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Ewing sarcoma (EwS) is an aggressive tumor that affects adolescents and young adults. EwS is defined by a chromosomal translocation, EWSR1-FLI1 being the most common, that causes genome reprogramming through remodeling of enhancers. Here, we describe an unexpected function of RING1B, which is highly expressed in EwS. While retaining its repressive activity at Polycomb developmental regulated genes, RING1B colocalizes with EWSR1-FLI1 at active enhancers. We demonstrate that RING1B is necessary for the expression of key EWSR1-FLI1 targets by facilitating oncogene recruitment to their enhancers. Knockdown of RING1B impairs growth of tumor xenografts and expression of genes regulated by EWSR1-FLI1 bound enhancers. Pharmacological inhibition of AURKB with AZD1152 increases H2Aub levels causing down-regulation of RING1B/EWSR1-FLI1 common targets. Our findings demonstrate that RING1B is a critical modulator of EWSR1-FLI1–induced chromatin remodeling, and its inhibition is a potential therapeutic strategy for the treatment of these tumors.

Chemotherapy is one of the most established and effective treatments for almost all types of cancer. However, the elevated toxicity due to the non-tumor-associated effects, development of secondary malignancies, infertility, radiation-induced fibrosis and resistance to treatment limit the effectiveness and safety of treatment. In addition, these multiple factors significantly impact quality of life. Over the last decades, our increased understanding of cancer epigenetics has led to new therapeutic approaches and the promise of improved patient outcomes. Epigenetic alterations are commonly found in cancer, especially the increased expression and activity of histone deacetylases (HDACs). Dysregulation of HDACs are critical to the development and progression of the majority of tumors. Hence, HDACs inhibitors (HDACis) were developed and now represent a very promising treatment strategy. The use of HDACis as monotherapy has shown very positive pre-clinical results, but clinical trials have had only limited success. However, combinatorial regimens with other cancer drugs have shown synergistic effects both in pre-clinical and clinical studies. At the same time, these combinations have enhanced the efficacy, reduced the toxicity and tumor resistance to therapy. In this review, we will examine examples of HDACis used in combination with other cancer drugs and highlight the synergistic effects observed in recent preclinical and clinical studies.

SNAI2 overexpression appears to be associated with poor prognosis in breast cancer, yet it remains unclear in which breast cancer subtypes this occurs. Here we show that excess SNAI2 is associated with a poor prognosis of luminal B HER2+/ERBB2+ breast cancers in which SNAI2 expression in the stroma but not the epithelium correlates with tumor proliferation. To determine how stromal SNAI2 might influence HER2+ tumor behavior, Snai2-deficient mice were crossed with a mouse line carrying the ErbB2/Neu protooncogene to generate HER2+/ERBB2+ breast cancer. Tumors generated in this model expressed SNAI2 in the stroma but not the epithelium, allowing for the role of stromal SNAI2 to be studied without interference from the epithelial compartment. The absence of SNAI2 in the stroma of HER2+/ERBB2+ tumors is associated with: (i) lower levels of cyclin D1 (CCND1) and reduced tumor epithelium proliferation; (ii) higher levels of AKT and a lower incidence of metastasis; (iii) lower levels of angiopoietin-2 (ANGPT2), and more necrosis. Together, these results indicate that the loss of SNAI2 in cancer-associated fibroblasts limits the production of some cytokines, which influences AKT/ERK tumor signaling and subsequent proliferative and metastatic capacity of ERBB2+ breast cancer cells. Accordingly, SNAI2 expression in the stroma enhanced the tumorigenicity of luminal B HER2+/ERBB2+ breast cancers. This work emphasizes the importance of stromal SNAI2 in breast cancer progression and patients' prognosis.

