17 research outputs found

    MALAT1 as a Regulator of the Androgen-Dependent Choline Kinase A Gene in the Metabolic Rewiring of Prostate Cancer

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    Simple Summary Despite the rapid advance in cancer therapies, treatment-resistant relapse remains a significant challenge in cancer treatment. Acquired resistance arises during or after treatment administration, and is usually the main contributor to relapse. For example, prostate cancer, the most frequent type of cancer in the elderly male population, frequently develops into aggressive forms resistant to chemical and hormonal therapies. In this condition, the so-called "cholinic phenotype" that is characterized by the overexpression of choline kinase alpha (CHKA) and increased phosphocholine levels leads to aberrant lipid metabolism. Our work demonstrates that CHKA, which is necessary for membrane phospholipid synthesis, is a target of the long non-coding RNA MALAT1. This study helps to further decipher how MALAT1 affects the regulation of crucial phospholipid/sphingolipid metabolic enzymes, as well as how the androgen receptor pathway is involved in MALAT1-dependent transcriptional regulation. Background. Choline kinase alpha (CHKA), an essential gene in phospholipid metabolism, is among the modulated MALAT1-targeted transcripts in advanced and metastatic prostate cancer (PCa). Methods. We analyzed CHKA mRNA by qPCR upon MALAT1 targeting in PCa cells, which is characterized by high dose-responsiveness to the androgen receptor (AR) and its variants. Metabolome analysis of MALAT1-depleted cells was performed by quantitative High-resolution 1 H-Nuclear Magnetic Resonance (NMR) spectroscopy. In addition, CHKA genomic regions were evaluated by chromatin immunoprecipitation (ChIP) in order to assess MALAT1-dependent histone-tail modifications and AR recruitment. Results. In MALAT1-depleted cells, the decrease of CHKA gene expression was associated with reduced total choline-containing metabolites compared to controls, particularly phosphocholine (PCho). Upon MALAT1 targeting a significant increase in repressive histone modifications was observed at the CHKA intron-2, encompassing relevant AR binding sites. Combining of MALAT1 targeting with androgen treatment prevented MALAT1-dependent CHKA silencing in androgen-responsive (LNCaP) cells, while it did not in hormone-refractory cells (22RV1 cells). Moreover, AR nuclear translocation and its activation were detected by confocal microscopy analysis and ChIP upon MALAT1 targeting or androgen treatment. Conclusions. These findings support the role of MALAT1 as a CHKA activator through putative association with the liganded or unliganded AR, unveiling its targeting as a therapeutic option from a metabolic rewiring perspective

    Young transgenic hMTH1 mice are protected against dietary fat-induced metabolic stress—implications for enhanced longevity

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    hMTH1 protects against mutation during oxidative stress. It degrades 8-oxodGTP to exclude potentially mutagenic oxidized guanine from DNA. hMTH1 expression is linked to ageing. Its downregulation in cultured cells accelerates RAS-induced senescence, and its overexpression in hMTH1-Tg mice extends lifespan. In this study, we analysed the effects of a brief (5 weeks) high-fat diet challenge (HFD) in young (2 months old) and adult (7 months old) wild-type (WT) and hMTH1-Tg mice. We report that at 2 months, hMTH1 overexpression ameliorated HFD-induced weight gain, changes in liver metabolism related to mitochondrial dysfunction and oxidative stress. It prevented DNA damage as quantified by a comet assay. At 7 months old, these HFD-induced effects were less severe and hMTH1-Tg and WT mice responded similarly. hMTH1 overexpression conferred lifelong protection against micronucleus induction, however. Since the canonical activity of hMTH1 is mutation prevention, we conclude that hMTH1 protects young mice against HFD by reducing genome instability during the early period of rapid growth and maximal gene expression. hMTH1 protection is redundant in the largely non-growing, differentiated tissues of adult mice. In hMTH1-Tg mice, expression of a less heavily mutated genome throughout life provides a plausible explanation for their extended longevity

