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Inhibitors of the mechanistic target of rapamycin (mTOR) are currently used to treat advanced metastatic breast cancer. However, whether an aggressive phenotype is sustained through adaptation or resistance to mTOR inhibition remains unknown. Here, complementary studies in human tumors, cancer models and cell lines reveal transcriptional reprogramming that supports metastasis in response to mTOR inhibition. This cancer feature is driven by EVI1 and SOX9. EVI1 functionally cooperates with and positively regulates SOX9, and promotes the transcriptional upregulation of key mTOR pathway components (REHB and RAPTOR) and of lung metastasis mediators (FSCN1 and SPARC). The expression of EVI1 and SOX9 is associated with stem cell-like and metastasis signatures, and their depletion impairs the metastatic potential of breast cancer cells. These results establish the mechanistic link between resistance to mTOR inhibition and cancer metastatic potential, thus enhancing our understanding of mTOR targeting failure.

Malignant progression in cancer requires populations of tumor-initiating cells (TICs) endowed with unlimited self renewal, survival under stress, and establishment of distant metastases. Additionally, the acquisition of invasive properties driven by epithelial-mesenchymal transition (EMT) is critical for the evolution of neoplastic cells into fully metastatic populations. Here, we characterize 2 human cellular models derived from prostate and bladder cancer cell lines to better understand the relationship between TIC and EMT programs in local invasiveness and distant metastasis. The model tumor subpopulations that expressed a strong epithelial gene program were enriched in highly metastatic TICs, while a second subpopulation with stable mesenchymal traits was impoverished in TICs. Constitutive overexpression of the transcription factor Snai1 in the epithelial/TIC-enriched populations engaged a mesenchymal gene program and suppressed their self renewal and metastatic phenotypes. Conversely, knockdown of EMT factors in the mesenchymal-like prostate cancer cell subpopulation caused a gain in epithelial features and properties of TICs. Both tumor cell subpopulations cooperated so that the nonmetastatic mesenchymal-like prostate cancer subpopulation enhanced the in vitro invasiveness of the metastatic epithelial subpopulation and, in vivo, promoted the escape of the latter from primary implantation sites and accelerated their metastatic colonization. Our models provide new insights into how dynamic interactions among epithelial, self-renewal, and mesenchymal gene programs determine the plasticity of epithelial TICs. [ABSTRACT FROM AUTHOR]

Prostate cancer exhibits high prevalence and accounts for a high number of cancer-related deaths. The discovery and characterization of molecular determinants of aggressive prostate cancer represents an active area of research. The Immediate Early Response (IER) family of genes, which regulate Protein Phosphatase 2A (PP2A) activity, has emerged among the factors that influence cancer biology. Here, we show that the less studied member of this family, Immediate Early Response 5 like (IER5L), is upregulated in aggressive prostate cancer. Interestingly, the upregulation of IER5L expression exhibits a robust association with metastatic disease in prostate and is recapitulated in other cancer types. In line with this observation, IER5L silencing reduces foci formation, migration and invasion ability in a variety of human and murine prostate cancer cell lines. In vivo, using zebrafish and immunocompromised mouse models, we demonstrate that IER5L-silencing reduces prostate cancer tumor growth, dissemination, and metastasis. Mechanistically, we characterize the transcriptomic and proteomic landscapes of IER5L-silenced cells. This approach allowed us to identify DNA replication and monomeric G protein regulators as downstream programs of IER5L through a pathway that is consistent with the regulation of PP2A. In sum, we report the alteration of IER5L in prostate cancer and beyond and provide biological and molecular evidence of its contribution to tumor aggressiveness., (© 2024. The Author(s).)

MAF amplification increases the risk of breast cancer (BCa) metastasis through mechanisms that are still poorly understood yet have important clinical implications. Oestrogen-receptor-positive (ER + ) BCa requires oestrogen for both growth and metastasis, albeit by ill-known mechanisms. Here we integrate proteomics, transcriptomics, epigenomics, chromatin accessibility and functional assays from human and syngeneic mouse BCa models to show that MAF directly interacts with oestrogen receptor alpha (ERα), thereby promoting a unique chromatin landscape that favours metastatic spread. We identify metastasis-promoting genes that are de novo licensed following oestrogen exposure in a MAF-dependent manner. The histone demethylase KDM1A is key to the epigenomic remodelling that facilitates the expression of the pro-metastatic MAF/oestrogen-driven gene expression program, and loss of KDM1A activity prevents this metastasis. We have thus determined that the molecular basis underlying MAF/oestrogen-mediated metastasis requires genetic, epigenetic and hormone signals from the systemic environment, which influence the ability of BCa cells to metastasize., (© 2023. The Author(s).)

A substantial proportion of cancer patients do not benefit from platinum-based chemotherapy (CT) due to the emergence of drug resistance. Here, we apply elemental imaging to the mapping of CT biodistribution after therapy in residual colorectal cancer and achieve a comprehensive analysis of the genetic program induced by oxaliplatin-based CT in the tumor microenvironment. We show that oxaliplatin is largely retained by cancer-associated fibroblasts (CAFs) long time after the treatment ceased. We determine that CT accumulation in CAFs intensifies TGF-beta activity, leading to the production of multiple factors enhancing cancer aggressiveness. We establish periostin as a stromal marker of chemotherapeutic activity intrinsically upregulated in consensus molecular subtype 4 (CMS4) tumors and highly expressed before and/or after treatment in patients unresponsive to therapy. Collectively, our study underscores the ability of CT-retaining CAFs to support cancer progression and resistance to treatment., (© 2023. The Author(s).)

