Publications by authors named "Massimiliano Bonafè"

102 Publications

Detection and Investigation of Extracellular Vesicles in Serum and Urine Supernatant of Prostate Cancer Patients.

Diagnostics (Basel) 2021 Mar 8;11(3). Epub 2021 Mar 8.

Biosciences Laboratory, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Via P. Maroncelli 40, 47014 Meldola, Italy.

Prostate Cancer (PCa) is one of the most frequently identified urological cancers. PCa patients are often over-diagnosed due to still not highly specific diagnostic methods. The need for more accurate diagnostic tools to prevent overestimated diagnosis and unnecessary treatment of patients with non-malignant conditions is clear, and new markers and methods are strongly desirable. Extracellular vesicles (EVs) hold great promises as liquid biopsy-based markers. Despite the biological and technical issues present in their detection and study, these particles can be found highly abundantly in the biofluid and encompass a wealth of macromolecules that have been reported to be related to many physiological and pathological processes, including cancer onset, metastasis spreading, and treatment resistance. The present study aims to perform a technical feasibility study to develop a new workflow for investigating EVs from several biological sources. Serum and urinary supernatant EVs of PCa, benign prostatic hyperplasia (BPH) patients, and healthy donors were isolated and investigated by a fast, easily performable, and cost-effective cytofluorimetric approach for a multiplex detection of 37 EV-antigens. We also observed significant alterations in serum and urinary supernatant EVs potentially related to BPH and PCa, suggesting a potential clinical application of this workflow.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/diagnostics11030466DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7998238PMC
March 2021

TP53 drives abscopal effect by secretion of senescence-associated molecular signals in non-small cell lung cancer.

J Exp Clin Cancer Res 2021 Mar 5;40(1):89. Epub 2021 Mar 5.

Department of Medical Physics, IRCCS University Hospital of Bologna, via Massarenti 9, 40138, Bologna, Italy.

Background: Recent developments in abscopal effect strongly support the use of radiotherapy for the treatment of metastatic disease. However, deeper understanding of the molecular mechanisms underlying the abscopal effect are required to best benefit a larger proportion of patients with metastasis. Several groups including ours, reported the involvement of wild-type (wt) p53 in radiation-induced abscopal effects, however very little is known on the role of wtp53 dependent molecular mechanisms.

Methods: We investigated through in vivo and in vitro approaches how wtp53 orchestrates radiation-induced abscopal effects. Wtp53 bearing (A549) and p53-null (H1299) NSCLC lines were xenotransplanted in nude mice, and cultured in 2D monolayers and 3D tumor spheroids. Extracellular vesicles (EVs) were isolated from medium cell culture by ultracentrifugation protocol followed by Nanoparticle Tracking Analysis. Gene expression was evaluated by RT-Real Time, digital qRT-PCR, and dot blot technique. Protein levels were determined by immunohistochemistry, confocal anlysis, western blot techniques, and immunoassay.

Results: We demonstrated that single high-dose irradiation (20 Gy) induces significant tumor growth inhibition in contralateral non-irradiated (NIR) A549 xenograft tumors but not in NIR p53-null H1299 or p53-silenced A549 (A549sh/p53) xenografts. We further demonstrates that irradiation of A549 cells in vitro induces a senescence-associated secretory phenotype (SASP) producing extracellular vesicles (EVs) expressing CD63 and carrying DNA:RNA hybrids and LINE-1 retrotransposon. IR-A549 EVs also hamper the colony-forming capability of recipient NIR A549 cells, induce senescent phenotype, nuclear expression of DNA:RNA hybrids, and M1 macrophage polarization.

Conclusions: In our models, we demonstrate that high radiation dose in wtp53 tumors induce the onset of SASP and secretion of CD63+ EVs loaded with DNA:RNA hybrids and LINE-1 retrotransposons that convey senescence messages out of the irradiation field triggering abscopal effect in NIR tumors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13046-021-01883-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7934399PMC
March 2021

Urinary Metabolic Biomarkers in Cancer Patients: An Overview.

Methods Mol Biol 2021 ;2292:203-212

Department of Biological and Environmental Sciences and Technologies, University of Salento, Lecce, Italy.

The pathogenesis of cancer involves multiple molecular alterations at the level of genome, epigenome, and stromal environment, resulting in several deregulated signal transduction pathways. Metabolites are not only end products of gene and protein expression but also a consequence of the mutual relationship between the genome and the internal environment. Considering that metabolites serve as a comprehensive chemical fingerprint of cell metabolism, metabolomics is emerging as the method able to discover metabolite biomarkers that can be developed for early cancer detection, prognosis, and response to treatment. Urine represents a noninvasive source, available and rich in metabolites, useful for cancer diagnosis, prognosis, and treatment monitoring. In this chapter, we reported the main published evidences on urinary metabolic biomarkers in the studied cancers related to hepatopancreatic and urinary tract with the aim at discussing their promising role in clinical practice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-0716-1354-2_18DOI Listing
April 2021

Interleukin-6 neutralization ameliorates symptoms in prematurely aged mice.

Aging Cell 2021 01 3;20(1):e13285. Epub 2021 Jan 3.

CNR Institute of Molecular Genetics "Luigi Luca Cavalli-Sforza", Unit of Bologna, Bologna, Italy.

