The p30 isoform of CEBPA uncovers a silent enhancer to drive the expression of the tumor promotive factor CD73 in CEBPA mutant AML

CEBPA is a key hematopoietic transcription factor (TF), found mutated in 5-14% of all acute myeloid leukemia (AML) cases, but the direct molecular ramifications of this driver mutation remains elusive. To investigate CEBPA-mutant AML, we compared patient aberrant genetic programs with changes in a precise mouse model (Lp30) expressing only the cancer-prevalent truncated CEBPA, p30, and identified a stringent cross-species AML program. Small-scale ChIP-seq methodology revealed aberrantly activated enhancers, exclusively occupied by CEBPA in leukemia. One cancer-enhancer upstream of Nt5e, encoding CD73, was physically and functionally linked to this conserved AML gene, and could be activated by CEBPA. Targeting of CD73-adenosine signaling increased survival in AML transplanted mice. Our data thus indicate a first-in-class link between a TF cancer driver mutation and a druggable, direct transcriptional target. Overall design: RNA-seq of samples of human healthy donors vs CEBPA-biallelic AML patients was compared to RNA-seq of mouse WT versus CEBPA-biallelic AML model Lp30. ChIP-seq of CEBPA, Histone marks H3K27ac and H3K4me1 in mouse WT versus CEBPA-biallelic AML model Lp30. ChIP-seq of histone mark H3K27ac in samples of human healthy donors versus mono-allelic and biallelic CEBPA-mut AML. All samples are rigorously FACS isolated GMP phenotypic cells, in vivo material. RNA-seq, different shRNA constructs, Nt5e-KD versus scramble-KD in mouse Lp30 CEBPA-biallelic model cells in tissue culture.