chain variable region sequence is a VL1-51 germline sequence.
-
58. The humanized antibody or binding fragment thereof of claim 56, wherein the human lambda light chain variable region sequence is derived from a VL1-51 germline sequence.
-
59. The humanized antibody or binding fragment thereof of claim 58, wherein the VL1-51 germline sequence comprises a CDR1 comprising Ile29Val and Asn32Gly substitution based on Kabat numbering.
-
60. The humanized antibody or binding fragment thereof of claim 58, wherein the VL1-51 germline sequence comprises a CDR2 comprising a substitution of DNN to GDT.
-
61. The humanized antibody or binding fragment thereof of claim 58, wherein the VL1-51 germline sequence comprises a CDR2 comprising a substitution of DNNKRP (SEQ ID NO:
- 471) to GDTSRA (SEQ ID NO;
472).
- 471) to GDTSRA (SEQ ID NO;
-
62. The humanized antibody or binding fragment thereof of claim 58, wherein the VL1-51 germline sequence comprises a S2A, T5N, P8S, A12G, A13S, and P14L substitution based on Kabat numbering.
-
63. The humanized antibody or binding fragment thereof of claim 58, wherein the VL1-51 germline sequence comprises a S2A, T5N, P8S, A12G, A13S, and P14L substitution based on Kabat numbering, and a CDR2 comprising a substitution of DNN to GDT.
-
64. The humanized antibody or binding fragment thereof of claim 1, wherein the humanized antibody or binding fragment thereof comprising the amino acid sequence set forth in SEQ ID NO:
- 440, 441, 442, 443, or 444.
-
65. A polynucleotide encoding the humanized antibody or binding fragment thereof of any one of claims 1-64.
-
68. A vector comprising the polynucleotide of any one of claims 65-67.
-
69. A host cell comprising the vector of claim 68.
-
2. The humanized antibody or binding fragment thereof of claim 1, wherein the ultralong CDR3 is 35 amino acids in length or longer, 40 amino acids in length or longer, 45 amino acids in length or longer, 50 amino acids in length or longer, 55 amino acids in length or longer, or 60 amino acids in length or longer.
-
66. A polynucleotide encoding a humanized antibody or binding fragment thereof that comprises an ultralong CDR3.
-
67. A polynucleotide encoding a humanized variable region that comprises an ultralong CDR3.
-
70. A nucleic acid library comprising a plurality of polynucleotides comprising sequences coding for humanized antibodies or binding fragments thereof, wherein the antibodies or binding fragments thereof comprise an ultralong CDR3.
-
71. A library of humanized antibodies or binding fragments thereof, wherein the antibodies or binding fragments thereof comprise an ultralong CDR3.
- View Dependent Claims (72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133)
-
72. The library of claim 71, wherein the ultralong CDR3 is 35 amino acids in length or longer, 40 amino acids in length or longer, 45 amino acids in length or longer, 50 amino acids in length or longer, 55 amino acids in length or longer, or 60 amino acids in length or longer.
-
73. The library of claim 72, wherein the ultralong CDR3 is 35 amino acids in length or longer.
-
74. The library of claim 71, wherein the ultralong CDR3 comprises 3 or more cysteine residues, 4 or more cysteine residues, 5 or more cysteine residues, 6 or more cysteine residues, 7 or more cysteine residues, 8 or more cysteine residues, 9 or more cysteine residues, 10 or more cysteine residues, 11 or more cysteine residues, or 12 or more cysteine residues.
-
75. The library of claim 74, wherein the ultralong CDR3 comprises 3 or more cysteine residues.
-
76. The library of claim 71, wherein the antibodies or binding fragments thereof comprise a cysteine motif.
-
77. The library of claim 76, wherein the cysteine motif is selected from the group consisting of:
-
78. The library of claim 76, wherein the cysteine motif is selected from the group consisting of:
-
79. The library of claim 71, wherein the ultralong CDR3 comprises 2 to 6 disulfide bonds.
-
80. The library of claim 71, wherein the ultralong CDR3 comprises SEQ ID NO:
- 40 or a derivative thereof.
-
81. The library of claim 71, wherein the ultralong CDR3 comprises amino acid residues 3-6 of any of one SEQ ID NO:
- 1-4.
-
82. The library of claim 71, wherein the ultralong CDR3 comprises a non-human DH or a derivative thereof.
-
83. The library of claim 82, wherein the non-human DH is SEQ ID NO:
- 5, SEQ ID NO;
6, SEQ ID NO;
7, SEQ ID NO;
8, SEQ ID NO;
9, SEQ ID NO;
10, SEQ ID NO;
11, or SEQ ID NO;
12
- 5, SEQ ID NO;
-
84. The library of claim 71, wherein the ultralong CDR3 comprises a JH sequence or a derivative thereof.
-
85. The library of claim 84, wherein the JH sequence is SEQ ID NO:
- 13, SEQ ID NO;
14, SEQ ID NO;
15, SEQ ID NO;
16, or SEQ ID NO;
17.
- 13, SEQ ID NO;
-
86. The library of claim 71, wherein the ultralong CDR3 comprises:
-
a non-human VH sequence or a derivative thereof; a non-human DH sequence or a derivative thereof; and
/ora JH sequence or derivative thereof.
-
-
87. The library of claim 86, wherein the ultralong CDR3 comprises an additional amino acid sequence comprising two to six amino acid residues or more positioned between the VH sequence and the DH sequence.
-
88. The library of claim 87, wherein the additional amino acid sequence is selected from the group consisting of:
- IR, IF, SEQ ID NO;
18, SEQ ID NO;
19, SEQ ID NO;
20 or SEQ ID NO;
21.
- IR, IF, SEQ ID NO;
-
89. The library of claim 71, wherein the ultralong CDR3 comprises a sequence derived from or based on SEQ ID NO:
- 22, SEQ ID NO;
23, SEQ ID NO;
24, SEQ ID NO;
25, SEQ ID NO;
26, SEQ ID NO;
27, or SEQ ID NO;
28.
- 22, SEQ ID NO;
-
90. The library of claim 71, wherein the ultralong CDR3 comprises a non-bovine sequence or a non-antibody sequence.
-
91. The library of claim 90, wherein the non-antibody sequence is a synthetic sequence.
-
92. The library of claim 90, wherein the non-antibody sequence is a cytokine sequence, a lymphokine sequence, a chemokine sequence, a growth factor sequence, a hormone sequence, or a toxin sequence.
-
93. The library of claim 90, wherein the non-antibody sequence is an IL-8 sequence, an IL-21 sequence, an SDF-1 (alpha) sequence, a somatostatin sequence, a chlorotoxin sequence, a Pro-TxII sequence, or a ziconotide sequence.