Hutchinson-Gilford progeria syndrome (HGPS) causes premature aging in children, with adipose tissue, skin and bone deterioration, and cardiovascular impairment. In HGPS cells and mouse models, high levels of interleukin-6, an inflammatory cytokine linked to aging processes, have been detected. Here, we show that inhibition of interleukin-6 activity by tocilizumab, a neutralizing antibody raised against interleukin-6 receptors, counteracts progeroid features in both HGPS fibroblasts and Lmna progeroid mice. Tocilizumab treatment limits the accumulation of progerin, the toxic protein produced in HGPS cells, rescues nuclear envelope and chromatin abnormalities, and attenuates the hyperactivated DNA damage response. In vivo administration of tocilizumab reduces aortic lesions and adipose tissue dystrophy, delays the onset of lipodystrophy and kyphosis, avoids motor impairment, and preserves a good quality of life in progeroid mice. This work identifies tocilizumab as a valuable tool in HGPS therapy and, speculatively, in the treatment of a variety of aging-related disorders.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/acel.13285DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7811841PMC
January 2021

The role of extracellular DNA in COVID-19: Clues from inflamm-aging.

Ageing Res Rev 2021 03 13;66:101234. Epub 2020 Dec 13.

Department of Experimental and Diagnostic Medicine, University of Bologna, Italy. Electronic address:

Epidemiological data convey severe prognosis and high mortality rate for COVID-19 in elderly men affected by age-related diseases. These subjects develop local and systemic hyper-inflammation, which are associated with thrombotic complications and multi-organ failure. Therefore, understanding SARS-CoV-2 induced hyper-inflammation in elderly men is a pressing need. Here we focus on the role of extracellular DNA, mainly mitochondrial DNA (mtDNA) and telomeric DNA (telDNA) in the modulation of systemic inflammation in these subjects. In particular, extracellular mtDNA is regarded as a powerful trigger of the inflammatory response. On the contrary, extracellular telDNA pool is estimated to be capable of inhibiting a variety of inflammatory pathways. In turn, we underpin that telDNA reservoir is progressively depleted during aging, and that it is scarcer in men than in women. We propose that an increase in extracellular mtDNA, concomitant with the reduction of the anti-inflammatory telDNA reservoir may explain hyper-inflammation in elderly male affected by COVID-19. This scenario is reminiscent of inflamm-aging, the portmanteau word that depicts how aging and aging related diseases are intimately linked to inflammation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.arr.2020.101234DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7833688PMC
March 2021

Long-term exposure of human endothelial cells to metformin modulates miRNAs and isomiRs.

Sci Rep 2020 12 11;10(1):21782. Epub 2020 Dec 11.

Department of Clinical and Molecular Sciences, Università Politecnica Delle Marche, Via Tronto 10/A, 60126, Ancona, Italy.

Increasing evidence suggest that the glucose-lowering drug metformin exerts a valuable anti-senescence role. The ability of metformin to affect the biogenesis of selected microRNAs (miRNAs) was recently suggested. MicroRNA isoforms (isomiRs) are distinct variations of miRNA sequences, harboring addition or deletion of one or more nucleotides at the 5' and/or 3' ends of the canonical miRNA sequence. We performed a comprehensive analysis of miRNA and isomiR profile in human endothelial cells undergoing replicative senescence in presence of metformin. Metformin treatment was associated with the differential expression of 27 miRNAs (including miR-100-5p, -125b-5p, -654-3p, -217 and -216a-3p/5p). IsomiR analysis revealed that almost 40% of the total miRNA pool was composed by non-canonical sequences. Metformin significantly affects the relative abundance of 133 isomiRs, including the non-canonical forms of the aforementioned miRNAs. Pathway enrichment analysis suggested that pathways associated with proliferation and nutrient sensing are modulated by metformin-regulated miRNAs and that some of the regulated isomiRs (e.g. the 5' miR-217 isomiR) are endowed with alternative seed sequences and share less than half of the predicted targets with the canonical form. Our results show that metformin reshapes the senescence-associated miRNA/isomiR patterns of endothelial cells, thus expanding our insight into the cell senescence molecular machinery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-78871-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7732983PMC
December 2020

Decreased serum levels of the inflammaging marker miR-146a are associated with clinical non-response to tocilizumab in COVID-19 patients.

Mech Ageing Dev 2021 01 8;193:111413. Epub 2020 Dec 8.

Department of Clinical and Molecular Sciences, Università Politecnica delle Marche, Ancona, Italy; Center of Clinical Pathology and Innovative Therapy, IRCCS INRCA, Ancona, Italy. Electronic address:

Current COVID-19 pandemic poses an unprecedented threat to global health and healthcare systems. The most amount of the death toll is accounted by old people affected by age-related diseases that develop a hyper-inflammatory syndrome. In this regard, we hypothesized that COVID-19 severity may be linked to inflammaging. Here, we examined 30 serum samples from patients enrolled in the clinical trial NCT04315480 assessing the clinical response to a single-dose intravenous infusion of the anti-IL-6 receptor drug Tocilizumab (TCZ) in COVID-19 patients with multifocal interstitial pneumonia. In these serum samples, as well as in 29 age- and gender-matched healthy control subjects, we assessed a set of microRNAs that regulate inflammaging, i.e. miR-146a-5p, miR-21-5p, and miR-126-3p, which were quantified by RT-PCR and Droplet Digital PCR. We showed that COVID-19 patients who did not respond to TCZ have lower serum levels of miR-146a-5p after the treatment (p = 0.007). Among non-responders, those with the lowest serum levels of miR-146a-5p experienced the most adverse outcome (p = 0.008). Our data show that a blood-based biomarker, such as miR-146a-5p, can provide clues about the molecular link between inflammaging and COVID-19 clinical course, thus allowing to better understand the use of biologic drug armory against this worldwide health threat.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.mad.2020.111413DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7722494PMC
January 2021

Inflamm-aging: Why older men are the most susceptible to SARS-CoV-2 complicated outcomes.

