Development of a human organotypic tumor invasion model of cancer cell line MDA-MB-231 in Precision-Cut Lung Slices (PCLS)

Abstract

Cancer such as breast carcinoma is a major public health problem worldwide. The actual cause of death is the formation of metastasis which often occurs in the lung. Among the cell types present in the tumor microenvironment, macrophages have been proven to be the dominant leukocyte population with high macrophage density correlating to poor patient prognosis. Thus, these cells are of high interest as targets for cancer therapeutics, making them a valuable research topic. We utilize an innovative organotypic tumor invasion model, using living human Precision-Cut Lung Slices (PCLS) and cancer cells to focus on the local immunological response during early metastasis formation. An AdGFP-transduced human breast cancer cell line MDA-MB-231 was added to human PCLS over a period of one week. Viability assays such as LIVE/DEAD® staining and LDH measurements were performed to assure intact human tissue throughout the experimental procedure. Tissue immune staining methods against CD68 and Ki67 were used to visualize locations of macrophages and proliferating cells, respectively. Immune response and neoangiogenesis were determined by cytokines IL-10 and IL-1beta and tumor markers VEGF and GM-CSF. An in vitro neoangiogenesis assay was performed to observe whether the tissue is able to attract endothelial cells through a BME-coated membrane. Surrounding parenchyma remains viable during infection and invasion with cancer cells. Colocalization of CD68-positive macrophages and MDA-MB-231 was found during the entire invasion period, visualizing expansion and proliferation of the cancer cells within the organotypic model. The VEGF/GM-CSF release correlated with the MDA-MB-231 growth curves. HUVEC cell invasion shows a 5-fold increase in the PCLS that had been treated with MDA-MB-231 in relation to untreated controls. Here we mimic cancer cell proliferation and immune responses in the native microenvironment of human lung tissue which will be used for testing of anti-tumor drugs in the near future