-
94. The library of claim 90, wherein the non-antibody sequence is any one of SEQ ID NOS:
- 475-481.
-
95. The library of claim 71, wherein the ultralong CDR3 comprises a X1X2X3X4X5 motif, wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q).
-
96. The library of claim 95, wherein the X1X2X3X4X5 motif is TTVHQ (SEQ ID NO:
- 153), TSVHQ (SEQ ID NO;
154), SSVTQ (SEQ ID NO;
155), STVHQ (SEQ ID NO;
156), ATVRQ (SEQ ID NO;
157), TTVYQ (SEQ ID NO;
158), SPVHQ (SEQ ID NO;
159), ATVYQ (SEQ ID NO;
160), TAVYQ (SEQ ID NO;
161), TNVHQ (SEQ ID NO;
162), ATVHQ (SEQ ID NO;
163), STVYQ (SEQ ID NO;
164), TIVHQ (SEQ ID NO;
165), AIVYQ (SEQ ID NO;
166), TTVFQ (SEQ ID NO;
167), AAVFQ (SEQ ID NO;
168), GTVHQ (SEQ ID NO;
169), ASVHQ (SEQ ID NO;
170), TAVFQ (SEQ ID NO;
171), ATVFQ (SEQ ID NO;
172), AAAHQ (SEQ ID NO;
173), VVVYQ (SEQ ID NO;
174), GTVFQ (SEQ ID NO;
175), TAVHQ (SEQ ID NO;
176), ITVHQ (SEQ ID NO;
177), ITAHQ (SEQ ID NO;
178), VTVHQ (SEQ ID NO;
179);
AAVHQ (SEQ ID NO;
180), GTVYQ (SEQ ID NO;
181), TTVLQ (SEQ ID NO;
182), TTTHQ (SEQ ID NO;
183), or TTDYQ (SEQ ID NO;
184).
- 153), TSVHQ (SEQ ID NO;
-
97. The library of claim 71, wherein the ultralong CDR3 comprises a CX1X2X3X4X5 motif.
-
98. The library of claim 97, wherein the CX1X2X3X4X5 motif is CTTVHQ (SEQ ID NO:
- 185), CTSVHQ (SEQ ID NO;
186), CSSVTQ (SEQ ID NO;
187), CSTVHQ (SEQ ID NO;
188), CATVRQ (SEQ ID NO;
189), CTTVYQ (SEQ ID NO;
190), CSPVHQ (SEQ ID NO;
191), CATVYQ (SEQ ID NO;
192), CTAVYQ (SEQ ID NO;
193), CTNVHQ (SEQ ID NO;
194), CATVHQ (SEQ ID NO;
195), CSTVYQ (SEQ ID NO;
196), CTIVHQ (SEQ ID NO;
197), CAIVYQ (SEQ ID NO;
198), CTTVFQ (SEQ ID NO;
199), CAAVFQ (SEQ ID NO;
200), CGTVHQ (SEQ ID NO;
201), CASVHQ (SEQ ID NO;
202), CTAVFQ (SEQ ID NO;
203), CATVFQ (SEQ ID NO;
204), CAAAHQ (SEQ ID NO;
205), CVVVYQ (SEQ ID NO;
206), CGTVFQ (SEQ ID NO;
207), CTAVHQ (SEQ ID NO;
208), CITVHQ (SEQ ID NO;
209), CITAHQ (SEQ ID NO;
210), CVTVHQ (SEQ ID NO;
211);
CAAVHQ (SEQ ID NO;
212), CGTVYQ (SEQ ID NO;
213), CTTVLQ (SEQ ID NO;
214), CTTTHQ (SEQ ID NO;
215), or CTTDYQ (SEQ ID NO;
216).
- 185), CTSVHQ (SEQ ID NO;
-
99. The library of claim 71, wherein the ultralong CDR3 comprises a (XaXb)z motif, wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of:
- tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
-
100. The library of claim 99, wherein the (XaXb)z motif is CYTYNYEF (SEQ ID NO:
- 217), HYTYTYDF (SEQ ID NO;
218), HYTYTYEW (SEQ ID NO;
219), KHRYTYEW (SEQ ID NO;
220), NYIYKYSF (SEQ ID NO;
221), PYIYTYQF (SEQ ID NO;
222), SFTYTYEW (SEQ ID NO;
223), SYIYIYQW (SEQ ID NO;
224), SYNYTYSW (SEQ ID NO;
225), SYSYSYEY (SEQ ID NO;
226), SYTYNYDF (SEQ ID NO;
227), SYTYNYEW (SEQ ID NO;
228), SYTYNYQF (SEQ ID NO;
229), SYVWTHNF (SEQ ID NO;
230), TYKYVYEW (SEQ ID NO;
231), TYTYTYEF (SEQ ID NO;
232), TYTYTYEW (SEQ ID NO;
233), VFTYTYEF (SEQ ID NO;
234), AYTYEW (SEQ ID NO;
235), DYIYTY (SEQ ID NO;
236), IHSYEF (SEQ ID NO;
237), SFTYEF (SEQ ID NO;
238), SHSYEF (SEQ ID NO;
239), THTYEF (SEQ ID NO;
240), TWTYEF (SEQ ID NO;
241), TYNYEW (SEQ ID NO;
242), TYSYEF (SEQ ID NO;
243), TYSYEH (SEQ ID NO;
244), TYTYDF (SEQ ID NO;
245), TYTYEF (SEQ ID NO;
246), TYTYEW (SEQ ID NO;
247), AYEF (SEQ ID NO;
248), AYSF (SEQ ID NO;
249), AYSY (SEQ ID NO;
250), CYSF (SEQ ID NO;
251), DYTY (SEQ ID NO;
252), KYEH (SEQ ID NO;
253), KYEW (SEQ ID NO;
254), MYEF (SEQ ID NO;
255), NWIY (SEQ ID NO;
256), NYDY (SEQ ID NO;
257), NYQW (SEQ ID NO;
258), NYSF (SEQ ID NO;
259), PYEW (SEQ ID NO;
260), RYNW (SEQ ID NO;
261), RYTY (SEQ ID NO;
262), SYEF (SEQ ID NO;
263), SYEH (SEQ ID NO;
264), SYEW (SEQ ID NO;
265), SYKW (SEQ ID NO;
266), SYTY (SEQ ID NO;
267), TYDF (SEQ ID NO;
268), TYEF (SEQ ID NO;
269), TYEW (SEQ ID NO;
270), TYQW (SEQ ID NO;
271), TYTY (SEQ ID NO;
272), or VYEW (SEQ ID NO;
273).