Cytokine Growth Factor Rev 2020 06 3;53:33-37. Epub 2020 May 3.

Department of Clinical and Molecular Sciences, DISCLIMO, Università Politecnica delle Marche, Ancona, Italy; Center of Clinical Pathology and Innovative Therapy, IRCCS INRCA, Ancona, Italy.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is characterized by a high mortality of elderly men with age-related comorbidities. In most of these patients, uncontrolled local and systemic hyperinflammation induces severe and often lethal outcomes. The aging process is characterized by the gradual development of a chronic subclinical systemic inflammation (inflamm-aging) and by acquired immune system impairment (immune senescence). Here, we advance the hypothesis that four well-recognized features of aging contribute to the disproportionate SARS-CoV-2 mortality suffered by elderly men: i. the presence of subclinical systemic inflammation without overt disease, ii. a blunted acquired immune system and type I interferon response due to the chronic inflammation; iii. the downregulation of ACE2 (i.e. the SARS-CoV-2 receptor); and iv. accelerated biological aging. The high mortality rate of SARS-CoV-2 infection suggests that clarification of the mechanisms of inflamm-aging and immune senescence can help combat not only age-related disorders but also SARS-CoV-2 infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cytogfr.2020.04.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7252014PMC
June 2020

Pleiotropic effects of polyphenols on glucose and lipid metabolism: Focus on clinical trials.

Ageing Res Rev 2020 08 23;61:101074. Epub 2020 Apr 23.

Department of Clinical and Molecular Sciences, DISCLIMO, Università Politecnica delle Marche, Ancona, Italy.

Epidemiological evidence from observational studies suggests that dietary polyphenols (PPs) - phytochemicals found in a variety of plant-based foods - can reduce the risk of developing type 2 diabetes mellitus (T2DM). Clinical trials have also indicated that PPs may help manage the two key features of T2DM, hyperglycemia and dyslipidemia. Since the incidence of T2DM is dramatically increasing worldwide, identifying food-based approaches that can reduce the risk of developing it and help manage its main risk factors in early-stage disease has clinical and socioeconomic relevance. After a brief overview of current epidemiological data on the incidence of T2DM in individuals consuming PP-rich diets, we review the evidence from clinical trials investigating PP-enriched foods and/or PP-based nutraceutical compounds, report their main results, and highlight the knowledge gaps that should be bridged to enhance our understanding of the role of PPs in T2DM development and management.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.arr.2020.101074DOI Listing
August 2020

Cauliflower Mosaic Virus TAV, a Plant Virus Protein That Functions like Ribonuclease H1 and is Cytotoxic to Glioma Cells.

Biomed Res Int 2020 16;2020:7465242. Epub 2020 Mar 16.

Scientific Directorate, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, 47014 Meldola, FC, Italy.

Recent comparisons between plant and animal viruses reveal many common principles that underlie how all viruses express their genetic material, amplify their genomes, and link virion assembly with replication. Cauliflower mosaic virus (CaMV) is not infectious for human beings. Here, we show that CaMV transactivator/viroplasmin protein (TAV) shares sequence similarity with and behaves like the human ribonuclease H1 (RNase H1) in reducing DNA/RNA hybrids detected with S9.6 antibody in HEK293T cells. We showed that TAV is clearly expressed in the cytosol and in the nuclei of transiently transfected human cells, similar to its distribution in plants. TAV also showed remarkable cytotoxic effects in U251 human glioma cells in vitro. These characteristics pave the way for future analysis on the use of the plant virus protein TAV, as an alternative to human RNAse H1 during gene therapy in human cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1155/2020/7465242DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7102451PMC
December 2020

Small extracellular vesicles deliver miR-21 and miR-217 as pro-senescence effectors to endothelial cells.

J Extracell Vesicles 2020 18;9(1):1725285. Epub 2020 Feb 18.

Department of Clinical and Molecular Sciences, Università Politecnica delle Marche, Ancona, Italy.

The role of epigenetics in endothelial cell senescence is a cutting-edge topic in ageing research. However, little is known of the relative contribution to pro-senescence signal propagation provided by microRNAs shuttled by extracellular vesicles (EVs) released from senescent cells. Analysis of microRNA and DNA methylation profiles in non-senescent (control) and senescent (SEN) human umbilical vein endothelial cells (HUVECs), and microRNA profiling of their cognate small EVs (sEVs) and large EVs demonstrated that SEN cells released a significantly greater sEV number than control cells. sEVs were enriched in miR-21-5p and miR-217, which target DNMT1 and SIRT1. Treatment of control cells with SEN sEVs induced a miR-21/miR-217-related impairment of DNMT1-SIRT1 expression, the reduction of proliferation markers, the acquisition of a senescent phenotype and a partial demethylation of the locus encoding for miR-21. MicroRNA profiling of sEVs from plasma of healthy subjects aged 40-100 years showed an inverse U-shaped age-related trend for miR-21-5p, consistent with senescence-associated biomarker profiles. Our findings suggest that miR-21-5p/miR-217 carried by SEN sEVs spread pro-senescence signals, affecting DNA methylation and cell replication.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/20013078.2020.1725285DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7048230PMC
February 2020

Exploiting the telomere machinery to put the brakes on inflamm-aging.

