The emerging role of FKBP51 in melanoma: characterization of a promising molecular target for innovative therapies

Melanoma is a malignant skin cancer with high metastatic potential. The number of melanoma cases worldwide is increasing faster than any other cancer. The prognosis is bad in the advanced stages of the disease due to resistance to conventional anti-cancer treatments. Patients diagnosed with advanced stage melanoma continue to pose a significant challenge for clinicians. Although many therapeutic regimens for metastatic melanoma have been tested, very few achieve response rates greater than 25%. There is much hope for targeted therapies and promising agents include those that act on apoptosis-regulating molecules. Very recently, we have identified that an immunophilin, namely FK506 binding protein (FKBP) 51 controls response of melanoma to DNA damaging agents, such as anthracyclines compounds. The aim of this study was to investigate the possible role of FKBP51 in the control of response to ionizing radiation (IR) in malignant melanoma. FKBP51-silenced cells showed reduced clonogenic potential after irradiation compared with non-silenced cells. After IR, we observed apoptosis in FKBP51-silenced cells and autophagy in non-silenced cells. FKBP51 was required for the activation of Rx-induced NF-kappaB, which in turn inhibited apoptosis by stimulating X-linked inhibitor of apoptosis protein and promoting authophagymediated Bax degradation. Using a tumor-xenograft mouse model, the in vivo pretreatment of tumors with FKBP51-siRNA provoked massive apoptosis after irradiation. Immunohistochemical analysis of 10 normal skin samples and 80 malignant cutaneous melanomas showed that FKBP51 is a marker of melanocyte malignancy, correlating with vertical growth phase and lesion thickness. Moreover, we provided evidence that FKBP51 targeting radiosensitizes cancer stem/initiating cells. Finally, we expanded the study to other types of cancer, including ovarian, pancreatic, prostate, colon, breast and lung cancer and found a strong correlation between the protein expression and malignity of the tumoral lesions. In conclusion, our study identifies a possible molecular target for radiosensitizing therapeutic strategies against malignant melanoma and a potential novel tumoral biomarker

Tumour prevention and tumour progression: a dual role for statins?☆

www.sciencedirect.com The use of statins is essential for the treatment of hyperlipidemia as well as for the primary and secondary prevention of coronary artery disease and strokes. Statins decrease low-density lipoprotein (LDL) cholesterol levels by inhibiting 3-hydroxy-3-methyl-glutaryl-CoA HMG-CoA reductase (HMGCR). HMGCR in turn catalyses the conversion of HMG-CoA into mevalonic acid, an important intermediate metabolite in hepatic cholesterol biosynthesis. Statins exert pleiotropic effects independent of their cholesterol-lowering activity, which are in part mediated by mevalonic acid, a precursor of the isoprenoid intermediates farnesyl and geranyl-geranyl pyrophosphate [1]. These compounds participate in the post-translational modification of intracellular G-proteins, such as Rho, Rac, and Ras. In turn, it is well-known that G-proteins drive signalling pathways that are widely involved in carcinogenesis [2], and their inhibition by statins has been proven to efficiently impair the growth of several tumours [3]. Furthermore, statins inhibit cellular matrix metalloproteinases and nuclear factor-kB (NFkB) transcription factors that are, additionally, often deregulated in cancer. A large body of studies exists that support the efficacy of statins against several cancer types [3]. For this reason, these compounds has been proposed as adjuvant options in cancer therapies and in cancer prevention. Currently, recommendations and guidelines on statin use in malignancies do not exist; in addition, to date, results from randomised controlled trials are inconclusive regarding the question of whether cancer treatment may benefit from statins

FKBP51 in cancer

Overview sulla nostra ricerca condotta negli ultimi 10 anni, sul ruolo della immunofillina FKBP51 nel cancro, e particolarmente nel melanoma. Introduzione della nostra recente identificazione di una variante di splicing della proteina, fino al 2015 sconosciuta

The effect of FK506 on transforming growth factor beta signaling and apoptosis in chronic lymphocytic leukemia B cells.