- 217), HYTYTYDF (SEQ ID NO;
-
101. The library of claim 99, wherein the (XaXb), motif is YXYXYX.
-
102. The library of claim 71, wherein the ultralong CDR3 comprises a X1X2X3X4X5Xn motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q), and wherein n is 27-54. -
103. The library of claim 71, wherein the ultralong CDR3 comprises Xn(XaXb)z motif,
wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), wherein n is 27-54, and wherein z is 1-4.
-
104. The library of claim 71, wherein the ultralong CDR3 comprises a X1X2X3X4X5Xn(XaXb)z motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), wherein X5 is glutamine (Q), Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), wherein n is 27-54, and wherein z is 1-4.
-
105. The library of claim 104, wherein the X1X2X3X4X5 motif is TTVHQ (SEQ ID NO:
- 153) or TSVHQ (SEQ ID NO;
154), and wherein the (XaXb)z motif is YXYXYX.
- 153) or TSVHQ (SEQ ID NO;
-
106. The library of claim 71, wherein the ultralong CDR3 comprises:
-
a CX1X2X3X4X5 motif, wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q), a cysteine motif selected from the group consisting of;
CX10CX5CX5CXCX7C (SEQ ID NO;
41), CX10CX6CX5CXCX15C (SEQ ID NO;
42), CX11CXCX5C (SEQ ID NO;
43), CX11CX5CX5CXCX7C (SEQ ID NO;
44), CX10CX6CX5CXCX13C (SEQ ID NO;
45), CX10CX5CXCX4CX8C (SEQ ID NO;
46), CX10CX6CX6CXCX7C (SEQ ID NO;
47), CX10CX4CX7CXCX8C (SEQ ID NO;
48), CX10CX4CX7CXCX7C (SEQ ID NO;
49), CX13CX8CX8C (SEQ ID NO;
50), CX10CX6CX5CXCX7C (SEQ ID NO;
51), CX10CX5CX5C (SEQ ID NO;
52), CX10CX5CX6CXCX7C (SEQ ID NO;
53), CX10CX6CX5CX7CX9C (SEQ ID NO;
54), CX9CX7CX5CXCX7C (SEQ ID NO;
55), CX10CX6CX5CXCX9C (SEQ ID NO;
56), CX10CXCX4CX5CX11C (SEQ ID NO;
57), CX7CX3CX6CX5CXCX5CX10C (SEQ ID NO;
58), CX10CXCX4CX5CXCX2CX3C (SEQ ID NO;
59), CX16CX5CXC (SEQ ID NO;
60), CX6CX4CXCX4CX5C (SEQ ID NO;
61), CX11CX4CX5CX6CX3C (SEQ ID NO;
62), CX8CX2CX6CX5C (SEQ ID NO;
63), CX10CX5CX5CXCX10C (SEQ ID NO;
64), CX10CXCX6CX4CXC (SEQ ID NO;
65), CX10CX5CX5CXCX2C (SEQ ID NO;
66), CX14CX2CX3CXCXC (SEQ ID NO;
67), CX15CX5CXC (SEQ ID NO;
68), CX4CX6CX9CX2CX11C (SEQ ID NO;
69), CX6CX4CX5CX5CX12C (SEQ ID NO;
70), CX7CX3CXCXCX4CX5CX9C (SEQ ID NO;
71), CX10CX6CX5C (SEQ ID NO;
72), CX7CX3CX5CX5CX9C (SEQ ID NO;
73), CX7CX5CXCX2C (SEQ ID NO;
74), CX10CXCX6C (SEQ ID NO;
75), CX10CX3CX3CX5CX7CXCX6C (SEQ ID NO;
76), CX10CX4CX5CX12CX2C (SEQ ID NO;
77), CX12CX4CX5CXCXCX9CX3C (SEQ ID NO;
78), CX12CX4CX5CX12CX2C (SEQ ID NO;
79), CX10CX6CX5CXCX11C (SEQ ID NO;
80), CX16CX5CXCXCX14C (SEQ ID NO;
81), CX10CX5CXCX8CX6C (SEQ ID NO;
82), CX12CX4CX5CX8CX2C (SEQ ID NO;
83), CX12CX5CX5CXCX8C (SEQ ID NO;
84), CX10CX6CX5CXCX4CXCX9C (SEQ ID NO;
85), CX11CX4CX5CX8CX2C (SEQ ID NO;
86), CX10CX6CX5CX8CX2C (SEQ ID NO;
87), CX10CX6CX5CXCX8C (SEQ ID NO;
88), CX10CX6CX5CXCX3CX8CX2C (SEQ ID NO;
89), CX10CX6CX5CX3CX8C (SEQ ID NO;
90), CX10CX6CX5CXCX2CX6CX5C (SEQ ID NO;
91), CX7CX6CX3CX3CX9C (SEQ ID NO;
92), CX9CX8CX5CX6CX5C (SEQ ID NO;
93), CX10CX2CX2CX7CXCX11CX5C (SEQ ID NO;
94), and CX10CX6CX5CXCX2CX8CX4C (SEQ ID NO;
95), anda (XaXb)z motif, Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of;
tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
-
-
107. The library of claim 71, wherein the ultralong CDR3 comprises:
-
a CX1X2X3X4X5 motif, wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q); a cysteine motif selected from the group consisting of;
wherein the cysteine motif is selected from the group consisting of;
CCX3CXCX3CX2CCXCX5CX9CX5CXC (SEQ ID NO;
96), CX6CX2CX5CX4CCXCX4CX6CXC (SEQ ID NO;
97), CX7CXCX5CX4CCCX4CX6CXC (SEQ ID NO;
98), CX9CX3CXCX2CXCCCX6CX4C (SEQ ID NO;
99), CX5CX3CXCX4CX4CCX10CX2CC (SEQ ID NO;
100), CX5CXCX1CXCX3CCX3CX4CX10C (SEQ ID NO;
101), CX9CCCX3CX4CCCX5CX6C (SEQ ID NO;
102), CCX8CX5CX4CX3CX4CCXCX1C (SEQ ID NO;
103), CCX6CCX5CCCX4CX4CX12C (SEQ ID NO;
104), CX6CX2CX3CCCX4CX5CX3CX3C (SEQ ID NO;
105), CX3CX5CX6CX4CCXCX5CX4CXC (SEQ ID NO;
106), CX4CX4CCX4CX4CXCX11CX2CXC (SEQ ID NO;
107), CX5CX2CCX5CX4CCX3CCX7C (SEQ ID NO;
108), CX5CX5CX3CX2CXCCX4CX7CXC (SEQ ID NO;
109), CX3CX7CX3CX4CCXCX2CX5CX2C (SEQ ID NO;
110), CX9CX3CXCX4CCX5CCCX6C (SEQ ID NO;
111), CX9CX3CXCX2CXCCX6CX3CX3C (SEQ ID NO;
112), CX8CCXCX3CCX3CXCX3CX4C (SEQ ID NO;
113), CX9CCX4CX2CXCCXCX4CX3C (SEQ ID NO;
114), CX10CXCX3CX2CXCCX4CX5CXC (SEQ ID NO;
115), CX9CXCX3CX2CXCCX4CX5CXC (SEQ ID NO;
116), CX6CCXCX5CX4CCXCX5CX2C (SEQ ID NO;
117), CX6CCXCX3CXCCX3CX4CC (SEQ ID NO;
118), CX6CCXCX3CXCX2CXCX4CX8C (SEQ ID NO;
119), CX4CX2CCX3CXCX4CCX2CX3C (SEQ ID NO;
