Enhanced mitochondrial fission suppresses signaling and metastasis in triple-negative breast cancer.

BACKGROUND: Mitochondrial dynamics underlies malignant transformation, cancer progression, and response to treatment. Current research presents conflicting evidence for functions of mitochondrial fission and fusion in tumor progression. Here, we investigated how mitochondrial fission and fusion states regulate underlying processes of cancer progression and metastasis in triple-negative breast cancer (TNBC). METHODS: We enforced mitochondrial fission and fusion states through chemical or genetic approaches and measured migration and invasion of TNBC cells in 2D and 3D in vitro models. We also utilized kinase translocation reporters (KTRs) to identify single cell effects of mitochondrial state on signaling cascades, PI3K/Akt/mTOR and Ras/Raf/MEK/ERK, commonly activated in TNBC. Furthermore, we determined effects of fission and fusion states on metastasis, bone destruction, and signaling in mouse models of breast cancer. RESULTS: Enforcing mitochondrial fission through chemical or genetic approaches inhibited migration, invasion, and metastasis in TNBC. Breast cancer cells with predominantly fissioned mitochondria exhibited reduced activation of Akt and ERK both in vitro and in mouse models of breast cancer. Treatment with leflunomide, a potent activator of mitochondrial fusion proteins, overcame inhibitory effects of fission on migration, signaling, and metastasis. Mining existing datasets for breast cancer revealed that increased expression of genes associated with mitochondrial fission correlated with improved survival in human breast cancer. CONCLUSIONS: In TNBC, mitochondrial fission inhibits cellular processes and signaling pathways associated with cancer progression and metastasis. These data suggest that therapies driving mitochondrial fission may benefit patients with breast cancer.

Enhanced mitochondrial fission inhibits triple-negative breast cancer cell migration through an ROS-dependent mechanism.

Mitochondria produce reactive oxygen species (ROS), which function in signal transduction. Mitochondrial dynamics, encompassing morphological shifts between fission and fusion, can directly impact ROS levels in cancer cells. In this study, we identified an ROS-dependent mechanism for how enhanced mitochondrial fission inhibits triple negative breast cancer (TNBC) cell migration. We found that enforcing mitochondrial fission in TNBC resulted in an increase in intracellular ROS levels and reduced cell migration and the formation of actin-rich migratory structures. Consistent with mitochondrial fission, increasing ROS levels in cells inhibited cell migration. Conversely, reducing ROS levels with either a global or mitochondrially targeted scavenger overcame the inhibitory effects of mitochondrial fission. Mechanistically, we found that the ROS sensitive SHP-1/2 phosphatases partially regulate inhibitory effects of mitochondrial fission on TNBC migration. Overall, our work reveals the inhibitory effects of ROS in TNBC and supports mitochondrial dynamics as a potential therapeutic target for cancer.

Bone Marrow Mesenchymal Stem Cells Induce Metabolic Plasticity in Estrogen Receptor-Positive Breast Cancer.

Cancer cells reprogram energy metabolism through metabolic plasticity, adapting ATP-generating pathways in response to treatment or microenvironmental changes. Such adaptations enable cancer cells to resist standard therapy. We employed a coculture model of estrogen receptor-positive (ER+) breast cancer and mesenchymal stem cells (MSC) to model interactions of cancer cells with stromal microenvironments. Using single-cell endogenous and engineered biosensors for cellular metabolism, coculture with MSCs increased oxidative phosphorylation, intracellular ATP, and resistance of cancer cells to standard therapies. Cocultured cancer cells had increased MCT4, a lactate transporter, and were sensitive to the MCT1/4 inhibitor syrosingopine. Combining syrosingopine with fulvestrant, a selective estrogen receptor degrading drug, overcame resistance of ER+ breast cancer cells in coculture with MSCs. Treatment with antiestrogenic therapy increased metabolic plasticity and maintained intracellular ATP levels, while MCT1/4 inhibition successfully limited metabolic transitions and decreased ATP levels. Furthermore, MCT1/4 inhibition decreased heterogenous metabolic treatment responses versus antiestrogenic therapy. These data establish MSCs as a mediator of cancer cell metabolic plasticity and suggest metabolic interventions as a promising strategy to treat ER+ breast cancer and overcome resistance to standard clinical therapies. IMPLICATIONS: This study reveals how MSCs reprogram metabolism of ER+ breast cancer cells and point to MCT4 as potential therapeutic target to overcome resistance to antiestrogen drugs.

Ultrasound-Induced Mechanical Compaction in Acoustically Responsive Scaffolds Promotes Spatiotemporally Modulated Signaling in Triple Negative Breast Cancer.