The presence of the chimeric EWSR1-FLI1 oncoprotein is the main and initiating event defining Ewing sarcoma (ES). The dysregulation of epigenomic and proteomic homeostasis induced by the oncoprotein contributes to a wide variety of events involved in oncogenesis and tumor progression. Attempts at studying the effects of EWSR1-FLI1 in non-tumor cells to understand the mechanisms underlying sarcomagenesis have been unsuccessful to date, as ectopic expression of EWSR1-FLI1 blocks cell cycle progression and induces apoptosis in the tested cell lines. Therefore, it is essential to find a permissive cell type for EWSR1-FLI1 expression that allows its endogenous molecular functions to be studied. Here we have demonstrated that HeLa cell lines are permissive to EWSR1-FLI1 ectopic expression, and that our model substantially recapitulates the endogenous activity of the EWSR1-FLI1 fusion protein. This model could contribute to better understanding ES sarcomagenesis by helping to understand the molecular mechanisms induced by the EWSR1-FLI1 oncoprotein.

[Purpose] Epigenetic regulation is crucial in mammalian development and maintenance of tissue-cell specific functions. Perturbation of epigenetic balance may lead to alterations in gene expression, resulting in cellular transformation and malignancy. Previous studies in Ewing sarcoma (ES) have shown that the Nucleosome Remodeling Deacetylase (NuRD) complex binds directly to EWS-FLI1 oncoprotein and modulates its transcriptional activity. The role of EWS-FLI1 as a driver of proliferation and transformation in ES is widely known, but the effect of epigenetic drugs on fusion activity remains poorly described. The present study evaluated the combination effects of the histone deacetylases inhibitor suberoylanilide hydroxamic acid (SAHA) and Lysine-specific demethylase1 inhibitor (HCI-2509) on different biological functions in ES and in comparison to monotherapy treatments., [Results] The study of proliferation and cell viability showed a synergistic effect in most ES cell lines analyzed. An enhanced effect was also observed in the induction of apoptosis, together with accumulation of cells in G1 phase and a blockage of the migratory capacity of ES cell lines. Treatment, either in monotherapy or in combination, caused a significant decrease of EWS-FLI1 mRNA and protein levels and this effect is mediated in part by fusion gene promoter regulation. The anti-tumor effect of this combination was confirmed in patient-derived xenograft mouse models, in which only the combination treatment led to a statistically significant decrease in tumor volume., [Conclusions] The combination of SAHA and HCI-2509 is proposed as a novel treatment strategy for ES patients to inhibit the essential driver of this sarcoma and tumor growth.

The data presented in this article are related to the research paper entitled “The biological age linked to oxidative stress modifies breast cancer aggressiveness” (M.M. Sáez-Freire, A. Blanco-Gómez, S. Castillo-Lluva, A. Gómez-Vecino, J.M. Galvis-Jiménez, C. Martín-Seisdedos, M. Isidoro-García, L. Hontecillas-Prieto, M.B. García-Cenador, F.J. García-Criado, M.C. Patino-Alonso, P. Galindo-Villardón, J.H. Mao, C. Prieto, A. Castellanos-Martín, L. Kaderali, J. Pérez-Losada). The data shown were obtained from a population of transgenic mice, MMTV-Erbb2/Neu, with different susceptibility to breast cancer and a mixed genetic background generated by backcrossing. It was observed that the aggressiveness of breast cancer negatively correlates with age, being lower in chronologically old mice, similar to what occurs in humans. Given that oxidative stress is associated with tumour susceptibility and the degree of aging, the association between the aggressiveness of breast cancer and multiple intermediate phenotypes directly or indirectly related to oxidative stress was studied. Using a mathematical model, we defined biological age and the degree of aging as the difference between biological and chronological ages. As a result, we observed that biologically old mice predominated among those that developed the disease early on, that is, those that were chronologically young. We then identified the specific and common genetic components of Quantitative Trait loci or QTL associated with different evolution of breast cancer, the intermediate phenotypes related to oxidative stress studied, the biological age and the degree of aging. Lastly, we showed that the expression pattern in the livers of biologically old mice were enriched in signalling pathways related to inflammation and response to infections; whereas the biologically young mice exhibited enriched pathways related to mitochondrial activity. For the explanation and discussion of these data refer to the research article cit

