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Mitochondrial dynamics orchestrate many essential cellular functions, including metabolism, which is instrumental in promoting cancer growth and metastatic progression. However, how mitochondrial dynamics influences metastatic progression remains poorly understood. Here, we show that breast cancer cells with low metastatic potential exhibit a more fused mitochondrial network compared to highly metastatic cells. To study the impact of mitochondrial dynamics on metastasis, we promoted mitochondrial elongation in metastatic breast cancer cells by individual genetic deletion of three key regulators of mitochondrial fission (Drp1, Fis1, Mff) or by pharmacological intervention with leflunomide. Omics analyses revealed that mitochondrial elongation causes substantial alterations in metabolic pathways and processes related to cell adhesion. In vivo, enhanced mitochondrial elongation by loss of mitochondrial fission mediators or treatment with leflunomide notably reduced metastasis formation. Furthermore, the transcriptomic signature associated with elongated mitochondria correlated with improved clinical outcome in patients with breast cancer. Overall, our findings highlight mitochondrial dynamics as a potential therapeutic target in breast cancer.

Metabolic rewiring is essential for tumor growth and progression to metastatic disease, yet little is known regarding how cancer cells modify their acquired metabolic programs in response to different metastatic microenvironments. We have previously shown that liver-metastatic breast cancer cells adopt an intrinsic metabolic program characterized by increased HIF-1α activity and dependence on glycolysis. Here, we confirm by in vivo stable isotope tracing analysis (SITA) that liver-metastatic breast cancer cells retain a glycolytic profile when grown as mammary tumors or liver metastases. However, hepatic metastases exhibit unique metabolic adaptations including elevated expression of genes involved in glutathione (GSH) biosynthesis and reactive oxygen species (ROS) detoxification when compared to mammary tumors. Accordingly, breast-cancer-liver-metastases exhibited enhanced de novo GSH synthesis. Confirming their increased capacity to mitigate ROS-mediated damage, liver metastases display reduced levels of 8-Oxo-2'-deoxyguanosine. Depletion of the catalytic subunit of the rate-limiting enzyme in glutathione biosynthesis, glutamate-cysteine ligase (GCLC), strongly reduced the capacity of breast cancer cells to form liver metastases, supporting the importance of these distinct metabolic adaptations. Loss of GCLC also affected the early steps of the metastatic cascade, leading to decreased numbers of circulating tumor cells (CTCs) and impaired metastasis to the liver and the lungs. Altogether, our results indicate that GSH metabolism could be targeted to prevent the dissemination of breast cancer cells., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: Compared to minimally invasive brain metastases (MI BrM), highly invasive (HI) lesions form abundant contacts with cells in the peritumoral brain parenchyma and are associated with poor prognosis. Reactive astrocytes (RAs) labeled by phosphorylated STAT3 (pSTAT3) have recently emerged as a promising therapeutic target for BrM. Here, we explore whether the BrM invasion pattern is influenced by pSTAT3+ RAs and may serve as a predictive biomarker for STAT3 inhibition., Methods: We used immunohistochemistry to identify pSTAT3+ RAs in HI and MI human and patient-derived xenograft (PDX) BrM. Using PDX, syngeneic, and transgenic mouse models of HI and MI BrM, we assessed how pharmacological STAT3 inhibition or RA-specific STAT3 genetic ablation affected BrM growth in vivo. Cancer cell invasion was modeled in vitro using a brain slice-tumor co-culture assay. We performed single-cell RNA sequencing of human BrM and adjacent brain tissue., Results: RAs expressing pSTAT3 are situated at the brain-tumor interface and drive BrM invasive growth. HI BrM invasion pattern was associated with delayed growth in the context of STAT3 inhibition or genetic ablation. We demonstrate that pSTAT3+ RAs secrete Chitinase 3-like-1 (CHI3L1), which is a known STAT3 transcriptional target. Furthermore, single-cell RNA sequencing identified CHI3L1-expressing RAs in human HI BrM. STAT3 activation, or recombinant CHI3L1 alone, induced cancer cell invasion into the brain parenchyma using a brain slice-tumor plug co-culture assay., Conclusions: Together, these data