In the last four decades, treatment of oestrogen receptor positive (ER+) breast cancer (BCa), has focused on targeting the estrogenic receptor signaling pathway. This signaling function is pivotal to sustain cell proliferation. Tamoxifen, a competitive inhibitor of oestrogen, has played a major role in therapeutics. However, primary and acquired resistance to hormone blockade occurs in a large subset of these cancers, and new approaches are urgently needed. Aromatase inhibitors and receptor degraders were approved and alternatively used. Yet, resistance appears in the metastatic setting. Here we report the design and synthesis of a series of proteolysis targeting chimeras (PROTACs) that induce the degradation of estrogen receptor alpha in breast cancer MCF-7 (ER+) cells at nanomolar concentration. Using a warhead based on 4-hydroxytamoxifen, bifunctional degraders recruiting either cereblon or the Von Hippel Lindau E3 ligases were synthesized. Our efforts resulted in the discovery of TVHL-1, a potent ERα degrader (DC 50 : 4.5 nM) that we envisage as a useful tool for biological study and a platform for potential therapeutics., 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 © 2022. Published by Elsevier Masson SAS.)

About 50% of human epidermal growth factor receptor 2 (HER2)+ breast cancer patients do not benefit from HER2-targeted therapy and almost 20% of them relapse after treatment. Here, we conduct a detailed analysis of two independent cohorts of HER2+ breast cancer patients treated with trastuzumab to elucidate the mechanisms of resistance to anti-HER2 monoclonal antibodies. In addition, we develop a fully humanized immunocompetent model of HER2+ breast cancer recapitulating ex vivo the biological processes that associate with patients' response to treatment. Thanks to these two approaches, we uncover a population of TGF-beta-activated cancer-associated fibroblasts (CAF) specific from tumors resistant to therapy. The presence of this cellular subset related to previously described myofibroblastic (CAF-S1) and podoplanin+ CAF subtypes in breast cancer associates with low IL2 activity. Correspondingly, we find that stroma-targeted stimulation of IL2 pathway in unresponsive tumors restores trastuzumab anti-cancer efficiency. Overall, our study underscores the therapeutic potential of exploiting the tumor microenvironment to identify and overcome mechanisms of resistance to anti-cancer treatment., (© 2022. The Author(s).)

Genetic studies suggest that sequential dissemination from a primary metastasis, usually at the bone, is a major route of metastatic progression in early, radically resected cancer. Disseminated tumor cells (DTCs) can likely infiltrate but not grow, and may remain dormant once disseminated for extended intervals (from months to decades). The stationary nature of DTCs prevents them from being successfully treated as an asymptomatic residual disease in the adjuvant setting; critically, they can eventually relapse, adapt, and develop therapy resistance, causing incurable overt metastasis. Metastatic lesions usually first appear in one tissue, which invigorates metastatic cells for further dissemination to other organs, with a fatal outcome. Clinical and genetic data now indicate that metastatic lesions in one organ can seed secondary metastases in other organs: in other words, metastasis arising from metastasis. Herein we discuss recent insight into metastasis cell dormancy mechanisms, survival, communication with the local microenvironment, and eventual changes that endow DTCs with the capacity to expand and colonize to other metastatic sites., Competing Interests: Declaration of interests R.R.G. declares shares of Inbiomotion and has patents and patents pending related to MAF biomarker work., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Ligand-dependent corepressor (LCOR) mediates normal and malignant breast stem cell differentiation. Cancer stem cells (CSCs) generate phenotypic heterogeneity and drive therapy resistance, yet their role in immunotherapy is poorly understood. Here we show that immune-checkpoint blockade (ICB) therapy selects for LCOR low CSCs with reduced antigen processing/presentation machinery (APM) driving immune escape and ICB resistance in triple-negative breast cancer (TNBC). We unveil an unexpected function of LCOR as a master transcriptional activator of APM genes binding to IFN-stimulated response elements (ISREs) in an IFN signaling-independent manner. Through genetic modification of LCOR expression, we demonstrate its central role in modulation of tumor immunogenicity and ICB responsiveness. In TNBC, LCOR associates with ICB clinical response. Importantly, extracellular vesicle (EV) Lcor-messenger RNA therapy in combination with anti-PD-L1 overcame resistance and eradicated breast cancer metastasis in preclinical models. Collectively, these data support LCOR as a promising target for enhancement of ICB efficacy in TNBC, by boosting of tumor APM independently of IFN., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Background: The Adjuvant Zoledronic Acid (ZA) study in early breast cancer (AZURE) showed correlation between a nonamplified MAF gene in the primary tumor and benefit from adjuvant ZA. Adverse ZA outcomes occurred in MAF-amplified patients. NSABP B-34 is a validation study., Methods: A retrospective analysis of MAF gene status in NSABP B-34 was performed. Eligible patients were randomly assigned to standard adjuvant systemic treatment plus 3 years oral clodronate (1600 mg/daily) or placebo. Tumors were tested for MAF gene amplification and analyzed for their relationship to clodronate for disease-free survival (DFS) and overall survival (OS) in MAF nonamplified patients. All statistical tests were 2-sided ., Results: MAF status was assessed in 2533 available primary tumor samples from 3311 patients. Of these, 37 withdrew consent; in 77 samples, no tumor was found; 536 assays did not meet quality standards, leaving 1883 (77.8%) evaluable for MAF assay by fluorescence in situ hybridization (947 from placebo and 936 from clodronate arms). At 5 years, in MAF nonamplified patients receiving clodronate, DFS improved by 30% (hazard ratio = 0.70, 95% confidence interval = 0.51 to 0.94; P  =   .02). OS improved at 5 years (hazard ratio = 0.59, 95% confidence interval = 0.37 to 0.93; P  =   .02) remaining statistically significant for clodronate throughout study follow-up. Conversely, adjuvant clodronate in women with MAF -amplified tumors was not associated with benefit but rather possible harm in some subgroups. Association between MAF status and menopausal status was not seen., Conclusions: Nonamplified MAF showed statistically significant benefits (DFS and OS) with oral clodronate, supporting validation of the AZURE study., (© The Author(s) 2021. Published by Oxford University Press.)

