Targeting Iron Homeostasis As a Therapeutic Strategy in Multiple Myeloma (2022)

Multiple myeloma (MM) is a hematological cancer characterized by the accumulation of malignant plasma cells (PCs) in the bone marrow (BM). Despite the survival improvement provided by current treatments, the majority of patients relapse and eventually become resistant to all treatments. Drug resistance and relapse are the main current challenges in the treatment of MM and development of new therapeutic interventions is a pressing need (Pinto et al., 2020 Cancers). Abnormal iron homeostasis is implicated in tumorigenesis and the progression of several cancers. Ironomycin is a small synthetic derivative of salinomycin that induces DNA damage and non-apoptotic cell death by sequestering iron in lysosomes (Mai et al., 2017 Nat Chem). Preclinical studies in Acute Myeloid Leukemia (Garciaz et al., 2022 Cancer Discovery) and Diffuse Large B-Cell Lymphoma (Devin et al., 2022 Can Res) have revealed the therapeutic interest of ironomycin in treating hematological malignancies. Here, we have evaluated the effect of ironomycin on MM cells, alone and in combination with standard anti-myeloma treatments. A list of 63 genes related to iron metabolism in cancer was defined. Using Maxstat R function, we showed that 6 genes out of the 63 investigated have a prognostic value in a cohort of newly diagnosed MM patients (n=345). Based on these prognostic genes, we created a gene expression profile (GEP)-based risk score as the sum of the beta coefficients weighted by ±1 according to the patient signal above or below the probe set Maxstat value as previously reported (Herviou et al., 2018 Clinical Epigenetics). This "Iron Score” (IS) was significantly associated with high-risk myeloma (OS and EFS) in 3 independent cohorts of newly diagnosed MM patients (TT2 cohort (N=345), HM cohort (N=206) and TT3 cohort (N=158)), as well as in a cohort of 188 patients at relapse treated with bortezomib monotherapy (Mulligan cohort N=188). GSEA analysis revealed that high-risk MM patients defined by iron score are significantly enriched in genes involved in cell cycle, TFRC targets and hypoxia (p < 0.001). We analyzed the therapeutic interest of ironomycin in 19 human myeloma cell lines (HMCLs). Ironomycin inhibits MM cell growth at nanomolar concentrations compared to other iron chelators. Ironomycin caused a significant cell growth inhibition mediated by cell cycle arrest and triggering of apoptosis, without significant ferroptosis induction. Moreover, cell proliferation arrest correlated with increased DNA damage and caspases activation, and a metabolic swift towards a glycolytic or quiescent metabolism. Iron could also play a role in transcriptional regulation of cancer cells mediated by epigenetic reprogramming. Iron dependent modifications of histone or DNA methylation have been shown to play a significant role in solid cancers. Ironomycin also induced a downregulation of major oncogenes of MM cells including MMSET, c-MYC and CCND1. Furthermore, ironomycin treatment resulted in an epigenetic dysregulation with H3K27ac, H3K4me3 and H3K9me3 downregulation. With major importance, we validated the therapeutic interest of ironomycin in primary MM cells from patients (n=5) without significant toxicity for non-tumor cells from the microenvironment. Furthermore, ironomycin presented a low toxicity on hematopoietic progenitors (CFU assays, n=5) compared to conventional treatment. We tested the therapeutic interest to combine ironomycin with conventional drugs used in MM treatment. The combination of ironomycin with IMiDs (lenalidomide and pomalidomide) had a synergistic effect on cell growth arrest and the activation of apoptotic mechanisms, associated with a marked reduction of several targets of IMiDs including Cereblon (CRBN), MYC, IRF4, IKZF1 and IKZF3. Our data demonstrate that targeting iron homeostasis using ironomycin could be of therapeutic interest in high-risk MM defined by the IS, especially in combination with IMiDs.