Publications by authors named "Michele Cioffi"

65 Publications

Calcium signaling induces a partial EMT.

EMBO Rep 2021 Sep 29;22(9):e51872. Epub 2021 Jul 29.

Abramson Family Cancer Research Institute and Department of Medicine, University of Pennsylvania, Philadelphia, PA, USA.

Epithelial plasticity, or epithelial-to-mesenchymal transition (EMT), is a well-recognized form of cellular plasticity, which endows tumor cells with invasive properties and alters their sensitivity to various agents, thus representing a major challenge to cancer therapy. It is increasingly accepted that carcinoma cells exist along a continuum of hybrid epithelial-mesenchymal (E-M) states and that cells exhibiting such partial EMT (P-EMT) states have greater metastatic competence than those characterized by either extreme (E or M). We described recently a P-EMT program operating in vivo by which carcinoma cells lose their epithelial state through post-translational programs. Here, we investigate the underlying mechanisms and report that prolonged calcium signaling induces a P-EMT characterized by the internalization of membrane-associated E-cadherin (ECAD) and other epithelial proteins as well as an increase in cellular migration and invasion. Signaling through Gαq-associated G-protein-coupled receptors (GPCRs) recapitulates these effects, which operate through the downstream activation of calmodulin-Camk2b signaling. These results implicate calcium signaling as a trigger for the acquisition of hybrid/partial epithelial-mesenchymal states in carcinoma cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.15252/embr.202051872DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8419705PMC
September 2021

Extracellular vesicle- and particle-mediated communication shapes innate and adaptive immune responses.

J Exp Med 2021 Aug 28;218(8). Epub 2021 Jun 28.

Children's Cancer and Blood Foundation Laboratories, Departments of Pediatrics and Cell and Developmental Biology, Drukier Institute for Children's Health, Meyer Cancer Center, Weill Cornell Medicine, New York, NY.

Intercellular communication among immune cells is vital for the coordination of proper immune responses. Extracellular vesicles and particles (EVPs) act as messengers in intercellular communication, with important consequences for target cell and organ physiology in both health and disease. Under normal physiological conditions, immune cell-derived EVPs participate in immune responses by regulating innate and adaptive immune responses. EVPs play a major role in antigen presentation and immune activation. On the other hand, immune cell-derived EVPs exert immunosuppressive and regulatory effects. Consequently, EVPs may contribute to pathological conditions, such as autoimmune and inflammatory diseases, graft rejection, and cancer progression and metastasis. Here, we provide an overview of the role of EVPs in immune homeostasis and pathophysiology, with a particular focus on their contribution to innate and adaptive immunity and their potential use for immunotherapies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1084/jem.20202579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8241538PMC
August 2021

Hereditary Prostate Cancer: Genes Related, Target Therapy and Prevention.

Int J Mol Sci 2021 Apr 4;22(7). Epub 2021 Apr 4.

Department of Precision Medicine, University of Campania "Luigi Vanvitelli", Via L. De Crecchio, 80138 Naples, Italy.

Prostate cancer (PCa) is globally the second most diagnosed cancer type and the most common cause of cancer-related deaths in men. Family history of PCa, hereditary breast and ovarian cancer (HBOC) and Lynch syndromes (LS), are among the most important risk factors compared to age, race, ethnicity and environmental factors for PCa development. Hereditary prostate cancer (HPCa) has the highest heritability of any major cancer in men. The proportion of PCa attributable to hereditary factors has been estimated in the range of 5-15%. To date, the genes more consistently associated to HPCa susceptibility include mismatch repair (MMR) genes (, , , and ) and homologous recombination genes (, , , ). Additional genes are also recommended to be integrated into specific research, including , and . Importantly, and mutated patients potentially benefit from Poly (ADP-ribose) polymerase PARP inhibitors, through a mechanism of synthetic lethality, causing selective tumor cell cytotoxicity in cell lines. Moreover, the detection of germline alterations in MMR genes has therapeutic implications, as it may help to predict immunotherapy benefits. Here, we discuss the current knowledge of the genetic basis for inherited predisposition to PCa, the potential target therapy, and the role of active surveillance as a management strategy for patients with low-risk PCa. Finally, the current PCa guideline recommendations are reviewed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/ijms22073753DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8038462PMC
April 2021

Prevalence of mutations in BRCA and MMR genes in patients affected with hereditary endometrial cancer.

Med Oncol 2021 Jan 23;38(2):13. Epub 2021 Jan 23.

Department of Precision Medicine, University of Campania "Luigi Vanvitelli", 80138, Naples, Italy.

Endometrial cancer (EC) is the fifth most common cancer in women from developed countries, accounting for 4.8% of new cases and 2.1% of deaths. The genetic basis for the familial risk of endometrial cancer has not been completely defined. Mostly, hereditary EC is part of two syndromes as Lynch syndrome (LS) and Hereditary Breast and Ovarian Cancer syndrome (HBOC). LS is the prototypical hereditary cancer syndrome in EC and accounts for 2-6% of all endometrial cancers. This disease is caused by autosomal dominant mutations in DNA mismatch repair (MMR) genes. Patients carrying a germline mutation in one of the MMR genes have a cumulative lifetime risk to develop EC of 20-70%. HBOC is an autosomal dominantly inherited disease, which mostly predisposes to breast and ovarian cancers, but it can be also associated with other malignancies. HBOC results from germline mutations in BRCA1/2 genes. The aim of this study was to determine the mutational status of a cohort of 40 EC patients, 19 belonging to families with LS and 21 to HBOC. Mutation analysis of MLH1, MSH2, BRCA1 and BRCA2 genes showed pathogenic variants in 17/40 (42.5%) patients. Out of 19 patients belonging to LS families, 8 (42.1%) showed a pathogenic variant. Out of 21 patients belonging to HBOC families, 9 (42.8%) showed a pathogenic variant. 1/21 (4.8%) patient report 1 variant of unknown significance (UV), c.599 C > T (p.T200I), in BRCA2. Moreover, in 1/21 (4.8%) patient we identified a novel missense variant in BRCA2, c.9541A > T (p.Met3181Leu). Mutational analysis was extended to family members, both healthy and cancer affected, of mutated patients; all the tested relatives affected with cancer displayed the pathogenic variant. Our data suggest that patients with hereditary EC have a high percentage of mutations in the LS and HBOC main susceptibility genes; therefore, the surveillance for EC, already indicated in LS patients, should also be recommended for patients with HBOC.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s12032-021-01454-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7826304PMC
January 2021

Cell-free DNA (cfDNA) and Exosome Profiling from a Year-Long Human Spaceflight Reveals Circulating Biomarkers.

iScience 2020 Dec 25;23(12):101844. Epub 2020 Nov 25.