Background Loss of response to transforming growth factor-beta (TGF-beta) is thought to contribute to the progression of chronic lymphocytic leukemia. Recent findings of over-activation of the TGF-beta signal in FKBP12-knockout mouse prompted us to investigate whether FK506, the canonical ligand of FKBP, can activate the TGF-beta signal in chronic lymphocytic leukemia. Design and Methods We studied 62 chronic lymphocytic leukemia samples from patients with Rai/Binet stage 0 to 4 disease. The TGF-beta signal was investigated by western blotting and flow cytometry. The levels of Bcl2-family members and death-associated-protein kinase were also investigated by western blotting, whereas apoptosis was studied in flow cytometry. Down-modulation of FKBP12 was obtained by gene silencing with short interfering RNA. Results Twenty-two out of 62 chronic lymphocytic leukemia samples were sensitive to TGF-beta-induced apoptosis. All but two of the responsive samples underwent apoptosis also when cultured with FK506, but not with cyclosporine. Thirteen samples that were not sensitive to TGF-beta were sensitive to FK506. Overall, response to FK506 occurred in 33 samples. FK506 induced Smad2 phosphorylation and nuclear translocation. Accordingly, death-associated-protein kinase, a transcriptional target of Smad, was induced. At the same time, Bcl-2 and Bcl-xL levels decreased whereas the levels of Bim and Bmf increased. A loss of mitochondrial membrane potential preceded caspase activation and cell death. FK506 removed FKBP12 from its binding to the TGF-beta-receptor. FKBP12 release activated the receptor-kinase activity as suggested by the enhanced levels of phospho-Smad found in cells depleted of FKBP12. Conclusions Our study shows that most chronic lymphocytic leukemia cells escape the homeostatic control of TGF-beta and that FK506 restores the TGF-beta signal in a proportion of non-responsive samples. We demonstrated that FK506 activates TGF-beta receptor I kinase activity in chronic lymphocytic leukemia, which transduces apoptosis by a mitochondrial-dependent pathway

Geophysical multidisciplinary investigation of the structure of an unstable flank: the NE sector of Mt. Etna.

Mount Etna is characterized by a complex regional tectonics with a N-S compression related to the Africa – Europe convergence that interacts with a WNW-ESE extension associated to the Malta Escarpment. A general eastward motion is present in the eastern flank. Although the existence of these phenomena is overt, the geometry of the sliding sector is still debated. The non-uniqueness of the geophysical inverse models and the different limitations in resolution and sensitivity of each technique spurred us to undertake, in the frame of the MEDiterranean Supersites Volcanoes (MED-SUV) project, a joint interpretation of independent data in order to better constrain the results. Seismic data come from the network run by the Istituto Nazionale di Geofisica e Vulcanologia (INGV) – Osservatorio Etneo, Sezione di Catania. The relocated seismicity defines two main seismogenic volumes in the NE sector of the volcano: the first cluster is related to the known Pernicana Fault system, while the second one is located southwards, beneath the northern wall of the Valle del Bove. The resistivity models come from a MT survey carried out on the eastern flank of the volcano and consisting of thirty broad-band soundings along N-S and NW-SE oriented profiles. The resistivity modeling of MT profiles reveal three major layers in a resistive-conductive-resistive sequence. A low resistivity volume is clearly identified on the NE flank of the volcano, between The Pernicana fault and the northern wall of the Valle del Bove. Ground deformation studies (GPS and InSAR) revealed the segmentation of the unstable flank and define the NE sector as the most mobile one; this sector is perfectly bounded by the two seismic clusters and corresponds to the low resistivity volume. The sliding surface modeled by ground deformation data inversions well matches in depth with a resistivity transition and with two seismogenic layers

Neuropilin 1mediates keratinocyte growth factor signaling in adipose-derived stem cells: potential involvement in adipogenesis