With most cancer-related deaths resulting from metastasis, the development of new therapeutic approaches against metastatic colorectal cancer (mCRC) is essential to increasing patient survival. The metabolic adaptations that support mCRC remain undefined and their elucidation is crucial to identify potential therapeutic targets. Here, we employed a strategy for the rational identification of targetable metabolic vulnerabilities. This strategy involved first a thorough metabolic characterisation of same-patient-derived cell lines from primary colon adenocarcinoma (SW480), its lymph node metastasis (SW620) and a liver metastatic derivative (SW620-LiM2), and second, using a novel multi-omics integration workflow, identification of metabolic vulnerabilities specific to the metastatic cell lines. We discovered that the metastatic cell lines are selectively vulnerable to the inhibition of cystine import and folate metabolism, two key pathways in redox homeostasis. Specifically, we identified the system xCT and MTHFD1 genes as potential therapeutic targets, both individually and combined, for combating mCRC.

Metastasis is the main cause of death for cancer patients, but our ability to improve clinical outcome first requires a better understanding of the dynamics, cellular mechanisms, and kinetics of metastasis. In prostate cancer (PCa), metastatic tumor cells preferentially colonize to bone. However, a lack of applicable mouse models has limited our ability to study this process accurately. Here, we describe a strategy to bypass this limitation: human PCa cells are injected into immunodeficient mice (at tibia, the left ventricle of heart and the iliac artery). Using this novel technique, the metastatic capabilities of these human PCa cells (e.g., colonization and proliferation potential) can be analyzed in bone with an in vivo imaging system.

Carcinoma development in colorectal cancer is driven by genetic alterations in numerous signaling pathways. Alterations in the RAS-ERK1/2 pathway are associated with the shortest overall survival for patients after diagnosis of colorectal cancer metastatic disease, yet how RAS-ERK signaling regulates colorectal cancer metastasis remains unknown. In this study, we used an unbiased screening approach based on selection of highly liver metastatic colorectal cancer cells in vivo to determine genes associated with metastasis. From this, an ERK1/2-controlled metastatic gene set (EMGS) was defined. EMGS was associated with increased recurrence and reduced survival in patients with colorectal cancer tumors. Higher levels of EMGS expression were detected in the colorectal cancer subsets consensus molecular subtype (CMS)1 and CMS4. ANGPT2 and CXCR4 , two genes within the EMGS, were subjected to gain-of-function and loss-of-function studies in several colorectal cancer cell lines and then tested in clinical samples. The RAS-ERK1/2 axis controlled expression of the cytokine ANGPT2 and the cytokine receptor CXCR4 in colorectal cancer cells, which facilitated development of liver but not lung metastases, suggesting that ANGPT2 and CXCR4 are important for metastatic outgrowth in the liver. CXCR4 controlled the expression of cytokines IL10 and CXCL1, providing evidence for a causal role of IL10 in supporting liver colonization. In summary, these studies demonstrate that amplification of ERK1/2 signaling in KRAS -mutated colorectal cancer cells affects the cytokine milieu of the tumors, possibly affecting tumor-stroma interactions and favoring liver metastasis formation. SIGNIFICANCE: These findings identify amplified ERK1/2 signaling in KRAS -mutated colorectal cancer cells as a driver of tumor-stroma interactions that favor formation of metastases in the liver., (©2020 American Association for Cancer Research.)

Gene dosage is a key defining factor to understand cancer pathogenesis and progression, which requires the development of experimental models that aid better deconstruction of the disease. Here, we model an aggressive form of prostate cancer and show the unconventional association of LKB1 dosage to prostate tumorigenesis. Whereas loss of Lkb1 alone in the murine prostate epithelium was inconsequential for tumorigenesis, its combination with an oncogenic insult, illustrated by Pten heterozygosity, elicited lethal metastatic prostate cancer. Despite the low frequency of LKB1 deletion in patients, this event was significantly enriched in lung metastasis. Modeling the role of LKB1 in cellular systems revealed that the residual activity retained in a reported kinase-dead form, LKB1K78I, was sufficient to hamper tumor aggressiveness and metastatic dissemination. Our data suggest that prostate cells can function normally with low activity of LKB1, whereas its complete absence influences prostate cancer pathogenesis and dissemination., Competing Interests: Disclosures: Dr. Graupera reported, "the Graupera laboratory has research agreements with ArQule and Venthera. None of those have a relationship with cancer or LKB1." Dr. Gomis reported shares of Inbiomotion (<$10,000). No other disclosures were reported., (© 2020 Hermanova et al.)

Organogenesis is directed by coordinated cell proliferation and differentiation programs. The hierarchical networks of transcription factors driving mammary gland development and function have been widely studied. However, the contribution of posttranscriptional gene expression reprogramming remains largely unexplored. The 3' untranslated regions of messenger RNAs (mRNAs) contain combinatorial ensembles of cis-regulatory elements that define transcript-specific regulation of protein synthesis through their cognate RNA binding proteins. We analyze the contribution of the RNA binding cytoplasmic polyadenylation element-binding (CPEB) protein family, which collectively regulate mRNA translation for about 30% of the genome. We find that CPEB2 is required for the integration of hormonal signaling by controlling the protein expression from a subset of ER/PR- regulated transcripts. Furthermore, CPEB2 is critical for the development of ER-positive breast tumors. This work uncovers a previously unknown gene expression regulation level in breast morphogenesis and tumorigenesis, coordinating sequential transcriptional and posttranscriptional layers of gene expression regulation., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

