Inhibition of the Growth of Breast Cancer-Associated Brain Tumors by the Osteocyte-Derived Conditioned Medium

Abstract

The brain is a common site of metastasis from advanced breast cancer but few effective treatments are available. We examined a therapeutic option with a conditioned medium (CM), focusing on the role of Lrp5 and β-catenin in Wnt signaling, and IL1ra in osteocytes. Osteocytes presented the innate anti-tumor effect and the overexpression of the above genes strengthened their action. In a mouse model, the injection of their CM inhibited mammary tumors and tumor-driven osteolysis. Importantly, Lrp5- and/or IL1ra-overexpressing osteocytes or the local administration of β-catenin-overexpressing CM markedly inhibited brain tumors. In the transport analysis, tumor-suppressing factors in CM were shown to diffuse through the skull. Mechanistically, the CM with overexpression of the above genes downregulated oncogenic genes such as MMP9, Runx2, TGFβ, and Snail in breast cancer cells. Also, the CM with β-catenin overexpression downregulated CXCL1 and CXCL5 and upregulated tumor suppressors such as LIMA1, DSP, p53, and TRAIL in breast cancer cells. Notably, whole-genome proteomics revealed that histone H4 was enriched in CM and acted as an atypical tumor suppressor. Lrp5-overexpressing MSCs were also shown to act as anti-tumor agents. Collectively, this study demonstrated the therapeutic role of engineered CM in brain tumors and the tumor-suppressing action of extracellular histone H4. The result sheds light on the potential CM-based therapy for breast cancer-associated brain metastases in a minimally invasive manner.

Generation of the tumor-suppressive secretome from tumor cells

“…One such multi-tasking protein is histone H4 that packs DNA in the nucleus and stimulates innate immune responses outside of the cell 49 . We previously reported that extracellular histone H4 served as a tumor suppressor 50 . Another moonlighting example is a high mobility group box 1 (HMGB1) protein.…”

“…In the generation of iTS cells from osteocytes, we have previously shown that the overexpression of β-catenin and Lrp5 but not Lrp6 promoted the anti-tumor capabilities and the protection of tumor-invased bone 1 . In osteocytes and MSCs, the overexpression of multiple pro-tumorigenic genes and the administration of tumor-promoting compounds have been shown to generate iTSC that significantly inhibited the progression of adjacent tumors 1 - 3 . Further analyses are needed to elucidate the requirement for creating effective iTS CM from varing tumor and non-tumor cells, considering types of target cancer cells, as well as pathways and genes to be activated and overexpressed.…”

“…We named those osteocytes and MSCs induced tumor-suppressing cells (iTSCs). Cell culture and preclinical studies have revealed that iTSCs generate anti-tumor secretomes and their complete and partial conditioned medium (CM) inhibits the progression of tumor cells in vitro , ex vivo and in vivo 1 - 3 . The question herein was whether the anti-tumor secretomes can be generated not only from non-tumor cells but also from tumor cells.…”

Conversion of Osteoclasts into Bone-Protective, Tumor-Suppressing Cells

“…While these agents significantly improve the quality of life in patients with bone metastasis and the risk of bone fracture, their efficacy is in many cases insufficient to prevent tumor-induced bone loss and eliminate cancer cells from the bone. To examine the possibility of developing a novel treatment option, we herein examined a recent technology of induced tumor-suppressing cells (iTSCs) that was successfully applied to osteocytes, mesenchymal stem cells (MSCs), and tumor cells [ 10 , 11 , 12 , 13 ]. In the iTSC technology, a dichotomous role of oncogenic signaling in tumor promotion as well as in tumor suppression is utilized.…”

“…For instance, it is well known that the activation of Wnt signaling in tumor cells by the overexpression of β-catenin or Lrp5, or the application of a pharmacological Wnt activator, promotes tumor progression [ 13 ]. However, our previous studies have shown that Wnt-activated osteocytes, MSCs, and tumor cells produce tumor-suppressing secretomes, and their conditioned medium (CM) serves as a potent tumor-suppressing agent [ 10 , 11 , 13 ]. The question herein is whether it is possible to convert osteoclasts into iTSCs and generate bone-protective, tumor-suppressing secretomes by activating oncogenic signaling.…”