    The over-expression of human hydrolase hMTH1 modulates metabolism and fat composition in mice exposed to high fat diet: a MRI and MRS study

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    The over-expression of human hydrolase hMTH1 modulates metabolism and fatcomposition in mice exposed to high fat diet: a MRI and MRS study Synopsis Oxidative stress is implicated in cancer, neurodegeneration and aging. hMTH1 is a hydrolase able to remove oxidized precursors from nucleotide’spool, thus avoiding oxidative nucleic acids damage. Overexpression of hMTH1 in mice is protective against oxidative damage, neurodegenerationand prolongs life span. Our study showed that the overexpression of hMTH1 in mice fed with high fat diet (HFD), a dietary regimen linked toinflammation, is associated with increased brown interscapular fat (linked to protection from obesity) and with reduced perivesical fat volume(indicator of poor cardiovascular outcomes) up to four weeks. These effects seem to be reversed by prolonging HFD. Introduction The role of the oxidative stress in the pathogenesis of cancer, neurodegeneration and aging is well established. The human MutT homologue(hMTH1) is a hydrolase able to protect nucleic acids from oxidative damage, by avoiding the incorporation of oxidized precursors in both DNA andRNA. Transgenic mice, which overexpress the human MTH1 gene (hMTH1‐Tg) are protected from neurodegeneration and motor impairment andare characterized by a decreased oxidative DNA damage, a prolonged life-span and an enhanced exploratory behavior. It has been shown thathigh fat diet is one of the most frequent environmental trigger, able to cause chronic inflammation which characterizes obesity and metabolicsyndrome. Aims Aims of this study were to understand if the the oxidative DNA damage protection mediated by the over-expression of hMTH1 is able to counteractthe metabolic alterations and the inflammation induced by the exposure to a high-fat diet (HFD, 45% of fat) for 33 weeks. Methods Male C57bl6 mice 10 wk old, wild-type (wt) and hMTH1-Tg, were fed with HFD for 33 weeks (two groups of 5 mice each). Body weight, oxidative DNAdamage/repair (by Single Cell Gel Electrophoresis assay) and other clinic parameters were measured. At 4, 11 (or 16), 22 and 33 weeks of HFDfeeding and two months after the end the HFD exposure (recovery time), animals underwent to MRI and MRS. Brain metabolism, interscapularbrown fat and liver fat were assessed by MRS. The volume of perivesical fat was assessed by MRI. Experiments were performed on a VARIAN Inovasystem operating at 4.7T with a transmitter volume RF coil actively decoupled from the receiver surface coil (RAPID Biomedical, Rimpar, Germany).1H localised MR spectra were collected from the hippocampus (HIP) and prefrontal cortex (PFC) using a PRESS sequence, according to a quantitativeprotocol. A STEAM sequence was used in the interscapular brown fat and liver for water to lipid ratio determination. T1-weighted MRI wasperformed to quantify the volume of visceral fat depot (a risk factor for metabolic dysfunction) in the four different mice groups. Repeatedmeasurements ANOVA was used for statistical comparisons (significance at p<0.05). Serum Metabolomics was carried out on Bruker Avanceoperating at 9.4 T spectrometer, by using standard presaturation pulse sequence and spin echo Carr-Purcell-Meiboom-Gill 1D sequence (CPMG)according to Beckonert protocol on intact serum. Results During HFD regimen, a significant increase in the accumulation of oxidative DNA damage has been observed in wt mice (repeated measurementsANOVA: genotype effect, p=0.02), as shown in Figure 1a). Notably, hMTH1-Tg mice resulted to be protected both in basal and HFD conditions, albeita comparable weight gain has been observed (Figure 1b). Alterations in brain metabolite concentrations have been detected in both PFC and HIP atbasal level and during the HFD regimen. Quantitative results are shown in Figure 2. In the interscapular fat we observed an increase in thewater/lipid signals ratio (which corresponds to increased brown adipose tissue, BAT) in the hMTH1-tg mice group which is maintained up to 4 weeksof HFD regimen (figure 3a and b). 1H MRS of the liver revealed a reduced amount of fat for the hMTH1-tg mice after 4 weeks of HFD stimulus(p<0.05) which is no longer observed at late times, as shown in Figure 3c and d. T1-weighted MRI of the abdomen revealed a slightly reducedamount of visceral fat for the hMTH1-tg mice at basal level but an increase in the hMTH1-tg animals at late stages after the beginning of the HFDstimulus, as shown in Figure 4a and b. Analyses at the end of HFD (33 weeks) and during the recovery time are in progress. Metabolomics analyseson serum samples showed that the overexpression of hMHT1 induced a tight control of glucose and lipid homeostasis metabolism as compared towt-mice in HFD (figure 5a and b). In particular, we found significant changes in glucose, and low-density lipoprotein/very low density lipoprotein(LDL/VLDL) signals following HFD in wt mice. Despite the stimulation of a high-fat diet, MTH1 mice are not able to alter blood glucose levels, nor toincrease the levels of VLDL / LDL in circulation. Discussion and conclusions Differences have been detected between wt and hMTH1-Tg mice before the beginning of HFD, suggesting a crosstalk between genomic instabilityand metabolic dysfunction. Brain metabolism alteration highlights a direct effect on brain functionality. In spite of similar body weight increase,nuclear oxidative DNA damage is lower in hMTH1-Tg than in wt at all time points, while adipose organ extension and composition maintain thebasal differences between wt and hMTH1-Tg mice only up to 4 weeks of oxidative stimulus. The protective role of hMTH1 against oxidative damageis associated with an increase in BAT (which may provide protection from obesity ) and with a reduced abdominal perivesical fat volume (which isconsidered an independent indicator of poor cardiovascular outcomes ). The protective effect observed in the hMTH1 mice up to 4 weeks seems tobe reversed by a prolonged HFD exposition, suggesting a not-obvious link between HFD and oxidative damage modulation by hMTH1. Acknowledgements We thank the Italian National Institute of Health for financial support References 1. De Luca G et al PLoS Genet 2008;4:e1000266. 2. De Luca G et al Aging Cell 2013;12:695-705. 3. Canese R et al NMR Biomed 2012;25:632-642. 4. Beckonert O et al. Nat Protoc 2007; 2:2692–2703 5. Matsuita M et al, International Journal of Obesity 2014; 38,812–817. 6. Powell-Wiley et al, Circulation 2021;43:e984

