Deregulation Of Signaling Pathways Research Articles

Protein Phosphatase 1 Regulates Human Cytomegalovirus Protein Translation by Restraining AMPK Signaling

Human cytomegalovirus (HCMV) carries the human protein phosphatase 1 (PP1) and other human proteins important for protein translation in its tegument layer for a rapid supply upon infection. However, the biological relevance behind PP1 incorporation and its role during infection is unclear. Additionally, PP1 is a difficult molecular target due to its promiscuity and similarities between the catalytic domain of multiple phosphatases. In this study, we circumvented these shortcomings by using 1E7-03, a small molecule protein–protein interaction inhibitor, as a molecular tool of noncatalytic PP1 inhibition. 1E7-03 treatment of human fibroblasts severely impaired HCMV replication and viral protein translation. More specifically, PP1 inhibition led to the deregulation of metabolic signaling pathways starting at very early time points post-infection. This effect was at least partly mediated by the prevention of AMP-activated protein kinase dephosphorylation, leading to elongation factor 2 hyperphosphorylation and reduced translation rates. These findings reveal an important mechanism of PP1 for lytic HCMV infection.

Mitogen signal-associated pathways, energy metabolism regulation, and mediation of tumor immunogenicity play essential roles in the cellular response of malignant pleural mesotheliomas to platinum-based treatment: a retrospective study.

BackgroundMalignant pleural mesothelioma (MPM) is a rare malignant tumor associated with asbestos exposure, with infaust prognosis and overall survival below 20 months in treated patients. Platinum is still the backbone of the chemotherapy protocols, and the reasons for the rather poor efficacy of platinum compounds in MPM remain largely unknown. Therefore, we aimed to analyze differences in key signaling pathways and biological mechanisms in therapy-naïve samples and samples after chemotherapy in order to evaluate the effect of platinum-based chemotherapy.MethodsThe study cohort comprised 24 MPM tumor specimens, 12 from therapy-naïve and 12 from patients after platinum-based therapy. Tumor samples were screened using the NanoString nCounter platform for digital gene expression analysis with an appurtenant custom-designed panel comprising a total of 366 mRNAs covering the most important tumor signaling pathways. Significant pathway associations were identified by gene set enrichment analysis using the WEB-based GEne SeT AnaLysis Toolkit (WebGestalt)ResultsWe have found reduced activity of TNF (normalized enrichment score: 2.03), IL-17 (normalized enrichment score: 1.93), MAPK (normalized enrichment score: 1.51), and relaxin signaling pathways (normalized enrichment score: 1.42) in the samples obtained after platinum-based therapy. In contrast, AMPK (normalized enrichment score: –1.58), mTOR (normalized enrichment score: –1.50), Wnt (normalized enrichment score: –1.38), and longevity regulating pathway (normalized enrichment score: –1.31) showed significantly elevated expression in the same samples.ConclusionsWe could identify deregulated signaling pathways due to a directed cellular response to platinum-induced cell stress. Our results are paving the ground for a better understanding of cellular responses and escape mechanisms, carrying a high potential for improved clinical management of patients with MPM.

Gene expression analysis in EBV-infected ataxia-telangiectasia cell lines by RNA-sequencing reveals protein synthesis defect and immune abnormalities

BackgroundEpstein–Barr virus (EBV) targets B-cells where it establishes a latent infection. EBV can transform B-cells in vitro and is recognized as an oncogenic virus, especially in the setting of immune compromise. Indeed, immunodeficient patients may fail to control chronic EBV infection, leading to the development EBV-driven lymphoid malignancies. Ataxia telangiectasia (AT) is a primary immune deficiency caused by mutations in the ATM gene, involved in the repair of double-strand breaks. Patients with AT are at high risk of developing cancers, mostly B-cell lymphoid malignancies, most of which being EBV-related. Aside from immune deficiency secondary to AT, loss of ATM function could also hinder the control of the virus within B-cells, favoring lymphomagenesis in AT patients.ResultsWe used RNA sequencing on lymphoblastoid cell lines derived from patients with AT and healthy donors to analyze and compare both cellular and viral gene expression. We found numerous deregulated signaling pathways involving transcription, translation, oncogenesis and immune regulation. Specifically, the translational defect was confirmed in vitro, suggesting that the pathogenesis of AT may also involve a ribosomal defect. Concomitant analysis of viral gene expression did not reveal significant differential gene expression, however, analysis of EBV interactome suggests that the viral latency genes EBNA-3A, EBNA-3C and LMP1 may be disrupted in LCL from AT patients.ConclusionOur data support the notion that ATM deficiency deregulates cellular gene expression possibly disrupting interactions with EBV latent genes, promoting the oncogenic potential of the virus. These preliminary findings provide a new step towards the understanding of EBV regulation and of AT pathogenesis.

