Alanoud Aldossry

Breast cancer is the most frequently diagnosed malignancy among women worldwide, with the luminal A subtype being the most prevalent. Several studies have reported a complex interplay between breast cancer cells and the local microbiome; however, the mechanisms by which tumor cell-secreted factors influence bacterial biological properties remain insufficiently explored. In this study, we established an in vitro model that partially recapitulates the luminal A breast cancer microenvironment by exposing three breast-associated bacterial species, Pseudomonas aeruginosa, Enterococcus faecalis, and Escherichia coli, to conditioned media (CM) derived from MCF-7 (tumor) or MCF-10A (non-tumor control) cell lines. A combination of complementary approaches, including ultrastructural morphological assessment, biofilm formation assays, antimicrobial susceptibility testing, and virulence gene abundance profiling by genomic qPCR, was employed to reveal distinct tumor-microbiota interactions. Exposure to MCF-7 CM induced dose-dependent structural alterations in P. aeruginosa and E. faecalis, with pronounced membrane blebbing and structural disruption in E. faecalis. Biofilm formation was differentially modulated in a species- and concentration-dependent manner, with a persistent increase observed in E. coli. Antibiotic susceptibility profiles were selectively altered in E. faecalis, which displayed increased sensitivity to vancomycin, penicillin, and imipenem, along with decreased sensitivity to chloramphenicol. P. aeruginosa exhibited increased sensitivity to imipenem along with reduced sensitivity to meropenem and gentamicin, whereas no significant changes were observed in E. coli. qPCR analyses demonstrated that MCF-7 CM was associated with enrichment of multiple virulence-associated genes (e.g., lasB, exoS, pilB, plcH, fsrC, esp, fimH, and papG), reflecting enhanced pathogenic and adhesive potential. Collectively, these findings suggest that luminal A breast cancer-derived factors can reprogram microbial phenotypes in a species-specific manner, providing mechanistic insight into breast tumor-microbiome crosstalk and a platform to explore microbiome-targeted interventions.

Breast cancer (BC) is the most frequently diagnosed malignancy in women worldwide. Despite therapeutic advances, disease relapse and metastasis remain major challenges and drivers of mortality. Fascin, an actin-bundling protein, promotes BC progression by enhancing drug resistance. However, the role of fascin in proliferation, a hallmark of cancer, and the underlying mechanism remain poorly elucidated. In this study, bioinformatics analysis of publicly available BC datasets, gene manipulation (gain and loss of function) in BC cell lines, flow cytometry, Western blots, and a real-time cell analyzer (RTCA) were employed to assess the role of fascin in proliferation. The clinical relevance of bioinformatics data and in vitro findings was assessed in BC patient samples using immunohistochemistry. FSCN1 expression exhibited a significant correlation with proliferation signature scores in BC datasets. Ectopic expression of fascin in fascin-negative SK-BR-3 and its silencing in fascin-positive MDA-MB-231 BC cell lines demonstrated its direct role in driving proliferation. In-depth bioinformatics analyses revealed a significant correlation between FSCN1 and the cell cycle signature score, particularly the G1-S signature score gene set. Indeed, fascin accelerated the cell cycle progression of synchronized cells from the G to S phase. Mechanistically, fascin upregulated nuclear SKP2 levels and reduced p27 expression—important G1-S cell cycle checkpoint regulators. Immunohistochemistry in samples from 68 patients demonstrated significant correlations between fascin and Ki-67 expression, in addition to SKP2 overexpression and p27 downregulation. Collectively, these data demonstrate the role of fascin as a driver of the G1-S-phase transition during cell cycle proliferation, thereby revealing new opportunities for targeted therapeutic intervention.

Omics technologies provide useful tools for the identification of novel biomarkers in many diseases, including breast cancer, which is the most diagnosed cancer in women worldwide. We and others have reported a central role for the actin-bundling protein (fascin) in regulating breast cancer disease progression at different levels. However, whether fascin expression promotes metabolic molecules that could predict disease progression has not been fully elucidated. Here, fascin expression was manipulated via knockdown (fascin^KD+NORF) and rescue (fascin^KD+FORF) in the naturally fascin-positive (fascin^pos+NORF) MDA-MB-231 breast cancer cells. Whether fascin dysregulates metabolic profiles that are associated with disease progression was assessed using untargeted metabolomics analyses via liquid chromatography–mass spectrometry. Overall, 12,226 metabolic features were detected in the tested cell pellets. Fascin^pos+NORF cell pellets showed 2510 and 3804 significantly dysregulated metabolites compared to their fascin^KD+NORF counterparts. Fascin rescue (fascin^KD+FORF) revealed 2710 significantly dysregulated cellular metabolites compared to fascin^KD+NORF counterparts. A total of 101 overlapped cellular metabolites between fascin^KD+FORF and fascin^pos+NORF were significantly dysregulated in the fascin^KD+NORF cells. Analysis of the significantly dysregulated metabolites by fascin expression revealed their involvement in the metabolism of sphingolipid, phenylalanine, tyrosine, and tryptophan biosynthesis, and pantothenate and CoA biosynthesis, which are critical pathways for breast cancer progression. Our findings of fascin-mediated alteration of metabolic pathways could be used as putative poor prognostic biomarkers and highlight other underlying mechanisms of fascin contribution to breast cancer progression.