The incidence of breast cancer increases with age until menopause, and breast cancer is more aggressive in younger women. The existence of epidemiological links between breast cancer and aging indicates that both processes share some common mechanisms of development. Oxidative stress is associated with both cancer susceptibility and aging. Here we observed that ERBB2-positive breast cancer, which developed in genetically heterogeneous ERBB2-positive transgenic mice generated by a backcross, is more aggressive in chronologically younger than in older mice (differentiated by the median survival of the cohort that was 79 weeks), similar to what occurs in humans. In this cohort, we estimated the oxidative biological age using a mathematical model that integrated several subphenotypes directly or indirectly related to oxidative stress. The model selected the serum levels of HDL-cholesterol and magnesium and total AKT1 and glutathione concentrations in the liver. The grade of aging was calculated as the difference between the predicted biological age and the chronological age. This comparison permitted the identification of biologically younger and older mice compared with their chronological age. Interestingly, biologically older mice developed more aggressive breast cancer than the biologically younger mice. Genomic regions on chromosomes 2 and 15 linked to the grade of oxidative aging were identified. The levels of expression of Zbp1 located on chromosome 2, a gene related to necroptosis and inflammation, positively correlated with the grade of aging and tumour aggressiveness. Moreover, the pattern of gene expression of genes linked to the inflammation and the response to infection pathways was enriched in the livers of biologically old mice. This study shows part of the complex interactions between breast cancer and aging.

Wilms tumor (WT) is an embryonal malignant neoplasm of the kidney that accounts for 6–7% of all childhood cancers. WT seems to derive from multipotent embryonic renal stem cells that have failed to differentiate properly. Since mechanisms underlying WT tumorigenesis remain largely unknown, the aim of this study was to explore the expression of embryonic stem cell (ESC) markers in samples of WT patients after chemotherapy treatment SIOP protocol, as the gene expression patterns of ESC are like those of most cancer cells. We found that expression of ESC markers is heterogeneous, and depends on histological WT components. Interestingly, among ESC markers, HMGA2 was expressed significantly stronger in the blastemal component than in the stromal and the normal kidney. Moreover, two subsets of patients of WT blastemal type were identified, depending on the expression levels of HMGA2. High HMGA2 expression levels were significantly associated with a higher proliferation rate (p=0.0345) and worse patient prognosis (p=0.0289). The expression of HMGA2 was a stage-independent factor of clinical outcome in blastemal WT patients. Our multivariate analyses demonstrated the association between LIN28B– LET7A–HMGA2 expression, and the positive correlation between HMGA2 and SLUG expression (p=0.0358) in blastemal WT components. In addition, patients with a poor prognosis and high HMGA2 expression presented high levels of MDR3 (multidrug resistance transporter). Our findings suggest that HMGA2 plays a prominent role in the pathogenesis of a subset of blastemal WT, strongly associated with relapse and resistance to chemotherapy.

Wilms tumour (WT) is the most common renal tumour in children. Most WT patients respond to chemotherapy, but subsets of tumours develop resistance to chemotherapeutic agents, which is a major obstacle in their successful treatment. Multidrug resistance transporters play a crucial role in the development of resistance in cancer due to the efflux of anticancer agents out of cells. The aim of this study was to explore several human multidrug resistance transporters in 46 WT and 40 nonneoplastic control tissues (normal kidney) from patients selected after chemotherapy treatment SIOP 93–01, SIOP 2001. Our data showed that the majority of the studied multidrug resistance transporters were downregulated or unchanged between tumours and control tissues. However, BCRP1, MDR3 and MRP1 were upregulated in tumours versus control tissues. MDR3 and MRP1 overexpression correlated with highrisk tumours (SIOP classification) (p = 0.0022 and p < 0.0001, respectively) and the time of disease-free survival was significantly shorter in patients with high transcript levels of MDR3 (p = 0.0359). MDR3 and MRP1 play a role in drug resistance in WT treatment, probably by alteration of an unspecific drug excretion system. Besides, within the blastemal subtype, we observed patients with low MDR3 expression were significantly associated with a better outcome than patients with high MDR3 expression. We could define two types of blastemal WT associated with different disease outcomes, enabling the stratification of blastemal WT patients based on the expression levels of the multidrug resistance transporter MDR3.