120), CX3CX5CX3CCCX4CX9C (SEQ ID NO;
121), CCX9CX3CXCCX3CX5C (SEQ ID NO;
122), CX9CX2CX3CX4CCX4CX5C (SEQ ID NO;
123), CX9CX7CX4CCXCX7CX3C (SEQ ID NO;
124), CX9CX3CCCX10CX2CX3C (SEQ ID NO;
125), CX3CX5CX5CX4CCX10CX6C (SEQ ID NO;
126), CX9CX5CX4CCXCX5CX4C (SEQ ID NO;
127), CX7CXCX6CX4CCCX10C (SEQ ID NO;
128), CX8CX2CX4CCX4CX3CX3C (SEQ ID NO;
129), CX7CX5CXCX4CCX7CX4C (SEQ ID NO;
130), CX11CX3CX4CCCX8CX2C (SEQ ID NO;
131), CX2CX3CX4CCX4CX5CX15C (SEQ ID NO;
132), CX9CX5CX4CCX7C (SEQ ID NO;
133), CX9CX7CX3CX2CX6C (SEQ ID NO;
134), CX9CX5CX4CCX14C (SEQ ID NO;
135), CX9CX5CX4CCX8C (SEQ ID NO;
136), CX9CX6CX4CCXC (SEQ ID NO;
137), CX5CCX7CX4CX12 (SEQ ID NO;
138), CX10CX3CX4CCX4C (SEQ ID NO;
139), CX9CX4CCX5CX4C (SEQ ID NO;
140), CX10CX3CX4CX7CXC (SEQ ID NO;
141), CX7CX7CX2CX2CX3C (SEQ ID NO;
142), CX9CX4CX4CCX6C (SEQ ID NO;
143), CX7CXCX3CXCX6C (SEQ ID NO;
144), CX7CXCX4CXCX4C (SEQ ID NO;
145), CX9CX5CX4C (SEQ ID NO;
146), CX3CX6CX8C (SEQ ID NO;
147), CX10CXCX4C (SEQ ID NO;
148), CX10CCX4C (SEQ ID NO;
149), CX15C (SEQ ID NO;
150), CX10C (SEQ ID NO;
151), and CX9C (SEQ ID NO;
152); anda (XaXb)z motif, wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of;
tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
-
-
108. The library of claim 90, wherein the ultralong CDR3 comprises an additional sequence that is a linker.
-
109. The library of claim 108, wherein the linker is linked to a C-terminus, a N-terminus, or both C-terminus and N-terminus of the non-antibody sequence.
-
110. The library of claim 71, wherein the ultralong CDR3 is a ruminant CDR3.
-
111. The library of claim 110, wherein the ruminant is a cow.
-
112. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprises a human heavy chain germline sequence that is a VH4 family member germline sequence.
-
113. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprises a VH4-34 germline sequence.
-
114. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprises a human heavy chain germline sequence that is derived from VH4-34.
-
115. The library of claim 114, wherein the VH4-34 germline sequence comprises a CDR1 comprising a Gly31Asp and Tyr32Lys substitution based on Kabat numbering.
-
116. The library of claim 114, wherein the VH4-34 germline sequence comprises a CDR2 comprising a Glu50Ser substitution based on Kabat numbering.
-
117. The library of claim 114, wherein the VH4-34 germline sequence comprises a CDR1 comprising Ala23Thr, Val24Ala, Tyr25Ser, Gly27Phe, Phe29Leu, Gly31Asp, Tyr32Lys, Tyr33Ala, Trp34Val, and Ser35Gly substitutions, and a CDR2 comprising Ile48Leu, Glu50Ser, Asn52Asp, His53Thr, Ser54Gly, Ser56Asn, and Asn58Gly substitutions, based on Kabat numbering.
-
118. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:
- 470.
-
119. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:
- 461, 462, 463, 464, 465, 466, 467, 468, or 469; and
the amino acid sequence set forth in SEQ ID NO;
470.
- 461, 462, 463, 464, 465, 466, 467, 468, or 469; and
-
120. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:
- 461, 462, 463, 464, 465, 466, 467, 468, or 469;
the amino acid sequence set forth in SEQ ID NO;
470; and
a peptide sequence, wherein the peptide sequence is positioned between SEQ ID NO;
461, 462, 463, 464, 465, 466, 467, 468, or 4697, and SEQ ID NO;
470.
- 461, 462, 463, 464, 465, 466, 467, 468, or 469;
-
121. The library of claim 120, wherein the peptide sequence is a bovine sequence, a non-bovine sequence, an antibody sequence, or a non-antibody sequence.
-
122. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprises a light chain variable region sequence that is a lambda light chain variable region sequence.
-
123. The library of claim 122, wherein the lambda light chain variable region sequence is a bovine lambda light chain variable region sequence.
-
124. The library of claim 122, wherein the lambda light chain variable region sequence is a human lambda light chain variable region sequence.
-
125. The library of claim 124, wherein the human lambda light chain variable region sequence is VL1-51.
-
126. The library of claim 124, wherein the human lambda light chain variable region sequence is derived from VL1-51.
-
127. The library of claim 126, wherein the VL1-51 germline sequence comprises a CDR1 comprising Ile29Val and Asn32Gly substitution based on Kabat numbering.
-
128. The library of claim 126, wherein the VL1-51 germline sequence comprises a CDR2 comprising a substitution of DNN to GDT.