Nancy E. and Peter C. Meinig School of Biomedical Engineering, Cornell University, Ithaca, NY, USA.

Liquid biopsies based on cell-free DNA (cfDNA) or exosomes provide a noninvasive approach to monitor human health and disease but have not been utilized for astronauts. Here, we profile cfDNA characteristics, including fragment size, cellular deconvolution, and nucleosome positioning, in an astronaut during a year-long mission on the International Space Station, compared to his identical twin on Earth and healthy donors. We observed a significant increase in the proportion of cell-free mitochondrial DNA (cf-mtDNA) inflight, and analysis of post-flight exosomes in plasma revealed a 30-fold increase in circulating exosomes and patient-specific protein cargo (including brain-derived peptides) after the year-long mission. This longitudinal analysis of astronaut cfDNA during spaceflight and the exosome profiles highlights their utility for astronaut health monitoring, as well as cf-mtDNA levels as a potential biomarker for physiological stress or immune system responses related to microgravity, radiation exposure, and the other unique environmental conditions of spaceflight.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.isci.2020.101844DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7756145PMC
December 2020

Five Italian Families with Two Mutations in Genes.

Genes (Basel) 2020 12 3;11(12). Epub 2020 Dec 3.

Department of Precision Medicine, University of Campania "Luigi Vanvitelli", Via L. De Crecchio, 80138 Naples, Italy.

Double heterozygosity (DH) in and genes and double mutation (DM) in or are extremely rare events in the general population, and few cases have been reported worldwide so far. Here, we describe five probands, all women, with breast and/or ovarian cancer and their families. Particularly, we identified two probands with DH in the genes with a frequency of 0.3% and three probands with DM in the gene with a frequency of 0.5%. The DH c.547+2T>A (IVS8+2T>A)/ c.2830A>T (p.Lys944Ter) and c.3752_3755GTCT (p.Ser1253fs)/ c.425+2T>C (IVS4+2T>C) have not been described together so far. The DM in , c.631G>A (p.Val211Ile) and c.7008-2A>T (IVS13-2A>T), found in three unrelated probands, was previously reported in further unrelated patients. Due to its peculiarity, it is likely that both pathogenic variants descend from a common ancestor and, therefore, are founder mutations. Interestingly, analyzing the tumor types occurring in DH and DM families, we observed ovarian cancer only in DH families, probably due to the presence in DH patients of pathogenic variants, which predispose one more to ovarian cancer onset. Furthermore, male breast cancer and pancreatic cancer ensued in families with DM but not with DH. These data confirm that pathogenic variants have greater penetrance to develop breast cancer in men and are associated with an increased risk of pancreatic cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/genes11121451DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761639PMC
December 2020

Extracellular Vesicle and Particle Biomarkers Define Multiple Human Cancers.

Cell 2020 08 13;182(4):1044-1061.e18. Epub 2020 Aug 13.

School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan.

There is an unmet clinical need for improved tissue and liquid biopsy tools for cancer detection. We investigated the proteomic profile of extracellular vesicles and particles (EVPs) in 426 human samples from tissue explants (TEs), plasma, and other bodily fluids. Among traditional exosome markers, CD9, HSPA8, ALIX, and HSP90AB1 represent pan-EVP markers, while ACTB, MSN, and RAP1B are novel pan-EVP markers. To confirm that EVPs are ideal diagnostic tools, we analyzed proteomes of TE- (n = 151) and plasma-derived (n = 120) EVPs. Comparison of TE EVPs identified proteins (e.g., VCAN, TNC, and THBS2) that distinguish tumors from normal tissues with 90% sensitivity/94% specificity. Machine-learning classification of plasma-derived EVP cargo, including immunoglobulins, revealed 95% sensitivity/90% specificity in detecting cancer. Finally, we defined a panel of tumor-type-specific EVP proteins in TEs and plasma, which can classify tumors of unknown primary origin. Thus, EVP proteins can serve as reliable biomarkers for cancer detection and determining cancer type.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cell.2020.07.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7522766PMC
August 2020

A Human Pluripotent Stem Cell-based Platform to Study SARS-CoV-2 Tropism and Model Virus Infection in Human Cells and Organoids.

Cell Stem Cell 2020 07 19;27(1):125-136.e7. Epub 2020 Jun 19.

The Pritzker School of Molecular Engineering, the Ben May Department for Cancer Research, the University of Chicago, IL, USA.

SARS-CoV-2 has caused the COVID-19 pandemic. There is an urgent need for physiological models to study SARS-CoV-2 infection using human disease-relevant cells. COVID-19 pathophysiology includes respiratory failure but involves other organ systems including gut, liver, heart, and pancreas. We present an experimental platform comprised of cell and organoid derivatives from human pluripotent stem cells (hPSCs). A Spike-enabled pseudo-entry virus infects pancreatic endocrine cells, liver organoids, cardiomyocytes, and dopaminergic neurons. Recent clinical studies show a strong association with COVID-19 and diabetes. We find that human pancreatic beta cells and liver organoids are highly permissive to SARS-CoV-2 infection, further validated using adult primary human islets and adult hepatocyte and cholangiocyte organoids. SARS-CoV-2 infection caused striking expression of chemokines, as also seen in primary human COVID-19 pulmonary autopsy samples. hPSC-derived cells/organoids provide valuable models for understanding the cellular responses of human tissues to SARS-CoV-2 infection and for disease modeling of COVID-19.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.stem.2020.06.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7303620PMC
July 2020

Double mutation of APC and BRCA1 in an Italian family.

Cancer Genet 2020 06 28;244:32-35. Epub 2020 Apr 28.

Department of Precision Medicine, University of Campania "Luigi Vanvitelli", Via L. De Crecchio, 7, 80138 Naples, Italy; U.O.C. Clinical and Molecular Pathology, A.O.U. University of Campania "Luigi Vanvitelli", 80138 Naples, Italy.