The HER2-enriched (HER2-E) subtype within HER2-positive (HER2+) breast cancer is highly addicted to the HER2 pathway. However, ∼20-60% of HER2+/HER2-E tumors do not achieve a complete response following anti-HER2 therapies. Here we evaluate gene expression data before, during and after neoadjuvant treatment with lapatinib and trastuzumab in HER2+/HER2-E tumors of the PAMELA trial and breast cancer cell lines. Our results reveal that dual HER2 blockade in HER2-E disease induces a low-proliferative Luminal A phenotype both in patient's tumors and in vitro models. These biological changes are more evident in hormone receptor-positive (HR+) disease compared to HR-negative disease. Interestingly, increasing the luminal phenotype with anti-HER2 therapy increased sensitivity to CDK4/6 inhibition. Finally, discontinuation of HER2-targeted therapy in vitro, or acquired resistance to anti-HER2 therapy, leads to restoration of the original HER2-E phenotype. Our findings support the use of maintenance anti-HER2 therapy and the therapeutic exploitation of subtype switching with CDK4/6 inhibition.

Bone metastasis is present in a high percentage of breast cancer (BCa) patients with distant disease, especially in those with the estrogen receptor-positive (ER + ) subtype. Most cells that escape primary tumors are unable to establish metastatic lesions, which suggests that target organ microenvironments are hostile for tumor cells. This implies that BCa cells must achieve a process of speciation to adapt to the new conditions imposed in the new organ. Bone has unique characteristics that can be exploited by cancer cells: it undergoes constant remodeling and comprises diverse environments (including osteogenic, perivascular, and hematopoietic stem cell niches). This allows colonizing cells to take advantage of numerous adhesion molecules, matrix proteins, and soluble factors that facilitate homing, survival, and, eventually, metastatic outgrowth. However, in most cases, metastatic lesions enter into a latency state that can last months, years, or even decades, before forming a clinically detectable macrometastasis. This dormant state challenges the effectiveness of adjuvant chemotherapy. Detecting which tumors are more prone to metastasize to bone and developing new specific therapies that target bone metastasis represent urgent clinical needs. Here, we review the biological mechanisms of BCa bone metastasis and provide the latest options of treatments and predictive markers that are currently in clinical use or are being tested in clinical assays. © 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland., (© 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.)

TP53INP2 positively regulates autophagy by binding to Atg8 proteins. Here, we uncover a novel role of TP53INP2 in death-receptor signaling. TP53INP2 sensitizes cells to apoptosis induced by death receptor ligands. In keeping with this, TP53INP2 deficiency in cultured cells or mouse livers protects against death receptor-induced apoptosis. TP53INP2 binds caspase-8 and the ubiquitin ligase TRAF6, thereby promoting the ubiquitination and activation of caspase-8 by TRAF6. We have defined a TRAF6-interacting motif (TIM) and a ubiquitin-interacting motif in TP53INP2, enabling it to function as a scaffold bridging already ubiquitinated caspase-8 to TRAF6 for further polyubiquitination of caspase-8. Mutations of key TIM residues in TP53INP2 abrogate its interaction with TRAF6 and caspase-8, and subsequently reduce levels of death receptor-induced apoptosis. A screen of cancer cell lines showed that those with higher protein levels of TP53INP2 are more prone to TRAIL-induced apoptosis, making TP53INP2 a potential predictive marker of cancer cell responsiveness to TRAIL treatment. These findings uncover a novel mechanism for the regulation of caspase-8 ubiquitination and reveal TP53INP2 as an important regulator of the death receptor pathway., (© 2019 The Authors.)

With the advent of OMICs technologies, both individual research groups and consortia have spear-headed the characterization of human samples of multiple pathophysiologic origins, resulting in thousands of archived genomes and transcriptomes. Although a variety of web tools are now available to extract information from OMICs data, their utility has been limited by the capacity of nonbioinformatician researchers to exploit the information. To address this problem, we have developed CANCERTOOL, a web-based interface that aims to overcome the major limitations of public transcriptomics dataset analysis for highly prevalent types of cancer (breast, prostate, lung, and colorectal). CANCERTOOL provides rapid and comprehensive visualization of gene expression data for the gene(s) of interest in well-annotated cancer datasets. This visualization is accompanied by generation of reports customized to the interest of the researcher (e.g., editable figures, detailed statistical analyses, and access to raw data for reanalysis). It also carries out gene-to-gene correlations in multiple datasets at the same time or using preset patient groups. Finally, this new tool solves the time-consuming task of performing functional enrichment analysis with gene sets of interest using up to 11 different databases at the same time. Collectively, CANCERTOOL represents a simple and freely accessible interface to interrogate well-annotated datasets and obtain publishable representations that can contribute to refinement and guidance of cancer-related investigations at all levels of hypotheses and design. Significance: In order to facilitate access of research groups without bioinformatics support to public transcriptomics data, we have developed a free online tool with an easy-to-use interface that allows researchers to obtain quality information in a readily publishable format. Cancer Res; 78(21); 6320-8. ©2018 AACR ., (©2018 American Association for Cancer Research.)

In the version of this Article originally published, the boxes framing the two plots in Fig. 1g were misaligned from the axes due to a technical error. This has now been corrected in all versions of the Article.