-
129. The library of claim 126, wherein the VL1-51 germline sequence comprises a CDR2 comprising a substitution of DNNKRP (SEQ ID NO:
- 471) to GDTSRA (SEQ ID NO;
472).
- 471) to GDTSRA (SEQ ID NO;
-
130. The library of claim 126, wherein the VL1-51 germline sequence comprises a S2A, T5N, P8S, A12G, A13S, and P14L substitution based on Kabat numbering.
-
131. The library of claim 126, wherein the VL1-51 germline sequence comprises a S2A, T5N, P8S, A12G, A13S, and P14L substitution based on Kabat numbering, and a CDR2 comprising a substitution of DNN to GDT.
-
132. The library of claim 71, wherein the humanized antibody or binding fragment thereof comprising the amino acid sequence set forth in SEQ ID NO:
- 440, 441, 442, 443, or 444.
-
133. The library of any one of claims 71-132, wherein the humanized antibodies or binding fragments thereof are present in a spatially addressed format.
-
72. The library of claim 71, wherein the ultralong CDR3 is 35 amino acids in length or longer, 40 amino acids in length or longer, 45 amino acids in length or longer, 50 amino acids in length or longer, 55 amino acids in length or longer, or 60 amino acids in length or longer.
-
134. A method of humanizing an antibody variable region comprising the step of genetically combining a nucleic acid sequence encoding a non-human ultralong CDR3 with a nucleic acid sequence encoding a human variable region framework (FR) sequence.
-
135. A method of generating a library of humanized antibodies that comprises an ultralong CDR3, the method comprising:
-
a.) combining a nucleic acid sequence encoding an ultralong CDR3 with a nucleic acid sequence encoding a human variable region framework (FR) sequence to produce nucleic acids encoding for humanized antibodies that comprises an ultralong CDR3; and b.) expressing the nucleic acids encoding for humanized antibodies that comprises an ultralong CDR3 to generate a library of humanized antibodies that comprises an ultralong CDR3.
-
-
136. A method of generating a library of humanized antibodies or binding fragments thereof comprising an ultralong CDR3 that comprises a non-antibody sequence, the method comprising:
-
a.) combining a nucleic acid sequence encoding an ultralong CDR3, a nucleic acid sequence encoding a human variable region framework (FR) sequence, and a nucleic acid sequence encoding a non-antibody sequence to produce nucleic acids encoding humanized antibodies or binding fragments thereof comprising an ultralong CDR3 and a non-antibody sequence, and b.) expressing the nucleic acids encoding humanized antibodies or binding fragments thereof comprising an ultralong CDR3 and a non-antibody sequence to generate a library of humanized antibodies or binding fragments thereof comprising an ultralong CDR3 and a non-antibody sequence.
-
-
137. A library of humanized antibodies or binding fragments thereof comprising an ultralong CDR3 that comprises a non-antibody sequence.
-
138. A method of generating a library of humanized antibodies or binding fragments thereof comprising an ultralong CDR3 that comprises a cysteine motif, the method comprising:
-
a.) combining a human variable region framework (FR) sequence, and a nucleic acid sequence encoding an ultralong CDR3 and a cysteine motif; b.) introducing one or more nucleotide changes to the nucleic acid sequence encoding one or more amino acid residues that are positioned between one or more cysteine residues in the cysteine motif for nucleotides encoding different amino acid residues to produce nucleic acids encoding humanized antibodies or binding fragments thereof comprising an ultralong CDR3 and a cysteine motif with one or more nucleotide changes introduced between one or more cysteine residues in the cysteine domain; and c.) expressing the nucleic acids encoding humanized antibodies or binding fragments thereof comprising an ultralong CDR3 and a cysteine motif with one or more nucleotide changes introduced between one or more cysteine residues in the cysteine domain to generate a library of humanized antibodies or binding fragments thereof comprising an ultralong CDR3 and a cysteine motif with one or more amino acid changes introduced between one or more cysteine residues in the cysteine domain.
-
-
139. A library of humanized antibodies or binding fragments thereof comprising an ultralong CDR3 that comprises a cysteine motif, wherein the antibodies or binding fragments comprise one or more substitutions of amino acid residues that are positioned between cysteine residues in the cysteine motif.
-
140. A method of generating a library of humanized antibodies or binding fragments thereof comprising a bovine ultralong CDR3, the method comprising:
-
a.) combining a nucleic acid sequence encoding a human variable region framework (FR) sequence and a nucleic acid encoding a bovine ultralong CDR3, and b.) expressing the nucleic acids encoding a human variable region framework (FR) sequence and a nucleic acid encoding a bovine ultralong CDR3 to generate a library of humanized antibodies or binding fragments thereof comprising a bovine ultralong CDR3.
-
-
141. A library of humanized antibodies or binding fragments thereof comprising a bovine ultralong CDR3.
-
142. A CDR3 scaffold comprising a X1X2X3X4X5 motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q). - View Dependent Claims (143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179)
-
143. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold is 35 amino acids in length or longer, 40 amino acids in length or longer, 45 amino acids in length or longer, 50 amino acids in length or longer, 55 amino acids in length or longer, or 60 amino acids in length or longer.
-
144. The CDR3 scaffold of claim 143, wherein the CDR3 scaffold is 35 amino acids in length or longer.
-
145. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises 3 or more cysteine residues, 4 or more cysteine residues, 5 or more cysteine residues, 6 or more cysteine residues, 7 or more cysteine residues, 8 or more cysteine residues, 9 or more cysteine residues, 10 or more cysteine residues, 11 or more cysteine residues, or 12 or more cysteine residues.
-
146. The CDR3 scaffold of claim 145, wherein the CDR3 scaffold comprises 3 or more cysteine residues.
-
147. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a cysteine motif.
