Means and methods for producing high affinity antibodies
The invention provides means and methods for modulating the occurrence of somatic hypermutations in antibody producing plasmablast-like B-cells.
1. A method for increasing the occurrence of somatic hypermutations in an antibody producing plasmablast-like B-cell, the method comprising:
introducing an exogenous nucleic acid molecule encoding Bcl-6 into a B cell, wherein said B cell is a memory B cell, and introducing an exogenous anti-apoptotic nucleic acid molecule into said B cell, thus generating an antibody producing plasmablast-like B-cell with functional AID activity; and
demonstrating an increased incidence of somatic hypermutations resulting from said functional AID activity.
2. The method according to claim 1 , wherein said anti-apoptotic nucleic acid molecule comprises a BCL2 family gene.
3. The method according to claim 1 , further comprising: providing said B-cell with IL21 and CD40L.
4. The method according to claim 2 , wherein the BCL2 family gene is Bcl-xL or Mcl 1.
5. The method according to claim 1 , wherein the increased incidence of somatic hypermutations is demonstrated by analyzing the VH gene of said B cell for the presence of mutations.
6. A method according to claim 5 , further comprising: analyzing the VH and VL sequences of said B cell or said antibody to determine the immunoglobulin residues important for antigen binding.
7. The method according to claim 5 , further comprising at least one of:
selecting immunoglobulins that harbor mutations that were not present in the B cell before transduction with Bcl-6 and the anti-apoptotic nucleic acid;
selecting one or more immunoglobulin variants having a higher affinity for a ligand that is recognized by the immunoglobulin, or
selecting a variant that is more stable than other variants.