Radium 223 induces transient functional bone marrow toxicity

Radium 223 (Rad223) is a bone-seeking, alpha-particle-emitting radionuclide approved for the treatment of patients with metastatic castration resistant prostate cancer and is currently being tested in a variety of clinical trials for primary and metastatic cancers to bone. Clinical evaluation of Rad223 hematologic safety showed a significantly increased rate of neutropenia and thrombocytopenia in patients, hinting at myelosuppression as side effect. In this study we present a preclinical approach to investigate the consequences of Rad223 treatment on bone marrow biology. Rad223 accumulated in bones and induced confined radiation damage, followed by progressive replacement of the impaired areas with adipocyte infiltration, as monitored by three-dimensional multiphoton microscopy, ex vivo. Flow cytometry and single cell transcriptomic analyses on bone marrow hematopoietic populations revealed transient, non-specific Rad223-mediated cytotoxicity on resident populations, including stem, progenitor and mature leukocytes. This was paralleled by a significant decrease of white blood cells and platelets in peripheral blood, which was overcome within 40 days post-treatment. Rad223 exposure did not impair full hematopoietic reconstitution, suggesting that the bone marrow function is not permanently hampered. Our results provide a comprehensive explanation of Rad223 reversible effects on bone marrow cells and exclude long-term myelotoxicity, supporting its safety during treatment of patients at both early and late disease stages. Overall design: Bone marrow was collected at d0, d4, d11, d40 from Rad223 treatment. After sorting for CD117+ cells from 3 mice/timepoint, each timepoint was tagged with oligo-tagged antibodies to uniquely label cells from distinct samples (https://cite-seq.com/cell-hashing/). Samples were pooled and single-cell RNA-seq libraries were prepared using Single Cell 3' Reagent Kits v.2: Chromium Single Cell 3' Library & Gel Bead Kit v.2, PN-120237; Single Cell 3' Chip Kit v.2 PN-120236; and i7 Multiplex Kit PN-120262 (10x Genomics) following the Single Cell 3' Reagent Kits v.2 User Guide (Manual Part no. CG00052 Rev A). Libraries were run on an Illumina NovaSeq 6000 system as 150-bp paired-end reads, one full lane per sample.