Breast cancer is the second leading cause of cancer-related death among women. Here we report a role for the protein kinase p38α in coordinating the DNA damage response and limiting chromosome instability during breast tumor progression, and identify the DNA repair regulator CtIP as a p38α substrate. Accordingly, decreased p38α signaling results in impaired ATR activation and homologous recombination repair, with concomitant increases in replication stress, DNA damage, and chromosome instability, leading to cancer cell death and tumor regression. Moreover, we show that pharmacological inhibition of p38α potentiates the effects of taxanes by boosting chromosome instability in murine models and patient-derived xenografts, suggesting the potential interest of combining p38α inhibitors with chemotherapeutic drugs that induce chromosome instability., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Gene expression profiling has had a considerable impact on our understanding ofbreastcancer biology. Duringthelast decade, 4 intrinsic molecular subtypes of breast cancer (Luminal A, Luminal B, HER2-enriched [HER2-E] and Basal-like) have been identified and intensively studied. In this article, we review and discuss the clinical implications of the 2 non-luminal subtypes (i.e. HER2-E and Basal-like) identified within hormone receptor (HR)-positive disease. After reviewing 32 studies for a total of 13,091 samples, ∼8% and ∼ 15% of early and metastatic HR+/HER2-negative breast cancer, respectively, were found to be non-luminal. Clinically, HR+/HER2-negative/non-luminal subtypes have been associated with estrogen independence, chemo-sensitivity, resistance to CDK4/6 inhibition and poor outcome. Interestingly, EGFR/HER2 tyrosine kinase inhibition might be of value in the HR+/HER2-negative/HER2-E subtype. Finally, the HER2-E subtype within HR+/HER2 + disease represents ∼ 30% and has been associated with anti-HER2 sensitivity, chemo-sensitivity and resistance to CDK4/6 inhibition. In the upcoming years, retrospective and prospective clinical trials evaluating both biomarkers should lead to improvements in patient outcomes., (Copyright © 2018 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

For many patients with breast cancer, symptomatic bone metastases appear after years of latency. How micrometastatic lesions remain dormant and undetectable before initiating colonization is unclear. Here, we describe a mechanism involved in bone metastatic latency of oestrogen receptor-positive (ER + ) breast cancer. Using an in vivo genome-wide short hairpin RNA screening, we identified the kinase MSK1 as an important regulator of metastatic dormancy in breast cancer. In patients with ER + breast cancer, low MSK1 expression associates with early metastasis. We show that MSK1 downregulation impairs the differentiation of breast cancer cells, increasing their bone homing and growth capacities. MSK1 controls the expression of genes required for luminal cell differentiation, including the GATA3 and FOXA1 transcription factors, by modulating their promoter chromatin status. Our results indicate that MSK1 prevents metastatic progression of ER + breast cancer, suggesting that stratifying patients with breast cancer as high or low risk for early relapse based on MSK1 expression could improve prognosis.

Mammary stem and progenitor cells are essential for mammary gland homeostasis and are also candidates for cells of origin of mammary tumors. Here, we have investigated the function of the protein kinase p38α in the mammary gland using mice that delete this protein in the luminal epithelial cells. We show that p38α regulates the fate of luminal progenitor cells through modulation of the transcription factor RUNX1, an important controller of the estrogen receptor-positive cell lineage. We also provide evidence that the regulation of RUNX1 by p38α probably involves the kinase MSK1, which phosphorylates histone H3 at the RUNX1 promoter. Moreover, using a mouse model for breast cancer initiated by luminal cells, we show that p38α downregulation in mammary epithelial cells reduces tumor burden, which correlates with decreased numbers of tumor-initiating cells. Collectively, our results define a key role for p38α in luminal progenitor cell fate that affects mammary tumor formation., (Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.)

Transforming growth factor-β (TGF-β) induces a pleiotropic pathway that is modulated by the cellular context and its integration with other signaling pathways. In cancer, the pleiotropic reaction to TGF-β leads to a diverse and varied set of gene responses that range from cytostatic and apoptotic tumor-suppressive ones in early stage tumors, to proliferative, invasive, angiogenic, and oncogenic ones in advanced cancer. Here, we review the knowledge accumulated about the molecular mechanisms involved in the dual response to TGF-β in cancer, and how tumor cells evolve to evade the tumor-suppressive responses of this signaling pathway and then hijack the signal, converting it into an oncogenic factor. Only through the detailed study of this complexity can the suitability of the TGF-β pathway as a therapeutic target against cancer be evaluated., (Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.)

