Diffuse large B cell lymphomas (DLBCLs) arise from proliferating B cells transiting different stages of the germinal center reaction. In activated B cell DLBCLs (ABC-DLBCLs), a class of DLBCLs that respond poorly to current therapies, chromosomal translocations and amplification lead to constitutive expression of the B cell lymphoma 6 (BCL6) oncogene. The role of BCL6 in maintaining these lymphomas has not been investigated. Here, we designed small-molecule inhibitors that display higher affinity for BCL6 than its endogenous corepressor ligands to evaluate their therapeutic efficacy for targeting ABC-DLBCL. We used an in silico drug design functional-group mapping approach called SILCS to create a specific BCL6 inhibitor called FX1 that has 10-fold greater potency than endogenous corepressors and binds an essential region of the BCL6 lateral groove. 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. Low doses of FX1 induced regression of established tumors in mice bearing DLBCL xenografts. Furthermore, FX1 suppressed ABC-DLBCL cells in vitro and in vivo, as well as primary human ABC-DLBCL specimens ex vivo. These findings indicate that ABC-DLBCL is a BCL6-dependent disease that can be targeted by rationally designed inhibitors that exceed the binding affinity of natural BCL6 ligands.

BCL6 was initially discovered as an oncogene in B-cell lymphomas, where it drives the malignant phenotype by repressing proliferation and DNA damage checkpoints and blocking B-cell terminal differentiation. BCL6 mediates its effects by binding to hundreds of target genes, and then repressing these genes by recruiting several different chromatin modifying corepressor complexes. Structural characterization of BCL6-corepressor complexes suggested that BCL6 might be a druggable target. Accordingly a number of compounds have been designed to bind to BCL6 and block corepressor recruitment. These compounds, based on peptide or small molecule scaffolds, can potently block BCL6 repression of target genes and kill lymphoma cells. In the case of diffuse large B-cell lymphomas (DLBCLs), BCL6 inhibitors are equally effective in suppressing both the GCB- and the more aggressive ABC-DLBCL subtypes, both of which require BCL6 to maintain their survival. In addition, BCL6 is implicated in an expanding scope of hematologic and solid tumors. These include but are not limited to B-acute lymphoblastic leukemia, chronic myeloid leukemia, breast cancer and non-small cell lung cancer. BCL6 inhibitors have been shown to exert potent effects against these tumor types. Moreover mechanism based combinations of BCL6 inhibitors with other agents has yielded synergistic and often quite dramatic activity. Hence there is a compelling case to accelerate development of BCL6 targeted therapies for translation to the clinical setting.

Summary Although the BCL6 transcriptional repressor is frequently expressed in human follicular lymphomas (FL), its biological role in this disease remains unknown. Herein we comprehensively identify the set of gene promoters directly targeted by BCL6 in primary human FLs. We noted that BCL6 binds and represses NOTCH2 and Notch pathway genes. Moreover, BCL6 and NOTCH2 pathway gene expression is inversely correlated in FL. Notably BCL6 up-regulation is associated with repression of Notch2 and its target genes in primary human and murine germinal center cells. Repression of Notch2 is an essential function of BCL6 in FL and GC B-cells since inducible expression of Notch2 abrogated GC formation in mice and kills FL cells. Indeed BCL6-targeting compounds or gene silencing leads to the induction of NOTCH2 activity and compromises survival of FL cells whereas NOTCH2 depletion or pathway antagonists rescue FL cells from such effects. Moreover, BCL6 inhibitors induced NOTCH2 expression and suppressed growth of human FL xenografts in vivo and primary human FL specimens ex vivo. These studies suggest that established FLs are thus dependent on BCL6 through its suppression of NOTCH2.

The innate abilities of cancer stem cells (CSCs), such as multi-drug resistance, drug efflux, quiescence and ionizing radiation tolerance, protect them from most traditional chemotherapeutics. As a result, this small subpopulation of persistent cells leads to more aggressive and chemoresistant cancers, causing tumour relapse and metastasis. This subpopulation is differentiated from the bulk tumour population through a wide variety of surface markers expressed on the cell surface. Recent developments in nanomedicine and targeting delivery methods have given rise to new possibilities for specifically targeting these markers and preferentially eliminating CSCs. Herein, we first summarize the range of surface markers identifying CSC populations in a variety of cancers; then, we discuss recent attempts to actively target CSCs and their niches using liposomal, nanoparticle, carbon nanotube and viral formulations.

Rationale: The BCL6 oncogene is constitutively activated by chromosomal translocations and amplification in ABC-DLBCLs, a class of DLBCLs that respond poorly to current therapies. Yet the role of BCL6 in maintaining these lymphomas has not been investigated. BCL6 mediates its effects by recruiting corepressors to an extended groove motif. Development of effective BCL6 inhibitors requires compounds exceeding the binding affinity of these corepressors. Objectives: To design small molecule inhibitors with superior potency vs. endogenous BCL6 ligands for unmet putative therapeutic needs such as targeting ABC-DLBCL. Findings: We used an in silico drug design functional-group mapping approach called SILCS to create a specific BCL6 inhibitor with 10-fold greater potency than endogenous corepressors. The compound, called FX1, binds in such a way as to occupy an essential region of the BCL6 lateral groove. FX1 disrupts BCL6 repression complex formation, reactivates BCL6 target genes, and mimics the phenotype of mice engineered to express BCL6 with lateral groove mutations. This compound eradicated established DLBCLs xenografts at low doses. Most strikingly, FX1 suppressed ABC-DLBCL cells as well as primary human ABC-DLBCL specimens ex vivo. Conclusions: ABC-DLBCL is a BCL6 dependent disease that can be targeted by novel inhibitors able to exceed the binding affinity of natural BCL6 ligands. Citation Format: Mariano Gonzalo Cardenas, Wenbo Yu, Wendy Beguelin, Teater R. Matthew, Huimin Geng, Rebecca Goldstein, Erin Oswald, Katerina Hatzi, Shao-Ning Yang, Joanna Cohen, Rita Shaknovich, Kenno Vanommeslaeghe, Huimin Cheng, Dongdong Liang, Hyoje Cho, Joshua Abbott, Wayne Tam, John P. Leonard, Leandro Cerchietti, Tomasz Cierpicki, Fengtian Xue, Alexander D. MacKerell, Jr., Ari Melnick. Therapeutic targeting of GCB- and ABC-DLBCLS by rationally designed BCL6 inhibitors. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Targeting the Vulnerabilities of Cancer; May 16-19, 2016; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(1_Suppl):Abstract nr A11.