PhD Scientific Days 2021

PO_I_P: Pathology and Oncology I. Posters

Autophagy-induction Derived Non-apoptotic Cell Death in Human Breast Cancer Cell Lines

Text of the abstract

Introduction
Metabolic remodeling is a characteristic of cancer cells as a response to treatments. Besides promoting cell growth and survival, the activation of mTOR pathway can also lead to developing resistance mechanism or metastasis formation. In addition, autophagy has a double-edged role as it can either maintain cellular homeostasis or stimulate tumorigenesis in stress.
Aims
As doxycycline (antibiotic agent) has an off-target effect on mitochondrial processes, we aimed to study whether its combination with rapamycin (mTOR inhibitor) is potentially more effective in treatments.
Methods
Rapamycin + doxycycline was tested in several cell lines in vitro, and the mechanism of action was examined using human breast cancer models. Alamar blue and sulforhodamine B tests were applied and the impact of long-term mono- and combination treatments on cell and tumor growth was also monitored in in vitro and in vivo. The induced cell death was detected by flow cytometry and caspase-3 activity measurements. mTOR-, autophagy-, and necroptosis-related proteins were analyzed by WES, Western blot and IHC. Treatment-induced morphological alterations were examined by fluoresce and transmission electron microscopy.
Results
Rapamycin + doxycycline combination could decrease the cell proliferation capacity in the majority of the investigated cell lines in vitro. Continuous rapamycin + doxycycline addition could reduce cell and tumor growth significantly. The studied combination did not cause apoptosis, necrosis or necroptosis, but altered the expressions of LC3 and p62 which were associated with autophagy induction. Lowered intensity of MitoTracker and TOM20 staining suggested that mitophagy could be induced after rapamycin + doxycycline treatment. In parallel, rapamycin + doxycycline combination increased the sign of autophagy (mitophagy)-related alterations at tissue level.
Conclusion
A novel therapeutic strategy could be to co-target mitochondria and mTOR pathway to influence metabolic rewiring and inhibit metabolic adaptation in tumor cells.
Funding
NKFI-FK-128404, National Bionics program-ED_17-1-2017-0009, STIA-KFI2020, VEKOP-2.3.2-16-2016-00002, OTKA120237, PPD-222/2018

University and Doctoral School

Semmelweis University, Doctoral School of Pathological Sciences