Repeat enhancers as drivers of ETV6-RUNX1+ and ETV6-RUNX1-like B-lymphoblastic leukemia

PROJECT SUMMARY Genetic subtypes of B-cell acute lymphoblastic leukemia (B-ALL) bearing aberrations of the ETV6 gene, including the ETV6-RUNX1 fusion gene and inactivating mutations of ETV6, are collectively among the most common subtypes of B-ALL in children. However, our understanding of the biological basis of this common disease has stalled, in part because mouse models have failed to recapitulate many aspects of the human disease biology. We have identified a novel mechanism that directly ties aberrations in ETV6 function to the cancer-specific gene expression programs that define ETV6-altered subtypes of B-ALL. We have shown that the ETV6 transcriptional repressor binds to microsatellite repeats with the sequence ‘GGAA’ in B-ALL with intact ETV6. In B-ALL with defective ETV6 function, repression of these repeats is lost. These repeats are bound by the ETS activator ERG and other unidentified factors, take on an active enhancer-like state, and directly activate the expression of long-identified ETV6-RUNX1 signature genes, including oncogenic pathway regulators. Because GGAA repeat locations are not conserved between humans and mice, we believe that these specific regulatory relationships can only be studied in human cells. In this project, we will use innovative methods to study the activating mechanisms of GGAA repeat enhancers and their specific effects on leukemia fitness in human leukemia cell lines, samples, and xenografts. We will provide direct evidence for GGAA repeat enhancer activation as the causative mechanism for the subtype- defining gene expression signature of ETV6-RUNX1-like B-ALL, a rare but aggressive B-ALL subtype. We will identify how pervasive repeat enhancer activation affects global and local chromatin interactions, and identify epigenetic barriers to de novo repeat enhancer activation. We will use a proteomic approach with engineered repeat-binding proteins to identify the factors that cooperate in the setting of ETV6 loss to directly activate GGAA repeat enhancers in B-ALL. Together, these investigations will provide answers to critical questions about the biology of this common childhood cancer, opening new potential opportunities for therapeutic de-intensification or prevention.