Formation of stable small cell number three-dimensional ovarian cancer spheroids using hanging drop arrays for preclinical drug sensitivity assays.
BACKGROUND
Ovarian cancer grows and metastasizes from multicellular spheroidal aggregates within the ascites fluid. Multicellular tumor spheroids are therefore physiologically significant 3D in vitro models for ovarian cancer research. Conventional hanging drop cultures require high starting cell numbers, and are tedious for long-term maintenance. In this study, we generate stable, uniform multicellular spheroids using very small number of ovarian cancer cells in a novel 384 well hanging drop array platform.


METHODS
We used novel tumor spheroid platform and two ovarian cancer cell lines (A2780 and OVCAR3) to demonstrate the stable incorporation of as few as 10 cells into a single spheroid.


RESULTS
Spheroids had uniform geometry, with projected areas (42.60×10(3)μm-475.22×10(3)μm(2) for A2780 spheroids and 37.24×10(3)μm(2)-281.01×10(3)μm(2) for OVCAR3 spheroids) that varied as a function of the initial cell seeding density. Phalloidin and nuclear stains indicated cells formed tightly packed spheroids with demarcated boundaries and cell-cell interaction within spheroids. Cells within spheroids demonstrated over 85% viability. 3D tumor spheroids demonstrated greater resistance (70-80% viability) to cisplatin chemotherapy compared to 2D cultures (30-50% viability).


CONCLUSIONS
Ovarian cancer spheroids can be generated from limited cell numbers in high throughput 384 well plates with high viability. Spheroids demonstrate therapeutic resistance relative to cells in traditional 2D culture. Stable incorporation of low cell numbers is advantageous when translating this research to rare patient-derived cells. This system can be used to understand ovarian cancer spheroid biology, as well as carry out preclinical drug sensitivity assays.
Department of Materials Science Engineering, University of Michigan, Ann Arbor, USA.
Shreya
S
Raghavan
Department of Materials Science Engineering, University of Michigan, Ann Arbor, USA.
Maria R
MR
Ward
Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA.
Katelyn R
KR
Rowley
Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA.
Rachel M
RM
Wold
Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA; Macromolecular Science and Engineering, University of Michigan, Ann Arbor, USA.
Shuichi
S
Takayama
Department of Obstetrics and Gynecology - Gynecologic Oncology, University of Michigan, Ann Arbor, USA; Department of Internal Medicine - Hematology/Oncology, , University of Michigan, Ann Arbor, USA.
Ronald J
RJ
Buckanovich
Department of Materials Science Engineering, University of Michigan, Ann Arbor, USA; Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA; Macromolecular Science and Engineering, University of Michigan, Ann Arbor, USA. Electronic address: mehtagee@umich.edu.
Geeta
G
Mehta
D045744:Cell Line, Tumor; D004354:Drug Screening Assays, Antitumor / Q000379:methods; D005260:Female; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D010051:Ovarian Neoplasms / Q000188:drug therapy* / Q000473:pathology*; D018874:Spheroids, Cellular
3D culture; High throughput; Multicellular tumor spheroids; Ovarian cancer; Preclinical drug testing