Tumor-Endothelial Cell Three-dimensional Spheroids: New Aspects to Enhance Radiation and Drug Therapeutics

Abstract

Classic cancer research for several decades has focused on understanding the biology of tumor cells in vitro. However, extending these findings to in vivo settings has been impeded owing to limited insights on the impact of microenvironment on tumor cells. We hypothesized that tumor cell biology and treatment response would be more informative when done in the presence of stromal components, like endothelial cells, which exist in the tumor microenvironment. To that end, we have developed a system to grow three-dimensional cultures of GFP-4T1 mouse mammary tumor and 2H11 murine endothelial cells in hanging drops of medium in vitro. The presence of 2H11 endothelial cells in these three-dimensional cocultures was found to sensitize 4T1-GFP tumor cells to chemotherapy (Taxol) and, at the same time, protect cells from ionizing radiation. These spheroidal cultures can also be implanted into the dorsal skinfold window chamber of mice for fluorescence imaging of vascularization and disease progression/treatment response. We observed rapid neovascularization of the tumor-endothelial spheroids in comparison to tumor spheroids grown in nude mice. Molecular analysis revealed pronounced up-regulation of several proangiogenic factors in the tumor tissue derived from the tumor-endothelial spheroids compared with tumor-only spheroids. Furthermore, the rate of tumor growth from tumor-endothelial spheroids in mice was faster than the tumor cell-only spheroids, resulting in greater metastasis to the lung. This three-dimensional coculture model presents an improved way to investigate more pertinent aspects of the therapeutic potential for radiation and/or chemotherapy alone and in combination with antiangiogenic agents.

3D tumor tissue analogs and their orthotopic implants for understanding tumor-targeting of microenvironment-responsive nanosized chemotherapy and radiation

“…4T1-mcherry tumor cells, C166-GFP endothelial cells and murine embryonic fibroblasts (MEF) were used to generated single or multicell type 3D cultures in “hanging drops” of media (Dulbecco modified Eagle medium with 10% fetal bovine serum and antibiotic mix) as previously described 3 and in supplementary information. The TTA were sized using a phospho imager and TTA of equal size were used in all experimental studies.…”

“…TTA harvested at day 8 post-treatment and the tumor tissues harvested 24 hr post treatment were homogenized using a 21-gauge syringe in lysis buffer as described earlier 3 . Insoluble material were removed by centrifugation (15 min,12000g).…”

“…There is a critical need for novel tumor models for investigating multimodal treatment strategies such as those utilizing nanoparticles and radiation. Several studies, including our own, have demonstrated that genetic alterations of tumor cells are not the sole driving force behind tumor development and that tumor growth, metastasis and response to treatment are intimately controlled by the tumor microenvironment 1-3 . Tumor models incorporating aspects of the tumor microenvironment may explain and predict why many therapies do not reach the expected level of activity in the patient.…”

The role of photodynamic therapy in overcoming cancer drug resistance

“…Endothelial cells, for example, are emerging as increasingly important drivers of tumor biology and response to treatment, including radiation therapy and chemotherapy 80,81 . EMT status is an additional critical determinant of the response to chemotherapy and targeted inhibitors 11–14 .…”

Chemotaxis-driven assembly of endothelial barrier in a tumor-on-a-chip platform

“…Other efforts in the area include encapsulation of tumor spheroids within biomaterials to mimic the extracellular matrix environment 8–11 . 3D in vitro systems incorporating stromal and vascular cells have also been developed 12–14 . In a recent study, George and colleagues have created vascularized tumor spheroids where the endothelial cells incorporated within the spheroids formed vascular networks in presence of stromal fibroblasts 15 .…”