Background: Adjuvant use of bisphosphonates can reduce the incidence of bone metastases in early breast cancer. Recurrence and survival seem to be improved only in postmenopausal patients, but the underlying mechanisms remain unclear. We investigated whether MAF amplification (a biomarker for bone metastasis) in primary tumours could predict the treatment outcomes of adjuvant zoledronic acid., Methods: The study population included patients enrolled in the international, open-label, randomised, controlled, phase 3 AZURE trial at eligible UK sites who had stage II or III breast cancer and who gave consent for use of their primary tumour samples. Patients were randomly assigned (1:1) to receive standard adjuvant systemic therapy alone (control group) or with zoledronic acid every 3-4 weeks for six doses, then every 3-6 months until the end of 5 years. Minimisation took into account the number of involved axillary lymph nodes, clinical tumour stage, oestrogen-receptor status, type and timing of systemic therapy, menopausal status, statin use, and treating centre. The primary endpoint was disease-free survival; the secondary endpoint, invasive-disease-free survival, was the primary disease endpoint for the analysis in this report. MAF amplification was assessed by fluorescence in-situ hybridisation of two cores of breast tumour tissue in a microarray, done in a central laboratory by technicians unaware of treatment assignment. We used multivariate analyses to assess disease outcomes by intention to treat. We also assessed interactions between MAF-positive status and menopausal status on efficacy of zoledronic acid. The AZURE trial is registered with the International Standard Randomised Controlled Trial Registry, number ISRCTN79831382., Findings: 1739 AZURE patients contributed primary tumour samples, of whom 865 (50%) had two assessable cores (445 in the control groups and 420 in the zoledronic acid group). 184 (21%) tumours were MAF positive (85 in the control groups and 99 in the zoledronic acid group) and the remaining tumours were MAF negative. At a median follow-up of 84·6 months (IQR 72·0-95·8), MAF status was not prognostic for invasive-disease-free survival in the control group (MAF-positive vs MAF-negative: hazard ratio [HR] 0·92, 95% CI 0·59-1·41), but was in the zoledronic acid group (0·52, 0·36-0·75). In patients with MAF-negative tumours, zoledronic acid was associated with higher invasive-disease-free survival than was control treatment (HR 0·74, 95% CI 0·56-0·98), but not in patients who had MAF-positive tumours. Additionally, among 121 patients not postmenopausal at randomisation with MAF-positive tumours, zoledronic acid was associated with lower invasive-disease-free survival (HR 2·47, 95% CI 1·23-4·97) and overall survival (2·27, 95% CI 1·04-4·93) than control treatment., Interpretation: MAF status can predict likelihood of benefit from adjuvant zoledronic acid and merits further investigation as a potential companion diagnostic., Funding: Novartis Global and Inbiomotion., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Biological changes that occur during metastatic progression of breast cancer are still incompletely characterized. In this study, we compared intrinsic molecular subtypes and gene expression in 123 paired primary and metastatic tissues from breast cancer patients. Intrinsic subtype was identified using a PAM50 classifier and χ 2 tests determined the differences in variable distribution. The rate of subtype conversion was 0% in basal-like tumors, 23.1% in HER2-enriched (HER2-E) tumors, 30.0% in luminal B tumors, and 55.3% in luminal A tumors. In 40.2% of cases, luminal A tumors converted to luminal B tumors, whereas in 14.3% of cases luminal A and B tumors converted to HER2-E tumors. We identified 47 genes that were expressed differentially in metastatic versus primary disease. Metastatic tumors were enriched for proliferation-related and migration-related genes and diminished for luminal-related genes. Expression of proliferation-related genes were better at predicting overall survival in metastatic disease (OSmet) when analyzed in metastatic tissue rather than primary tissue. In contrast, a basal-like gene expression signature was better at predicting OSmet in primary disease compared with metastatic tissue. We observed correlations between time to tumor relapse and the magnitude of changes of proliferation, luminal B, or HER2-E signatures in metastatic versus primary disease. Although the intrinsic subtype was largely maintained during metastatic progression, luminal/HER2-negative tumors acquired a luminal B or HER2-E profile during metastatic progression, likely reflecting tumor evolution or acquisition of estrogen independence. Overall, our analysis revealed the value of stratifying gene expression by both cancer subtype and tissue type, providing clinicians more refined tools to evaluate prognosis and treatment. Cancer Res; 77(9); 2213-21. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Background: Activated protein C/endothelial protein C receptor (APC/EPCR) axis is physiologically involved in anticoagulant and cytoprotective activities in endothelial cells. Emerging evidence indicates that EPCR also plays a role in breast stemness and human tumorigenesis. Yet, its contribution to breast cancer progression and metastasis has not been elucidated., Methods: Transcriptomic status of EPCR was examined in a cohort of 286 breast cancer patients. Cell growth kinetics was evaluated in control and EPCR and SPARC/osteonectin, Cwcv, and kazal-like domains proteoglycan (SPOCK1/testican 1) silenced breast cancer cells in 2D, 3D, and in co-culture conditions. Orthotopic tumor growth and lung and osseous metastases were evaluated in several human and murine xenograft breast cancer models. Tumor-stroma interactions were further studied in vivo by immunohistochemistry and flow cytometry. An EPCR-induced gene signature was identified by microarray analysis., Results: Analysis of a cohort of breast cancer patients revealed an association of high EPCR levels with adverse clinical outcome. Interestingly, EPCR knockdown did not affect cell growth kinetics in 2D but significantly reduced cell growth in 3D cultures. Using several human and murine xenograft breast cancer models, we showed that EPCR silencing reduced primary tumor growth and secondary outgrowths at metastatic sites, including the skeleton and the lungs. Interestingly, these effects were independent of APC ligand stimulation in vitro and in vivo. Transcriptomic analysis of EPCR-silenced tumors unveiled an effect mediated by matricellular secreted proteoglycan SPOCK1/testican 1. Interestingly, SPOCK1 silencing suppressed in vitro 3D growth. Moreover, SPOCK1 ablation severely decreased orthotopic tumor growth and reduced bone metastatic osteolytic tumors. High SPOCK1 levels were also associated with poor clinical outcome in a subset breast cancer patients. Our results suggest that EPCR through SPOCK1 confers a cell growth advantage in 3D promoting breast tumorigenesis and metastasis., Conclusions: EPCR represents a clinically relevant factor associated with poor outcome and a novel vulnerability to develop combination therapies for breast cancer patients.

Metastasis is the primary cause of death in cancer patients and current treatments fail to provide durable responses. Efforts to treat metastatic disease are hindered by the fact that metastatic cells often remain dormant for prolonged intervals of years, or even decades. Tumor dormancy reflects the capability of disseminated tumor cells (DTCs), or micrometastases, to evade treatment and remain at low numbers after primary tumor resection. Unfortunately, dormant cells will eventually produce overt metastasis. Innovations are needed to understand metastatic dormancy and improve cancer detection and treatment. Currently, few models exist that faithfully recapitulate metastatic dormancy and metastasis to clinically relevant tissues, such as the bone. Herein, we discuss recent advances describing genetic cell-autonomous and systemic or local changes in the microenvironment that have been shown to endow DTCs with properties to survive and eventually colonize distant organs., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)

Patient stratification has been instrumental for the success of targeted therapies in breast cancer. However, the molecular basis of metastatic breast cancer and its therapeutic vulnerabilities remain poorly understood. Here we show that PML is a novel target in aggressive breast cancer. The acquisition of aggressiveness and metastatic features in breast tumours is accompanied by the elevated PML expression and enhanced sensitivity to its inhibition. Interestingly, we find that STAT3 is responsible, at least in part, for the transcriptional upregulation of PML in breast cancer. Moreover, PML targeting hampers breast cancer initiation and metastatic seeding. Mechanistically, this biological activity relies on the regulation of the stem cell gene SOX9 through interaction of PML with its promoter region. Altogether, we identify a novel pathway sustaining breast cancer aggressiveness that can be therapeutically exploited in combination with PML-based stratification.

