Epigenetik yeniden programlama ile MLL-AF9 lösemilerini hedefleyen yeni tedavi yöntemleri

Leukemia is a highly complex disorder of blood and bone marrow and characterized by inhibition of differentiation during hematopoiesis which leads to abnormal cell proliferation. Mixed lineage leukemia (MLL) is a form of acute leukemia that represents poor prognosis and due to chromosomal translocation, resulting in a hyperactive MLL fusion protein. MLL leukemias are largely based on epigenetic irregulations rather than genomic instability. A transcription factor (AF9) that fuses with MLL plays a role in the uncontrolled growth of acute monocytic leukemia. Chromatin modifying enzymes are aberrantly expressed in leukemias and targeting these regulators such as the Lysine-specific demethylase (LSD1) and Histone deacetylase (HDAC) has been considered as a novel treatment modality. In MLL-AF9 leukemias, recruitment of these enzymes due to translocation results in the activation of several genes that inhibit differentiation and lead uncontrolled cell proliferation. In this study, novel compounds that synthesized to inhibit LSD1, HDAC6, and both LSD1& HDAC6 (Dual Inhibitor) are characterized. We showed that the compounds inhibit target enzymes by in vitro enzyme activity tests, we also showed that the compounds engage with the target enzymes in the cell by cellular thermal shift assay (CETSA). The toxic effects of the compounds on leukemia cells were shown with ATP-dependent cell viability tests. Increased histone methylation and acetylation levels after inhibition of target enzymes were indicated with Western blot. Increased expression of downstream genes due to enzyme inhibition was showed with RT-qPCR. The synergistic effect of the inhibitors with the therapeutic drugs used in the clinic was investigated. We showed that the death response could be increased with a combination of Doxorubicin and our epigenetic inhibitors. Finally, RNA-seq analysis revealed the molecular mechanism of Doxorubicin synergy. Defects in epigenetic pathways involving increased expression levels or abnormal patterns of activity are one of the key drivers of cell proliferation in cancer. Targeting MLL-AF9 is an attractive strategy for therapeutic intervention in patients with this genetic translocation. Since our inhibitors are more effective on MLL-AF9 leukemias, we hope that our study will result in the characterization of targeted drugs for use in leukemia therapy.