reveal that pSTAT3+ RA-derived CHI3L1 is associated with BrM invasion, implicating STAT3 and CHI3L1 as clinically relevant therapeutic targets for the treatment of HI BrM., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Transmembrane glycoprotein NMB (GPNMB) is a prognostic marker of poor outcome in patients with triple-negative breast cancer (TNBC). Glembatumumab Vedotin, an antibody drug conjugate targeting GPNMB, exhibits variable efficacy against GPNMB-positive metastatic TNBC as a single agent. We show that GPNMB levels increase in response to standard-of-care and experimental therapies for multiple breast cancer subtypes. While these therapeutic stressors induce GPNMB expression through differential engagement of the MiTF family of transcription factors, not all are capable of increasing GPNMB cell-surface localization required for Glembatumumab Vedotin inhibition. Using a FACS-based genetic screen, we discovered that suppression of heat shock protein 90 (HSP90) concomitantly increases GPNMB expression and cell-surface localization. Mechanistically, HSP90 inhibition resulted in lysosomal dispersion towards the cell periphery and fusion with the plasma membrane, which delivers GPNMB to the cell surface. Finally, treatment with HSP90 inhibitors sensitizes breast cancers to Glembatumumab Vedotin in vivo, suggesting that combination of HSP90 inhibitors and Glembatumumab Vedotin may be a viable treatment strategy for patients with metastatic TNBC., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Growing tumors exist in metabolically compromised environments that require activation of multiple pathways to scavenge nutrients to support accelerated rates of growth. The folliculin (FLCN) tumor suppressor complex (FLCN, FNIP1, FNIP2) is implicated in the regulation of energy homeostasis via 2 metabolic master kinases: AMPK and mTORC1. Loss-of-function mutations of the FLCN tumor suppressor complex have only been reported in renal tumors in patients with the rare Birt-Hogg-Dube syndrome. Here, we revealed that FLCN, FNIP1, and FNIP2 are downregulated in many human cancers, including poor-prognosis invasive basal-like breast carcinomas where AMPK and TFE3 targets are activated compared with the luminal, less aggressive subtypes. FLCN loss in luminal breast cancer promoted tumor growth through TFE3 activation and subsequent induction of several pathways, including autophagy, lysosomal biogenesis, aerobic glycolysis, and angiogenesis. Strikingly, induction of aerobic glycolysis and angiogenesis in FLCN-deficient cells was dictated by the activation of the PGC-1α/HIF-1α pathway, which we showed to be TFE3 dependent, directly linking TFE3 to Warburg metabolic reprogramming and angiogenesis. Conversely, FLCN overexpression in invasive basal-like breast cancer models attenuated TFE3 nuclear localization, TFE3-dependent transcriptional activity, and tumor growth. These findings support a general role of a deregulated FLCN/TFE3 tumor suppressor pathway in human cancers.

Non-alcoholic fatty liver disease (NAFLD) is the most frequent liver disease worldwide and can progress to non-alcoholic steatohepatitis (NASH), which is characterized by triglyceride accumulation, inflammation, and fibrosis. No pharmacological agents are currently approved to treat these conditions, but it is clear now that modulation of lipid synthesis and autophagy are key biological mechanisms that could help reduce or prevent these liver diseases. The folliculin (FLCN) protein has been recently identified as a central regulatory node governing whole body energy homeostasis, and we hypothesized that FLCN regulates highly metabolic tissues like the liver. We thus generated a liver specific Flcn knockout mouse model to study its role in liver disease progression. Using the methionine- and choline-deficient diet to mimic liver fibrosis, we demonstrate that loss of Flcn reduced triglyceride accumulation, fibrosis, and inflammation in mice. In this aggressive liver disease setting, loss of Flcn led to activation of transcription factors TFEB and TFE3 to promote autophagy, promoting the degradation of intracellular lipid stores, ultimately resulting in reduced hepatocellular damage and inflammation. Hence, the activity of FLCN could be a promising target for small molecule drugs to treat liver fibrosis by specifically activating autophagy. Collectively, these results show an unexpected role for Flcn in fatty liver disease progression and highlight new potential treatment strategies., (© 2021. The Author(s).)