Familial adenomatous polyposis (FAP) is a rare genetic disorder caused mainly by monoallelic mutations of APC gene. The hereditary breast and ovarian cancer (HBOC) syndrome is an autosomal dominantly inherited disease, which mostly predisposes to breast and ovarian cancers as a result of germline mutations in BRCA1 or BRCA2 genes. In a family, mutations in two cancer susceptibility genes are extremely rare. We studied a family with a case of a 46 years-old woman affected with FAP and ovarian cancer. The patient was affected with profuse FAP since the age of 18 years and a serous ovarian cancer was diagnosed at the age of 45 years. She reported other FAP cases and one case of breast cancer in maternal family. Initially, she was tested for FAP predisposition with mutational analysis of APC gene that revealed the presence of a frameshift mutation, c.3927_3931delAAAGA (p.Glu1309AspfsX4). The presence of ovarian cancer in the patient and of a breast cancer case in the maternal family, suggested an extended analysis to HBOC susceptibility genes that led to the detection of a frameshift mutation, c.3756_3759delGTCT (p.Ser1253Argfs), in BRCA1 gene. The genetic analysis was extended also to family members. The occurrence of the double mutation in APC and BRCA1 genes in the patient was responsible for the onset of FAP and ovarian cancer respectively. The genetic counselling in hereditary cancer with a careful analysis of the pedigree allows identifying the gene to be analyzed. The development of multi-gene panels testing for cancer predisposition, with next generation sequencing (NGS), may reveal mutations in genes of high and moderate penetrance for cancer, although at a low frequency and allows diagnosing a possible double heterozygosity. This enables an adjusted treatment for the affected patient and is critical as it allows initiation of early risk-reducing measures for identified mutation carriers among family members.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cancergen.2020.04.074DOI Listing
June 2020

BRCA and PALB2 mutations in a cohort of male breast cancer with one bilateral case.

Eur J Med Genet 2020 Jun 11;63(6):103883. Epub 2020 Feb 11.

Department of Precision Medicine, University of Campania "Luigi Vanvitelli", 80138, Naples, Italy; U.O.C. Clinical and Molecular Pathology, A.O.U. University of Campania "Luigi Vanvitelli", 80138, Naples, Italy.

Introduction: Male Breast Cancer (MBC) is a rare disease, about 1% of all breast cancers worldwide and less than 1% of cancers occurring in men. The bilateral male breast cancer (bMBC) is extremely rare. Germline mutations of BRCA1/BRCA2 genes are associated with a significantly increased risk of cancer in MBC; the role of PALB2 remains to be clarified. Our main goal was to provide contribution on characterization of BRCA1/BRCA2 and PALB2 mutations in MBC patients.

Methods: We observed 28 MBC cases; one of them was a bMBC. Screening for BRCA1, BRCA2 and PALB2 genes was performed on all 28 MBC patients. Mutational analysis was extended to family members of mutated patients.

Results: In our study, the MBC incidence was 5.2% and for bMBC was 3.6%. Mutation analysis showed pathogenic mutations in 11/28 (39.3%) patients; 2/28 (7.1%) displayed a mutation in BRCA1, 8/28 (28.6%) in BRCA2 and 1/28 (3.6%) in PALB2. Out of 11 mutated patients, one (9.1%) reported a double mutation in BRCA2. Personal history of other cancers was reported in 2/28 (7.1%) patients affected by bladder cancer. A first/second degree family history of breast/ovarian and other cancers occurred in 23/28 (82.1%) patients.

Conclusion: Our findings indicate BRCA2 as the main MBC susceptibility gene and describe an increased risk of bMBC and bladder cancer in mutated patients. The identification of mutations in MBC susceptibility genes supports the usage of oncology prevention programs in affected patients and their relatives carrying the mutation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejmg.2020.103883DOI Listing
June 2020

Inflammatory Cytokines and SIRT1 Levels in Subcutaneous Abdominal Fat: Relationship With Cardiac Performance in Overweight Pre-diabetics Patients.

Front Physiol 2018 21;9:1030. Epub 2018 Aug 21.

Department of Medical, Surgical, Neurological, Metabolic and Aging Sciences, University of Campania "Luigi Vanvitelli", Naples, Italy.

In obese patients the superficial adipose tissue expresses cytokines, and sirtuins, that may affect myocardial function. In this study, we investigated the effect of metformin therapy added to a hypocaloric diet on the inflammatory pattern and cardiac performance (MPI) in obese patients with pre-diabetic condition. Fifty-eight obese patients that were enrolled for abdominoplastic surgery were divided into patients with pre-diabetic condition (n 40) and normo-glycemic patients (n18). Patients with pre-diabetic condition were randomly assigned to metformin therapy added to a hypocaloric diet (group 1, n 20) or to a hypocaloric diet therapy alone (group 2, n20). Patients with normo-glycemic condition were assigned to a hypocaloric diet therapy. During enrollment, obese patients with a pre-diabetic condition (group 1 and 2) presented higher glucose values, lower values of insulin, and higher values of the homeostasis model for the assessment of insulin resistance (HOMA-IR) than obese patients with normo-glycemic condition(group 3). In addition, they had higher values of C Reactive protein (CRP), interleukin 6 (IL6), and lower values of sirtuin 1(SIRT1). In the 12th month of the follow-up, metformin therapy induced in patients with pre-diabetic condition (group 1) a significant reduction of glucose values, HOMA-IR, and inflammatory markers such as CRP (1.04 ± 0.48 vs. 0.49 ± 0.02 mmol/L, < 0.05), IL6 (4.22 ± 0.45 vs. 3.33 ± 0.34 pg/ml, < 0.05), TNFα (6.95 ± 0.59 vs. 5.15 ± 0.44 pg/ml, < 0.05), and Nitrotyrosine (5,214 ± 0,702 vs. 2,151 ± 0,351 nmol/l, < 0.05). This was associated with a significant reduction of Intima-media thickness (1.01 ± 0.15 vs. 0.86 ± 0.15 mm, < 0.05), Septum (14 ± 2.5 vs. 10.5 ± 2 mm, < 0.05), Posterior wall (11 ± 1.5 vs. 8 ± 1 mm, < 0.05), LV mass (192.5 ± 49.5 vs. 133.2 ± 37.6 g, < 0.05) and of MPI (0.58 ± 0.03 vs. 0.38 ± 0.02, < 0.05). At 12 months of follow-up, group 2 experienced only a reduction of cholesterol (4.15 ± 0.94 vs. 4.51 ± 0.88 mmol/L, < 0.05) and triglycerides (1.71 ± 1.18 vs. 1.83 ± 0.54 mmol/L, < 0.05). At 12 months of follow-up, group 3 experienced a significant reduction of inflammatory markers, and also of echographic parameters, associated with amelioration of myocardial performance. To date, IL6 expression was related to higher values of left ventricle mass (-value 0.272, -value 0.039), and to higher IMT (-value 0.272, -value 0.039), such as those observed for CRP (-value 0.308, -value 0.021), for glucose blood values (-value 0.449, -value 0.001), and for HOMA-IR (-value 0.366, -value 0.005). An inverse correlation was found between subcutaneous fat expression of SIRT1 and myocardial performance index (-value-0.236, -value 0.002). In obese patients with pre-diabetic condition a metformin therapy may reduce inflammation and oxidative stress, and this may be associated with the amelioration of the cardiac performance. Clinical research trial number: NCT03439592.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fphys.2018.01030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6111758PMC
August 2018

and Analysis in FAP Patients: A Novel Mutation in Gene and Genotype-Phenotype Correlation.

