Breast Cancer Lecture 3.doc

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Western University
Anatomy and Cell Biology
Anatomy and Cell Biology 4461B
Geordie Shepherd

LP Lecture 6: Breast Cancer Translational Models of Breast Cancer Criteria for a Cancer Cell Line • • Altered cytomorphology Increased growth • • Reduced serum dependency • Increased clonogenicity • Anchorage independent growth • Alterations in ploidy • Immortality • Tumorigenic in nude mice • Majority of breast cancer research uses established cell lines • MCF-7 • Most widely used line • From a pleural effusion • Most derived from metastases • Ascites • Pleural effusions • Biased towards the most advanced types of cancers • Primary Cell Culture • Derived directly from a tumor • Explants • Mixed cell populations grow out from a chunk of tissue • Often overgrown with fibroblasts • Culture of individual cells • Spillage technique • Cut tumor and collect cells that spill out • Enzymatic dispersal and degradation of stroma • Sedimentation • Enzymatic dispersal followed by centrifugation • In Vitro Assays Designed to specifically examine the hallmarks of cancer • • Few are specific to breast cancer Can modify the microenvironment • • Oxygen • ECM components • Measure proliferation and apoptosis • In response to drugs or genetic modifications • Metric in high throughput screens • Measure tumorigenicity • Anchorage independent growth • Measure alterations in protein and gene expression • Correlative studies • Can assess processes key to tumor formation and metastasis • ex. self renewal (tumorsphere assays) • ex. cellular migration • 3D Organotypic Assays • When grown in 3D, in Matrigel (basement membrane like preparation) breast epithelial cells will form hollowed out acinar-like structures • Very aggressive cancer cells form invasive tumor like structures • Can assess how well differentiated a tumor population is • Can assess which drugs/genetic modifications can alter morphology and aggressive- ness of cancer cell line • Animal Models • Better representation of tumor than cells • Has stromal cells • Is 3D • Next step after in vitro observations • Sometimes changes will be seen in the animal model without corresponding to in vitro effects • May be due to combinatorial alterations, perhaps in the stroma • Xenografts • Uses SCID (T-cell and B-cell deficient) • NUDE (T-cell deficient) • Orthotopic sites to better mimic tumor environment; insertion into mammary fat pad • Tumor growth can be measured by • Calipers • Small animal imaging • Luminescence • MRI • Fluorescence • ER+ cells require estrogen supplementation • Can use primary cells, usually in a matrix; better recapitulates tumor of origin • Can do serial dilutions to test cancer stem cells/cancer initiating fractions • Can implant tumors made in vitro with human cancer cells, stromal cells and endothe- lial cells • Metastasis is rare but can occur • Variants that metastasize to lymph nodes • Experimental metastasis assays • Cancer cells injected into the blood stream • Intr
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