Structural analysis of the complex formed between the ansamycin antibiotic rifabutin and the BCL6 BTB-POZ domain

BCL6 is a zinc finger transcriptional repressor that is over-expressed due to chromosomal translocations, or other abnormalities, in ~30-40% of the aggressive disease diffuse large B-cell lymphoma. BCL6 accomplishes its effects through the recruitment of co-repressors to the BTB-POZ domain of BCL6, which is a critical interaction for both a normal immune response and lymphomagenesis. Peptides or small molecule inhibitors, which prevent the association of the SMRT co-repressor with BCL6, abolish BCL6 function by attenuating its transcriptional repression. However, these agents are not yet suitable for clinical practice and there is a need to develop high-affinity and cell permeable BCL6 inhibitors. In order to discover compounds, which have the potential to be developed into BCL6 inhibitors, a natural product library was screened, and it was found that the ansamycin antibiotic, rifamycin SV, had the ability to inhibit BCL6 transcriptional repression. NMR spectroscopy confirmed a direct interaction between rifamycin SV and the BTBPOZ domain of BCL6. In addition to rifamycin SV, NMR was used to screen other members of the ansamycin family for potential binding to BCL6. The rifamycin SV derivative, rifabutin, was also shown to interact with the BCL6 BTB-POZ domain. A 2.3Å X-ray crystal structure of the BCL6-rifabutin complex revealed that rifabutin occupies a shallow pocket within the lateral groove, which is similar to that occupied by the SMRT binding peptide and 79-6, the previously described small molecule inhibitor. Further work employing artificial peptides showed the importance of interactions between specific residues of SMRT and the BCL6 BTB-POZ domain. The data presented in this thesis demonstrates a potentially druggable site on the BCL6 BTBPOZ domain and a unique approach to developing a structure activity relationship for a compound that will form the basis of a therapeutically useful BCL6 inhibitor.