Identification of novel drug targets in neurotransmitter signalling pathways to eradicate CML stem cells

Abstract

In chronic myeloid leukaemia (CML), BCR-ABL tyrosine kinase inhibitors (TKI) fail to eliminate leukaemia stem cells (LSC) which can serve as a reservoir to drive relapse, TKI resistance and promote disease progression. Therefore, identification of pathways/targets that promote LSC survival is essential for the development of curative therapies. Previous transcriptomics analysis from my laboratory pointed to neurotransmitter pathways as being aberrantly expressed in LSC compared to normal haemopoietic stem cells. It was therefore hypothesised that LSCs may require neurotransmitter pathways to maintain their stem cell potential. To identify drug-able targets within these pathways, compound screens were per-formed across BCR-ABL-/+ cell lines and primary CML CD34+ cells using 658 compounds known to modulate neurotransmitter signalling, many of which are used clinically to treat neuro-psychiatric disorders. Hits selective for BCR-ABL-expressing cells were further eval-uated in phenotypic assays using primary CML CD34+ versus non-CML CD34+ cells. 3-CPMT, a selective dopamine reuptake inhibitor (SDRI), and paroxetine, a clinical grade se-lective serotonin reuptake inhibitor (SSRI), that target SLC6A3 and SLC6A4 respectively, emerged as compounds that selectively reduced CML CD34+ cells and inhibited CFC out-puts. It was subsequently shown that LSC and CML cell lines express 3-4-fold higher levels of DAT and SERT than their non-CML counterparts by using FACs analysis, with expres-sion differences for DAT and SERT reaching significance. Targeting of CML cells with 3-CPMT or paroxetine +/- nilotinib inhibits their targets as well as number of known LSC survival factors including PRKCH, ERK1/2, p38 MAPK, c-Myc and Foxo3a. Furthermore, it was demonstrated that paroxetine in combination with nilotinib is highly effective at erad-icating LSC compared to nilotinib alone in a patient-derived xenografts as well as in a trans-genic mouse model of CML. In conclusion, pre-clinical data presented in this thesis suggests that repurposing drugs that target neurotransmitter transporters may represent a novel way to target LSC in CML patients.