Ageing Res Rev 2020 05 14;59:101027. Epub 2020 Feb 14.

Department of Clinical and Molecular Sciences, Università Politecnica delle Marche, Ancona, Italy; Center of Clinical Pathology and Innovative Therapy, IRCCS INRCA, Ancona, Italy.

Telomere shortening accompanies mammalian aging in vivo, and the burden of senescent cells with short telomeres and a senescence-associated secretory phenotype (SASP) increases with aging. The release into the cytoplasm and the extracellular vesicle-mediated intercellular exchange of telomeric TTAGGG repeats could exert an anti-inflammatory activity by preventing the activation of the misplaced nucleic acid-sensing pathway. Many pharmacological and genetic strategies have been developed to prevent telomere shortening or to achieve telomere elongation. Recently, it was demonstrated that telomere elongation can be obtained - without genetic manipulation - by culturing mice embryonic stem cells into appropriate media. Based on this observation, we hypothesize that environmental factors could affect the initial length of telomeres by modulating the activity of telomerase during the early stages of pregnancy. Therefore, organisms with longer telomeres could exploit the anti-inflammatory activity of telomeric sequences over an extended time span, eventually delaying the development and progression of age-related diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.arr.2020.101027DOI Listing
May 2020

Ribosomal DNA instability: An evolutionary conserved fuel for inflammaging.

Ageing Res Rev 2020 03 9;58:101018. Epub 2020 Jan 9.

Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Italy; Center for Applied Biomedical Research, CRBA, S. Orsola-Malpighi, University Hospital, Bologna, Italy. Electronic address:

Across eukaryotes, ribosomal DNA (rDNA) loci are characterized by intrinsic genomic instability due to their repetitive nature and their base composition that facilitate DNA double strand breaks and RNA:DNA hybrids formation. In the yeast, ribosomal DNA instability affects lifespan via the formation of extrachromosomal rDNA circles (ERC) that accrue into aged cells. In humans, rDNA instability has long been reported in a variety of progeric syndromes caused by the dysfunction of DNA helicases, but its role in physiological aging and longevity still needs to be clarified. Here we propose that rDNA instability leads to the activation of innate immunity and inflammation via the interaction with the cytoplasmic DNA sensing machinery. Owing to the recent clarified role of cytoplasmic DNA in the pro-inflammatory phenotype of senescent cells, we hypothesize that the accrual of rDNA derived molecules (i.e. ERC and RNA:DNA hybrids) may have a role in aging by contributing to inflammaging i.e. the systemic pro-inflammatory drift that associates with the onset of geriatric syndromes and age related dysfunctions in humans.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.arr.2020.101018DOI Listing
March 2020

HPV DNA Associates With Breast Cancer Malignancy and It Is Transferred to Breast Cancer Stromal Cells by Extracellular Vesicles.

Front Oncol 2019 16;9:860. Epub 2019 Sep 16.

Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.

A causal link between Human Papillomavirus (HPV) and breast cancer (BC) remains controversial. In spite of this, the observation that HPV DNA is over-represented in the Triple Negative (TN) BC has been reported. Here we remark the high prevalence of HPV DNA (44.4%) in aggressive BC subtypes (TN and HER2+) in a population of 273 Italian women and we convey the presence of HPV DNA in the epithelial and stromal compartments by . As previously reported, we also found that serum derived-extracellular vesicles (EVs) from BC affected patients contain HPV DNA. Interestingly, in one TNBC patient, the same HPV DNA type was detected in the serum-derived EVs, cervical and BC tissue samples. Then, we report that HPV DNA can be transferred by EVs to recipient BC stromal cells that show an activated phenotype (e.g., CD44, IL6 expression) and an enhanced capability to sustain mammospheres (MS) formation. These data suggest that HPV DNA vehiculated by EVs is a potential trigger for BC niche aggressiveness.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fonc.2019.00860DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6756191PMC
September 2019

NMR-Based Metabolomic Approach Tracks Potential Serum Biomarkers of Disease Progression in Patients with Type 2 Diabetes Mellitus.

J Clin Med 2019 May 21;8(5). Epub 2019 May 21.

Department of Biological and Environmental Sciences and Technologies, University of Salento, via Monteroni, 73100 Lecce, Italy.

Type 2 diabetes mellitus (T2DM) is a metabolic disorder characterized by chronic hyperglycemia associated with alterations in carbohydrate, lipid, and protein metabolism. The prognosis of T2DM patients is highly dependent on the development of complications, and therefore the identification of biomarkers of T2DM progression, with minimally invasive techniques, is a huge need. In the present study, we applied a H-Nuclear Magnetic Resonance (H-NMR)-based metabolomic approach coupled with multivariate data analysis to identify serum metabolite profiles associated with T2DM development and progression. To perform this, we compared the serum metabolome of non-diabetic subjects, treatment-naïve non-complicated T2DM patients, and T2DM patients with complications in insulin monotherapy. Our analysis revealed a significant reduction of alanine, glutamine, glutamate, leucine, lysine, methionine, tyrosine, and phenylalanine in T2DM patients with respect to non-diabetic subjects. Moreover, isoleucine, leucine, lysine, tyrosine, and valine levels distinguished complicated patients from patients without complications. Overall, the metabolic pathway analysis suggested that branched-chain amino acid (BCAA) metabolism is significantly compromised in T2DM patients with complications, while perturbation in the metabolism of gluconeogenic amino acids other than BCAAs characterizes both early and advanced T2DM stages. In conclusion, we identified a metabolic serum signature associated with T2DM stages. These data could be integrated with clinical characteristics to build a composite T2DM/complications risk score to be validated in a prospective cohort.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/jcm8050720DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6571571PMC
May 2019

Evaluation of Androgen Receptor in Relation to Estrogen Receptor (AR/ER) and Progesterone Receptor (AR/PgR): A New Must in Breast Cancer?