Genes (Basel) 2018 Jun 27;9(7). Epub 2018 Jun 27.

Department of Precision Medicine, University of Campania "Luigi Vanvitelli", 80138 Naples, Italy.

and genes are mutated in 70⁻90% and 10⁻30% of familial adenomatous polyposis cases (FAP) respectively. An association between mutation localization and FAP clinical phenotype is reported. The aims of this study were to determine and mutational status in a small cohort of FAP patients and to evaluate the genotype-phenotype correlation in mutated patients. Here, we report the identification of a novel germline mutation, c.510_511insA. Overall, mutational analysis showed pathogenic mutations in 6/10 patients: 5/10 in and 1/10 in . Additionally, we found three variants of unknown significance in gene that showed no evidence of possible splicing defects by in silico analysis. Molecular analysis was also extended to family members of mutated patients. A genotype-phenotype correlation was observed for colonic signs whereas a variation of disease onset age was revealed for the same mutation. Moreover, we found an intrafamilial variability of FAP onset age. Regarding extracolonic manifestations, the development of desmoid tumors was related to surgery and not to mutation position, while a genotype-phenotype correspondence was observed for the onset of thyroid or gastric cancer. These findings can be useful in association to clinical data for early surveillance and suitable treatment of FAP patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/genes9070322DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6071208PMC
June 2018

The miR-25-93-106b cluster regulates tumor metastasis and immune evasion via modulation of CXCL12 and PD-L1.

Oncotarget 2017 Mar;8(13):21609-21625

Stem Cells & Cancer Group, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.

The stromal microenvironment controls response to injury and inflammation, and is also an important determinant of cancer cell behavior. However, our understanding of its modulation by miRNA (miR) and their respective targets is still sparse. Here, we identified the miR-25-93-106b cluster and two new target genes as critical drivers for metastasis and immune evasion of cancer cells. Using miR-25-93-106b knockout mice or antagomiRs, we demonstrated regulation of the production of the chemoattractant CXCL12 controlling bone marrow metastasis. Moreover, we identified the immune checkpoint PD-L1 (CD274) as a novel miR-93/106b target playing a central role in diminishing tumor immunity. Eventually, upregulation of miR-93 and miR-106b via miR-mimics or treatment with an epigenetic reader domain (BET) inhibitor resulted in diminished expression of CXCL12 and PD-L1. These data suggest a potential new therapeutic rationale for use of BET inhibitors for dual targeting of cancers with strong immunosuppressive and metastatic phenotypes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.15450DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5400610PMC
March 2017

MiR-93 Controls Adiposity via Inhibition of Sirt7 and Tbx3.

Cell Rep 2015 Sep 28;12(10):1594-605. Epub 2015 Aug 28.

Stem Cells & Cancer Group, Spanish National Cancer Research Centre (CNIO), Madrid 28028, Spain; Stem Cells in Cancer & Ageing, Barts Cancer Institute, Queen Mary University of London, London EC1M 6BQ, UK. Electronic address:

Conquering obesity has become a major socioeconomic challenge. Here, we show that reduced expression of the miR-25-93-106b cluster, or miR-93 alone, increases fat mass and, subsequently, insulin resistance. Mechanistically, we discovered an intricate interplay between enhanced adipocyte precursor turnover and increased adipogenesis. First, miR-93 controls Tbx3, thereby limiting self-renewal in early adipocyte precursors. Second, miR-93 inhibits the metabolic target Sirt7, which we identified as a major driver of in vivo adipogenesis via induction of differentiation and maturation of early adipocyte precursors. Using mouse parabiosis, obesity in mir-25-93-106b(-/-) mice could be rescued by restoring levels of circulating miRNA and subsequent inhibition of Tbx3 and Sirt7. Downregulation of miR-93 also occurred in obese ob/ob mice, and this phenocopy of mir-25-93-106b(-/-) was partially reversible with injection of miR-93 mimics. Our data establish miR-93 as a negative regulator of adipogenesis and a potential therapeutic option for obesity and the metabolic syndrome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2015.08.006DOI Listing
September 2015

Studying Pancreatic Cancer Stem Cell Characteristics for Developing New Treatment Strategies.

J Vis Exp 2015 Jun 20(100):e52801. Epub 2015 Jun 20.

Stem Cells & Cancer Group, Molecular Pathology Program, Spanish National Cancer Research Center; Center for Stem Cells in Cancer & Ageing, Barts Cancer Institute, Queen Mary University of London;

Pancreatic ductal adenocarcinoma (PDAC) contains a subset of exclusively tumorigenic cancer stem cells (CSCs) which have been shown to drive tumor initiation, metastasis and resistance to radio- and chemotherapy. Here we describe a specific methodology for culturing primary human pancreatic CSCs as tumor spheres in anchorage-independent conditions. Cells are grown in serum-free, non-adherent conditions in order to enrich for CSCs while their more differentiated progenies do not survive and proliferate during the initial phase following seeding of single cells. This assay can be used to estimate the percentage of CSCs present in a population of tumor cells. Both size (which can range from 35 to 250 micrometers) and number of tumor spheres formed represents CSC activity harbored in either bulk populations of cultured cancer cells or freshly harvested and digested tumors. Using this assay, we recently found that metformin selectively ablates pancreatic CSCs; a finding that was subsequently further corroborated by demonstrating diminished expression of pluripotency-associated genes/surface markers and reduced in vivo tumorigenicity of metformin-treated cells. As the final step for preclinical development we treated mice bearing established tumors with metformin and found significantly prolonged survival. Clinical studies testing the use of metformin in patients with PDAC are currently underway (e.g., NCT01210911, NCT01167738, and NCT01488552). Mechanistically, we found that metformin induces a fatal energy crisis in CSCs by enhancing reactive oxygen species (ROS) production and reducing mitochondrial transmembrane potential. In contrast, non-CSCs were not eliminated by metformin treatment, but rather underwent reversible cell cycle arrest. Therefore, our study serves as a successful example for the potential of in vitro sphere formation as a screening tool to identify compounds that potentially target CSCs, but this technique will require further in vitro and in vivo validation to eliminate false discoveries.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3791/52801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4544916PMC
June 2015

Identification of a distinct population of CD133(+)CXCR4(+) cancer stem cells in ovarian cancer.

Sci Rep 2015 May 28;5:10357. Epub 2015 May 28.

Molecular Immunology and Immuneregulation, Istituto Nazionale per lo Studio e la Cura dei Tumori "Fondazione Giovanni Pascale" IRCCS-ITALIA. Via Mariano Semmola 80131, Naples, Italy.

