Doxorubicin treatment accelerates bone loss in preclinical breast cancer bone metastasis model.

Five-week old female Balb/C mice received either 5 mg/Kg doxorubicin (i.p. once weekly for three weeks) or PBS and microCT of tibiae were performed. For orthotopic breast cancer bone metastasis model, another set of four-week old female Balb/C mice were injected in the left #4 mammary fat pad using a bone-tropic 4T1 cell line and upon tumor development (palpable size after 1 week) mice were treated with either PBS or doxorubicin (5 mg/Kg doxorubicin, i.p., once weekly for three weeks) and microCT and histology of tibiae were performed. (A) Schematic representation of orthotopic injection of 4T1 cells in mammary fat pad. (B) Histology of mouse tibia showing tumor cells in the bone. (C) Representative microCT images (3D reconstruction) of tibiae collected from non-tumor bearing and tumor-bearing Balb/C mice after three weeks of treatment and (D) Quantification of average BV/TV showing further decrease in trabecular bone volume in tumor-bearing mice upon doxorubicin treatment, compared to non-tumor bearing mice. (E) Average BV/TV were assessed using microCT analysis of tibiae collected from 4 week old athymic nude mice mouse treated with either vehicle or doxorubicin showing loss of trabecular bone volume (P = 0.02). (F) Representative X-ray images showing bone loss upon doxorubicin treatment, compared to control. Quantification of (G) osteolytic lesion area and (H) osteolytic lesion numbers in tumor-bearing Balb/C mice receiving 4T1 cell via cardiac injection and treated with either PBS or doxorubicin (5 mg/Kg) once per week for three weeks. (I) Ratlaps ELISA showing significant increase (P<0.05) in bone resorption upon doxorubicin treatment in non-tumor bearing mice. Statistical analysis was performed using student’s T–test. P<0.05 was considered significant. At least 5 mice were used in each group for these experiments.