Deferasirox Induces Apoptosis in Mantle Cell Lymphoma in a Mechanism Involving Cyclin-D1, p53 and the Ribonucleotide Reductase

Mantle cell lymphoma (MCL) is a difficult to treat B-cell malignancy characterized by a rapid progressive course and poor prognosis. Despite advances in conventional and novel agents such as bortezomib, lenalidomide and ibrutinib, many MCL patients are still subject to rapid relapses of the disease, responsible for the relatively short overall survival. MCL is characterized by the t(11,14) translocation, resulting in cyclin-D1 (CD1) overexpression and dysregulation of the cell cycle. Recently, it has been demonstrated that depletion of CD1 results in cell cycle arrest, apoptosis and cell death of MCL cells thus labeling CD1 as a therapeutic target of this aggressive malignancy. Accumulating findings indicate that not only overexpressed CD1 but also deregulated signaling pathways such as PI3K/AKT/mTOR and Wnt are accountable for the tumorigenesis of MCL and for the maintenance of its proliferative and survival capacity. We thus believe that MCL treatment should target multiple deregulated signaling pathways and key molecules. Iron chelators have emerged as a novel strategy to treat tumors due to their capacity to target a multitude of key molecules, signaling pathways and cellular processes. Yet, the application of common iron chelators is limited due to their low bioavailability and high toxicities. Deferasirox (DFX), a clinically approved iron chelator applied for the treatment of iron overload, in contrast to the majority of iron chelators, is highly bioavailable and exhibits little to no side effects. To date, DFX is reported as being an orally effective antitumor agent against solid tumors by various mechanisms. In our project, we examined the in vitro impact of DFX on three MCL cell lines; Jeko-1, REC-1 and Z138; aiming thereby to resolve its molecular mode of action. DFX inhibited the growth of the MCL cell lines with 50% inhibitory concentrations (IC50) of 8.07±1.08µM, 1.8±1.03µM and 1.52±1.54µM for Jeko-1, REC-1 and Z138 cell lines, respectively. Moreover, we demonstrated that DFX treatment trigged a rapid and intense (~2-fold) production of reactive oxygen species (ROS), induced severe DNA damage and arrested cells at the G1 phase within 6 hrs. Ultimately, a 72h treatment of 10µM DFX led to apoptosis in 60% to 86% of the MCL cells. Most importantly, we found DFX to substantially reduce CD1 protein levels in a dose- and time-dependent manner. The depletion of CD1 levels was accompanied with its phosphorylation at Thr-286 (pCD1), a phosphorylation known to sentence CD1 to proteasomal degradation. In agreement with previous studies that have shown a correlation between ROS induction, CD1-phosphorylation, CD1 depletion and DNA damage, we found CD1 levels to negatively correlate with increasing DNA damage levels following DFX treatment.Interestingly, the efficacy of DFX treatment was dependent on the p53-status of the respective cell line. The wild-type p53 expressing lines, REC-1 and Z138, were found to be ~5-fold more sensitive to DFX than the p53 negative Jeko-1 cell line. In an attempt to understand this intriguing finding, we found DFX treatment to post-translationally elevate and stabilize p53 levels, most likely by depleting the intracellular heme pool which serves as a negative p53-regulator. Furthermore, we showed that DFX plays an important role in the attenuation of the iron dependent ribonucleotide reductase enzyme (RR); the rate limiting enzyme in dNTP biosynthesis and thus the key enzyme in DNA synthesis, cell cycle regulation and proliferation. DFX inhibited the enzymatic activity of RR with an IC50 of ~25 µM; which is ~20-fold more effective than other established inhibitors such as deferoxamine.In conclusion and pending further analysis, we demonstrated a potent impact of DFX on MCL cells. The vigorous anti-tumoral activity of DFX is most likely mediated via its ability to induce oxidative stress, to inhibit the RR enzyme and to eliminate the main oncogene in MCL pathogenesis; CD1. The depleting of intracellular iron/heme pools and the modulation of p53 only intensify the effect of DFX. The ability to inhibit the RR activity qualifies DFX as a sensitizer to existing DNA damaging radio- and chemotherapy options of MCL. DFX possesses the unique capacity to affect a multitude of cellular mechanisms in an auto-potentiating manner that propose this clinically established molecule as a potential therapeutic option for mantle cell lymphoma. DisclosuresNo relevant conflicts of interest to declare.