CD133 and CXCR4 were evaluated in the NCI-60 cell lines to identify cancer stem cell rich populations. Screening revealed that, ovarian OVCAR-3, -4 and -5 and colon cancer HT-29, HCT-116 and SW620 over expressed both proteins. We aimed to isolate cells with stem cell features sorting the cells expressing CXCR4(+)CD133(+) within ovarian cancer cell lines. The sorted population CD133(+)CXCR4(+) demonstrated the highest efficiency in sphere formation in OVCAR-3, OVCAR-4 and OVCAR-5 cells. Moreover OCT4, SOX2, KLF4 and NANOG were highly expressed in CD133(+)CXCR4(+) sorted OVCAR-5 cells. Most strikingly CXCR4(+)CD133(+) sorted OVCAR-5 and -4 cells formed the highest number of tumors when inoculated in nude mice compared to CD133(-)CXCR4(-), CD133(+)CXCR4(-), CD133(-)CXCR4(+) cells. CXCR4(+)CD133(+) OVCAR-5 cells were resistant to cisplatin, overexpressed the ABCG2 surface drug transporter and migrated toward the CXCR4 ligand, CXCL12. Moreover, when human ovarian cancer cells were isolated from 37 primary ovarian cancer, an extremely variable level of CXCR4 and CD133 expression was detected. Thus, in human ovarian cancer cells CXCR4 and CD133 expression identified a discrete population with stem cell properties that regulated tumor development and chemo resistance. This cell population represents a potential therapeutic target.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/srep10357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4650662PMC
May 2015

The miR-17-92 cluster counteracts quiescence and chemoresistance in a distinct subpopulation of pancreatic cancer stem cells.

Gut 2015 Dec 17;64(12):1936-48. Epub 2015 Apr 17.

Stem Cells & Cancer Group, CNIO, Madrid, Spain Barts Cancer Institute, Centre for Stem Cells in Cancer & Ageing, Queen Mary University of London, London, UK.

Objective: Cancer stem cells (CSCs) represent the root of many solid cancers including pancreatic ductal adenocarcinoma, are highly chemoresistant and represent the cellular source for disease relapse. However the mechanisms involved in these processes still need to be fully elucidated. Understanding the mechanisms implicated in chemoresistance and metastasis of pancreatic cancer is critical to improving patient outcomes.

Design: Micro-RNA (miRNA) expression analyses were performed to identify functionally defining epigenetic signatures in pancreatic CSC-enriched sphere-derived cells and gemcitabine-resistant pancreatic CSCs.

Results: We found the miR-17-92 cluster to be downregulated in chemoresistant CSCs versus non-CSCs and demonstrate its crucial relevance for CSC biology. In particular, overexpression of miR-17-92 reduced CSC self-renewal capacity, in vivo tumourigenicity and chemoresistance by targeting multiple NODAL/ACTIVIN/TGF-β1 signalling cascade members as well as directly inhibiting the downstream targets p21, p57 and TBX3. Overexpression of miR-17-92 translated into increased CSC proliferation and their eventual exhaustion via downregulation of p21 and p57. Finally, the translational impact of our findings could be confirmed in preclinical models for pancreatic cancer.

Conclusions: Our findings therefore identify the miR-17-92 cluster as a functionally determining family of miRNAs in CSCs, and highlight the putative potential of developing modulators of this cluster to overcome drug resistance in pancreatic CSCs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1136/gutjnl-2014-308470DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4680182PMC
December 2015

Microenvironmental hCAP-18/LL-37 promotes pancreatic ductal adenocarcinoma by activating its cancer stem cell compartment.

Gut 2015 Dec 3;64(12):1921-35. Epub 2015 Apr 3.

Stem Cells and Cancer Group, Molecular Pathology Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain Centre for Stem Cells in Cancer & Ageing, Barts Cancer Institute, Queen Mary University of London, London, UK.

Objectives: The tumour stroma/microenvironment not only provides structural support for tumour development, but more importantly it provides cues to cancer stem cells (CSCs) that regulate their self-renewal and metastatic potential. This is certainly true for pancreatic ductal adenocarcinomas (PDAC), where tumour-associated fibroblasts, pancreatic stellate cells and immune cells create an abundant paracrine niche for CSCs via microenvironment-secreted factors. Thus understanding the role that tumour stroma cells play in PDAC development and CSC biology is of utmost importance.

Design: Microarray analyses, tumour microarray immunohistochemical assays, in vitro co-culture experiments, recombinant protein treatment approaches and in vivo intervention studies were performed to understand the role that the immunomodulatory cationic antimicrobial peptide 18/LL-37 (hCAP-18/LL-37) plays in PDAC biology.

Results: We found that hCAP-18/LL-37 was strongly expressed in the stroma of advanced primary and secondary PDAC tumours and is secreted by immune cells of the stroma (eg, tumour-associated macrophages) in response to tumour growth factor-β1 and particularly CSC-secreted Nodal/ActivinA. Treatment of pancreatic CSCs with recombinant LL-37 increased pluripotency-associated gene expression, self-renewal, invasion and tumourigenicity via formyl peptide receptor 2 (FPR2)- and P2X purinoceptor 7 receptor (P2X7R)-dependent mechanisms, which could be reversed by inhibiting these receptors. Importantly, in a genetically engineered mouse model of K-Ras-driven pancreatic tumourigenesis, we also showed that tumour formation was inhibited by either reconstituting these mice with bone marrow from cathelicidin-related antimicrobial peptide (ie, murine homologue of hCAP-18/LL-37) knockout mice or by pharmacologically inhibiting FPR2 and P2X7R.

Conclusions: Thus, hCAP-18/LL-37 represents a previously unrecognised PDAC microenvironment factor that plays a critical role in pancreatic CSC-mediated tumourigenesis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1136/gutjnl-2014-308935DOI Listing
December 2015

Inhibition of CD47 Effectively Targets Pancreatic Cancer Stem Cells via Dual Mechanisms.

Clin Cancer Res 2015 May 23;21(10):2325-37. Epub 2015 Feb 23.

Stem Cells and Cancer Group, Molecular Pathology Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain. Centre for Stem Cells in Cancer and Ageing, Barts Cancer Institute, A CR-UK Centre of Excellence, Queen Mary University of London, United Kingdom.

Purpose: Pancreatic ductal adenocarcinoma (PDAC) is a cancer of the exocrine pancreas with unmet medical need and is strongly promoted by tumor-associated macrophages (TAM). The presence of TAMs is associated with poor clinical outcome, and their overall role, therefore, appears to be protumorigenic. The "don't eat me" signal CD47 on cancer cells communicates to the signal regulatory protein-α on macrophages and prevents their phagocytosis. Thus, inhibition of CD47 may offer a new opportunity to turn TAMs against PDAC cells, including cancer stem cells (CSC), as the exclusively tumorigenic population.

