Evaluation of the Activity of Monensin and Its Analogs for Modulation of Stem-like Cell Functionality in 2D and 3D Breast Cancer Models

Monensin (MON) is a polyether ionophore
antibiotic
of natural origin and is an FDA-approved drug for veterinary use.
Recent studies have highlighted its potential anticancer activity
in various in vitro and in vivo models.
In this study, we evaluated the anti-breast cancer activity of MON and 37 synthetic analog compounds using cell monolayer
and organoid models. Through a mini-ring cell viability assay, several
compounds were identified that were more potent and selective against
breast cancer cells compared to non-cancerous cells, surpassing the
activity of parent MON. MON and these compounds
induced significant DNA fragmentation, reduced cell migration, and
downregulated SOX2 expression. Furthermore, MON and the
most potent analog, compound 12, reduced the percentage
of CD44+/CD24–/low stem-like cells and
diminished colony formation properties. Proteomics analyses revealed
that several pathways, including extracellular matrix organization,
were significantly dysregulated by MON and compound 12 in breast cancer cells. Among these, TIMP2, a protein associated
with the suppression of tumor growth and metastasis, was identified
as one of the most prominently upregulated proteins by MON and compound 12 in MDA-MB-231 cells. This finding was
also validated in other breast cancer and melanoma cell lines. To
simulate breast cancer metastasis to the brain, a human hybrid organoid
system: tumor in brain organoid (HOSTBO) model was developed. MON and compound 12 significantly reduced Ki-67
expression within the HOSTBOs, and compound 12 significantly
downregulated SOX2 expression. Collectively, MON and
compound 12 significantly reduced the proliferation of
breast cancer stem-like cells in the organoid models, inhibited their
migration, and dysregulated markers associated with stemness, demonstrating
their potential as anti-metastatic agents and warranting further clinical
development.