Induction of hypersignaling as a therapeutic approach for treatment of BCR-ABL1 positive Acute Lymphoblastic Leukemia (ALL) cells

Current therapy approaches for tyrosine kinase driven leukemia including Ph+ ALL and CML are almost entirely focused on the development of more potent tyrosine kinase inhibitors (TKI). The ultimate goal in this approach is to reduce the oncogenic signaling below a minimum threshold that is required for the survival of leukemia cells. Despite the successful results obtained so far, this approach has its own drawbacks. Unfortunately, TKI treatment is not very effective in Ph+ ALL patients. Furthermore, in CML patients it can lead to the appearance of resistant tumors that are not sensitive to therapy anymore. Therefore, new strategies for killing the Ph+ leukemia independent of kinase inhibition are necessary.;In chapter two of this thesis, we focused on the mitogen-activated protein kinase (MAPK) pathway. By the analysis of gene expression changes and evaluation of protein levels, we found that DUSP6 and SPRY2, two negative feedback regulators of the MAPK pathway, are highly expressed in human B cell lineage Ph + ALL, while they are not expressed in B cell progenitor cells. Interestingly, in contrast to BCR-ABL1 kinase driven leukemia, non kinase-driven B cell Non-Hodgkin-Lymphoma (B-NHL) lack expression of these genes. To study the function of DUSP6 and SPRY2 in a genetic mouse model of Ph+ like leukemia, we transduced bone marrow pre-B cells from DUSP6-/-, SPRY2fl/fl mice and respective wildtype controls with retroviral BCR-ABL1. Defects in either of these two negative feedback mediators caused profound signaling imbalances in BCR-ABL1 leukemia cells and resulted in drastic negative consequences for the cells.;To test whether negative feedback signaling molecules represent a potential target for pharmacological inhibition in the treatment of kinase driven leukemia, we tested the DUSP6 small molecule inhibitor 2-benzylidene-3-(cyclohexylamino)-1-Indanone hydrochloride (BCI). BCI induced massive accumulation of ROS and subsequent cell death in a panel of patient-derived cases of Ph+ ALL, including two cases with T315I mutation. In vivo and ex vivo studies showed that BCI can increase the survival of NOD/SCID recipient mice transplanted with patient-derived Ph+ ALL cells with T315I mutation.;In chapter three, we turn to the inhibitory phosphatases in Ph + ALL cells. We found that three key inhibitory phosphatases (INPP5D/SHIP1, PTEN and PTPN6/SHP1) are expressed at high levels in these cells. INPP5D and PTEN negatively regulate AKT mediated signaling by dephosphorylating phosphatidylinositol-3,4,5-trisphosphate, at 5- and 3-phosphates, respectively. In addition, PTPN6 and INPP5D can both be recruited to the ITIM motifs of various inhibitory surface receptors and negatively regulate activation signals from tyrosine kinases and activating receptors.;Studying B cell lineage BCR-ABL1 transformed Inpp5dfl/fl, Ptenfl/fl and Ptpn6fl/fl leukemia cells in vivo, we observed that deletion of these inhibitory phosphatases resulted in prolonged survival of leukemia transplant recipient mice. In addition, using small molecule inhibitors of INPP5D (3AC) and PTEN (VO-OHpic) recapitulated the genetic findings and killed patient-derived leukemia cells carrying the T315I mutant BCR-ABL1.;In summary, our studies identify negative feedback regulators of the MAPK pathway (including DUSP6 and SPRY2) and inhibitory phosphatases such as PTEN, INPP5D and PTPN6 as novel therapeutic targets in BCR-ABL1 positive B cell lineage leukemia. Pharmacological blockade of these molecules represents a fundamentally novel and powerful approach to increase oncogenic signaling beyond a tolerable threshold, thus causing excessive accumulation of ROS, crosstalk between different signaling pathways and subsequent cell death. (Abstract shortened by UMI.).