Tumour cell-secreted factors skew infiltrating immune cells towards a tumour-supporting phenotype, expressing pro-tumourigenic mediators. However, the influence of lipocalin-2 (Lcn2) on the metastatic cascade in the tumour micro-environment is still not clearly defined. Here, we explored the role of stroma-derived, especially macrophage-released, Lcn2 in breast cancer progression. Knockdown studies and neutralizing antibody approaches showed that Lcn2 contributes to the early events of metastasis in vitro. The release of Lcn2 from macrophages induced an epithelial-mesenchymal transition programme in MCF-7 breast cancer cells and enhanced local migration as well as invasion into the extracellular matrix, using a three-dimensioanl (3D) spheroid model. Moreover, a global Lcn2 deficiency attenuated breast cancer metastasis in both the MMTV-PyMT breast cancer model and a xenograft model inoculating MCF-7 cells pretreated with supernatants from wild-type and Lcn2-knockdown macrophages. To dissect the role of stroma-derived Lcn2, we employed an orthotopic mammary tumour mouse model. Implantation of wild-type PyMT tumour cells into Lcn2-deficient mice left primary mammary tumour formation unaltered, but specifically reduced tumour cell dissemination into the lung. We conclude that stroma-secreted Lcn2 promotes metastasis in vitro and in vivo, thereby contributing to tumour progression. Our study highlights the tumourigenic potential of stroma-released Lcn2 and suggests Lcn2 as a putative therapeutic target. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Cellular transformation and cancer progression is accompanied by changes in the metabolic landscape. Master co-regulators of metabolism orchestrate the modulation of multiple metabolic pathways through transcriptional programs, and hence constitute a probabilistically parsimonious mechanism for general metabolic rewiring. Here we show that the transcriptional co-activator peroxisome proliferator-activated receptor gamma co-activator 1α (PGC1α) suppresses prostate cancer progression and metastasis. A metabolic co-regulator data mining analysis unveiled that PGC1α is downregulated in prostate cancer and associated with disease progression. Using genetically engineered mouse models and xenografts, we demonstrated that PGC1α opposes prostate cancer progression and metastasis. Mechanistically, the use of integrative metabolomics and transcriptomics revealed that PGC1α activates an oestrogen-related receptor alpha (ERRα)-dependent transcriptional program to elicit a catabolic state and metastasis suppression. Importantly, a signature based on the PGC1α-ERRα pathway exhibited prognostic potential in prostate cancer, thus uncovering the relevance of monitoring and manipulating this pathway for prostate cancer stratification and treatment.

The mechanisms that allow breast cancer (BCa) cells to metabolically sustain rapid growth are poorly understood. Here we report that BCa cells are dependent on a mechanism to supply precursors for intracellular lipid production derived from extracellular sources and that the endothelial lipase (LIPG) fulfils this function. LIPG expression allows the import of lipid precursors, thereby contributing to BCa proliferation. LIPG stands out as an essential component of the lipid metabolic adaptations that BCa cells, and not normal tissue, must undergo to support high proliferation rates. LIPG is ubiquitously and highly expressed under the control of FoxA1 or FoxA2 in all BCa subtypes. The downregulation of either LIPG or FoxA in transformed cells results in decreased proliferation and impaired synthesis of intracellular lipids.

Drug delivery across the blood-brain barrier (BBB) is a formidable challenge for therapies targeting the central nervous system. Although BBB shuttle peptides enhance transport into the brain non-invasively, their application is partly limited by lability to proteases. The present study proposes the use of cyclic peptides derived from venoms as an affordable way to circumvent this drawback. Apamin, a neurotoxin from bee venom, was minimized by reducing its complexity, toxicity, and immunogenicity, while preserving brain targeting, active transport, and protease resistance. Among the analogues designed, the monocyclic lactam-bridged peptidomimetic MiniAp-4 was the most permeable. This molecule is capable of translocating proteins and nanoparticles in a human-cell-based BBB model. Furthermore, MiniAp-4 can efficiently deliver a cargo across the BBB into the brain parenchyma of mice., (© 2015 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA. This is an open access article under the terms of the Creative Commons Attribution Non-Commercial NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.)

Background: There are currently no biomarkers for early breast cancer patient populations at risk of bone metastasis. Identification of mediators of bone metastasis could be of clinical interest., Methods: A de novo unbiased screening approach based on selection of highly bone metastatic breast cancer cells in vivo was used to determine copy number aberrations (CNAs) associated with bone metastasis. The CNAs associated with bone metastasis were examined in independent primary breast cancer datasets with annotated clinical follow-up. The MAF gene encoded within the CNA associated with bone metastasis was subjected to gain and loss of function validation in breast cancer cells (MCF7, T47D, ZR-75, and 4T1), its downstream mechanism validated, and tested in clinical samples. A multivariable Cox cause-specific hazard model with competing events (death) was used to test the association between 16q23 or MAF and bone metastasis. All statistical tests were two-sided., Results: 16q23 gain CNA encoding the transcription factor MAF mediates breast cancer bone metastasis through the control of PTHrP. 16q23 gain (hazard ratio (HR) for bone metastasis = 14.5, 95% confidence interval (CI) = 6.4 to 32.9, P < .001) as well as MAF overexpression (HR for bone metastasis = 2.5, 95% CI = 1.7 to 3.8, P < .001) in primary breast tumors were specifically associated with risk of metastasis to bone but not to other organs., Conclusions: These results suggest that MAF is a mediator of breast cancer bone metastasis. 16q23 gain or MAF protein overexpression in tumors may help to select patients at risk of bone relapse., (© The Author 2015. Published by Oxford University Press.)