Adipogenesis is regulated by a complex network of molecules, including fibroblast growth factors. Keratinocyte growth factor (KGF) has been previously reported to promote proliferation on rat preadipocytes, although the expression of its specific receptor, FGFR2-IIIb/KGFR, is not actually detected in mesenchymal cells. Here, we demonstrate that human adipose-derived stem cells (ASCs) show increased expression of KGF during adipogenic differentiation, especially in the early steps. Moreover, KGF is able to induce transient activation of ERK and p38 MAPK pathways in these cells. Furthermore, KGF promotes ASC differentiation and supports the activation of differentiation pathways, such as those of PI3K/Akt and the retinoblastoma protein (Rb). Notably, we observed only a low amount of FGFR2-IIIb in ASCs, which seems not to be responsible for KGF activity. Here, we demonstrate for the first time that Neuropilin 1 (NRP1), a transmembrane glycoprotein expressed in ASCs acting as a coreceptor for some growth factors, is responsible for KGF-dependent pathway activation in these cells. Our study contributes to clarify the molecular bases of human adipogenesis, demonstrating a role of KGF in the early steps of this process, and points out a role of NRP1 as a previously unknown mediator of KGF action in ASCs

Silencing of Keratinocyte Growth Factor Receptor Restores 5-Fluorouracil and Tamoxifen Efficacy on Responsive Cancer Cells

BACKGROUND: Keratinocyte growth factor receptor (KGFR) is a splice variant of the FGFR2 gene expressed in epithelial cells. Activation of KGFR is a key factor in the regulation of physiological processes in epithelial cells such as proliferation, differentiation and wound healing. Alterations of KGFR signaling have been linked to the pathogenesis of different epithelial tumors. It has been also hypothesized that its specific ligand, KGF, might contribute to the development of resistance to 5-fluorouracil (5-FU) in epithelial cancers and tamoxifen in estrogen-positive breast cancers. METHODOLOGY/PRINCIPAL FINDINGS: Small interfering RNA was transfected into a human keratinocyte cell line (HaCaT), a breast cancer derived cell line (MCF-7) and a keratinocyte primary culture (KCs) to induce selective downregulation of KGFR expression. A strong and highly specific reduction of KGFR expression was observed at both RNA (reduction = 75.7%, P = 0.009) and protein level. KGFR silenced cells showed a reduced responsiveness to KGF treatment as assessed by measuring proliferation rate (14.2% versus 39.0% of the control cells, P<0.001) and cell migration (24.6% versus 96.4% of the control cells, P = 0.009). In mock-transfected MCF-7 cells, KGF counteracts the capacity of 5-FU to inhibit cell proliferation, whereas in KGFR silenced cells KGF weakly interferes with 5-FU antiproliferative effect (11.2% versus 28.4% of the control cells, P = 0.002). The capacity of 5-FU to induce cell death is abrogated by co-treatment with KGF, whereas in KGFR silenced cells 5-FU efficiently induces cell death even combined to KGF, as determined by evaluating cell viability. Similarly, the capacity of tamoxifen to inhibit MCF-7 and KCs proliferation is highly reduced by KGF treatment and is completely restored in KGFR silenced cells (12.3% versus 45.5% of the control cells, P<0.001). CONCLUSIONS/SIGNIFICANCE: These findings suggest that selective inhibition of the KGF/KGFR pathway may provide a useful tool to ameliorate the efficacy of the therapeutic strategies for certain epithelial tumors

Gastrointestinal symptoms in children: Primary care and specialist interface.