Cell migration is central to many developmental, physiologic and pathological processes, including cancer progression. The Ral GTPases (RalA and RalB) which act down-stream the Ras oncogenes, are key players in the coordination between membrane trafficking and actin polymerization. A major direct effector of Ral, the exocyst complex, works in polarized exocytosis and is at the center of multiple protein-protein interactions that support cell migration by promoting protrusion formation, front-rear polarization, and extra-cellular matrix degradation. In this review we describe the recent advancements in deciphering the molecular mechanisms underlying this role of Ral via exocyst on cell migration. Among others, we will discuss the recently identified cross-talk between Ral and Rac1 pathways: exocyst binds to a negative regulator (the RacGAP SH3BP1) and to the major effector (the Wave Regulatory Complex, WRC) of Rac1, the master regulator of protrusions. Next challenge will be to better characterize the dynamics in space and in time of these molecular interplays, to better understand the pleiotropic functions of Ral in both normal and cancer cells.

GPNMB has emerged as an immunomodulator and an important positive mediator of tumor progression and metastasis in numerous solid cancers. Tumor intrinsic GPNMB-mediated effects on cellular signaling, coupled with the ability of GPNMB to influence the primary tumor and metastatic microenvironments in a non-cell autonomous fashion, combine to augment malignant cancer phenotypes. In addition, GPNMB is often overexpressed in a variety of cancers, making it an attractive therapeutic target. In this regard, glembatumumab vedotin, an antibody-drug conjugate (ADC) that targets GPNMB, is currently in clinical trials as a single agent in multiple cancers. In this review, we will describe the physiological functions of GPNMB in normal tissues and summarize the processes through which GPNMB augments tumor growth and metastasis. We will review the pre-clinical and clinical development of glembatumumab vedotin, evaluate on-going clinical trials, explore emerging opportunities for this agent in new disease indications and discuss exciting possibilities for this ADC in the context of combination therapies., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Purpose: To determine if BRAF and/or MEK inhibitor-induced GPNMB expression renders melanomas sensitive to CDX-011, an antibody-drug conjugate targeting GPNMB., Experimental Design: The Cancer Genome Atlas melanoma dataset was interrogated for a panel of MITF-regulated melanosomal differentiation antigens, including GPNMB. BRAF-mutant melanoma cell lines treated with BRAF or MEK inhibitors were assessed for GPNMB expression by RT-qPCR, immunoblot, and FACS analyses. Transient siRNA-mediated knockdown approaches were used to determine if MITF is requirement for treatment-induced GPNMB upregulation. GPNMB expression was analyzed in serial biopsies and serum samples from patients with melanoma taken before, during, and after disease progression on MAPK inhibitor treatment. Subcutaneous injections were performed to test the efficacy of MAPK inhibitors alone, CDX-011 alone, or their combination in suppressing melanoma growth., Results: A MITF-dependent melanosomal differentiation signature is associated with poor prognosis in patients with this disease. MITF is increased following BRAF and MEK inhibitor treatment and induces the expression of melanosomal differentiation genes, including GPNMB. GPNMB is expressed at the cell surface in MAPK inhibitor-treated melanoma cells and is also elevated in on-treatment versus pretreatment biopsies from melanoma patients receiving MAPK pathway inhibitors. Combining BRAF and/or MEK inhibitors with CDX-011, an antibody-drug conjugate targeting GPNMB, is effective in causing melanoma regression in preclinical animal models and delays the recurrent melanoma growth observed with MEK or BRAF/MEK inhibitor treatment alone., Conclusions: The combination of MAPK pathway inhibitors with an antibody-drug conjugate targeting GPNMB is an effective therapeutic option for patients with melanoma. Clin Cancer Res; 22(24); 6088-98. ©2016 AACR., (©2016 American Association for Cancer Research.)