Mammary cancer stem cells (MCSC) have been operationally defined as cells that re-form secondary tumors upon transplantation into immunodeficient mice. Building on this observation, it has also been suggested that MCSCs are responsible for metastasis as well as evasion and resistance to therapeutic treatments. MCSC reinitiating potential is usually tested by implantation of limited amounts of cells orthotopically or subcutaneously, yet this poorly recapitulates the metastatic niche where truly metastatic reinitiation will occur. Herein, we describe the implantation of small amounts of MCSC selected populations in the bone (intra tibiae injection) and the lung (intra thoracic injection) to test for their metastatic reinitiation capabilities.

In estrogen receptor-negative breast cancer patients, metastatic relapse usually occurs in the lung and is responsible for the fatal outcome of the disease. Thus, a better understanding of the biology of metastasis is needed. In particular, biomarkers to identify patients that are at risk of lung metastasis could open the avenue for new therapeutic opportunities. Here we characterize the biological activity of RARRES3, a new metastasis suppressor gene whose reduced expression in the primary breast tumors identifies a subgroup of patients more likely to develop lung metastasis. We show that RARRES3 downregulation engages metastasis-initiating capabilities by facilitating adhesion of the tumor cells to the lung parenchyma. In addition, impaired tumor cell differentiation due to the loss of RARRES3 phospholipase A1/A2 activity also contributes to lung metastasis. Our results establish RARRES3 downregulation as a potential biomarker to identify patients at high risk of lung metastasis who might benefit from a differentiation treatment in the adjuvant programme., (© 2014 The Authors. Published under the terms of the CC BY 4.0 license.)

The mechanisms that allow colon cancer cells to form liver and lung metastases, and whether KRAS mutation influences where and when metastasis occurs, are unknown. We provide clinical and molecular evidence showing that different MAPK signalling pathways are implicated in this process. Whereas ERK2 activation provides colon cancer cells with the ability to seed and colonize the liver, reduced p38 MAPK signalling endows cancer cells with the ability to form lung metastasis from previously established liver lesions. Downregulation of p38 MAPK signalling results in increased expression of the cytokine PTHLH, which contributes to colon cancer cell extravasation to the lung by inducing caspase-independent death in endothelial cells of the lung microvasculature. The concerted acquisition of metastatic traits in the colon cancer cells together with the sequential colonization of liver and lung highlights the importance of metastatic lesions as a platform for further dissemination.

Matrix metalloproteinases (MMPs) have been traditionally implicated in cancer progression because of their ability to degrade the extracellular matrix. However, some members of the MMP family have recently been identified as proteases with antitumor properties. Thus, it has been described that collagenase-2 (MMP-8) has a protective role in tumor and metastasis progression, but the molecular mechanisms underlying these effects are unknown. We show herein that Mmp8 expression causes a decrease in miR-21 levels that in turn leads to a reduction in tumor growth and lung metastasis formation by MDA-MB-231 (4175) breast cancer cells. By using both in vitro and in vivo models, we demonstrate that the mechanism responsible for these MMP-8 beneficial effects involves cleavage of decorin by MMP-8 and a subsequent reduction of transforming growth factor β (TGF-β) signaling that controls miR-21 levels. In addition, miR-21 downregulation induced by MMP-8 increases the levels of tumor suppressors such as programmed cell death 4, which may also contribute to the decrease in tumor formation and metastasis of breast cancer cells overexpressing this metalloproteinase. These findings reveal a new signaling pathway for cancer regulation controlled by MMP-8, and contribute to clarify the molecular mechanisms by which tumor-defying proteases may exert their protective function in cancer and metastasis.

Objective: To assess whether celecoxib, a selective cyclooxygenase-2 (COX-2) inhibitor with anti-cancer properties, has an inhibitory effect on tumour establishment and progression of prostate cancer (PCa) bone metastases., Materials and Methods: PC-3 stable luciferase-expressing cells were injected into male nude mice by intracardiac (i.c.) and intratibial (i.t.) injections, and the effect of celecoxib on bone metastases was then recorded using bioluminescence image analysis. In cases of chemoprevention, mice received 3 mg/kg celecoxib from 1 week before cell implantation until the end of the study, and to test the therapeutic effect, mice received celecoxib 1 week after cell implantation until the end of the study. Tumour tissue samples were histologically examined and COX-2 expression was quantified at the protein level., Results: Celecoxib significantly decreased cell viability and the proliferation of human PCa cells in vitro in a dose-dependent manner. Bone metastases were detected after i.c. injection in nude mice. Celecoxib (15 ppm) administered before i.c. injection did not inhibit the cellular metastatic potential, as the numbers of bone metastases were similar in both groups. However, celecoxib did decrease metastatic progression in the osseous environment when cells were injected directly into the tibia (P < 0.05). At the protein level, COX-2 expression was significantly decreased in the celecoxib treatment group (P < 0.01)., Conclusion: In a preclinical mice model, celecoxib administered orally at the standard human dose inhibits the progression of established PCa bone metastases., (© 2013 The Authors. BJU International © 2013 BJU International.)