Integrated Phosphoproteomic and N-Glycoproteomic Landscape of Cutaneous T-Cell Lymphoma Reveals Distinctive Pathogenetic Insights and Therapeutic Vulnerabilities

Background: A critical goal of precision oncology is the identification of deregulated signaling pathways that serve as disease-specific biomarkers and vulnerabilities that may be exploited as therapeutic targets. Cutaneous T cell lymphomas (CTCLs) are heterogeneous neoplasms with variable clinical outcomes. Whereas the genomic landscapes of CTCLs are being characterized, a large scale analysis of their proteomic signatures and post-translational modifications (PTM) has not been performed. In this study, we sought to gain insights into the pathogenesis of CTCLs using high mass accuracy tandem mass spectrometry (HMA-MS/MS)-based phosphoproteomics and glycoproteomics.Methods: CTCL-derived cell lines representative of Sézary syndrome (SS, HUT78), mycosis fungoides (MF, HH), and ALK-negative anaplastic large cell lymphoma (ALCL) with PCM1-JAK2 or NPM1-TYK2 translocations (Mac1, Mac2a, and Myla) were analyzed in technical and biologic triplicates to perform unbiased proteome characterization. Phosphoproteins were enriched using metal oxide affinity chromatography and immunoprecipitation using 3 anti-phosphotyrosine antibodies and N-glycoproteins were enriched using solid phase extraction and hydrazide resin and an Orbitrap Fusion or LTQ OrbitrapXL in-line with a Paradigm MS2 HPLC. Precise site-mapping of the PTMs was assigned and quantitated using Maxquant. Hierarchical clustering was performed using Perseus. Whole exome sequencing (WES) was performed using an Illumina HiSeq 2500. Functional studies using tyrosine kinase inhibitors were performed to assess selective vulnerabilities.Results: Phosphoproteomic analysis identified 1899 tyrosine (Y) phosphoproteins and 6029 serine/threonine (S/T) phosphoproteins. Unsupervised hierarchical clustering of the Y-phosphoproteome data accurately grouped the CTCLs to those with and without underlying tyrosine kinase translocations. Unique driver kinase phosphosignaling signatures were identified in MYLA (NPM1-TYK2) and MAC1/2 (PCM1-JAK2) representing the ALCL subgroup, as well as close similarity between the signatures of MF and SS-derived cells (HH and HUT78). Notably, phosphoproteomic analysis of HUT78 (SS) yielded high spectral counts of Y-phosphorylated residues at IL2RG p.Y325, JAK3 p.Y980/Y981, and STAT5 p.Y699 indicative of constitutive activation of the JAK/STAT5 pathway corroborated by WES showing activating mutations in JAK3 p.A573V and JAK1 p.Y654F.Beyond driver kinases, oncogenic kinase-specific substrates with distinct site-specific phosphorylated residues were identified for each disease class. Phosphoproteins selectively expressed in MF/SS, (46 p-S/T and 25 p-Y proteins) included members of the killer-cell immunoglobulin-like receptor (KIR) family implicated in immune modulation through immune tyrosine-based inhibitor or TYRO protein tyrosine kinase binding domains for signal transduction. Additionally, proteins not previously associated with MF/SS including NKG2-D type integral membrane protein family members were discriminative of this subgroup. Phosphoprotein members of lysine demethylases (KDM family) and Aryl Hydrocarbon Receptor families were distinctively identified in MF/SS cells, while components of the T-cell receptor signaling pathways were common in all groups of CTCLs. By comparison, N-glycoproteomic analysis identified 811 unique N-glycoproteins. A total of 51 N-glycoproteins were found solely in HUT78 of which 46 were novel in their specificity and included 13 with a role in cell-adhesion while N-glycoproteins unique to HH revealed 12 novel candidate biomarkers (e.g. MERTK, CD97 and SLC44A2). Integrative phosphoproteomic and N-glycoproteomic analysis strikingly revealed convergent pathways implicating a prominent role for the KIR family proteins in SS. Orthogonal immunophenotyping by flow cytometry has shown high sensitivity (96%) detection of KIR3DL2 in primary human SS cases. Significantly, anti-KIR3DL2 inhibition using a humanized antibody revealed activity against SS cells. By comparison, TYK2-driven CTCL (MYLA) was sensitive to inhibitors of TYK2 and JAK.Conclusions: Integrated phosphoproteomic and N-glycoproteomic analysis identifies known and novel candidate biomarkers, unique PTM signatures, and therapeutic vulnerabilities implicating disease-specific pathogenetic mechanisms in CTCL. DisclosuresNo relevant conflicts of interest to declare.