Experimental Design: We studied in vitro and in vivo the effects of CD47 inhibition on CSCs using a large set of primary pancreatic cancer (stem) cells as well as xenografts of primary human PDAC tissue.

Results: CD47 was highly expressed on CSCs, but not on other nonmalignant cells in the pancreas. Targeting CD47 efficiently enhanced phagocytosis of a representative set of primary human pancreatic cancer (stem) cells and, even more intriguingly, also directly induced their apoptosis in the absence of macrophages during long-term inhibition of CD47. In patient-derived xenograft models, CD47 targeting alone did not result in relevant slowing of tumor growth, but the addition of gemcitabine or Abraxane resulted in sustained tumor regression and prevention of disease relapse long after discontinuation of treatment.

Conclusions: These data are consistent with efficient in vivo targeting of CSCs, and strongly suggest that CD47 inhibition could be a novel adjuvant treatment strategy for PDAC independent of underlying and highly variable driver mutations.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1078-0432.CCR-14-1399DOI Listing
May 2015

Arg72Pro polymorphism of p53 may predict poor response to medical treatment in ulcerative colitis.

Ann Ital Chir 2014 Nov-Dec;85(6):513-7

Aim: Arg72Pro is a polymorphism commonly occurring in the proline-rich domain of Tp53. It can determine the development of different types of cancers, such as breast, lung, cervical, colorectal and hepatocellular carcinoma. Previous studies reported a correlation between Pro72 homozygosity and the clinical course of ulcerative colitis (UC). Our aim was to evaluate Arg72Pro genotype in patients who underwent proctocolectomy with ileo-pouch-anal anastomosis (IPAA) for UC compared with those who did not need surgery.

Material Of Study: The distribution of the different genotype of Arg72Pro was studied in 264 (234 medically treated [MT] and 30 IPAA) patients affected with UC observed between 2008 and 2011. IPAA patients underwent restorative proctocolectomy for refractory UC; MT ones were managed medically. Blood samples for genotyping were collected from all patients. Arg72Pro genotype analysis was carried out by polymerase chain reaction confronting two-pair primers (PCR-CTPP).

Results: In MT patients (n=234) Arg/Arg, Arg/Pro and Pro/Pro frequencies were 51.28%, 41.02% and 7.7%, respectively, while in IPAA patients (n=30) were 53.4%, 23.3% and 23.3%, respectively. A statistically significant association was found between Pro/Pro and need for surgery (p=<0.0059, χ =7.59).

Conclusion: Our results showed that the Pro/Pro genotype was higher in IPAA (23.3%) than in MT (7.7%) patients. In UC patients the proline homozygosity identifies likeliness to resist to any standard pharmacologic therapy. It could potentially identify patients who would benefit from early surgical treatment, thereby reducing the rate of emergency colectomies and complications related to them.

Key Words: Arg72Pro, Arg72Pro polymorphism, Ileopouch-anal anastomosis, IPAA, p53 polymorphism, Ulcerative colitis.
View Article and Find Full Text PDF

Download full-text PDF

Source
September 2017

Laboratory markers in ulcerative colitis: Current insights and future advances.

World J Gastrointest Pathophysiol 2015 Feb;6(1):13-22

Michele Cioffi, Antonella De Rosa, Rosalba Serao, Ilaria Picone, Maria Teresa Vietri, Department of Biochemistry, Biophysics and General Pathology, Second Medical School Naples, 80138 Naples, Italy.

Ulcerative colitis (UC) and Crohn's disease (CD) are the major forms of inflammatory bowel diseases (IBD) in man. Despite some common features, these forms can be distinguished by different genetic predisposition, risk factors and clinical, endoscopic and histological characteristics. The aetiology of both CD and UC remains unknown, but several evidences suggest that CD and perhaps UC are due to an excessive immune response directed against normal constituents of the intestinal bacterial flora. Tests sometimes invasive are routine for the diagnosis and care of patients with IBD. Diagnosis of UC is based on clinical symptoms combined with radiological and endoscopic investigations. The employment of non-invasive biomarkers is needed. These biomarkers have the potential to avoid invasive diagnostic tests that may result in discomfort and potential complications. The ability to determine the type, severity, prognosis and response to therapy of UC, using biomarkers has long been a goal of clinical researchers. We describe the biomarkers assessed in UC, with special reference to acute-phase proteins and serologic markers and thereafter, we describe the new biological markers and the biological markers could be developed in the future: (1) serum markers of acute phase response: The laboratory tests most used to measure the acute-phase proteins in clinical practice are the serum concentration of C-reactive protein and the erythrocyte sedimentation rate. Other biomarkers of inflammation in UC include platelet count, leukocyte count, and serum albumin and serum orosomucoid concentrations; (2) serologic markers/antibodies: In the last decades serological and immunologic biomarkers have been studied extensively in immunology and have been used in clinical practice to detect specific pathologies. In UC, the presence of these antibodies can aid as surrogate markers for the aberrant host immune response; and (3) future biomarkers: The development of biomarkers in UC will be very important in the future. The progress of molecular biology tools (microarrays, proteomics and nanotechnology) have revolutionised the field of the biomarker discovery. The advances in bioinformatics coupled with cross-disciplinary collaborations have greatly enhanced our ability to retrieve, characterize and analyse large amounts of data generated by the technological advances. The techniques available for biomarkers development are genomics (single nucleotide polymorphism genotyping, pharmacogenetics and gene expression analyses) and proteomics. In the future, the addition of new serological markers will add significant benefit. Correlating serologic markers with genotypes and clinical phenotypes should enhance our understanding of pathophysiology of UC.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4291/wjgp.v6.i1.13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4325297PMC
February 2015

Analysis of PALB2 in a cohort of Italian breast cancer patients: identification of a novel PALB2 truncating mutation.

