ABSTRACT
Renal cell carcinoma bone metastases (RCCBM) are typically osteolytic. We previously showed that BIGH3 (beta Ig-h3/TGFBI), secreted by 786-O renal cell carcinoma, plays a role in osteolytic bone lesion in RCCBM through inhibition of osteoblast (OSB) differentiation. To study this interaction, we employed three-dimensional (3D) hydrogels to coculture bone-derived 786-O (Bo-786) renal cell carcinoma cells with MC3T3-E1 pre-OSBs. Culturing pre-OSBs in the 3D hydrogels preserved their ability to differentiate into mature OSB; however, this process was decreased when pre-OSBs were cocultured with Bo-786 cells. Knockdown of BIGH3 in Bo-786 cells recovered OSB differentiation. Furthermore, treatment with bone morphogenetic protein 4, which stimulates OSB differentiation, or cabozantinib (CBZ), which inhibits VEGFR1 and MET tyrosine kinase activities, also increased OSB differentiation in the coculture. CBZ also inhibited pre-osteoclast RAW264.7 cell differentiation. Using RCCBM mouse models, we showed that CBZ inhibited Bo-786 tumor growth in bone. CBZ treatment also increased bone volume and OSB number, and decreased osteoclast number and blood vessel density. When tested in SN12PM6 renal cell carcinoma cells that have been transduced to overexpress BIGH3, CBZ also inhibited SN12PM6 tumor growth in bone. These observations suggest that enhancing OSB differentiation could be one of the therapeutic strategies for treating RCCBM that exhibit OSB inhibition characteristics, and that this 3D coculture system is an effective tool for screening osteoanabolic agents for further in vivo studies.
Subject(s)
Anilides/pharmacology , Bone Neoplasms/drug therapy , Carcinoma, Renal Cell/drug therapy , Cell Differentiation , Kidney Neoplasms/drug therapy , Osteoblasts/cytology , Osteolysis/drug therapy , Pyridines/pharmacology , Animals , Apoptosis , Bone Neoplasms/metabolism , Bone Neoplasms/secondary , Carcinoma, Renal Cell/metabolism , Carcinoma, Renal Cell/pathology , Cell Proliferation , Coculture Techniques , Humans , In Vitro Techniques , Kidney Neoplasms/metabolism , Kidney Neoplasms/pathology , Male , Mice , Mice, SCID , Osteoblasts/drug effects , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
Bone metastases from renal cell carcinoma (RCC) are typically lytic, destructive, and resistant to treatment regimens. Current in vitro models for studying metastasis introduce artifacts that limit their usefulness. Many features of tumors growing in bone are lost when human RCC cells are cultured in two-dimensional (2D) plastic substrata. In this study, we established that RCC spheroids, consisting of aggregates of cells, can be grown in a three-dimensional (3D) hyaluronate hydrogel-based culture system. The bone-derived human 786-O RCC subline proliferated and survived long term in these hydrogels. Additionally, RCC spheroids in 3D hydrogels demonstrated lower proliferation rates than their counterparts grown in 2D. Overall, gene expression patterns of RCC spheroids in 3D more closely mimicked those observed in vivo than did those of cells grown in 2D. Of particular importance, selected adhesion molecules, angiogenesis factors, and osteolytic factors that have been shown to be involved in RCC bone metastasis were found to be expressed at higher levels in 3D than in 2D cultures. We propose that the 3D culture system provides an improved platform for RCC bone metastasis studies compared with 2D systems.
Subject(s)
Bone Neoplasms/secondary , Carcinoma, Renal Cell/secondary , Cell Culture Techniques , Kidney Neoplasms/pathology , Angiogenic Proteins/genetics , Angiogenic Proteins/metabolism , Animals , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Bone Neoplasms/genetics , Bone Neoplasms/metabolism , Carcinoma, Renal Cell/genetics , Carcinoma, Renal Cell/metabolism , Cell Adhesion Molecules/genetics , Cell Adhesion Molecules/metabolism , Cell Proliferation , Cell Separation , Cell Shape , Cell Survival , Gene Expression Regulation, Neoplastic , Humans , Hyaluronic Acid/metabolism , Hydrogels , Kidney Neoplasms/genetics , Kidney Neoplasms/metabolism , Mice, SCID , Osteolysis/genetics , Osteolysis/metabolism , Osteolysis/pathology , Phenotype , RNA, Messenger/metabolism , Spheroids, Cellular , Time Factors , Tumor Cells, CulturedABSTRACT
The lack of effective therapies for bone metastatic prostate cancer (PCa) underscores the need for accurate models of the disease to enable the discovery of new therapeutic targets and to test drug sensitivities of individual tumors. To this end, the patient-derived xenograft (PDX) PCa model using immunocompromised mice was established to model the disease with greater fidelity than is possible with currently employed cell lines grown on tissue culture plastic. However, poorly adherent PDX tumor cells exhibit low viability in standard culture, making it difficult to manipulate these cells for subsequent controlled mechanistic studies. To overcome this challenge, we encapsulated PDX tumor cells within a three-dimensional hyaluronan-based hydrogel and demonstrated that the hydrogel maintains PDX cell viability with continued native androgen receptor expression. Furthermore, a differential sensitivity to docetaxel, a chemotherapeutic drug, was observed as compared to a traditional PCa cell line. These findings underscore the potential impact of this novel 3D PDX PCa model as a diagnostic platform for rapid drug evaluation and ultimately push personalized medicine toward clinical reality.