J Oncol 2019 14;2019:1393505. Epub 2019 Apr 14.

Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, Italy.

Steroid nuclear receptors are known to be involved in the regulation of epithelial-mesenchymal transition process with important roles in invasion and metastasis initiation. Androgen receptor (AR) has been extensively studied, but its role in relation to breast cancer patient prognosis remains to be clarified. AR/ER ratio has been reported to be an unfavorable prognostic marker in early primary breast cancer, but its role in the patients with advanced disease has to be cleared. We retrospectively analyzed ER, PgR, and AR expression on a case series of 159 specimens of primary BC samples by using immunohistochemistry and 89 patients of these had luminal tumors for which AR and ER expression and survival data were available. For twenty-four patients both primary and metastatic tumors were available. A significantly shorter overall survival was observed in primary tumors with AR/PgR ratio ≥ 1.54 (HR = 2.27; 95% CI 1.30-3.97; = 0.004). Similarly OS was significantly shorter when ER/PgR ratio ≥2 in primary tumors (HR = 1.89; 95% CI 1.10-3.24; = 0.021). The analysis of the 24 patients who had biomarker determinations both in primary tumors and metastasis showed a better OS when AR/ER ratio in the metastasis was ≥ 0.90 ( = 0.022). Patients with a high AR/ER ratio in primary tumor that remained high in the metastasis had better prognosis in terms of OS ( = 0.011). Despite we suggested that the ratios AR/ER and AR/PgR could be used to identify patients with different prognosis, their real value needs to be better clarified in different BC settings through prospective studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1155/2019/1393505DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6487115PMC
April 2019

Androgen receptor in breast cancer: A wolf in sheep's clothing? A lesson from prostate cancer.

Semin Cancer Biol 2020 02 16;60:132-137. Epub 2019 Apr 16.

Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, Italy. Electronic address:

The possibility that a receptor for androgen is expressed in Breast Cancer (BC) is fascinating given that the tumor is predominantly estrogen-dependent. The androgen receptor (AR) is emerging as a new marker and a potential new therapeutic target in the treatment of BC patients. The recent availability of selective AR inhibitors (e.g. bicalutamide, enzalutamide, apalutamide) approved for the treatment of prostate cancer has opened up the possibility to use them in BC patients whose tumors express AR. However, AR appears to have various functions according to the BC subtype, e.g. ER-positive or triple negative BC and the patient prognosis is different on the basis of the presence or absence of estrogen and progesterone receptors. Moreover, a different AR expression was seen according to the various ethnicities. Of note, in population at low economical income, the availability of anti-AR compounds at low cost could open the possibility to treat AR-positive triple negative BC that are highly present in these populations. Up to now, AR detection is not routinely performed in BC. The standardization of AR detection methods could render AR an easily detectable marker in primary BC and metastatic samples. Nevertheless, the overall concordance of 60% of AR expression in primary tumor and metastasis implies that a clinician who need the AR value to give anti-AR therapy should have the data on both the tumor materials. Following the comprehensive studies on prostate cancer the possibility to test AR on liquid biopsies suggest the use of this biomarker for a real-time disease monitoring. Finally, considering the possibility to treat patients with immune checkpoint inhibitors there is the need to know the relation between microenvironment and AR in BC.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.semcancer.2019.04.002DOI Listing
February 2020

Anticancer drug discovery using multicellular tumor spheroid models.

Expert Opin Drug Discov 2019 03 28;14(3):289-301. Epub 2019 Jan 28.

a Biosciences Laboratory , Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS , Meldola , Italy.

Introduction: Despite the increasing financial outlay on cancer research and drug discovery, many advanced cancers remain incurable. One possible strategy for increasing the approval rate of new anticancer drugs for use in clinical practice could be represented by three-dimensional (3D) tumor models on which to perform in vitro drug screening. There is a general consensus among the scientific community that 3D tumor models more closely recapitulate the complexity of tumor tissue architecture and biology than bi-dimensional cell cultures. In a 3D context, cells are connected to each other through tissue junctions and show proliferative and metabolic gradients that resemble the intricate milieu of organs and tumors. Areas covered: The present review focuses on available techniques for generating tumor spheroids and discusses current and future applications in the field of drug discovery. The article is based on literature obtained from PubMed. Expert opinion: Given the relative simplicity of spheroid models with respect to clinical tumors, we must be careful not to overestimate the reliability of their drug-response prediction capacity. The next challenge is to combine our knowledge of co-culture methodologies with high-content imaging and advanced microfluidic technologies to improve the readout and biomimetic potential of spheroid-based models.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/17460441.2019.1570129DOI Listing
March 2019

Prognostic Role of Circulating miRNAs in Early-Stage Non-Small Cell Lung Cancer.

J Clin Med 2019 Jan 23;8(2). Epub 2019 Jan 23.

General Pathology Unit, Department of Medicine, University of Perugia, 06132 Perugia, Italy.

