Abstract PR09: Single-cell transcriptional profiling of acute myeloid leukemia identifies self-renewing stem cells

Acute myeloid leukemia (AML) is a lethal cancer with a survival of less than 50%. Standard cytotoxic therapies frequently induce complete remission, but patients frequently relapse and die of their disease. Leukemia stem cells (LSCs) are the leukemia cells with self-renewal potential and ability to recapitulate the disease. Most anticancer therapies are designed to inhibit proliferation. Yet, in hematopoietic stem cells, the mechanisms of proliferation are distinct from self-renewal (Li et al. Nature 2013). Consequently, targeting proliferation may explain the failure of traditional chemotherapy to target LSCs and eradicate AML. Our goal is define the self-renewing LSCs in order to develop therapeutics that target them and eliminate AML relapse. We previously showed that activated NRAS (NRASG12V) facilitates self-renewal in the LSC-enriched subpopulation of a transgenic mouse model of AML (Mll-AF9/NRASG12V) (Sachs et al. Blood 2014; Kim et al. Blood 2009). We hypothesize that self-renewal capacity and the NRAS-activated pathways required for self-renewal are limited to a subpopulation of LSCs. We used single-cell RNA sequencing to identify the self-renewing cells among the LSC-enriched subgroup in this model (Mac1LowKit+Sca1+, “MKS”). We identified three discrete transcriptional profiles among the LSC-enriched subpopulation and found that that two of these profiles (Profile 1 and Profile 2) are NRASG12V-dependent. These two profiles can be differentiated by CD36 and CD69 expression. We sorted the MKS LSCs based on CD36 and CD69 expression. Sorted LSC subsets were transplanted into recipient mice to compare their ability to transfer leukemia as a measure of their self-renewal capacity. We found that MKS-CD36-CD69+ cells (consistent with Profile 1) rapidly transferred leukemia with high penetrance in 20 of 22 mice. In contrast, MKS-CD36+CD69- cells (Profile 2) failed transfer leukemia in most mice; only 2 of 25 of these mice developed AML (p < 0.004). In our previous work, we demonstrated that the NRASG12V-activated self-renewal gene expression profile that we identified in our murine model was expressed in human AML, suggesting that the gene expression behavior of LSCs from this model may recapitulate the gene expression behavior of human LSCs (Sachs et al. Blood 2014). In order to determine if the single-cell transcriptional profiles of our murine AML can be found in primary human AML precursors, we performed single-cell RNA sequencing on CD34+ human AML c