Abstract:
Three-dimensional cell culture models are increasingly adopted as preferred pre-clinical
drug testing platforms, as they circumvent limitations associated with traditional monolayer cell
cultures. However, many of these models are not fully characterized. This study aimed to characterize
a BT-20 triple-negative breast carcinoma spheroid model and assess its susceptibility to doxorubicin
in comparison to a monolayer model. Spheroids were developed using the liquid overlay method.
Phenotypic attributes were analyzed by characterizing changes in size, gross morphology, protein
content, metabolic activity, hypoxic status, and cell–cell junctions. The cytotoxic range of doxorubicin
in monolayers was determined using the sulforhodamine B assay, and the comparative effect of toxic
and sub-toxic concentrations was assessed in both spheroids and monolayers. Similar to the in vivo
microenvironment, spheroids had a heterogeneous spatial cytoarchitecture, inherent hypoxia and
strong adherens junctions. Doxorubicin induced dose-dependent cytotoxicity in monolayers (IC25:
130 nM, IC50: 320 nM and IC75: 1580 nM); however, these concentrations did not alter the spheroid
size or acid phosphatase activity. Only concentrations 6 M had any effect on spheroid integrity. In
comparison to monolayers, the BT-20 spheroid model has decreased sensitivity to doxorubicin and
could serve as a better model for susceptibility testing in triple-negative breast cancer.
Description:
DATA AVAILABILITY STATEMENT : All data created or analyzed during this study are available from the corresponding author upon request.
SUPPLEMENTARY MATERIALS : FIGURE S1: Dose–response curves showing the alteration of cell density, N = 5 biological repeats (A) and APH, N = 4 biological repeats (B) of monolayers treated with half-log dilutions of 32 M doxorubicin for 72 h. The IC50 was calculated using a non-linear regression curve fit (log[inhibitor] vs. response) with a robust fit.