Cancer cells continually sense and respond to mechanical cues from the extracellular matrix (ECM). Interaction with the ECM can alter intracellular signaling cascades, leading to changes in processes that promote cancer cell growth, migration, and survival. The present study used a recently developed composite hydrogel composed of a fibrin matrix and phase-shift emulsion, termed an acoustically responsive scaffold (ARS), to investigate effects of local mechanical properties on breast cancer cell signaling. Treatment of ARSs with focused ultrasound drives acoustic droplet vaporization (ADV) in a spatiotemporally controlled manner, inducing local compaction and stiffening of the fibrin matrix adjacent to the matrix-bubble interface. Combining ARSs and live single cell imaging of triple-negative breast cancer cells, it is discovered that both basal and growth-factor stimulated activities of protein kinase B (also known as Akt) and extracellular signal-regulated kinase (ERK), two major kinases driving cancer progression, negatively correlate with increasing distance from the ADV-induced bubble both in vitro and in a mouse model. Together, these data demonstrate that local changes in ECM compaction regulate Akt and ERK signaling in breast cancer and support further applications of the novel ARS technology to analyze spatial and temporal effects of ECM mechanics on cell signaling and cancer biology.

Multiparametric MRI to quantify disease and treatment response in mice with myeloproliferative neoplasms.

Histopathology, the standard method to assess BM in hematologic malignancies such as myeloproliferative neoplasms (MPNs), suffers from notable limitations in both research and clinical settings. BM biopsies in patients fail to detect disease heterogeneity, may yield a nondiagnostic sample, and cannot be repeated frequently in clinical oncology. Endpoint histopathology precludes monitoring disease progression and response to therapy in the same mouse over time, missing likely variations among mice. To overcome these shortcomings, we used MRI to measure changes in cellularity, macromolecular constituents, and fat versus hematopoietic cells in BM using diffusion-weighted imaging (DWI), magnetization transfer, and chemical shift-encoded fat imaging. Combining metrics from these imaging parameters revealed dynamic alterations in BM following myeloablative radiation and transplantation. In a mouse MPLW515L BM transplant model of MPN, MRI detected effects of a JAK2 inhibitor, ruxolitinib, within 5 days of initiating treatment and identified differing kinetics of treatment responses in subregions of the tibia. Histopathology validated the MRI results for BM composition and heterogeneity. Anatomic MRI scans also showed reductions in spleen volume during treatment. These findings establish an innovative, clinically translatable MRI approach to quantify spatial and temporal changes in BM in MPN.

Effects of iron modulation on mesenchymal stem cell-induced drug resistance in estrogen receptor-positive breast cancer.

Patients with estrogen receptor-positive (ER+) breast cancer, the most common subtype, remain at risk for lethal metastatic disease years after diagnosis. Recurrence arises partly because tumor cells in bone marrow become resistant to estrogen-targeted therapy. Here, we utilized a co-culture model of bone marrow mesenchymal stem cells (MSCs) and ER+ breast cancer cells to recapitulate interactions of cancer cells in bone marrow niches. ER+ breast cancer cells in direct contact with MSCs acquire cancer stem-like (CSC) phenotypes with increased resistance to standard antiestrogenic drugs. We confirmed that co-culture with MSCs increased labile iron in breast cancer cells, a phenotype associated with CSCs and disease progression. Clinically approved iron chelators and in-house lysosomal iron-targeting compounds restored sensitivity to antiestrogenic therapy. These findings establish iron modulation as a mechanism to reverse MSC-induced drug resistance and suggest iron modulation in combination with estrogen-targeted therapy as a promising, translatable strategy to treat ER+ breast cancer.

Targeting disseminated estrogen-receptor-positive breast cancer cells in bone marrow.

Estrogen receptor-positive (ER+) breast cancer can recur up to 20 years after initial diagnosis. Delayed recurrences arise from disseminated tumors cells (DTCs) in sites such as bone marrow that remain quiescent during endocrine therapy and subsequently proliferate to produce clinically detectable metastases. Identifying therapies that eliminate DTCs and/or effectively target cells transitioning to proliferation promises to reduce risk of recurrence. To tackle this problem, we utilized a 3D co-culture model incorporating ER+ breast cancer cells and bone marrow mesenchymal stem cells to represent DTCs in a bone marrow niche. 3D co-cultures maintained cancer cells in a quiescent, viable state as measured by both single-cell and population-scale imaging. Single-cell imaging methods for metabolism by fluorescence lifetime (FLIM) of NADH and signaling by kinases Akt and ERK revealed that breast cancer cells utilized oxidative phosphorylation and signaling by Akt to a greater extent both in 3D co-cultures and a mouse model of ER+ breast cancer cells in bone marrow. Using our 3D co-culture model, we discovered that combination therapies targeting oxidative phosphorylation via the thioredoxin reductase (TrxR) inhibitor, D9, and the Akt inhibitor, MK-2206, preferentially eliminated breast cancer cells without altering viability of bone marrow stromal cells. Treatment of mice with disseminated ER+ human breast cancer showed that D9 plus MK-2206 blocked formation of new metastases more effectively than tamoxifen. These data establish an integrated experimental system to investigate DTCs in bone marrow and identify combination therapy against metabolic and kinase targets as a promising approach to effectively target these cells and reduce risk of recurrence in breast cancer.