Bone Research, Published online: 5 April 2016; | doi:10.1038/boneres.2016.4

The therapeutic alpha-emitter ²²³Ra can be stably complexed in vivo, creating opportunities for the development of targeted radiopharmaceutical agents with this radionuclide.

Bone-metastatic, castration-resistant prostate cancer (bmCRPC) represents a lethal stage of the most common noncutaneous cancer in men. The recent introduction of Radium-223 dichloride, a bone-seeking alpha particle (α)–emitting radiopharmaceutical, demonstrates statistically significant survival benefit and palliative effect for bmCRPC patients. Clinical results have established safety and efficacy, yet questions remain regarding pharmacodynamics and dosing for optimized patient benefit. We elucidated the biodistribution of ²²³ Ra as well as interaction with the bone and tumor compartments in skeletally mature mice (C57Bl/6 and CD-1, n = 3–6) and metastasis models (LNCaP and PC3, n = 4). Differences in uptake were evaluated by µCT and histological investigation. Novel techniques were leveraged on whole-mount undecalcified cryosections to determine microdistribution of Radium-223. All statistical tests were two-sided. ²²³ Ra uptake in the bones (>30% injected activity per gram) at 24 hours was also accompanied by non-negligible remnant activity in the kidney (2.33% ± 0.36%), intestines (5.73% ± 2.04%), and spleen (10.5% ± 5.9%) Skeletal accumulation across strains did not correspond with bone volume or surface area but instead to local blood vessel density ( P = .04). Microdistribution analysis by autoradiography and α camera revealed targeting of the ossifying surfaces adjacent to the epiphyseal growth plate. In models of PCa metastasis, radioactivity does not localize directly within tumors but instead at the apposite bone surface. Osteoblastic and lytic lesions display similar intensity, which is comparable with uptake at sites of normal bone remodeling. Profiling the macro- and microdistribution of ²²³ Ra in healthy and diseased models has important implications to guide precision application of this emerging α-therapy approach for bmCRPC and other bone metastastic diseases.

Imaging Mass Cytometry (IMC) is an emerging multiplexed imaging technology for analyzing complex microenvironments using more than 40 molecularly-specific channels. However, this modality has unique data processing requirements, particularly for patient tissue specimens where signal-to-noise ratios for markers can be low, despite optimization, and pixel intensity artifacts can deteriorate image quality and downstream analysis. Here we demonstrate an automated content-aware pipeline, IMC-Denoise, to restore IMC images deploying a differential intensity map-based restoration (DIMR) algorithm for removing hot pixels and a self-supervised deep learning algorithm for shot noise image filtering (DeepSNiF). IMC-Denoise outperforms existing methods for adaptive hot pixel and background noise removal, with significant image quality improvement in modeled data and datasets from multiple pathologies. This includes in technically challenging human bone marrow; we achieve noise level reduction of 87% for a 5.6-fold higher contrast-to-noise ratio, and more accurate background noise removal with approximately 2 × improved F1 score. Our approach enhances manual gating and automated phenotyping with cell-scale downstream analyses. Verified by manual annotations, spatial and density analysis for targeted cell groups reveal subtle but significant differences of cell populations in diseased bone marrow. We anticipate that IMC-Denoise will provide similar benefits across mass cytometric applications to more deeply characterize complex tissue microenvironments.

Tumor progression to metastasis is not cancer cell autonomous, but rather involves the interplay of multiple cell types within the tumor microenvironment. Here we identify asporin (ASPN) as a novel, secreted mesenchymal stromal cell (MSC) factor in the tumor microenvironment that regulates metastatic development. MSCs expressed high levels of ASPN, which decreased following lineage differentiation. ASPN loss impaired MSC self-renewal and promoted terminal cell differentiation. Mechanistically, secreted ASPN bound to BMP-4 and restricted BMP-4–induced MSC differentiation prior to lineage commitment. ASPN expression was distinctly conserved between MSC and cancer-associated fibroblasts (CAF). ASPN expression in the tumor microenvironment broadly impacted multiple cell types. Prostate tumor allografts in ASPN-null mice had a reduced number of tumor-associated MSCs, fewer cancer stem cells, decreased tumor vasculature, and an increased percentage of infiltrating CD8⁺ T cells. ASPN-null mice also demonstrated a significant reduction in lung metastases compared with wild-type mice. These data establish a role for ASPN as a critical MSC factor that extensively affects the tumor microenvironment and induces metastatic progression. Significance: These findings show that asporin regulates key properties of mesenchymal stromal cells, including self-renewal and multipotency, and asporin expression by reactive stromal cells alters the tumor microenvironment and promotes metastatic progression.