Fam Cancer 2015 Sep;14(3):341-8

Department of Biochemistry, Biophysics and General Pathology, Second University of Naples, Via Luigi De Crecchio, 7, 80138, Naples, Italy,

PALB2 gene is mutated in about 1-2% of familial breast cancer as well as in 3-4% of familial pancreatic cancer cases. Few studies have reported mutations in Italian patients with breast or pancreatic cancer. We evaluate the occurrence of PALB2 mutations in Italian patients affected with hereditary breast and ovarian cancers and define the pathological significance of the putative allelic variants. We recruited 98 patients (F = 93, M = 5) affected with breast and/or ovarian cancer, negative for mutations in BRCA1 and BRCA2 (BRCAX). Genomic DNA was isolated from peripheral blood lymphocytes, PALB2 coding regions and adjacent intronic were sequenced; in silico predictions were carried out using prediction programs. Mutational analysis of PALB2 gene revealed the novel mutation c.1919C>A (p.S640X) in a 29 years old woman with breast cancer. The c.1919C>A (p.S640X) mutation causes the lack of C-terminus region inducing alteration of MORF4L1-PALB2 association and the lack of interaction of PALB2 with RAD51 and BRCA2. In addition, we identified two novel PALB2 variants, c.3047T>C (p.F1016S) and c.*146A>G. In silico analysis conducted for c.*146A>G indicates that this variant does not affect the splicing while c.3047T>C (p.F1016S) was predicted as damaging in three classifier algorithms. The proband carrier of c.3047T>C (p.F1016S) showed two breast cancer cases, two ovarian cancer cases and one pancreatic cancer in mother's family. c.3047T>C (p.F1016S) and c.*146A>G should be considered PALB2 UVs even though the genotype-phenotype correlation for these variants remains still unclear. Our findings indicate that the presence of PALB2 mutation should be routinely investigated in hereditary breast and ovarian cancers families since it could be of clinical relevance for clinical management.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10689-015-9786-zDOI Listing
September 2015

A novel PALB2 truncating mutation in an Italian family with male breast cancer.

Oncol Rep 2015 Mar 22;33(3):1243-7. Epub 2014 Dec 22.

Department of Biochemistry, Biophysics and General Pathology, School of Medicine, Second University of Naples, I-80138 Naples, Italy.

Male breast cancer (MBC) is a rare disease, accounting for ~1% of all breast cancer cases worldwide. Although other genes are also involved, predisposing genetic factors to MBC include germline mutations in the BRCA genes (BRCA2). Among the other genes, partner and localizer of BRCA2 (PALB2) is considered a moderate-penetrance breast cancer susceptibility gene that may also play a role in MBC predisposition. Thus, the aim of the present study was to determine the PALB2 gene status in 8 MBC cases selected from a cohort of 181 hereditary breast and/or ovarian cancer probands. We performed PALB2 mutational analysis by direct sequencing of 13 exons and adjacent intronic regions. This study showed the presence of a PALB2 truncating mutation in 1/8 (12.5%) cases. This novel mutation was named c.1285_1286delAinsTC (p.I429SfsX12) and is localized in exon 4 of PALB2, in the region encoding for the ChAM motif which is important for the efficient association of PALB2 to chromatin and for recruitment of the BRCA complex to accumulate RAD51 at double-strand break sites. Our findings indicate that PALB2 could be added to the list of breast cancer susceptibility genes also in families with MBC cases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3892/or.2014.3685DOI Listing
March 2015

Intracellular autofluorescence: a biomarker for epithelial cancer stem cells.

Nat Methods 2014 Nov 28;11(11):1161-9. Epub 2014 Sep 28.

1] Stem Cells and Cancer Group, Molecular Pathology Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain. [2] Centre for Stem Cells in Cancer &Ageing, Barts Cancer Institute, Queen Mary University of London, London, UK.

Cancer stem cells (CSCs) are thought to drive tumor growth, metastasis and chemoresistance. Although surface markers such as CD133 and CD44 have been successfully used to isolate CSCs, their expression is not exclusively linked to the CSC phenotype and is prone to environmental alteration. We identified cells with an autofluorescent subcellular compartment that exclusively showed CSC features across different human tumor types. Primary tumor-derived autofluorescent cells did not overlap with side-population (SP) cells, were enriched in sphere culture and during chemotherapy, strongly expressed pluripotency-associated genes, were highly metastatic and showed long-term in vivo tumorigenicity, even at the single-cell level. Autofluorescence was due to riboflavin accumulation in membrane-bounded cytoplasmic structures bearing ATP-dependent ABCG2 transporters. In summary, we identified and characterized an intrinsic autofluorescent phenotype in CSCs of diverse epithelial cancers and used this marker to isolate and characterize these cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nmeth.3112DOI Listing
November 2014

Chloroquine targets pancreatic cancer stem cells via inhibition of CXCR4 and hedgehog signaling.

Mol Cancer Ther 2014 Jul 30;13(7):1758-71. Epub 2014 Apr 30.

Authors' Affiliations: Stem Cells & Cancer Group, Molecular Pathology Programme, Spanish National Cancer Research Centre (CNIO); Barts Cancer Institute, Queen Mary University of London, London, United Kingdom

Pancreatic ductal adenocarcinoma is one of the deadliest carcinomas and is characterized by highly tumorigenic and metastatic cancer stem cells (CSC). CSCs evade available therapies, which preferentially target highly proliferative and more differentiated progenies, leaving behind CSCs as a putative source for disease relapse. Thus, to identify potentially more effective treatment regimens, we screened established and new compounds for their ability to eliminate CSCs in primary pancreatic cancer (stem) cells in vitro and corresponding patient-derived pancreatic cancer tissue xenografts in vivo. Intriguingly, we found that in vitro treatment with the antimalarial agent chloroquine significantly decreased CSCs, translating into diminished in vivo tumorigenicity and invasiveness in a large panel of pancreatic cancers. In vivo treatment in combination with gemcitabine was capable of more effectively eliminating established tumors and improved overall survival. The inhibitory effect of chloroquine was not related to inhibition of autophagy, but was due to inhibition of CXCL12/CXCR4 signaling, resulting in reduced phosphorylation of ERK and STAT3. Furthermore, chloroquine showed potent inhibition of hedgehog signaling by decreasing the production of Smoothened, translating into a significant reduction in sonic hedgehog-induced chemotaxis and downregulation of downstream targets in CSCs and the surrounding stroma. Our study demonstrates that via to date unreported effects, chloroquine is an effective adjuvant therapy to chemotherapy, offering more efficient tumor elimination and improved cure rates. Chloroquine should be further explored in the clinical setting as its success may help to more rapidly improve the poor prognosis of patients with pancreatic cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1535-7163.MCT-13-0948DOI Listing
July 2014

Clinical features of a new acid-labile subunit (IGFALS) heterozygous mutation: anthropometric and biochemical characterization and response to growth hormone administration.

Horm Res Paediatr 2014 12;81(1):67-72. Epub 2013 Dec 12.

Department of Woman, Child and General and Specialized Surgery, Seconda Università degli Studi di Napoli, Naples, Italy.

Background: Homozygous mutations in acid-labile subunit (IGFALS) gene result in short stature, very low circulating levels of acid-labile subunit (ALS), insulin growth factor 1 (IGF1) and insulin growth factor binding protein 3 (IGFBP3) and a poor response to growth hormone (GH). The impact of IGFALS mutations heterozygosity on growth is unknown.

