A rare subpopulation of dormant, drug resistant stem cells in all MIMICS cells of minimal residual disease

Acute lymphoblastic leukemia (ALL) represents a tumor disease with dismal prognosis upon relapse and novel treatment options are intensively desired especially targeting drug resistant cells. As major challenge, the subpopulation of drug resistant cells is rare and difficult to study. In the individualized mouse model, primary patients' ALL cells grow orthotopically in NSG mice as patient derived xenografts (PDX). Here, we developed the model further in order to be able to study the challenging rare subpopulation of drug resistant cells. ALL PDX cells were lentivirally transduced to generate what we called “genetically engineered PDX (GEPDX)” models in parallel to genetically engineered mouse models (GEMM). This model provides a novel platform for developing targeted therapies in ALL and was used here to express tags in PDX cells for purification of rare cell populations. In a first approach, we used dormancy as characteristic to identify drug resistant cells. GEPDX models were established where PDX cells express several transgenes allowing to enrich minor subpopulations from murine bone marrow. To identify resting PDX cells, PDX cells were labeled with the proliferation marker CFSE. Thereby, we could identify a rare subpopulation of ALL PDX cells which remained dormant over prolonged periods of time in mice; dormant cells revealed severe resistance against systemic drug treatment in mice. In a second approach, we performed prolonged chemotherapy of GEPDX models closely mimicking polychemotherapy of ALL patients. Remaining drug resistant cells at minimal residual disease (MRD) were isolated from murine bone marrow using their transgenes as anchors. Gene expression profiles of single RNA sequencing revealed that dormant and MRD cells shared major characteristics while both were clearly distinct from their cycling or drug sensitive counterparts. Taken together, complex in vivo modelling allowed direct studies on the enriched rare subpopulation of drug resistant patients ALL cells. Our approach will help characterizing these cells for developing future therapies to prevent ALL relapse.