Non-small cell lung cancer (NSCLC) is the primary cause of cancer-related death worldwide, with a low 5-year survival rate even in fully resected early-stage disease. Novel biomarkers to identify patients at higher risk of relapse are needed. We studied the prognostic value of 84 circulating microRNAs (miRNAs) in 182 patients with resected early-stage NSCLC (99 adenocarcinoma (ADC), 83 squamous cell carcinoma (SCC)) from whom peripheral blood samples were collected pre-surgery. miRNA expression was analyzed in relation to disease-free survival (DFS) and overall survival (OS). In univariable analyses, five miRNAs (miR-26a-5p, miR-126-3p, miR-130b-3p, miR-205-5p, and miR-21-5p) were significantly associated with DFS in SCC, and four (miR-130b-3p, miR-26a-5p, miR-126-3p, and miR-205-5p) remained significantly associated with OS. In ADC, miR-222-3p, miR-22-3p, and mir-93-5p were significantly associated with DFS, miR-22-3p remaining significant for OS. Given the high-dimensionality of the dataset, multivariable models were obtained using a regularized Cox regression including all miRNAs and clinical covariates. After adjustment for disease stage, only miR-126-3p showed an independent prognostic role, with higher values associated with longer DFS in SCC patients. With regard to ADC and OS, no miRNA remained significant in multivariable analysis. Further investigation into the role of miR-126 as a prognostic marker in early-stage NSCLC is warranted.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/jcm8020131DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6407000PMC
January 2019

Genomic stability, anti-inflammatory phenotype, and up-regulation of the RNAseH2 in cells from centenarians.

Cell Death Differ 2019 09 8;26(9):1845-1858. Epub 2019 Jan 8.

Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.

Current literature agrees on the notion that efficient DNA repair favors longevity across evolution. The DNA damage response machinery activates inflammation and type I interferon signaling. Both pathways play an acknowledged role in the pathogenesis of a variety of age-related diseases and are expected to be detrimental for human longevity. Here, we report on the anti-inflammatory molecular make-up of centenarian's fibroblasts (low levels of IL-6, type 1 interferon beta, and pro-inflammatory microRNAs), which is coupled with low level of DNA damage (measured by comet assay and histone-2AX activation) and preserved telomere length. In the same cells, high levels of the RNAseH2C enzyme subunit and low amounts of RNAseH2 substrates, i.e. cytoplasmic RNA:DNA hybrids are present. Moreover, RNAseH2C locus is hypo-methylated and RNAseH2C knock-down up-regulates IL-6 and type 1 interferon beta in centenarian's fibroblasts. Interestingly, RNAseH2C locus is hyper-methylated in vitro senescent cells and in tissues from atherosclerotic plaques and breast tumors. Finally, extracellular vesicles from centenarian's cells up-regulate RNAseH2C expression and dampen the pro-inflammatory phenotype of fibroblasts, myeloid, and cancer cells. These data suggest that centenarians are endowed with restrained DNA damage-induced inflammatory response, that may facilitate their escape from the deleterious effects of age-related chronic inflammation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41418-018-0255-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6748076PMC
September 2019

The telomere world and aging: Analytical challenges and future perspectives.

Ageing Res Rev 2019 03 4;50:27-42. Epub 2019 Jan 4.

Department of Clinical and Molecular Sciences, DISCLIMO, Università Politecnica delle Marche, Ancona, Italy; Center of Clinical Pathology and Innovative Therapy, IRCCS INRCA, Ancona, Italy. Electronic address:

Telomeres, the terminal nucleoprotein structures of eukaryotic chromosomes, play pleiotropic functions in cellular and organismal aging. Telomere length (TL) varies throughout life due to the influence of genetic factors and to a complex balancing between "shortening" and "elongation" signals. Telomerase, the only enzyme that can elongate a telomeric DNA chain, and telomeric repeat-containing RNA (TERRA), a long non-coding RNA involved in looping maintenance, play key roles in TL during life. Despite recent advances in the knowledge of TL, TERRA and telomerase activity (TA) biology and their measurement techniques, the experimental and theoretical issues involved raise a number of problems that should carefully be considered by researchers approaching the "telomere world". The increasing use of such parameters - hailed as promising clinically relevant biomarkers - has failed to be paralleled by the development of automated and standardized measurement technology. Consequently, associating given TL values to specific pathological conditions involves on the one hand technological issues and on the other clinical-biological issues related to the planning of clinically relevant association studies. Addressing these issues would help avoid major biases in association studies involving TL and a number of outcomes, especially those focusing on psychological and bio-behavioral variables. The main challenge in telomere research is the development of accurate and reliable measurement methods to achieve simple and sensitive TL, TERRA, and TA detection. The discovery of the localization of telomeres and TERRA in cellular and extracellular compartments had added an additional layer of complexity to the measurement of these age-related biomarkers. Since combined analysis of TL, TERRA and TA may well provide more exhaustive clinical information than a single parameter, we feel it is important for researchers in the various fields to become familiar with their most common measurement techniques and to be aware of the respective merits and drawbacks of these approaches.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.arr.2019.01.004DOI Listing
March 2019

Aberrant Metabolism in Hepatocellular Carcinoma Provides Diagnostic and Therapeutic Opportunities.

Oxid Med Cell Longev 2018 4;2018:7512159. Epub 2018 Nov 4.

Department of Biological and Environmental Sciences and Technologies, University of Salento, Lecce, Italy.

