In chronic myeloid leukemia, persistence of leukemia stem cells during tyrosine kinase inhibitor treatment is influenced by interaction with αβT cells

About This Grant

Survival of chronic myeloid leukemia (CML) patients with a sustained major molecular response to Bcr-abl- tyrosine kinase inhibitors (TKIs) may approach age matched controls. Attempts at therapy discontinuation are standard of care for such patients, and 40-50% sustain a therapy free remission (TFR). However, there are no clinical tools that reliably predict TFR duration for an individual. Prior single cell RNA-sequencing (scRNA-Seq) defined a TKI sensitive, proliferative subset of CML-leukemia stem cells (LSCs) and a relatively quiescent subset that persists during TKI treatment. Mechanisms for emergence of these subpopulations are unclear. In preliminary studies, we identified an interaction between CML-LSCs and αβT cells that predicted short TFR in human subjects or a murine CML model. This LSC subset and interacting T cells expanded during post TKI- discontinuation relapse; was not found with sustained TFR; and regressed with a second TKI-induced remission. We identified LSC-interacting T cells as CD8+αβTCR+. To study the impact on TFR, we isolated αβT cell/LSC doublets from mice in TKI-remission, and singlet LSCs from mice with or without doublets by flow cytometry for transplant. Without TKI treatment, recipients of LSC doublets rapidly relapsed, but recipients of single LSCs did not. Adding αβTCR or PDL1 antibody to TKI-treatment decreased doublets and lengthened TFR in recipients. In scRNA-Seq of isolated doublet vs singlet GFP+Lin-ckit+ cells, we found single LSC transcriptomes suggested cell cycle progression and resistance to apoptosis via the unfolded protein response, but doublet LSC transcriptomes suggested HSC quiescence and apoptosis resistance via IAPs. Expression of T cell suppressor genes was increased in doublet LSCs, including PDL1 and Ctla4 ligands, as were cognate proteins on T cells. We hypothesize that interaction of CML-LSC with αβT-cells favors LSC persistence during TKI treatment and provides a source of relapse post discontinuation. We also hypothesize this subset of CML-LSCs suppress αβT-cell immune surveillance, permitting LSC persistence. Conversely, we hypothesize proliferative single LSCs are the source of BCRABL mutations causing TKI-resistance. We will investigate this via 3 Aims; Aim 1: Determine the functional consequences of CML-LSC/αβT cell interaction and identify potential therapeutic targets. We will use a murine CML model to investigate potential mechanisms for interaction defined in preliminary data. Murine results will inform studies of T cell/LSC interactions in human CML cells. Aim 2: Identify mechanisms of LSC-persistence vs TKI-resistance in doublet vs singlet populations. Significance of molecular mechanisms that modulate apoptosis or proliferation in single LSCs vs those in doublets will be studied in mice. Results will be compared to human samples and explored in pre-clinical studies. Aim 3: Define influences of the T cell landscape on TFR and TKI-resistance. ScRNA-Seq data for doublet vs singlet CD8+αβTCR+ cells will be analyzed in humans and murine CML. T cell populations in CML with vs without doublets will be and compared to control subjects to identify influences on doublet formation.