Targeting dysregulated mitochondrial fission pathways in triple negative breast cancer therapy

[Speaker] Angelique Nyinawabera:1
[Co-author] Smiti Gupta:1, Karthikeyan Chandrabose:1, Amit K Tiwari:1
1:Pharmacology and Experimental Therapeutics, The University of Toledo, College of Pharmacy and Pharmaceutical Sciences, USA

 Triple-negative breast cancer (TNBC), is the most aggressive form of malignant mammary tumors that do not express three cell-surface receptors (estrogen receptor , progesterone receptor, and HER2/ERBB3). TNBC accounts for about 15 percent of breast cancer cases with >42,000 new cases each year and approximately 21,000 deaths. Currently, there are no targeted treatments available for TNBC. We found an interesting target associated with TNBC the mitochondrial fission protein GTPase dynamin-related protein-1 (Drp1), for which one inhibitor (mdivi-1) is under active investigation. However, mdivi-1 has shown inconsistent antineoplastic activity low potency and selectivity. This led us to use a 200,000-compound library of commercially available small molecules (Specs) in conjunction with the likely structure of the Drp1-mdivi-1 inhibitory complex resulting in the identification of twelve thieno-pyrimidin-4-yl-hydrazinylidene class of compounds that were highly potent and selective Drp1 inhibitors. In our preliminary work, we found one compound ANT4 with IC50 value of 180-220 nM that has up to 100-fold selectivity in different TNBC cells compared to normal breast cells and up to 30-fold compared to other normal cells. We discovered that ANT4 is a selective inhibitor of Drp1 and has a 100-fold greater affinity for the Drp1 complex. ANT4 inhibited TNBC colony formation, invasion, and metastasis 150-fold more potently than Mdivi-1. Interestingly, we discovered that ANT4, in nanomolar range, selectively induced a unique non-apoptotic necroptotic inducing (NANI) cell death in TNBC cells by increasing the expression of necroptotic cell death markers (RIP p-RIP MLKL p-MLKL). Further, ANT4 inhibited the expression of pro-apoptotic proteins and enhanced the expression of anti-apoptotic proteins. In addition ANT4 arrested the TNBC cells in S-phase without producing any significant nuclear condensation or release of reactive oxygen species (ROS). Further studies are underway to understand the Drp1 mediated NANI induction process in TNBC cells and to identify the structural requirements that are necessary for targeting cancers refractory to apoptosis. The findings of these studies will allow us to understand the biology of necroptosis-induced Drp1 inhibition and to develop this new class of small molecules for the treatment of TNBC patients.

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