Mic60‑low breast cancer cells activate epithelial-mesenchymal transition and support aggressive tumor growth

Background & Objectives : Breast cancer (BC) is the most frequent malignancy in women and a leading cause of cancer-related mortality worldwide. Despite advances in early detection and therapy, BC heterogeneity remains a major clinical challenge. Recent evidence highlights the role of mitochondria in sustaining BC aggressiveness and metabolic reprogramming. An shRNA screen identified Mic60 (also known as Mitofilin) as a mitochondrial gene whose reduced expression is associated with enhanced tumor cell invasion and altered proliferation. Mic60, part of the MICOS multiprotein complex, ensures cristae architecture, respiratory complex organization, and outer membrane biogenesis. Notably, Mic60 loss reduces mitochondrial fitness, contributing to aggressive disease. We investigated how Mic60 influences BC phenotype with regard to epithelial-mesenchymal transition (EMT) modulation in vitro and tumorigenic potential in vivo. Materials & Methods : Mic60 silencing was achieved using small interfering RNA (siRNA) and short hairpin RNA (shRNA) in human and murine BC cell lines. Gene expression was analyzed by RT‑qPCR, protein levels by Western blotting and flow cytometry. Migration and invasion were assessed in vitro using transwell assays with and without Matrigel, quantified by microscopy. In vivo tumorigenic capacity was evaluated in immunocompetent mice injected with syngeneic AT3 cells stably silenced for Mic60. Results : Mic60 silencing induced EMT master regulators (Twist, Snail, Lbx1, Zeb2) in both human and murine BC cell lines. This transcriptional increase was accompanied by elevated N‑cadherin and reduced E‑cadherin and EpCAM expression. Moreover, Mic60 silencing upregulated NF‑κB signaling, a key transcription factor linking inflammation, EMT, and cancer. Functionally, silenced cells exhibited enhanced migration and invasion in vitro. When injected into immunocompetent mice, AT3 Mic60‑silenced cells displayed accelerated tumor growth in vivo. Conclusions : Mic60 downmodulation activates EMT in BC cells, promoting migratory and invasive behavior. This phenotype is accompanied by NF‑κB pathway activation, linking Mic60 loss to pro‑tumorigenic signaling. In vivo, Mic60 depletion drives increased tumor growth, confirming the relevance of EMT activation in cancer progression. These findings extend previous observations on Mic60 in breast cancer, directly supporting its role in tumor progression and disease aggressiveness.