Wilms tumour (WT) is the most common renal tumour in children. Most WT patients respond to chemotherapy, but subsets of tumours develop resistance to chemotherapeutic agents, which is a major obstacle in their successful treatment. Multidrug resistance transporters play a crucial role in the development of resistance in cancer due to the efflux of anticancer agents out of cells. The aim of this study was to explore several human multidrug resistance transporters in 46 WT and 40 non-neoplastic control tissues (normal kidney) from patients selected after chemotherapy treatment SIOP 93–01, SIOP 2001. Our data showed that the majority of the studied multidrug resistance transporters were downregulated or unchanged between tumours and control tissues. However, BCRP1, MDR3 and MRP1 were upregulated in tumours versus control tissues. MDR3 and MRP1 overexpression correlated with high-risk tumours (SIOP classification) (p = 0.0022 and p < 0.0001, respectively) and the time of disease-free survival was significantly shorter in patients with high transcript levels of MDR3 (p = 0.0359). MDR3 and MRP1 play a role in drug resistance in WT treatment, probably by alteration of an unspecific drug excretion system. Besides, within the blastemal subtype, we observed patients with low MDR3 expression were significantly associated with a better outcome than patients with high MDR3 expression. We could define two types of blastemal WT associated with different disease outcomes, enabling the stratification of blastemal WT patients based on the expression levels of the multidrug resistance transporter MDR3.

Wilms tumor (WT) is an embryonal malignant neoplasm of the kidney that accounts for 6–7% of all childhood cancers. WT seems to derive from multipotent embryonic renal stem cells that have failed to differentiate properly. Since mechanisms underlying WT tumorigenesis remain largely unknown, the aim of this study was to explore the expression of embryonic stem cell (ESC) markers in samples of WT patients after chemotherapy treatment SIOP protocol, as the gene expression patterns of ESC are like those of most cancer cells. We found that expression of ESC markers is heterogeneous, and depends on histological WT components. Interestingly, among ESC markers, HMGA2 was expressed significantly stronger in the blastemal component than in the stromal and the normal kidney. Moreover, two subsets of patients of WT blastemal type were identified, depending on the expression levels of HMGA2. High HMGA2 expression levels were significantly associated with a higher proliferation rate (p=0.0345) and worse patient prognosis (p=0.0289). The expression of HMGA2 was a stage-independent factor of clinical outcome in blastemal WT patients. Our multivariate analyses demonstrated the association between LIN28B–LET7A–HMGA2 expression, and the positive correlation between HMGA2 and SLUG expression (p=0.0358) in blastemal WT components. In addition, patients with a poor prognosis and high HMGA2 expression presented high levels of MDR3 (multidrug resistance transporter). Our findings suggest that HMGA2 plays a prominent role in the pathogenesis of a subset of blastemal WT, strongly associated with relapse and resistance to chemotherapy.

Diseases of complex origin have a component of quantitative genetics that contributes to their susceptibility and phenotypic variability. However, after several studies, a major part of the genetic component of complex phenotypes has still not been found, a situation known as “missing heritability.” Although there have been many hypotheses put forward to explain the reasons for the missing heritability, its definitive causes remain unknown. Complex diseases are caused by multiple intermediate phenotypes involved in their pathogenesis and, very often, each one of these intermediate phenotypes also has a component of quantitative inheritance. Here we propose that at least part of the missing heritability can be explained by the genetic component of intermediate phenotypes that is not detectable at the level of the main complex trait. At the same time, the identification of the genetic component of intermediate phenotypes provides an opportunity to identify part of the missing heritability of complex diseases.