Patient And Methods: We describe a 10-year-old girl with severe short stature (height -3.2 SDS), heterozygous for a new IGFALS mutation.

Results: The girl showed low circulating IGF1, IGFBP3 and ALS levels and normal GH secretion. We found a novel heterozygous frameshift IGFALS mutation (c.1283delA, p.Gln428Argfs*14). Size-exclusion chromatography showed a reduction of the IGF1, IGFBP3 and ALS 150-kDa ternary complex (by about 55%) compared to a control. An IGF-1 generation test, with two different GH dosages, showed a good response in term of increase in IGF1 and in formation of the ternary complex at size-exclusion chromatography. Clinical response after 6 months of therapy with GH was satisfactory (height velocity increased from 3 to 8 cm/year).

Conclusion: We suggest that (1) heterozygous IGFALS mutations can be responsible for a subset of patients with severe short stature (below -2.5 SDS), low IGF1 (below -2 SDS) and normal GH secretion, and (2) the identification by IGFALS molecular screening of this subset of patients could help in the administration of the appropriate therapy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1159/000355017DOI Listing
January 2015

Metformin targets the metabolic achilles heel of human pancreatic cancer stem cells.

PLoS One 2013 18;8(10):e76518. Epub 2013 Oct 18.

Stem Cells & Cancer Group, Molecular Pathology Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.

Pancreatic ductal adenocarcinomas contain a subset of exclusively tumorigenic cancer stem cells (CSCs), which are capable of repopulating the entire heterogeneous cancer cell populations and are highly resistant to standard chemotherapy. Here we demonstrate that metformin selectively ablated pancreatic CSCs as evidenced by diminished expression of pluripotency-associated genes and CSC-associated surface markers. Subsequently, the ability of metformin-treated CSCs to clonally expand in vitro was irreversibly abrogated by inducing apoptosis. In contrast, non-CSCs preferentially responded by cell cycle arrest, but were not eliminated by metformin treatment. Mechanistically, metformin increased reactive oxygen species production in CSC and reduced their mitochondrial transmembrane potential. The subsequent induction of lethal energy crisis in CSCs was independent of AMPK/mTOR. Finally, in primary cancer tissue xenograft models metformin effectively reduced tumor burden and prevented disease progression; if combined with a stroma-targeting smoothened inhibitor for enhanced tissue penetration, while gemcitabine actually appeared dispensable.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0076518PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3799760PMC
August 2014

Double heterozygosity in the BRCA1 and BRCA2 genes in Italian family.

Clin Chem Lab Med 2013 Dec;51(12):2319-24

Background: Double heterozygosity (DH) is an extremely rare event in which both BRCA1 and BRCA2 are mutated simultaneously in a family. To date, few cases of DH have been reported, especially in Ashkenazi populations. In Italy some cases of DH have been reported. In this study, we have described an Italian family with double heterozygosity in the BRCA genes.

Methods: The proband is a 43-year-old woman with bilateral breast cancer. She presented two pathogenic mutations in both BRCA genes, IVS8+2T>A (c.547+2T>A;p.Gln148Aspfsx51) in BRCA1, K944X (c.2830A>T;p.Lys944X) in BRCA2 and a novel variant IVS4-57A>G (c.426-57A>G) in BRCA2, not previously described. Both mutations were inherited from the paternal lineage in the proband's family. We investigated all available members of this family and we identified other two family members with DH.

Results And Conclusions: Our observations support the hypothesis of a non-specific severe phenotype in DH carriers in terms of age of disease onset, cumulative lifetime risk and multiple primary tumours. Furthermore, our findings confirm that in order to identify all cases of DH, it is important not to limit the identification of mutations in a single gene, but extend the analysis to BRCA1 and BRCA2 and other breast cancer susceptibility genes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1515/cclm-2013-0263DOI Listing
December 2013

Identification of a novel in-frame deletion in BRCA2 and analysis of variants of BRCA1/2 in Italian patients affected with hereditary breast and ovarian cancer.

Clin Chem Lab Med 2012 Dec;50(12):2171-80

Dipartimento di Patologia Generale , Facoltà di Medicina e Chirurgia Seconda Università degli studi, Naples , Italy

Background: An estimated 5 % – 10 % of all breast cancers are due to an inherited predisposition and, out of these, about 30 % are caused by germline mutations of the BRCA1 and BRCA2 genes. The prevalence of germline mutations in theBRCA1 and BRCA2 genes varies among ethnic groups. The aims of this study are to evaluate deleterious mutations and genomic rearrangements in BRCA1/2 genes and the CHEK21100delC mutation in a cohort of Italian women affected with hereditary breast and/or ovarian cancer. In addition we clarify the effect of the novel variants identified in BRCA2 gene bymRNA analysis and prediction software.

Methods: We enrolled 103 consecutive Italian patients affected with hereditary breast and/or ovarian cancer, aged23 – 69 years.

Results: We found BRCA1/2 mutations in 15/103 probands(14.6 % ). Among these, a BRCA2 gene mutation has not been described previously. In addition, we identified five novel BRCA2 variants (S1341, IVS1-59t > c, IVS11-74insA, IVS12 + 74c > g and I1167V). No genomic BRCA1/2re arrangements or CHEK2 1100delC mutation was found in our patients. The novel BRCA2 mutation NS1742del(p.N1742_S1743del) was an in-frame 6 bp deletion that results in loss of two amino acids.

Conclusions: In silico analysis conducted for S1341, IVS1-59t > c, IVS11-74insA and IVS12 + 74c > g of BRCA2 predicted the variants as neutral and benign, whereas the results for I1167V was inconclusive. mRNA analysis for the novel BRCA2 intronic variant IVS11-74insA and the already published BRCA1 variant C197 shows that they have no effect on the splicing. These results are in agreement with in silico analysis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1515/cclm-2012-0154DOI Listing
December 2012

Immuno-targeting of pancreatic cancer stem cells: A new therapeutic strategy against a devastating disease?

Oncoimmunology 2012 Jul;1(4):560-562

Stem Cells & Cancer Group; Clinical Research Programme; Spanish National Cancer Research Centre (CNIO); Madrid, Spain.

Pancreatic cancer is a highly aggressive and deadly disease harboring a distinct population of cancer stem cells (CSC) that is not affected by conventional therapies. A new therapeutic approach using the EpCAM/CD3-bispecific antibody MT110 is capable of activating and redirecting cytotoxic T cells to eliminate primary human pancreatic cancer stem cells, which resulted in long-term survival of preclinical xenografts models.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3382890PMC
http://dx.doi.org/10.4161/onci.19368DOI Listing
July 2012
-->