Multicellular tumor spheroids are powerful in vitro models to perform preclinical chemosensitivity assays. We compare different methodologies to generate tumor spheroids in terms of resultant spheroid morphology, cellular arrangement and chemosensitivity. We used two cancer cell lines (MCF7 and OVCAR8) to generate spheroids using i) hanging drop array plates; ii) liquid overlay on ultra-low attachment plates; iii) liquid overlay on ultra-low attachment plates with rotating mixing (nutator plates). Analysis of spheroid morphometry indicated that cellular compaction was increased in spheroids generated on nutator and hanging drop array plates. Collagen staining also indicated higher compaction and remodeling in tumor spheroids on nutator and hanging drop arrays compared to conventional liquid overlay. Consequently, spheroids generated on nutator or hanging drop plates had increased chemoresistance to cisplatin treatment (20-60% viability) compared to spheroids on ultra low attachment plates (10-20% viability). Lastly, we used a mathematical model to demonstrate minimal changes in oxygen and cisplatin diffusion within experimentally generated spheroids. Our results demonstrate that in vitro methods of tumor spheroid generation result in varied cellular arrangement and chemosensitivity.

Breast cancer cells experience a range of shear stresses in the tumor microenvironment (TME). However most current in vitro three‐dimensional (3D) models fail to systematically probe the effects of this biophysical stimuli on cancer cell metastasis, proliferation, and chemoresistance. To investigate the roles of shear stress within the mammary and lung pleural effusion TME, a bioreactor capable of applying shear stress to cells within a 3D extracellular matrix was designed and characterized. Breast cancer cells were encapsulated within an interpenetrating network hydrogel and subjected to shear stress of 5.4 dynes cm−2 for 72 hr. Finite element modeling assessed shear stress profiles within the bioreactor. Cells exposed to shear stress had significantly higher cellular area and significantly lower circularity, indicating a motile phenotype. Stimulated cells were more proliferative than static controls and showed higher rates of chemoresistance to the anti‐neoplastic drug paclitaxel. Fluid shear stress‐induced significant upregulation of the PLAU gene and elevated urokinase activity was confirmed through zymography and activity assay. Overall, these results indicate that pulsatile shear stress promotes breast cancer cell proliferation, invasive potential, chemoresistance, and PLAU signaling.

Ovarian cancer is an extremely lethal gynecologic disease; with the high-grade serous subtype predominantly associated with poor survival rates. Lack of early diagnostic biomarkers and prevalence of post-treatment recurrence, present substantial challenges in treating ovarian cancers. These cancers are also characterized by a high degree of heterogeneity and protracted metastasis, further complicating treatment. Within the ovarian tumor microenvironment, cancer stem-like cells and mechanical stimuli are two underappreciated key elements that play a crucial role in facilitating these outcomes. In this review article, we highlight their roles in modulating ovarian cancer metastasis. Specifically, we outline the clinical relevance of cancer stem-like cells, and challenges associated with their identification and characterization and summarize the ways in which they modulate ovarian cancer metastasis. Further, we review the mechanical cues in the ovarian tumor microenvironment, including, tension, shear, compression and matrix stiffness, that influence (cancer stem-like cells and) metastasis in ovarian cancers. Lastly, we outline the challenges associated with probing these important modulators of ovarian cancer metastasis and provide suggestions for incorporating these cues in basic biology and translational research focused on metastasis. We conclude that future studies on ovarian cancer metastasis will benefit from the careful consideration of mechanical stimuli and cancer stem cells, ultimately allowing for the development of more effective therapies.

Perspective: The role of mechanobiology in the etiology of brain metastasis APL Bioengineering 2, 031801 (2018) Ovarian cancer remains a deadly diagnosis with an 85% recurrence rate and a 5-year survival rate of only 46%. The poor outlook of this disease has improved little over the past 50 years owing to the lack of early detection, chemoresistance and the complex tumor microenvironment. Within the peritoneal cavity, the presence of ascites stimulates ovarian tumors with shear stresses. The stiff environment found within the tumor extracellular matrix and the peritoneal membrane are also implicated in the metastatic potential and epithelial to mesenchymal transition (EMT) of ovarian cancer. Though these mechanical cues remain highly relevant to the understanding and treatment of ovarian cancers, our current knowledge of their biological processes and their clinical relevance is deeply lacking. Seminal studies on ovarian cancer mechanotransduction have demonstrated close ties between mechanotransduction and ovarian cancer chemoresistance, EMT, enhanced cancer stem cell populations, and metastasis. This review summarizes our current understanding of ovarian cancer mechanotransduction and the gaps in knowledge that exist. Future investigations on ovarian cancer mechanotransduction will greatly improve clinical outcomes via systematic studies that determine shear stress magnitude and its influence on ovarian cancer progression, metastasis, and treatment.

This report investigates the role of compressive stress on ovarian cancer in a 3D custom built bioreactor. Cells within the ovarian tumor microenvironment experience a range of compressive stimuli that contribute to mechanotransduction. As the ovarian tumor expands, cells are exposed to chronic load from hydrostatic pressure, displacement of surrounding cells, and growth induced stress. External dynamic stimuli have been correlated with an increase in metastasis, cancer stem cell marker expression, chemoresistance, and proliferation in a variety of cancers. However, how these compressive stimuli contribute to ovarian cancer progression is not fully understood. In this report, high grade serous ovarian cancer cell lines were encapsulated within an ECM mimicking hydrogel comprising of agarose and collagen type I, and stimulated with confined cyclic or static compressive stresses for 24 and 72 h. Compression stimulation resulted in a significant increase in proliferation, invasive morphology, and chemoresistance. Additionally, CDC42 was upregulated in compression stimulated conditions, and was necessary to drive increased proliferation and chemoresistance. Inhibition of CDC42 lead to significant decrease in proliferation, survival, and increased chemosensitivity. In summary, the dynamic in vitro 3D platform developed in this report, is ideal for understanding the influence of compressive stimuli, and can be widely applicable to any epithelial cancers. This work reinforces the critical need to consider compressive stimulation in basic cancer biology and therapeutic developments.