Development of clinically relevant in vivo metastasis models using human bone discs and breast cancer patient-derived xenografts

Background
Late stage breast cancer preferentially metastasises to bone, despite advances in targeted therapies this condition remains incurable. The lack of clinically relevant models for studying breast cancer metastasis to a human bone microenvironment has stunted the development of effective treatments for this condition. To address this problem we have developed humanised mouse models in which breast cancer PDX’s metastasise to human bone implants with low variability and high frequency.
Methods
To model the human bone environment, bone discs from femoral heads of patients undergoing hip replacement surgery were implanted subcutaneously into NOD/SCID mice. For metastasis studies, 7Patient-Derived Xenograft tumours (PDX: BB3RC32, ER+PR+HER2-; BB2RC08, ER+PR-ER2-; BB6RC37, ER-PR-HER2- and BB6RC39, ER+PR+HER2+), MDA-MB-231-luc2, T47D-luc2 or MCF7-Luc2 cells were injected into mammary ducts 6 and 9 and metastases monitored by luciferase imaging. Bone integrity, viability and vascularisation was assessed by uCT, calcine uptake and histomorphometry. Expression profiling of genes/proteins during different stages of metastasis were assessed by whole genome Affymetrix array, real time PCR and immunohistochemistry.
Results
Implantation of femoral bone provided a metabolically active, human specific, site for tumour cells to metastasise to. After four-weeks, bone implants were re-vascularised and demonstrated active bone remodelling (as evidenced by the presence of osteoclasts, osteoblasts and calcin uptake). Restricting bone implants to the use of subchondral bone and introduction of cancer cells via intraductal injection maximised metastasis to human bone implants. MDA-MB-231, cells specifically metastasised to human bone (70% metastases) whereas T47D, MCF7, BB3RC32, BB2RC08, and BB6RC37 cells metastasised to both human bone and mouse bones. Importantly, human bone was the preferred metastatic site especially from ER+ PDX (100% metastasis human bone compared with 20-75% to mouse bone), whereas ER-ve PDX developed metastases in 20% of human and 20% of mouse bone. Breast cancer cells used in this model underwent a series of molecular changes as they progressed from primary tumours to bone metastasis these included altered expression of molecules previously shown to be biomarkers for metastasis (IL-1B) and/or involved in osteomimickry S100A4, CTSK, SPP1 and RANK.
Conclusions
Our reliable humanised mouse models provide significant advancements towards more clinically relevant modelling of breast cancer bone metastasis.