[Background]: An essential question in cancer is why individuals with the same disease have different clinical outcomes. Progress toward a more personalized medicine in cancer patients requires taking into account the underlying heterogeneity at different molecular levels. [Results]: Here, we present a model in which there are complex interactions at different cellular and systemic levels that account for the heterogeneity of susceptibility to and evolution of ERBB2-positive breast cancers. Our model is based on our analyses of a cohort of mice that are characterized by heterogeneous susceptibility to ERBB2-positive breast cancers. Our analysis reveals that there are similarities between ERBB2 tumors in humans and those of backcross mice at clinical, genomic, expression, and signaling levels. We also show that mice that have tumors with intrinsically high levels of active AKT and ERK are more resistant to tumor metastasis. Our findings suggest for the first time that a site-specific phosphorylation at the serine 473 residue of AKT1 modifies the capacity for tumors to disseminate. Finally, we present two predictive models that can explain the heterogeneous behavior of the disease in the mouse population when we consider simultaneously certain genetic markers, liver cell signaling and serum biomarkers that are identified before the onset of the disease. [Conclusions]: Considering simultaneously tumor pathophenotypes and several molecular levels, we show the heterogeneous behavior of ERBB2-positive breast cancer in terms of disease progression. This and similar studies should help to better understand disease variability in patient populations.

Breast cancer is a major cause of mortality in women. The transcription factor SNAI2 has been implicated in the pathogenesis of several types of cancer, including breast cancer of basal origin. Here we show that SNAI2 is also important in the development of breast cancer of luminal origin in MMTV-ErbB2 mice. SNAI2 deficiency leads to longer latency and fewer luminal tumors, both of these being characteristics of pretumoral origin. These effects were associated with reduced proliferation and a decreased ability to generate mammospheres in normal mammary glands. However, the capacity to metastasize was not modified. Under conditions of increased ERBB2 oncogenic activity after pregnancy plus SNAI2 deficiency, both pretumoral defects - latency and tumor load - were compensated. However, the incidence of lung metastases was dramatically reduced. Furthermore, SNAI2 was required for proper postlactational involution of the breast. At 3 days post lactational involution, the mammary glands of Snai2-deficient mice exhibited lower levels of pSTAT3 and higher levels of pAKT1, resulting in decreased apoptosis. Abundant noninvoluted ducts were still present at 30 days post lactation, with a greater number of residual ERBB2+ cells. These results suggest that this defect in involution leads to an increase in the number of susceptible target cells for transformation, to the recovery of the capacity to generate mammospheres and to an increase in the number of tumors. Our work demonstrates the participation of SNAI2 in the pathogenesis of luminal breast cancer, and reveals an unexpected connection between the processes of postlactational involution and breast tumorigenesis in Snai2-null mutant mice.

Los tumores de mama y de pulmón son la causa principal de muerte por cáncer en el Mundo Occidental. La muerte viene, sobre todo, determinada por la diseminación a distancia o metástasis; aunque en el cáncer de pulmón también tienen relevancia las complicaciones debidas a la progresión local. Por ello, estamos interesados en comprender mejor los mecanismos moleculares y celulares que determinan la mala evolución de estos tumores. Con el fin de predecir la mala evolución de distintos tipos de cáncer, diversos laboratorios han descrito una serie de patrones de expresión génica con un fin pronóstico. Un problema no resuelto es identificar cuáles de esos genes, además de un valor pronóstico, están particularmente implicados en la patogenia tumoral y tendrían un interés terapéutico, por lo que serían dianas ideales para evitar la evolución de la enfermedad. Uno de esos genes, cuya sobreexpresión se ha implicado en el mal pronóstico y la progresión de diversos tipos de cáncer, incluidos mama y pulmón, es el gen SNAI2/SLUG. La proteína SNAI2/SLUG regula fundamentalmente procesos de transición epitelio-mesenquimal y de inhibición de la apoptosis. Por lo primero, la célula tumoral adquiere características mesenquimales con pérdida de adhesión y capacidad de movimiento, fenómenos que definen la metástasis. Por lo segundo, la célula tumoral no responde a las señales de muerte celular y, ello favorecería la iniciación tumoral, el crecimiento local, y la supervivencia de la célula tumoral lejos de su microambiente. Ello sugiere que el gen SNAI2/SLUG podría ser un candidato ideal para ser utilizado como diana terapeútica, cuya inhibición podría interferir con varias fases del desarrollo tumoral. Nuestro objetivo global fue evaluar in vivo la importancia del gen SNAI2/SLUG en el desarrollo y evolución de los cánceres de mama y pulmón, como evaluación indirecta de su interés como diana terapéutica. Para ello, se determinó el papel del gen Snai2/Slug en el inicio y progresión del cán