Coordination between membrane trafficking and actin polymerization is fundamental in cell migration, but a dynamic view of the underlying molecular mechanisms is still missing. The Rac1 GTPase controls actin polymerization at protrusions by interacting with its effector, the Wave regulatory complex (WRC). The exocyst complex, which functions in polarized exocytosis, has been involved in the regulation of cell motility. Here, we show a physical and functional connection between exocyst and WRC. Purified components of exocyst and WRC directly associate in vitro, and interactions interfaces are identified. The exocyst-WRC interaction is confirmed in cells by co-immunoprecipitation and is shown to occur independently of the Arp2/3 complex. Disruption of the exocyst-WRC interaction leads to impaired migration. By using time-lapse microscopy coupled to image correlation analysis, we visualized the trafficking of the WRC towards the front of the cell in nascent protrusions. The exocyst is necessary for WRC recruitment at the leading edge and for resulting cell edge movements. This direct link between the exocyst and WRC provides a new mechanistic insight into the spatio-temporal regulation of cell migration., (© 2016. Published by The Company of Biologists Ltd.)

RalA and RalB proteins are key mediators of oncogenic Ras signaling in human oncogenesis. Herein we investigated the mechanistic contribution of Ral proteins to invasion of lung cancer A549 cells after induction of epithelial-mesenchymal transition (EMT) with TGFβ. We show that TGFβ-induced EMT promotes dissemination of A549 cells in a 2/3D assay, independently of proteolysis, by activating the Rho/ROCK pathway which generates actomyosin-dependent contractility forces that actively remodel the extracellular matrix, as assessed by Traction Force microscopy. RalB, but not RalA, is required for matrix deformation and cell dissemination acting via the RhoGEF GEF-H1, which associates with the Exocyst complex, a major Ral effector. Indeed, uncoupling of the Exocyst subunit Sec5 from GEF-H1 impairs RhoA activation, generation of traction forces and cell dissemination. These results provide a novel molecular mechanism underlying the control of cell invasion by RalB via a cross-talk with the Rho pathway.

Characterizing the migration of a population of cells remains laborious and somewhat subjective. Advances in genetics and robotics allow researchers to perform many experiments in parallel, but analyzing the large sets of data remains a bottleneck. Here we describe a rapid, fully automated correlation-based method for cell migration analysis, compatible with standard video microscopy. This method allows for the computation of quantitative migration parameters via an extensive dynamic mapping of cell displacements.

The coordination of the several pathways involved in cell motility is poorly understood. Here, we identify SH3BP1, belonging to the RhoGAP family, as a partner of the exocyst complex and establish a physical and functional link between two motility-driving pathways, the Ral/exocyst and Rac signaling pathways. We show that SH3BP1 localizes together with the exocyst to the leading edge of motile cells and that SH3BP1 regulates cell migration via its GAP activity upon Rac1. SH3BP1 loss of function induces abnormally high Rac1 activity at the front, as visualized by in vivo biosensors, and disorganized and instable protrusions, as revealed by cell morphodynamics analysis. Consistently, constitutively active Rac1 mimics the phenotype of SH3BP1 depletion: slow migration and aberrant cell morphodynamics. Our finding that SH3BP1 downregulates Rac1 at the motile-cell front indicates that Rac1 inactivation in this location, as well as its activation by GEF proteins, is a fundamental requirement for cell motility., (Copyright © 2011 Elsevier Inc. All rights reserved.)