Hepatocellular carcinoma (HCC) accounts for over 80% of liver cancer cases and is highly malignant, recurrent, drug-resistant, and often diagnosed in the advanced stage. It is clear that early diagnosis and a better understanding of molecular mechanisms contributing to HCC progression is clinically urgent. Metabolic alterations clearly characterize HCC tumors. Numerous clinical parameters currently used to assess liver functions reflect changes in both enzyme activity and metabolites. Indeed, differences in glucose and acetate utilization are used as a valid clinical tool for stratifying patients with HCC. Moreover, increased serum lactate can distinguish HCC from normal subjects, and serum lactate dehydrogenase is used as a prognostic indicator for HCC patients under therapy. Currently, the emerging field of metabolomics that allows metabolite analysis in biological fluids is a powerful method for discovering new biomarkers. Several metabolic targets have been identified by metabolomics approaches, and these could be used as biomarkers in HCC. Moreover, the integration of different omics approaches could provide useful information on the metabolic pathways at the systems level. In this review, we provided an overview of the metabolic characteristics of HCC considering also the reciprocal influences between the metabolism of cancer cells and their microenvironment. Moreover, we also highlighted the interaction between hepatic metabolite production and their serum revelations through metabolomics researches.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1155/2018/7512159DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6247426PMC
January 2019

Inflammaging 2018: An update and a model.

Semin Immunol 2018 12 2;40:1-5. Epub 2018 Nov 2.

Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Italy; Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, Italy.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.smim.2018.10.008DOI Listing
December 2018

Intrabone transplant provides full stemness of cord blood stem cells with fast hematopoietic recovery and low GVHD rate: results from a prospective study.

Bone Marrow Transplant 2019 05 19;54(5):717-725. Epub 2018 Sep 19.

DIMES, Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.

Umbilical Cord Blood (UCB) represents a valid option for patients with hematopoietic malignancies lacking an HLA matched donor. To overcome the limitation of the low stem cell dose of UCB, the intrabone (IB) route has been proposed. We report the results of a prospective study on a poor-prognosis cohort of 23 patients receiving intrabone single UCB transplant (Clinicaltrials.gov NCT00886522). Cumulative incidence of hematological recovery at day 90 was 82 ± 9% (ANC > 0.5 × 10/L) and 70 ± 10% (platelet > 50 × 10/L) and correlated with CD34 + cells in the graft. NRM was 20 ±  9%. No severe aGVHD and only one extensive cGVHD occurred, with fast immune reconstitution. To test the hypothesis that the direct IB injection could affect the expression of stem cells regulatory pathways, CD34 + cells from BM aspirates at day + 10, + 20, + 30, processed in hypoxic conditions mimicking the BM-microenvironment (7%pO), were studied for the expression of c-Mpl, Notch1 and CXCR4. We found that the expression of c-Mpl in CD34 + cells at day + 10 significantly correlated with hematological recovery. In conclusion, IB-UCB transplant success is associated with low incidence of GVHD and high-speed platelet recovery; intrabone route may preserve full hematopoietic stemness by direct delivery of UCB stem cells into the hypoxic HSC niche.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41409-018-0335-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6760547PMC
May 2019

Changes in the biochemical taste of cytoplasmic and cell-free DNA are major fuels for inflamm-aging.

Semin Immunol 2018 12 15;40:6-16. Epub 2018 Sep 15.

Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna Italy; Bioscience Laboratory, Istituto Romagnolo per lo Studio e la Cura dei tumori IRST, IRCCS, Meldola, Forlì, Italy. Electronic address:

Inflamm-aging depicts the progressive activation of the innate immune system that accompanies human aging. Its role as a disease-predisposing condition has been proposed, but its molecular basis is still poorly understood. A wealth of literature conveys that, particularly upon stress, nuclear and mitochondrial genomes are released into the cytoplasmic and extracellular compartments. Cytoplasmic (cy) and cell-free (cf) DNA pools trigger inflammation and innate immunity at local and systemic level. In particular, cyDNA plays a crucial role in the phenomenon of cell senescence and in the cognate pro-inflammatory secretome. Here we propose that changes in a variety of biochemical characteristics "tastes" of cy- and cf-DNA (e.g. the amount of 8-oxo-deoxy-guanosine and 5-methyl-deoxy-cytosine, the proportion of DNA hybridized with RNA) potentially affect the capability of these DNA pools to ignite the innate immune system. We also underpin that telomeric sequences are major components of the cy/cfDNA payload. Telomere shortening, a hallmark of aging, causes the depletion of telomeric sequences in cy/cfDNA pool, thus unleashing their potential to exert an age-related activation of the innate immune system. Finally, we posit that various sources of DNA (extracellular vesicles, the commensal metagenome and food) contribute to the cy/cfDNA payloads. We speculate that changes in the biochemical "taste" of cy/cfDNA are major modifiers of inflamm-aging.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.smim.2018.08.003DOI Listing
December 2018

Immunosuppressive Treg cells acquire the phenotype of effector-T cells in chronic lymphocytic leukemia patients.

J Transl Med 2018 06 20;16(1):172. Epub 2018 Jun 20.

Scientific Directorate, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, Italy.

Background: In chronic lymphocytic leukemia (CLL) disease onset and progression are influenced by the behavior of specific CD4 T cell subsets, such as T regulatory cells (Tregs). Here, we focused on the phenotypic and functional characterization of Tregs in CLL patients to improve our understanding of the putative mechanism by which these cells combine immunosuppressive and effector-like properties.