    Metabolic reprogramming during spontaneus mammary tumor development and progression in HER2/neu transgenic mice: a role for endogenus type I Interferon

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    Metabolic reprogramming during spontaneous mammary tumor development and progression in HER2/neu transgenic mice: a role for endogenous type I Interferon Metabolic reprogramming was linked to the major hallmarks of cancer, including tumor development, progression and therapy resistance. Our study aims to identify, by non invasive in vivo MRS and ex vivo high resolution (HR)-MRS, the metabolic changes involved in spontaneous, oncogene-driven carcinogenesis occurring in Her2/neu transgenic mice, with particular focus on the role of endogenous type I IFN (IFN-I). Our group reported that the lack of endogenous IFN-I system significantly affects Her2/neu carcinogenesis (Castiello et al, 2018). Since this phenomenon was not related to the known immunomodulatory properties of these cytokines, we investigated whether the possible reshaping of metabolic pathways was involved. Both in vivo MRS and ex vivo high resolution (HR)-MRS revealed that normal (non-tumoral) mammary glands of mice lacking a functional endogenous IFN-I (IFNARI-/-) had increased fatty acids concentration with respect to the wild type counterpart. This result paralleled the observation of specific trascriptomic profiles in IFNARI-/- normal mammary glands, that exhibited decreased expression of genes involved in OXPHOS and mitochondrial activity and increased expression of Sterol regulatory element-binding proteins, a known regulator of fatty acids biosynthesis and an independent prognostic marker in breast cancer. Interestingly, a trend in fatty acids increase could be also observed in Her2+ early tumor lesions, but not in l bigger tumor masses (>500 um3) spontaneously developed in IFNARI-/- mice. HR-MRS analyses of aqueous metabolites revealed that, as expected, the metabolic fingerprint of Her2+ tumors lesions of all sizes was significantly different from normal mammary glands in both IFNARI+/+ and IFNARI-/- mice. Nevertheless, the impact of the lack of endogenous IFN-I could be more clearly observed in early stage Her2+ tumors. The alterations in myo-inositol and gluthathione concentrations observed in IFNARI-/- tumors suggest that endogenous IFN-I may exert its antitumor activity by affecting key metabolic regulators of tumor cells proliferation. Our study provide the first evidence that endogenous IFN-I is involved in metabolic pathways in both normal mammary glands and early phase breast cancer. Further research is needed to unravel the link between these IFN-related metabolic changes and the proliferation advantage of IFNARI-/- breast cancer cells