This paper develops scaling laws for plant roots of any arbitrary volume and branching configuration that maximize water uptake. Water uptake can occur along any part of the root network, and thus there is no branch-to-branch fluid conservation. Maximizing water uptake, therefore, involves balancing two flows that are inversely related: axial and radial conductivity. The scaling laws are tested against the root data of 1759 plants from 77 herbaceous species, and compared with those from the WBE model. I further discuss whether the scaling laws are invariant to soil water distribution. A summary of some of the results follows. (1) The optimal radius for a single root (no branches) scales with volume as r approximately volume(2/(8+a))(0

Far-red radiation (FR) reflected by green tissues is a key signal that plants use to detect the proximity of future competitors. Perception of increased levels of FR elicits a suite of responses collectively known as the shade-avoidance syndrome, which includes increased stem elongation, production of erect leaves, and reduced lateral branching. These responses improve the access to light for plants that occur in crowded populations. Responses to the proximity of competitors are known to affect the susceptibility to disease and predation in several organisms, including social animals. However, the impacts of warning signals of competition on the expression of defenses have not been explicitly investigated in plants. In the experiments reported here, we show that reflected FR induced a dramatic down-regulation of chemical defenses in wild tobacco (Nicotiana longiflora). FR altered the expression of several defense-related genes, inhibited the accumulation of herbivore-induced phenolic compounds, and augmented the performance of the specialist herbivore Manduca sexta. Complementary studies with tomato suggested that the effects of FR on defenses are mediated by the photoreceptor phytochrome B. The central implication of these results is that shade-intolerant species such as wild tobacco and tomato activate functional changes that affect their ability to cope with herbivore attack in response to phytochrome signals of future competition, even in the absence of real competition for resources. These findings suggest that competition overshadowed herbivory during the evolution of this group of species and add a new axis to the definition of the shade-avoidance syndrome.

Cannabis derivatives and alcohol are widely co-abused, particularly among adolescents. Since both ethanol and cannabinoids are known to impair learning and memory, the present study investigated in rats the effects of combined exposure to ethanol and delta(9)-tetrahydrocannabinol (THC) in a memory task, the object recognition test. The results of the present study provide evidence that ethanol, voluntarily ingested in alcohol-preferring rats, and THC, given by intraperitoneal injection, have a synergic action to impair object recognition, when a 15-min interval was adopted between the sample phase and the choice phase of the test. Neither voluntary ethanol ingestion nor 2 or 5 mg/kg of THC were able per se to modify object recognition in these experimental conditions, but when voluntary ethanol ingestion was combined with administration of these doses of THC object recognition was markedly impaired. THC impaired object recognition only at the dose of 10 mg/kg, when its administration was not combined with that of ethanol. The selective cannabinoid CB(1) receptor antagonist SR 141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1(2, 4-dichloro-phenyl)-4-methyl-1H-pyrazole carboxamide.HCl) at the dose of 1 mg/kg reversed the amnesic effect of THC, 10 mg/kg, suggesting that the effect is mediated by this receptor subtype. The synergism of ethanol and THC was not detected when an inter-trial interval of 1 min was adopted. The present findings are in keeping with the notion that Cannabis derivatives impair memory processes and provide evidence for a synergic action of THC and ethanol, thus emphasizing the risks consequent to their co-administration.