Aims Gastrointestinal symptoms and diseases represent one of the major reasons for paediatricians' requests for specialist consultations and hospital admissions. One fourth of annual medical consultations for children younger than 6 years can be attributed to gastrointestinal symptoms. High-quality guidelines have been validated worldwide to provide clinical recommendations and support healthcare providers' practice. Nevertheless, overall compliance to standards of care is unsatisfactory, and children with gastrointestinal symptoms frequently undergo expensive, useless specialist consultations and laboratory evaluations. The aim of this study is to review the main epidemiological and clinical aspects, together with management strategies, of the most common gastrointestinal symptoms in children, pointing out pitfalls and practical tips in primary care management, and providing correct indications for specialist consultations. Methods For this review, articles published in English from 2000 to January 2018 were identified from the PubMed/Medline (http://www.ncbi.nlm.nih.gov/pubmed/) database and selected on the basis of quality, relevance to the illness and importance in illustrating current management pathways. The search used the following keywords: gastrointestinal symptoms, functional gastrointestinal symptoms, children, primary care, specialist consultations and management. Particular emphasis was placed on evidence-based guidelines and high-quality studies. Results Functional gastrointestinal symptoms have a high impact on the quality of life of children and families and on healthcare costs. A complete medical history and clinical examination are often sufficient to guide the primary care provider in the diagnosis, further workup or referral to a paediatric gastroenterologist. Conclusion Paediatric gastroenterology outpatients' clinics are among the most crowded specialists, and functional gastrointestinal symptoms and disorders are the most frequent reason for counselling. The number of specialist consultations could be reduced if guidelines were applied in primary care settings

Neurophysiological monitoring in neonatal abstinence syndrome from cocaine

Introduction. Neonatal abstinence syndrome (NAS) in a newborn is a result of the sudden discontinuation of exposure to psychotropic drugs abused by the mother during pregnancy. Since forty decades, the standardized Finnegan Neonatal Abstinence Scoring Tool (FNAST) documents the infant withdrawal, and initiate the appropriate treatment regimen, when elevated scored are reported. Whereas FNAST is successfully applied for opioids NAS, in case of other psychotropic drugs and especially cocaine, the tool is not always efficacious or predictive. Methods. Continuous v-Electroencephalography (vEEG) provides particularly useful information about brain cortical functioning and evaluation of background activity in normal newborns. vEEG allows to properly study and identify clinical manifestations as physiological motor paroxysms, that disappear from birth to infant age in correlation with the neurological development. Due to its feature to be a non-invasive tool continuous vEEG monitoring could be used to describe some clinical manifestations and assess if they can be correlated to possible injuries in critical neonates as those exposed in utero to psychoactive drugs presenting NAS. Results. An example for the potential use of such methodology is discussed in a case of NAS due to prenatal exposure to cocaine as a complementary tool for the evaluation of behavioural state and clinical and neurological signs in newborns in utero exposed to psychoactive drugs, excluding epileptic phenomena. Discussion. Video-EEG recording could be considered an important and objective tool that allows the evaluation of behavioural state and clinical and neurological signs in newborns in utero exposed to psychoactive drugs and the neurophysiological definition of signs and symptoms, which cannot be evaluated by FNAST such as startles and its variability during subsequent days after birth, subclinical seizures or brain injuries

Fluid conduits and shallow-reservoir structure defined by geoelectrical tomography at the Nirano Salse (Italy)

Mud volcanoes are fluid escape structures allowing for surface venting of hydrocarbons (mostly gas but also liquid condensates and oils) and water–sediment slurries. For a better understanding of mud volcano dynamics, the characterization of the fluid dynamics within mud volcano conduits; the presence, extent, and depth of the fluid reservoirs; and the connection among aquifers, conduits, and mud reservoirs play a key role. To this aim, we performed a geoelectricalsurvey in the Nirano Salse Regional Nature Reserve, located at the edge of the northern Apennines (Fiorano Modenese, Italy), an area characterized by several active mud fluid vents. This study, for the first time, images the resistivity structure of the subsoil along two perpendicular cross sections down to a depth of 250 m. The electrical models show a clear difference between the northern and southern sectors of the area, where the latter hosts the main discontinuities. Shallow reservoirs, where fluid muds accumulate, are spatially associated with the main fault/fracture controlling the migration routes associated with surface venting and converge at depth towards a common clayey horizon. There is no evidence of a shallow mud caldera below the Nirano area. These findings represent a step forward in the comprehension of the Nirano Salse plumbing system and in pinpointing local site hazards, which promotes safer tourist access to the area along restricted routes