Elucidation of rutin's role in inducing caspase-dependent apoptosis via HPV-E6 and E7 down-regulation in cervical cancer HeLa cells.

Over the recent few years rutin has gained wider attention in exhibiting inhibitory potential against several oncotargets for inducing apoptotic and antiproliferative activity in several human cancer cells. Several deregulated signaling pathways are implicated in cancer pathogenesis. Therefore we have inclined our research towards exploring the anticancerous efficacy of a very potent phytocompound for modulating the incontinent expression of these two crucial E6 and E7 oncogenes. Further, inhibitory efficacy of rutin against human papillomavirus (HPV)-E6 and E7 oncoproteins in cervical cancer has not been elucidated yet. This research addresses the growth inhibitory efficacy of rutin against E6 and E7 oncoproteins in HeLa cells, which is known to inactivate several tumor suppressor proteins such as p53 and pRB. Rutin treatment exhibited reduced cell viability with increased cell accumulation in G0/G1 phase of cell cycle in HeLa cell lines. Additionally, rutin treatment has also led to down-regulation of E6 and E7 expression associated with an increased expression of p53 and pRB levels. This has further resulted in enhanced Bax expression and decreased Bcl-2 expression releasing cytochrome c into cytosol followed by caspase cascade activation with cleavage of caspase-3, caspase-8 and caspase-9. Further, in silico studies have also supported our in vitro findings by exhibiting significant binding energy against selected target oncoproteins. Therefore, our research findings might recommend rutin as one of the potent drug candidate in cervical cancer management via targeting two crucial oncoproteins associated with viral progression.

COVID-19-Associated Pneumonia: Radiobiological Insights.

The evolution of SARS-CoV-2 pneumonia to acute respiratory distress syndrome is linked to a virus-induced “cytokine storm”, associated with systemic inflammation, coagulopathies, endothelial damage, thrombo-inflammation, immune system deregulation and disruption of angiotensin converting enzyme signaling pathways. To date, the most promising therapeutic approaches in COVID-19 pandemic are linked to the development of vaccines. However, the fight against COVID-19 pandemic in the short and mid-term cannot only rely on vaccines strategies, in particular given the growing proportion of more contagious and more lethal variants among exposed population (the English, South African and Brazilian variants). As long as collective immunity is still not acquired, some patients will have severe forms of the disease. Therapeutic perspectives also rely on the implementation of strategies for the prevention of secondary complications resulting from vascular endothelial damage and from immune system deregulation, which contributes to acute respiratory distress and potentially to long term irreversible tissue damage. While the anti-inflammatory effects of low dose irradiation have been exploited for a long time in the clinics, few recent physiopathological and experimental data suggested the possibility to modulate the inflammatory storm related to COVID-19 pulmonary infection by exposing patients to ionizing radiation at very low doses. Despite level of evidence is only preliminary, these preclinical findings open therapeutic perspectives and are discussed in this article.

Recent Advances in Implantation-Based Genetic Modeling of Biliary Carcinogenesis in Mice.

