FX1

FX1 disrupted formation of the BCL6 repression complex, reactivated BCL6 target genes, and mimicked the phenotype of mice engineered to express BCL6 with corepressor binding site mutations. FX1 suppressed ABC-DLBCL cells in vitro and in vivo, as well as primary human ABC-DLBCL specimens ex vivo. FX1 is specific to BCL6 and binds with a greater affinity than the natural BCL6 ligand SMRT. FX1 almost invariantly induced significant derepression of these genes(BCL6 target genes CASP8, CD69, CXCR4, CDKN1A, and DUSP5) as compared with vehicle in 2 independent DLBCL cell lines. FX1 was more than 100-fold more powerful than the previous generation of BCL6 inhibitors represented by 79-6, and 300-fold more potent than the recently reported binding of the antibiotics rifamycin and rifabutin (KD ~1 mM)^[1].

Quantitative ChIP is performed in SUDHL-6 cells exposed to FX1 (black bars) or vehicle (white bars) in DLBCL cells using antibodies for BCL6, SMRT, BCOR, or IgG control to enrich for known BCL6 binding sites in the CD69, CXCR4, and DUSP5 loci, or a negative control region.

Low doses of FX1 induce regression of established tumors in mice bearing DLBCL xenografts. The half-life is estimated to be approximately 12 hours for FX1 in SCID mice. No signs of toxicity, inflammation, or infection are evident from H&E-stained sections of lung, gastrointestinal tract, heart, kidney, liver, spleen, and bone marrow of the fixed organs from mice treated with FX1 compared with vehicle. Peripheral blood counts and serum chemistry in FX1-treated mice are also examined and they remain within normal parameters. FX1 causes profound and significant suppression of DLBCLs in DLBCL xenografts, and indeed not only prevented growth of the xenografts but in addition causes these tumors to shrink from their initial volume. The maximal effect is already achieved by the lower 25 mg/kg dose. TUNEL and Ki-67 staining shows that FX1 also induced more apoptosis and growth arrest than 79-6, respectively^[1].