-
148. The CDR3 scaffold of claim 147, wherein the cysteine motif is selected from the group consisting of:
-
149. The CDR3 scaffold of claim 147, wherein the cysteine motif is selected from the group consisting of:
-
150. The CDR3 scaffold of claim 142, wherein CDR3 scaffold comprises 2 to 6 disulfide bonds.
-
151. The CDR3 scaffold of claim 142, wherein the X1X2X3X4X5 motif in the CDR3 scaffold is TTVHQ (SEQ ID NO:
- 153), TSVHQ (SEQ ID NO;
154), SSVTQ (SEQ ID NO;
155), STVHQ (SEQ ID NO;
156), ATVRQ (SEQ ID NO;
157), TTVYQ (SEQ ID NO;
158), SPVHQ (SEQ ID NO;
159), ATVYQ (SEQ ID NO;
160), TAVYQ (SEQ ID NO;
161), TNVHQ (SEQ ID NO;
162), ATVHQ (SEQ ID NO;
163), STVYQ (SEQ ID NO;
164), TIVHQ (SEQ ID NO;
165), AIVYQ (SEQ ID NO;
166), TTVFQ (SEQ ID NO;
167), AAVFQ (SEQ ID NO;
168), GTVHQ (SEQ ID NO;
169), ASVHQ (SEQ ID NO;
170), TAVFQ (SEQ ID NO;
171), ATVFQ (SEQ ID NO;
172), AAAHQ (SEQ ID NO;
173), VVVYQ (SEQ ID NO;
174), GTVFQ (SEQ ID NO;
175), TAVHQ (SEQ ID NO;
176), ITVHQ (SEQ ID NO;
177), ITAHQ (SEQ ID NO;
178), VTVHQ (SEQ ID NO;
179);
AAVHQ (SEQ ID NO;
180), GTVYQ (SEQ ID NO;
181), TTVLQ (SEQ ID NO;
182), TTTHQ (SEQ ID NO;
183), or TTDYQ (SEQ ID NO;
184).
- 153), TSVHQ (SEQ ID NO;
-
152. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a CX1X2X3X4X5 motif.
-
153. The CDR3 scaffold of claim 152, wherein the CX1X2X3X4X5 motif in the CDR3 scaffold is CTTVHQ (SEQ ID NO:
- 185), CTSVHQ (SEQ ID NO;
186), CSSVTQ (SEQ ID NO;
187), CSTVHQ (SEQ ID NO;
188), CATVRQ (SEQ ID NO;
189), CTTVYQ (SEQ ID NO;
190), CSPVHQ (SEQ ID NO;
191), CATVYQ (SEQ ID NO;
192), CTAVYQ (SEQ ID NO;
193), CTNVHQ (SEQ ID NO;
194), CATVHQ (SEQ ID NO;
195), CSTVYQ (SEQ ID NO;
196), CTIVHQ (SEQ ID NO;
197), CAIVYQ (SEQ ID NO;
198), CTTVFQ (SEQ ID NO;
199), CAAVFQ (SEQ ID NO;
200), CGTVHQ (SEQ ID NO;
201), CASVHQ (SEQ ID NO;
202), CTAVFQ (SEQ ID NO;
203), CATVFQ (SEQ ID NO;
204), CAAAHQ (SEQ ID NO;
205), CVVVYQ (SEQ ID NO;
206), CGTVFQ (SEQ ID NO;
207), CTAVHQ (SEQ ID NO;
208), CITVHQ (SEQ ID NO;
209), CITAHQ (SEQ ID NO;
210), CVTVHQ (SEQ ID NO;
211);
CAAVHQ (SEQ ID NO;
212), CGTVYQ (SEQ ID NO;
213), CTTVLQ (SEQ ID NO;
214), CTTTHQ (SEQ ID NO;
215), or CTTDYQ (SEQ ID NO;
216).
- 185), CTSVHQ (SEQ ID NO;
-
154. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a (XaXb)z motif,
wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
-
155. The CDR3 scaffold of claim 154, wherein the (XaXb)z motif in the CDR3 scaffold is CYTYNYEF (SEQ ID NO:
- 217), HYTYTYDF (SEQ ID NO;
218), HYTYTYEW (SEQ ID NO;
219), KHRYTYEW (SEQ ID NO;
220), NYIYKYSF (SEQ ID NO;
221), PYIYTYQF (SEQ ID NO;
222), SFTYTYEW (SEQ ID NO;
223), SYIYIYQW (SEQ ID NO;
224), SYNYTYSW (SEQ ID NO;
225), SYSYSYEY (SEQ ID NO;
226), SYTYNYDF (SEQ ID NO;
227), SYTYNYEW (SEQ ID NO;
228), SYTYNYQF (SEQ ID NO;
229), SYVWTHNF (SEQ ID NO;
230), TYKYVYEW (SEQ ID NO;
231), TYTYTYEF (SEQ ID NO;
232), TYTYTYEW (SEQ ID NO;
233), VFTYTYEF (SEQ ID NO;
234), AYTYEW (SEQ ID NO;
235), DYIYTY (SEQ ID NO;
236), IHSYEF (SEQ ID NO;
237), SFTYEF (SEQ ID NO;
238), SHSYEF (SEQ ID NO;
239), THTYEF (SEQ ID NO;
240), TWTYEF (SEQ ID NO;
241), TYNYEW (SEQ ID NO;
242), TYSYEF (SEQ ID NO;
243), TYSYEH (SEQ ID NO;
244), TYTYDF (SEQ ID NO;
245), TYTYEF (SEQ ID NO;
246), TYTYEW (SEQ ID NO;
247), AYEF (SEQ ID NO;
248), AYSF (SEQ ID NO;
249), AYSY (SEQ ID NO;
250), CYSF (SEQ ID NO;
251), DYTY (SEQ ID NO;
252), KYEH (SEQ ID NO;
253), KYEW (SEQ ID NO;
254), MYEF (SEQ ID NO;
255), NWIY (SEQ ID NO;
256), NYDY (SEQ ID NO;
257), NYQW (SEQ ID NO;
258), NYSF (SEQ ID NO;
259), PYEW (SEQ ID NO;
260), RYNW (SEQ ID NO;
261), RYTY (SEQ ID NO;
262), SYEF (SEQ ID NO;
263), SYEH (SEQ ID NO;
264), SYEW (SEQ ID NO;
265), SYKW (SEQ ID NO;
266), SYTY (SEQ ID NO;
267), TYDF (SEQ ID NO;
268), TYEF (SEQ ID NO;
269), TYEW (SEQ ID NO;
270), TYQW (SEQ ID NO;
271), TYTY (SEQ ID NO;
272), or VYEW (SEQ ID NO;
273).
- 217), HYTYTYDF (SEQ ID NO;
-
156. The CDR3 scaffold of claim 154, wherein the (XaXb)z motif in the CDR3 scaffold is YXYXYX.
-
157. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a X1X2X3X4X5Xn motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q), and wherein n is 27-54. -
158. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises Xn(XaXb)z motif,
wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), wherein n is 27-54, and wherein z is 1-4.
-
159. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a X1X2X3X4X5Xn(XaXb)z motif,
wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), wherein X5 is glutamine (Q), Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of: - tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), wherein n is 27-54, and wherein z is 1-4.
-
160. The CDR3 scaffold of claim 159, wherein the X1X2X3X4X5 motif in the CDR3 scaffold is TTVHQ (SEQ ID NO:
- 153) or TSVHQ (SEQ ID NO;
154), and wherein the (XaXb)z motif is YXYXYX.