    Changes in metabolic patterns during tumor development and progression in HER2/neu spontaneous breast cancer: a role for endogenous type I Interferon

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    Changes in metabolic patterns during tumor development and progression in HER2/neu spontaneous breast cancer: a role for endogenous type I Interferon In the last decades, cell metabolism emerged as a key factor involved in tumor development, progression and therapy resistance. Our group reported that the disruption of endogenous type I Interferon (IFN-I) system affects spontaneous Her2/neu-driven carcinogenesis through intrinsic control of cancer stem cells (Castiello et al, 2018). In order to investigate whether the possible remodeling of metabolic pathways was involved in this phenomenon, we applied non invasive in vivo MRS and ex vivo high resolution (HR)-MRS to analyze the changes in metabolic profiles occurring in healthy mammary glands and spontaneous tumors developed in Her2/ neu transgenic mice, with particular focus on IFN-I. Both in vivo MRS and ex vivo high resolution (HR)-MRS revealed that, in the absence of a functional endogenous IFN-I (IFNARI-/-), normal (non-tumoral) mammary glands had increased fatty acids concentration with respect to the wild type counterpart. This result paralleled the observation that IFNARI-/- normal mammary glands exhibited decreased expression of genes involved in OXPHOS and mitochondrial activity and increased expression of Sterol regulatory element-binding proteins, a known regulator of fatty acids biosynthesis. Interestingly, Her2+ early tumor lesions developed in IFNARI-/- mice also exhibited increased fatty acids content versus wild type mice. In contrast, bigger tumor masses (>500 um3) presented a more dysregulated and less consistent metabolic profile in both groups. As expected, the metabolic fingerprint of Her2+ tumors lesions of all sizes, assessed by HR-MRS analyses of aqueous metabolites, was significantly different from normal mammary glands in IFNARI+/+ and IFNARI-/- mice. Nevertheless, the impact of the lack of endogenous IFN-I could be observed in early stage Her2+ tumors. The alterations in myo-inositol and gluthathione concentrations observed in IFNARI-/- tumors suggest that endogenous IFN-I may exert its antitumor activity also by affecting key metabolic regulators of tumor cells proliferation. Our study provide the first evidence that endogenous IFN-I takes part in the regulation of some metabolic pathways occurring in normal mammary glands and affecting early phase of spontaneous breast cancer development. How these metabolic changes eventually provide a proliferation advantage to breast cancer cells is a matter of further investigation

    AKT-driven epithelial-mesenchymal transition is affected by copper bioavailability in HER2 negative breast cancer cells via a LOXL2-independent mechanism

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    The main mechanism underlying cancer dissemination is the epithelial to mesenchymal transition (EMT). This process is orchestrated by cytokines like TGFÎČ, involving "non-canonical" AKT- or STAT3-driven pathways. Recently, the alteration of copper homeostasis seems involved in the onset and progression of cancer
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