Focused libraries of genetic packages

• US 9,617,536 B2
• Filed: 08/10/2012
• Issued: 04/11/2017
• Est. Priority Date: 12/18/2000
• Status: Expired due to Term

• Current Assignee: Dyax Corporation (Shire plc)
• Original Assignee: Dyax Corporation (Shire plc)
• Inventors: Ladner, Robert Charles
• Primary Examiner(s): BOESEN, CHRISTIAN C
• Application Number:
  US13/571,661
• Publication Number:
  US 20120302463A1
• Time in Patent Office: 1,705 Days
• Field of Search: None
• CPC Class Codes: C07K 16/00 Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies antibodies with enzymatic activity, e.g. abzymes C12N9/0002
• CPC Class Codes: C07K 16/005 constructed by phage libraries
• CPC Class Codes: C07K 2317/565 Complementarity determining region [CDR]
• CPC Class Codes: C12N 15/1037 Screening libraries presented on the surface of microorganisms, e.g. phage display, E. coli display
• CPC Class Codes: C40B 40/02 Libraries contained in or displayed by microorganisms, e.g. bacteria or animal cells; Libraries contained in or displayed by vectors, e.g. plasmids; Libraries containing only microorganisms or vectors
• CPC Class Codes: C40B 40/08 Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries
• CPC Class Codes: C40B 40/10 Libraries containing peptides or polypeptides, or derivatives thereof

Abstract

Focused libraries of vectors or genetic packages that display, display and express, or comprise a member of a diverse family of antibody peptides, polypeptides or proteins and collectively display, display and express, or comprise at least a portion of the focused diversity of the family. The libraries have length and sequence diversities that mimic that found in native human antibodies.

Claims

1. A focused library of DNA plasmids or genetic packages comprising the DNA plasmids, the focused library comprising multiple different sets of variegated DNA molecules, each of which comprises a DNA sequence that encodes an antibody heavy chain variable domain having both frame work (FW) regions and complementary determining regions (CDR), wherein each antibody heavy chain variable domain comprises antibody heavy chain FW1, antibody heavy chain CDR1, antibody heavy chain FW2, antibody heavy chain CDR2, antibody heavy chain FW3, antibody heavy chain CDR3 and antibody heavy chain FW4 in a DNA molecule arranged in the orientation of FW1-CDR1-FW2-CDR2-FW3-CDR3-FW4, wherein each set of the variegated DNA molecules collectively encodes a plurality a different antibody heavy chain variable domains, which share a common heavy chain CDR3 motif selected from one of the following amino acid sequences:

xxxYCSSTSCxxx (SEQ ID NO;

     199), wherein each x independently is any amino acid or is absent;

xxxxYVWGSYxxx (SEQ ID NO;

200), wherein each x independently is any amino acid or is absent;

xxxxxxxSGYxxx (SEQ ID NO;

201), wherein each x independently is any amino acid or is absent;

xxxYDILTGYYxx (SEQ ID NO;

202), wherein each x independently is any amino acid or is absent;

xxxVVVPAAxxxx (SEQ ID NO;

203), wherein each x independently is any amino acid or is absent;

xxxYYDSSGYxx (SEQ ID NO;

204), wherein each x independently is any amino acid or is absent;

xxxDFWSGxxxx (SEQ ID NO;

205), wherein each x independently is any amino acid or is absent;

xxxTIFGVxxxx (SEQ ID NO;

206), wherein each x independently is any amino acid or is absent;

xxxxIVATxxxx (SEQ ID NO;

207), wherein each x independently is any amino acid or is absent;

xxx YGSGSYYx (SEQ ID NO;

208), wherein each x independently is any amino acid or is absent;

xxxxYSYGxxx (SEQ ID NO;

209), wherein each x independently is any amino acid or is absent;

xxxCSGxxCYx (SEQ ID NO;

210), wherein each x independently is any amino acid or is absent;

xxxxAAAGxxx (SEQ ID NO;

211), wherein each x independently is any amino acid or is absent;

xGxxxGGNxxx (SEQ ID NO;

212), wherein each x independently is any amino acid or is absent;

xxxSGSYxxx (SEQ ID NO;

213), wherein each x independently is any amino acid or is absent;

xxxSSSWxxx (SEQ ID NO;

214), wherein each x independently is any amino acid or is absent;

xxxxTTVTTx (SEQ ID NO;

215), wherein each x independently is any amino acid or is absent;

xxxC(SG)GDxCx (SEQ ID NO;

216), wherein each x independently is any amino acid or is absent;

xxx(IV)AVAGxx (SEQ ID NO;

217), wherein each x independently is any amino acid or is absent;

xxLWFGELxx (SEQ ID NO;

218), wherein each x independently is any amino acid or is absent;

GxxWLxxxxF (SEQ ID NO;

219), wherein each x independently is any amino acid or is absent;

xxxDTxMVxx (SEQ ID NO;

220), wherein each x independently is any amino acid or is absent;

xxxxxGExxx (SEQ ID NO;

221), wherein each x independently is any amino acid or is absent;

xxxxSGWxx (SEQ ID NO;

222), wherein each x independently is any amino acid or is absent;

xxxxGYNxx (SEQ ID NO;

223), wherein each x independently is any amino acid or is absent;

xxxVRGVxx (SEQ ID NO;

224), wherein each x independently is any amino acid or is absent;

xxxIAAxxx (SEQ ID NO;

225), wherein each x independently is any amino acid or is absent;

xxYxWNxxx (SEQ ID NO;

226), wherein each x independently is any amino acid or is absent;

xxxYGDxx (SEQ ID NO;

227), wherein each x independently is any amino acid or is absent;

xxVGATxx (SEQ ID NO;

228), wherein each x independently is any amino acid or is absent; and

xxxYSSSx (SEQ ID NO;

229), wherein each x independently is any amino acid or is absent;

wherein the variation within each set of the variegated DNA molecules, which collectively encodes the plurality of different antibody heavy chain variable domains, is due to variation in x in the residue(s) of the amino acid sequence CDR3 motif;

wherein the multiple different sets of the variegated DNA molecules collectively encode two or more amino acid sequences selected from SEQ ID NOs 199-229, and

wherein the variegated DNA molecules each comprise a human germline V_H gene as the FW1, FW2 and FW3 regions, and

the germline V_H gene being selected from the group consisting of 3-23, 4-34, 3-30, 3-30.3, and 4-30.1; and

wherein the diversity of the focused library is biased towards the natural diversity of heavy chain CDR3s of found in the human antibody family antibodies, both in sequence and in length.

2. The focused library of claim 1, wherein the library is a library of DNA plasmids.

3. The focused library of claim 1, wherein the library is a library of genetic packages comprising the DNA plasmids.

4. The focused library of DNA plasmids of claim 1, wherein the DNA plasmids are in yeast cells.

5. The focused library of claim 4, wherein the yeast cells display the plurality of antibody heavy chain variable regions.

6. The focused library of claim 1, wherein the DNA plasmids are yeast DNA plasmids.