- 153) or TSVHQ (SEQ ID NO;
-
161. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises:
-
a CX1X2X3X4X5 motif, wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q), a cysteine motif selected from the group consisting of;
CX10CX5CX5CXCX7C (SEQ ID NO;
41), CX10CX6CX5CXCX15C (SEQ ID NO;
42), CX11CXCX5C (SEQ ID NO;
43), CX11CX5CX5CXCX7C (SEQ ID NO;
44), CX10CX6CX5CXCX13C (SEQ ID NO;
45), CX10CX5CXCX4CX8C (SEQ ID NO;
46), CX10CX6CX6CXCX7C (SEQ ID NO;
47), CX10CX4CX7CXCX8C (SEQ ID NO;
48), CX10CX4CX7CXCX7C (SEQ ID NO;
49), CX13CX8CX8C (SEQ ID NO;
50), CX10CX6CX5CXCX7C (SEQ ID NO;
51), CX10CX5CX5C (SEQ ID NO;
52), CX10CX5CX6CXCX7C (SEQ ID NO;
53), CX10CX6CX5CX7CX9C (SEQ ID NO;
54), CX9CX7CX5CXCX7C (SEQ ID NO;
55), CX10CX6CX5CXCX9C (SEQ ID NO;
56), CX10CXCX4CX5CX11C (SEQ ID NO;
57), CX7CX3CX6CX5CXCX5CX10C (SEQ ID NO;
58), CX10CXCX4CX5CXCX2CX3C (SEQ ID NO;
59), CX16CX5CXC (SEQ ID NO;
60), CX6CX4CXCX4CX5C (SEQ ID NO;
61), CX11CX4CX5CX6CX3C (SEQ ID NO;
62), CX8CX2CX6CX5C (SEQ ID NO;
63), CX10CX5CX5CXCX10C (SEQ ID NO;
64), CX10CXCX6CX4CXC (SEQ ID NO;
65), CX10CX5CX5CXCX2C (SEQ ID NO;
66), CX14CX2CX3CXCXC (SEQ ID NO;
67), CX15CX5CXC (SEQ ID NO;
68), CX4CX6CX9CX2CX11C (SEQ ID NO;
69), CX6CX4CX5CX5CX12C (SEQ ID NO;
70), CX7CX3CXCXCX4CX5CX9C (SEQ ID NO;
71), CX10CX6CX5C (SEQ ID NO;
72), CX7CX3CX5CX5CX9C (SEQ ID NO;
73), CX7CX5CXCX2C (SEQ ID NO;
74), CX10CXCX6C (SEQ ID NO;
75), CX10CX3CX3CX5CX7CXCX6C (SEQ ID NO;
76), CX10CX4CX5CX12CX2C (SEQ ID NO;
77), CX12CX4CX5CXCXCX9CX3C (SEQ ID NO;
78), CX12CX4CX5CX12CX2C (SEQ ID NO;
79), CX10CX6CX5CXCX11C (SEQ ID NO;
80), CX16CX5CXCXCX14C (SEQ ID NO;
81), CX10CX5CXCX8CX6C (SEQ ID NO;
82), CX12CX4CX5CX8CX2C (SEQ ID NO;
83), CX12CX5CX5CXCX8C (SEQ ID NO;
84), CX10CX6CX5CXCX4CXCX9C (SEQ ID NO;
85), CX11CX4CX5CX8CX2C (SEQ ID NO;
86), CX10CX6CX5CX8CX2C (SEQ ID NO;
87), CX10CX6CX5CXCX8C (SEQ ID NO;
88), CX10CX6CX5CXCX3CX8CX2C (SEQ ID NO;
89), CX10CX6CX5CX3CX8C (SEQ ID NO;
90), CX10CX6CX5CXCX2CX6CX5C (SEQ ID NO;
91), CX7CX6CX3CX3CX9C (SEQ ID NO;
92), CX9CX8CX5CX6CX5C (SEQ ID NO;
93), CX10CX2CX2CX7CXCX11CX5C (SEQ ID NO;
94), and CX10CX6CX5CXCX2CX8CX4C (SEQ ID NO;
95), anda (XaXb)z motif, wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of;
tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
-
-
162. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises:
-
a CX1X2X3X4X5 motif, wherein X1 is threonine (T), glycine (G), alanine (A), serine (S), or valine (V), wherein X2 is serine (S), threonine (T), proline (P), isoleucine (I), alanine (A), valine (V), or asparagine (N), wherein X3 is valine (V), alanine (A), threonine (T), or aspartic acid (D), wherein X4 is histidine (H), threonine (T), arginine (R), tyrosine (Y), phenylalanine (F), or leucine (L), and wherein X5 is glutamine (Q); a cysteine motif selected from the group consisting of;
wherein the cysteine motif is selected from the group consisting of;
CCX3CXCX3CX2CCXCX5CX9CX5CXC (SEQ ID NO;
96), CX6CX2CX5CX4CCXCX4CX6CXC (SEQ ID NO;
97), CX7CXCX5CX4CCCX4CX6CXC (SEQ ID NO;
98), CX9CX3CXCX2CXCCCX6CX4C (SEQ ID NO;
99), CX5CX3CXCX4CX4CCX10CX2CC (SEQ ID NO;
100), CX5CXCX1CXCX3CCX3CX4CX10C (SEQ ID NO;
101), CX9CCCX3CX4CCCX5CX6C (SEQ ID NO;
102), CCX8CX5CX4CX3CX4CCXCX1C (SEQ ID NO;
103), CCX6CCX5CCCX4CX4CX12C (SEQ ID NO;
104), CX6CX2CX3CCCX4CX5CX3CX3C (SEQ ID NO;
105), CX3CX5CX6CX4CCXCX5CX4CXC (SEQ ID NO;
106), CX4CX4CCX4CX4CXCX11CX2CXC (SEQ ID NO;
107), CX5CX2CCX5CX4CCX3CCX7C (SEQ ID NO;
108), CX5CX5CX3CX2CXCCX4CX7CXC (SEQ ID NO;
109), CX3CX7CX3CX4CCXCX2CX5CX2C (SEQ ID NO;
110), CX9CX3CXCX4CCX5CCCX6C (SEQ ID NO;
111), CX9CX3CXCX2CXCCX6CX3CX3C (SEQ ID NO;
112), CX8CCXCX3CCX3CXCX3CX4C (SEQ ID NO;
113), CX9CCX4CX2CXCCXCX4CX3C (SEQ ID NO;
114), CX10CXCX3CX2CXCCX4CX5CXC (SEQ ID NO;
115), CX9CXCX3CX2CXCCX4CX5CXC (SEQ ID NO;
116), CX6CCXCX5CX4CCXCX5CX2C (SEQ ID NO;
117), CX6CCXCX3CXCCX3CX4CC (SEQ ID NO;
118), CX6CCXCX3CXCX2CXCX4CX8C (SEQ ID NO;
119), CX4CX2CCX3CXCX4CCX2CX3C (SEQ ID NO;
120), CX3CX5CX3CCCX4CX9C (SEQ ID NO;
121), CCX9CX3CXCCX3CX5C (SEQ ID NO;
122), CX9CX2CX3CX4CCX4CX5C (SEQ ID NO;
123), CX9CX7CX4CCXCX7CX3C (SEQ ID NO;
124), CX9CX3CCCX10CX2CX3C (SEQ ID NO;
125), CX3CX5CX5CX4CCX10CX6C (SEQ ID NO;
126), CX9CX5CX4CCXCX5CX4C (SEQ ID NO;
127), CX7CXCX6CX4CCCX10C (SEQ ID NO;
128), CX8CX2CX4CCX4CX3CX3C (SEQ ID NO;
129), CX7CX5CXCX4CCX7CX4C (SEQ ID NO;
130), CX11CX3CX4CCCX8CX2C (SEQ ID NO;
131), CX2CX3CX4CCX4CX5CX15C (SEQ ID NO;
