Exploring Mechanisms of Sensitivity and Resistance to Novel Therapeutic Targets in Aggressive Blood Malignancies.

Abstract

Chemotherapy is commonly used to induce cell death in diffuse large B-cell lymphoma (DLBCL) and acute myeloid leukaemia (AML), but not all patients respond well, and toxicities may be great; there remains a need for effective precision cancer medicines. Dihydroorotate dehydrogenase (DHODH) is a mitochondrial matrix enzyme involved in de novo pyrimidine synthesis, playing a crucial role in DNA and RNA synthesis. CRISPR studies have shown that DHODH is an essential gene in haematologic cells. BAY 2402234 is a highly specific DHODH inhibitor. My studies revealed that BAY 2402234 induced caspase-dependent apoptosis, uridine exogenous addition rescued cells from pyrimidine starvation and showed potential for synergy with BH3 mimetics in some DLBCL and AML cell lines. DHODHi are toxic and ineffective for cancer patients in clinical trials, so acquired resistance to BAY 2402234 was investigated in the OCI-LY19 DLBCL cell line, which is highly sensitive to DHODHi (IC50 of 5pM). In a derived cell line with 200,000-fold resistance, I identified a novel DHODH missense mutation (A58T), which was predicted to prevent the binding of BAY 2402234 but not to interfere with DHODH’s function. RNA-Seq experiments exhibited significant gene expression changes occurring in hallmark pathways with a specific emphasis on MYC OXPHOS and glycolytic targets. Additionally, elevated expression of PI3KCD was observed. Copanlisib a pan-PI3Ki synergised with BAY 2202234 in resistant OCI-LY19 cells, prompting its evaluation as a potential sensitiser. Collectively these findings provide insights for developing effective DHODHi combination therapies that might mitigate toxicities seen with DHODHi in vivo. I also investigated various approaches, such as a BCL-2 inhibitor, or other BH3 mimetics in combination with 14-3-3σ stabilisers FC-A and WR-1065, or MDM2 antagonist and inhibiting the B-cell receptor pathway using Tirabrutinib, CAL-101. DLBCL and AML cell lines showed diverse responses based on subtype and genetics, underlining the need for personalised treatments.