T cell acute leukaemia development and chemo-resistance are mediated by dynamic interactions with heterogeneous bone marrow microenvironments

It is widely accepted that complex interactions between cancer cells and their surrounding microenvironment contribute to disease development, chemo-resistance and disease relapse. In light of this observed interdependency, novel therapeutic interventions that target specific cancer stroma cell lineages and their interactions are being sought. To this end, we studied a mouse model of human T cell acute lymphoblastic leukaemia (T-ALL) and used intravital microscopy to monitor the progression of disease within the bone marrow at both the tissue-wide and single cell level over time. We observed highly dynamic cellular interactions and promiscuous distribution of leukaemia cells that migrated across, and proliferated within multiple bone marrow sub-compartments. Unexpectedly, this environment-agnostic behaviour was maintained throughout disease development, starting from the earliest bone marrow seeding to the response and resistance to chemotherapy. Our results reveal that T-ALL cells do not depend on specific bone marrow microenvironments for propagation of disease, nor for the selection of chemo-resistant clones, suggesting a stochastic mechanism underlies these processes. Yet, while T-ALL infiltration and progression are niche-independent, accumulated disease burden leads to rapid, selective remodelling of the endosteal space, resulting in a complete loss of mature osteoblastic cells whilst perivascular cells are maintained. This outcome leads to a shift in the balance of endogenous bone marrow stroma, towards a composition associated with less efficient haematopoietic stem cell function1. This novel, dynamic analysis of T-ALL interactions with the bone marrow microenvironment in vivo highlights that future therapeutic interventions should target cancer cell-intrinsic mechanisms, rather than signals from specific bone marrow stroma, in order to combat the invasion by and survival of chemo-resistant T-ALL cells. Notch-1 T-ALL tumours were generated as previously described in (Hawkins et al., PLoS One, 2014 - doi: 10.1371/journal.pone.0087376.).The T-ALL mouse model was generated by retroviral transduction of C57Bl/6 E14.5 fetal liver cells as described in (Hawkins et al., PLoS One, 2014 - doi: 10.1371/journal.pone.0087376.). Whole transduced populations were transplanted into 8-14 week old female C57Bl/6 recipient mice. T cell acute lymphoblastic leukemia cells were positive for the fluorescent protein (GFP or DsRed) indicating expression of the Notch-1 intracellular protein. These tumours were most frequently CD4+8+ as originally described in (Aster, J. C. et al. Essential roles for ankyrin repeat and transactivation domains in induction of T-cell leukemia by notch1. Molecular and cellular biology 20, 7505-7515 (2000). Disease was confirmed by flow cytometry and 2-photon imaging. Mice reconstituted with Notch transduced foetal livers were monitored daily for signs of leukaemia onset or other signs of ill health. Tumours were harvested from bone marrow once full infiltration was established as confirmed by peripheral blood and bone marrow flow cytometry as well as 2-photon imaging. For treated samples, dexamethasone (Dexamethasone Sodium Phosphate - Sigma) was administered daily I.V. - 15mg/kg for 7 days. Tumours were isolated purified from bone marrow by FACS sorting based on expression of fluorescent protein expression (GFP+ or DsRed+).