Single-Cell Analysis Uncovers Regulatory Program in Rare Leukemia

Identifying the causes of human diseases requires deconvolution of abnormal molecular phenotypes spanning DNA accessibility, gene expression and protein abundance. Mixed-phenotype acute leukemia exhibits features of both acute myeloid leukemia and acute lymphoblastic leukemia and, as such, is marked by features of multiple hematopoietic lineages.

Mixed phenotype acute leukemia is a very rare type of leukemia where more than one type of leukemia occurs at the same time. This can happen when a person has either: both acute lymphoblastic leukemia (ALL) blasts (cancer cells) and acute myeloblastic leukemia (AML) blasts at the same time or leukemic blasts that have features of both ALL and AML on the same cell.

Scientists at the Stanford University School of Medicine (Stanford, CA, USA) and their colleagues identified pathological molecular features of mixed-phenotype acute leukemia by first analyzing the single-cell transcriptomic and epigenetic profiles of healthy blood cells during their development. Once they established profiles of those healthy cells, they examined how the profiles of leukemic cells compared.

The team performed droplet-based cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) and single-cell assay for transposase-accessible chromatin by sequencing (scATAC-seq) of more than 35,000 healthy bone marrow and peripheral blood mononuclear cells. With this, they generated multi-omic maps of hematopoiesis. They validated the maps and found them to reflect the essential phenotypic, transcriptomic, and epigenetic features of blood development. They developed a framework to analyze signatures of hematopoietic development at the single-cell level. With this, they then sought to examine how those signatures differed between healthy and leukemic cells.

The team assayed thousands of single cells from mixed-phenotype acute leukemia (MPAL) samples using both CITE-seq and scATAC-seq and identified 4,616 genes that were differentially upregulated and 72,196 significantly upregulated peaks. They projected these single-cell analyses onto their hematopoietic maps to find epigenetic and gene expression diversity and grouped the cells into broad hematopoietic development compartments. They focused on the transcription factors that might regulate these leukemia programs and found that RUNX1 motifs were enriched among certain MPAL subsets.

RUNX1, they noted, is a frequently mutated gene in hematological malignancies, and they uncovered 732 genes regulated by a RUNX1-containing distal element in at least two MPAL subsets. Additionally, CD69 which has been linked to lymphocyte activation through JAK-STAT signaling and lymphocyte retention in lymphoid organs was differentially upregulated in nearly every MPAL subset. The authors concluded that their results demonstrate how integrative, multiomic analysis of single cells within the framework of normal development can reveal both distinct and shared molecular mechanisms of disease from patient samples. The study was published on December 2, 2019 in the journal Nature Biotechnology.

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Stanford University School of Medicine