Correlation between extent of osteolytic damage and metastatic burden of human breast cancer metastasis in nude mice: real-time PCR quantitation.
Orthotopic or intracardiac injection of human breast cancer cell lines into immunocompromised mice allows study of the molecular basis of breast cancer metastasis. We have established a quantitative real-time PCR approach to analyze metastatic spread of human breast cancer cells inoculated into nude mice via these routes. We employed MDA-MB-231 human breast cancer cells genetically tagged with a bacterial beta-galactosidase (Lac-Z) retroviral vector, enabling their detection by TaqMan real-time PCR. PCR detection was linear, specific, more sensitive than conventional PCR, and could be used to directly quantitative metastatic burden in bone and soft organs. Attesting to the sensitivity and specificity of the PCR detection strategy, as few as several hundred metastatic MDA-MB-231 cells were detectable in 100 microns segments of paraffin-embedded lung tissue, and only in samples adjacent to sections that scored positive by histological detection. Moreover, the measured real-time PCR metastatic burden in the bone environment (mouse hind-limbs, n = 48) displayed a high correlation to the degree of osteolytic damage observed by high resolution X-ray analysis (r2 = 0.972). Such a direct linear relationship to tumor burden and bone damage substantiates the so-called 'vicious cycle' hypothesis in which metastatic tumor cells promote the release of factors from the bone which continue to stimulate the tumor cells. The technique provides a useful tool for molecular and cellular analysis of human breast cancer metastasis to bone and soft organs, can easily be extended to other cell/marker/organ systems, and should also find application in preclinical assessment of anti-metastatic modalities.
VBCRC Breast Cancer Invasion and Metastasis Unit, St. Vincent's Institute of Medical Research, 41 Victoria Pde, Fitzroy, 3065, Australia.
Angus M
AM
Tester
Julie A
JA
Sharp
Nirada
N
Dhanesuan
Mark
M
Waltham
Erik W
EW
Thompson
D000818:Animals; D001857:Bone Matrix / Q000378:metabolism; D001859:Bone Neoplasms / Q000737:chemistry / Q000000981:diagnostic imaging / Q000556:secondary*; D001862:Bone Resorption / Q000209:etiology; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology*; D018270:Carcinoma, Ductal, Breast / Q000737:chemistry / Q000378:metabolism / Q000556:secondary*; D003199:Computer Systems; D016207:Cytokines / Q000378:metabolism; D004273:DNA, Neoplasm / Q000032:analysis; D005260:Female; D017930:Genes, Reporter; D006133:Growth Substances / Q000378:metabolism; D006321:Heart; D006801:Humans; D007267:Injections; D007763:Lac Operon; D008175:Lung Neoplasms / Q000737:chemistry / Q000556:secondary; D008321:Mammary Glands, Animal; D051379:Mice; D008819:Mice, Nude; D009363:Neoplasm Proteins / Q000378:metabolism; D009928:Organ Specificity; D010010:Osteoclasts / Q000502:physiology; D010014:Osteolysis / Q000000981:diagnostic imaging / Q000209:etiology* / Q000473:pathology; D019899:Paracrine Communication; D044162:Parathyroid Hormone-Related Protein; D016133:Polymerase Chain Reaction; D011506:Proteins / Q000378:metabolism; D011859:Radiography; D012680:Sensitivity and Specificity; D016212:Transforming Growth Factor beta / Q000378:metabolism; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000637:transplantation
D016207:Cytokines; D004273:DNA, Neoplasm; D006133:Growth Substances; D009363:Neoplasm Proteins; C470121:PTHLH protein, human; D044162:Parathyroid Hormone-Related Protein; D011506:Proteins; D016212:Transforming Growth Factor beta