Rationale: (1S,3aS)-8-(2,3,3,4,5,6-hexahydro-1H-phenalen-1-yl)-1-phenyl-1,3,8-triazaspiro[4.5]decan-4-one (Ro 64-6198), a non-peptidic agonist for the opioid receptor-like1 (ORL1) receptor, exhibits anxiolytic properties in stressful conditions., Objective: The present study was aimed at evaluating whether activation of ORL1 receptors by Ro 64-6198 may reverse the anorectic effect of restraint stress or intracerebroventricular (ICV) CRF injection., Methods: In body restraint experiments, 20-h food deprived rats were treated with intraperitoneal (IP) injection of Ro 64-6198 or vehicle. Ten minutes later, they were confined in cylindrical Plexiglas tubes for 60 min and then returned to their cage with food. In CRF experiments, 20-h food deprived rats were IP injected with Ro 64-6198 or vehicle. Ten minutes later, they received ICV CRF, 200 ng/rat or vehicle; food was offered after 20 min., Results: Intraperitoneal (IP) pretreatment with Ro 64-6198 reversed the hypophagic effect induced by both restraint or CRF; the effect was statistically significant at the three doses tested (0.3, 1.0 or 2.5 mg/kg). ICV administration of the selective ORL1 receptor antagonist [Nphe(1)]NC(1-13)NH(2)(two injections of 33 or 66 microg/rat) abolished the effect of Ro 64-6198 on CRF-induced anorexia. In freely feeding rats, Ro 64-6198 significantly increased feeding at 2.5, but not at 0.3 or 1.0 mg/kg; in food deprived rats, Ro 64-6198 (0.3 or 1.0 mg/kg) did not modify food intake. Thus, reversal of stress- and CRF-induced anorexia by Ro 64-6198 can be evoked at doses lower than those that are hyperphagic. Ro 64-6198 (1 or 2.5 mg/kg) did not modify the anorectic effect of E. coli lipopolysaccharide, suggesting that its effect is selective for stress- or CRF-induced anorexia. Lastly, the benzodiazepine diazepam was unable to reduce the anorectic effect of CRF at the anxiolytic dose of 0.3 mg/kg, and partially reduced it at the hyperphagic dose of 1 mg/kg., Conclusions: The results of this study show that the non-peptidic ORL1 receptor agonist Ro 64-6198 markedly and selectively inhibits the anorectic effect of stress and CRF, and provide evidence that this effect is mediated by ORL1 receptors. Thus, Ro 64-6198 may represent an interesting tool for treatment of stress-induced anorexia.

Eight ruminally cannulated steers (average initial weight 196 +/- 13 kg) were arranged in a replicated 4 x 4 Latin square. Steers had ad libitum access to chopped brome hay (Bromus inermus L. 9.9% CP) and one of four supplements. Supplements were fed to provide 195 g of CP.steer-1 x d-1 (equalized with soybean meal), and supplemental energy intake was similar among energy supplements. Supplemental treatments were control (CTRL; primarily soybean meal), barley (BAR), beet pulp (PULP), and corn (CORN). Amounts fed were 481, 1,285, 1,335, and 1,289 g of DM.steer-1 x d-1 for CTRL, BAR, PULP, and CORN, respectively. Individually stanchioned steers were allowed a 13-d adaptation period followed by an 8-d collection period. Chromic oxide was used as the indigestible flow marker. Forage intake was less (P < .10) for steers fed BAR, PULP, and CORN than for CTRL steers, but total intake (forage+supplement) did not differ among treatments. Total tract DM and OM digestibilities were greater (P < .10) for PULP- and CORN- than for CTRL- and BAR-supplemented steers. Digestibility of NDF was least (P < .10) for BAR, intermediate for CORN, and greatest for CTRL and PULP (44.0, 47.2, 50.1, and 52.1%, respectively). Apparent CP digestibility was least (P < .10) in BAR-(60.8), greatest in CTRL- (69.7), and intermediate in PULP- and CTRL- (64.0 and 64.4) supplemented steers. Ruminal DM fill was less (P < .10) in steers fed BAR and PULP than in CTRL steers. No differences (P < .10) were noted in DM and NDF rate of in situ degradation. Rate of forage CP in situ degradation was faster (P < .10) in CTRL- and PULP- than in CORN- and BAR-supplemented steers. Steers fed BAR and PULP supplements exhibited a lower overall pH response curve (P < .005) than did steers fed CTRL and CORN; this finding corresponded to increased (P < .005) response curves in PULP-fed steers for total VFA, acetate, and propionate compared with other treatments. These data indicate that energy supplementation decreased forage intake, altered ruminal fill, shifted VFA patterns, and changed in situ rate of forage CP degradation. Moreover, PULP maintained NDF digestibilities equal to CTRL levels, whereas CORN and BAR did not.