Methods: Peripheral blood mononuclear cells were isolated from newly diagnosed CLL patients (n = 25) and healthy volunteers (n = 25). The phenotypic and functional characterization of Tregs and their subsets was assessed by flow cytometry. In vitro analysis of TH1, TH2, TH17 and Tregs cytokines was evaluated by IFN-γ, IL-4, IL-17A and IL-10 secretion assays. The transcriptional profiling of 84 genes panel was evaluated by RT Profiler PCR Array. Statistical analysis was carried out using exact non parametric Mann-Whitney U test.

Results: In all CLL samples, we found a significant increase in the frequency of IL-10-secreting Tregs and Tregs subsets, a significant rise of TH2 IL-4 and TH17 IL-17A cells, and a higher percentage of IFN-γ/IL-10 and IL-4/IL-10 double-releasing CD4 T cells. In addition, we also observed the up-regulation of innate immunity genes and the down-regulation of adaptive immunity ones.

Conclusions: Our data show that Tregs switch towards an effector-like phenotype in CLL patients. This multifaceted behavior is accompanied by an altered cytokine profiling and transcriptional program of immune genes, leading to a dysfunction in immune response in the peripheral blood environment of CLL patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12967-018-1545-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6011245PMC
June 2018

Androgen Receptor Expression in Breast Cancer: What Differences Between Primary Tumor and Metastases?

Transl Oncol 2018 Aug 18;11(4):950-956. Epub 2018 Jun 18.

Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, FC, Italy.

Genomic studies have shown that the androgen receptor (AR) pathway plays an important role in some breast cancer subtypes. However few data are present on the concordance between AR expression in primary tumors and metastases. We investigated AR expression by using immunohistochemistry (IHC) in 164 primary tumors and 83 metastases, to explore its distribution in the different tumor subtypes and its concordance between the two sample types and according to sampling time. AR was more highly expressed in luminal A and B than HER2-positive and triple negative primary tumors. A similar distribution was found in metastases, and the concordance of AR expression between primary tumors and metastases was greater than 60%. No association between sampling time and AR expression was observed. We found a good concordance of AR expression between primary tumor and metastasis, but the variability remains high between the two types of specimens, regardless of the variation in sampling time. For this reason, if used for treatment decisions, AR evaluation should be repeated in each patient whenever a new biopsy is performed, as commonly done for the other breast cancer biomarkers.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.tranon.2018.05.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6036224PMC
August 2018

Age-related M1/M2 phenotype changes in circulating monocytes from healthy/unhealthy individuals.

Aging (Albany NY) 2018 Jun;10(6):1268-1280

Department of Experimental, Diagnostic and Specialty Medicine, DIMES, Alma Mater Studiorum, Bologna, Italy.

Macrophage polarization is a candidate biomarker of disease-related inflammatory status, but its modulation during aging has not been investigated. To do this, the M1/M2 profile was assessed by CD80/CD163 gating in classical (CD14CD16), intermediate (CD14CD16), and non-classical (CD14CD16) monocytes from 31 healthy subjects (CTRs) of different ages. Cytofluorimetric analysis showed a significantly different CD80/CD163 distribution in the three subsets, as more than 80% of classical and intermediate monocytes were CD80CD163, whereas most non-classical monocytes were CD80CD163 and CD163. Non-classical CD163 monocytes were significantly higher whereas classical CD163 and CD80CD163 monocytes significantly lower in older than younger CTRs (cut-off, 65 years), suggesting different age-related trends for M2 subsets. To establish whether an M1/M2 imbalance could be associated with disease, 21 patients with acute myocardial infarction (AMI) were compared with older CTRs. The AMI patients showed a significantly decreased proportion of CD163CD80 and an increased proportion of CD163 and CD163CD80 cells among classical monocytes, opposite trends to those observed in healthy aging. Moreover, a significantly greater proportion of intermediate and non-classical CD80 monocytes suggested a shift to a pro-inflammatory phenotype. Overall, CD163/CD80 cytofluorimetric characterization of circulating monocytes provides additional information about their polarization and could be an innovative tool to monitor aging.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/aging.101465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6046240PMC
June 2018

Anti-senescence compounds: A potential nutraceutical approach to healthy aging.

Ageing Res Rev 2018 Sep 6;46:14-31. Epub 2018 May 6.

Department of Clinical and Molecular Sciences, DISCLIMO, Università Politecnica delle Marche, Ancona, Italy; Center of Clinical Pathology and Innovative Therapy, INRCA-IRCCS National Institute, Ancona, Italy. Electronic address:

The desire of eternal youth seems to be as old as mankind. However, the increasing life expectancy experienced by populations in developed countries also involves a significantly increased incidence of the most common age-related diseases (ARDs). Senescent cells (SCs) have been identified as culprits of organismal aging. Their number rises with age and their senescence-associated secretory phenotype fuels the chronic, pro-inflammatory systemic state (inflammaging) that characterizes aging, impairing the regenerative ability of stem cells and increasing the risk of developing ARDs. A variegated class of molecules, including synthetic senolytic compounds and natural compounds contained in food, have been suggested to possess anti-senescence activity. Senolytics are attracting growing interest, and their safety and reliability as anti-senescence drugs are being assessed in human clinical trials. Notably, since SCs spread inflammation at the systemic level through pro-oxidant and pro-inflammatory signals, foods rich in polyphenols, which exert antioxidant and anti-inflammatory actions, have the potential to be harnessed as "anti-senescence foods" in a nutraceutical approach to healthier aging. We discuss the beneficial effects of polyphenol-rich foods in relation to the Mediterranean diet and the dietary habits of long-lived individuals, and examine their ability to modulate bacterial genera in the gut.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.arr.2018.05.001DOI Listing
September 2018