Single-cell Transcriptomic Analysis Identifies Senescent Osteocytes that Trigger Bone Destruction in Breast Cancer Metastasis.

Breast cancer bone metastases increase fracture risk and are a major cause of morbidity and mortality among women. Upon colonization by tumor cells, the bone microenvironment undergoes profound reprogramming to support cancer progression, which disrupts the balance between osteoclasts and osteoblasts and leads to bone lesions. A deeper understanding of the processes mediating this reprogramming could help develop interventions for treating patients with bone metastases. Here, we demonstrated that osteocytes in established breast cancer bone metastasis develop premature senescence and a distinctive senescence-associated secretory phenotype (SASP) that favors bone destruction. Single-cell RNA sequencing identified osteocytes from mice with breast cancer bone metastasis enriched in senescence, SASP markers, and pro-osteoclastogenic genes. Multiplex in situ hybridization and AI-assisted analysis depicted osteocytes with senescence-associated satellite distension, telomere dysfunction, and p16Ink4a expression in mice and patients with breast cancer bone metastasis. Breast cancer cells promoted osteocyte senescence and enhanced their osteoclastogenic potential in in vitro and ex vivo organ cultures. Clearance of senescent cells with senolytics suppressed bone resorption and preserved bone mass in mice with breast cancer bone metastasis. These results demonstrate that osteocytes undergo pathological reprogramming by breast cancer cells and identify osteocyte senescence as an initiating event triggering lytic bone disease in breast cancer metastases.

Single-cell Transcriptome Analysis Identifies Senescent Osteocytes as Contributors to Bone Destruction in Breast Cancer Metastasis.

Breast cancer bone metastases increase fracture risk and are a major cause of morbidity and mortality among women. Upon colonization by tumor cells, the bone microenvironment undergoes profound reprogramming to support cancer progression that disrupts the balance between osteoclasts and osteoblasts, leading to bone lesions. Whether such reprogramming affects matrix-embedded osteocytes remains poorly understood. Here, we demonstrate that osteocytes in breast cancer bone metastasis develop premature senescence and a distinctive senescence-associated secretory phenotype (SASP) that favors bone destruction. Single-cell RNA sequencing identified osteocytes from mice with breast cancer bone metastasis enriched in senescence and SASP markers and pro-osteoclastogenic genes. Using multiplex in situ hybridization and AI-assisted analysis, we detected osteocytes with senescence-associated distension of satellites, telomere dysfunction, and p16Ink4a expression in mice and patients with breast cancer bone metastasis. In vitro and ex vivo organ cultures showed that breast cancer cells promote osteocyte senescence and enhance their osteoclastogenic potential. Clearance of senescent cells with senolytics suppressed bone resorption and preserved bone mass in mice with breast cancer bone metastasis. These results demonstrate that osteocytes undergo pathological reprogramming by breast cancer cells and identify osteocyte senescence as an initiating event triggering bone destruction in breast cancer metastases.

The NAD salvage pathway in mesenchymal cells is indispensable for skeletal development in mice.

NAD is an essential co-factor for cellular energy metabolism and multiple other processes. Systemic NAD+ deficiency has been implicated in skeletal deformities during development in both humans and mice. NAD levels are maintained by multiple synthetic pathways but which ones are important in bone forming cells is unknown. Here, we generate mice with deletion of Nicotinamide Phosphoribosyltransferase (Nampt), a critical enzyme in the NAD salvage pathway, in all mesenchymal lineage cells of the limbs. At birth, NamptΔPrx1 exhibit dramatic limb shortening due to death of growth plate chondrocytes. Administration of the NAD precursor nicotinamide riboside during pregnancy prevents the majority of in utero defects. Depletion of NAD post-birth also promotes chondrocyte death, preventing further endochondral ossification and joint development. In contrast, osteoblast formation still occurs in knockout mice, in line with distinctly different microenvironments and reliance on redox reactions between chondrocytes and osteoblasts. These findings define a critical role for cell-autonomous NAD homeostasis during endochondral bone formation.