132), CX9CX5CX4CCX7C (SEQ ID NO;
133), CX9CX7CX3CX2CX6C (SEQ ID NO;
134), CX9CX5CX4CCX14C (SEQ ID NO;
135), CX9CX5CX4CCX8C (SEQ ID NO;
136), CX9CX6CX4CCXC (SEQ ID NO;
137), CX5CCX7CX4CX12 (SEQ ID NO;
138), CX10CX3CX4CCX4C (SEQ ID NO;
139), CX9CX4CCX5CX4C (SEQ ID NO;
140), CX10CX3CX4CX7CXC (SEQ ID NO;
141), CX7CX7CX2CX2CX3C (SEQ ID NO;
142), CX9CX4CX4CCX6C (SEQ ID NO;
143), CX7CXCX3CXCX6C (SEQ ID NO;
144), CX7CXCX4CXCX4C (SEQ ID NO;
145), CX9CX5CX4C (SEQ ID NO;
146), CX3CX6CX8C (SEQ ID NO;
147), CX10CXCX4C (SEQ ID NO;
148), CX10CCX4C (SEQ ID NO;
149), CX15C (SEQ ID NO;
150), CX10C (SEQ ID NO;
151), and CX9C (SEQ ID NO;
152); anda (XaXb)z motif, wherein Xa is any amino acid residue, Xb is an aromatic amino acid selected from the group consisting of;
tyrosine (Y), phenylalanine (F), tryptophan (W), and histidine (H), and wherein z is 1-4.
-
-
163. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises SEQ ID NO:
- 40 or a derivative thereof.
-
164. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises amino acid residues 3-6 of any of one SEQ ID NO:
- 1-4.
-
165. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a non-human DH or a derivative thereof.
-
166. The CDR3 scaffold of claim 165, wherein the non-human DH is SEQ ID NO:
- 5, SEQ ID NO;
6, SEQ ID NO;
7, SEQ ID NO;
8, SEQ ID NO;
9, SEQ ID NO;
10, SEQ ID NO;
11, or SEQ ID NO;
12
- 5, SEQ ID NO;
-
167. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a JH sequence or a derivative thereof.
-
168. The CDR3 scaffold of claim 167, wherein the JH sequence is SEQ ID NO:
- 13, SEQ ID NO;
14, SEQ ID NO;
15, SEQ ID NO;
16, or SEQ ID NO;
17.
- 13, SEQ ID NO;
-
169. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises:
-
a non-human VH sequence or a derivative thereof; a non-human DH sequence or a derivative thereof; and
/ora JH sequence or derivative thereof.
-
-
170. The CDR3 scaffold of claim 169, wherein the CDR3 scaffold comprises an additional amino acid sequence comprising two to six amino acid residues or more positioned between the VH sequence and the DH sequence.
-
171. The CDR3 scaffold of claim 170, wherein the additional amino acid sequence is selected from the group consisting of:
- IR, IF, SEQ ID NO;
18, SEQ ID NO;
19, SEQ ID NO;
20 or SEQ ID NO;
21.
- IR, IF, SEQ ID NO;
-
172. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold comprises a sequence derived from or based on SEQ ID NO:
- 22, SEQ ID NO;
23, SEQ ID NO;
24, SEQ ID NO;
25, SEQ ID NO;
26, SEQ ID NO;
27, or SEQ ID NO;
28.
- 22, SEQ ID NO;
-
173. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold is a ruminant CDR3 scaffold.
-
174. The CDR3 scaffold of claim 173, wherein the ruminant is a cow.
-
175. A library comprising the CDR3 scaffold of any one of claims 142-174.
-
176. A polynucleotide encoding the CDR3 scaffold of any one of claims 142-174.
-
177. A vector comprising the polynucleotide of claim 176.
-
178. A host cell comprising the vector of claim 177.
-
179. The host cell of claim 178, wherein the cell is a bacteria, virus, or bacteriophage.
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143. The CDR3 scaffold of claim 142, wherein the CDR3 scaffold is 35 amino acids in length or longer, 40 amino acids in length or longer, 45 amino acids in length or longer, 50 amino acids in length or longer, 55 amino acids in length or longer, or 60 amino acids in length or longer.
Specification
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Current AssigneeThe Scripps Research Institute
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Original AssigneeThe Scripps Research Institute
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InventorsSmider, Vaughn, Bazirgan, Omar A., Mao, Hongyuan Helen
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Application NumberUS14/371,170Publication NumberTime in Patent OfficeDaysField of SearchUS Class Current506/17CPC Class CodesA61P 35/00 Antineoplastic agentsC07K 14/5421 IL-8C07K 16/00 Immunoglobulins [IGs], e.g....C07K 16/005 constructed by phage librariesC07K 16/1081 Togaviridae, e.g. flaviviru...C07K 16/46 Hybrid immunoglobulins hybr...C07K 2317/20 characterized by taxonomic ...C07K 2317/24 containing regions, domains...C07K 2317/565 Complementarity determining...C07K 2318/10 Immunoglobulin or domain(s)...C07K 2319/00 Fusion polypeptideC07K 2319/33 fusions for targeting to sp...C40B 40/10 Libraries containing peptid...