Simple SummaryBiliary tract cancer (BTC) is often refractory to conventional therapeutics and is difficult to diagnose in the early stages. In addition, the pathogenesis of BTC is not fully understood, despite recent advances in cancer genome analysis. To address these issues, the development of fine disease models is critical for BTC. Although still limited in number, there are various platforms for genetic models of BTC owing to newly emerging technology. Among these, implantation-based models have recently drawn attention for their convenience, flexibility, and scalability. To highlight the relevance of this approach, we comprehensively summarize the advantages and disadvantages of BTC models developed using diverse approaches. Currently available research data on intra- and extrahepatic cholangiocarcinoma and gallbladder carcinoma are presented in this review. This information will likely help in selecting the optimal models for various applications and develop novel innovative models based on these technologies.Epithelial cells in the biliary system can develop refractory types of cancers, which are often associated with inflammation caused by viruses, parasites, stones, and chemicals. Genomic studies have revealed recurrent genetic changes and deregulated signaling pathways in biliary tract cancer (BTC). The causal roles have been at least partly clarified using various genetically engineered mice. Technical advances in Cre-LoxP technology, together with hydrodynamic tail injection, CRISPR/Cas9 technology, in vivo electroporation, and organoid culture have enabled more precise modeling of BTC. Organoid-based genetic modeling, combined with implantation in mice, has recently drawn attention as a means to accelerate the development of BTC models. Although each model may not perfectly mimic the disease, they can complement one another, or two different approaches can be integrated to establish a novel model. In addition, a comparison of the outcomes among these models with the same genotype provides mechanistic insights into the interplay between genetic alterations and the microenvironment in the pathogenesis of BTCs. Here, we review the current status of genetic models of BTCs in mice to provide information that facilitates the wise selection of models and to inform the future development of ideal disease models.

Halting the FGF/FGFR axis leads to antitumor activity in Waldenström macroglobulinemia by silencing MYD88

The human fibroblast growth factor/fibroblast growth factor receptor (FGF/FGFR) axis deregulation is largely involved in supporting the pathogenesis of hematologic malignancies, including Waldenström macroglobulinemia (WM). WM is still an incurable disease, and patients succumb because of disease progression. Therefore, novel therapeutics designed to specifically target deregulated signaling pathways in WM are required. We aimed to investigate the role of FGF/FGFR system blockade in WM by using a pan-FGF trap molecule (NSC12). Wide-transcriptome profiling confirmed inhibition of FGFR signaling in NSC12-treated WM cells; unveiling a significant inhibition of MYD88 was also confirmed at the protein level. Importantly, the NSC12-dependent silencing of MYD88 was functionally active, as it led to inhibition of MYD88-driven pathways, such as BTK and SYK, as well as the MYD88-downstream target HCK. Of note, both canonical and noncanonical NF-κB cascades were downregulated in WM cells upon NSC12 treatment. Functional sequelae exerted by NSC12 in WM cells were studied, demonstrating significant inhibition of WM cell growth, induction of WM cell apoptosis, halting MAPK, JAK/STAT3, and PI3K-Akt pathways. Importantly, NSC12 exerted an anti-WM effect even in the presence of bone marrow microenvironment, both in vitro and in vivo. Our studies provide the evidence for using NSC12 as a specific FGF/FGFR system inhibitor, thus representing a novel therapeutic strategy in WM.

Mesenchymal stem cells as professional actors in gastrointestinal cancer therapy: From Naïve to genetically modified.

Considering the high incidence and mortality rate of gastrointestinal cancers (GIs) worldwide and partial success of the current available GI cancer treatments, there is a necessity to discover more effective approaches in cancer therapy. The failure in conventional therapies seems to be related to the resistance of cancer cells to chemotherapy, inability to target tumor cells especially in metastatic cancers, deficient drug concentrations in tumor sites, and unfavorable effects on pivotal non-malignant bodily tissues following systemic administration. In this context, we need an appropriate carrier for the delivery of therapeutic agents specifically to the GI cancer site. Mesenchymal stem cells (MSCs), a prominent cell-based strategy for cancer treatment, overcome various cancer therapy limitations and could be used as vehicles to deliver many anticancer agents such as therapeutic genes (DNA or interference RNA), oncolytic viruses, and chemotherapeutic or nanoparticle drugs. Moreover, secreted molecules of unmodified MSCs lead to deregulation of several proteins and different signaling pathways eradicating cancer cells. In the present review, at first, we overview the characteristics and utility of MSCs in cancer therapy, secondly, we discuss the application of naïve MSCs and utilization of MSCs in the delivery of therapeutic agents in GI cancer therapy and, finally, more information about harnessing of genetically modified MSCs in GI cancer treatment will be presented.

Transgenic mice overexpressing the LH receptor in the female reproductive system spontaneously develop endometrial tumour masses

The receptor for the luteinizing hormone (LH-R) is aberrantly over expressed in cancers of the reproductive system. To uncover whether LH-R over expression has a causative role in cancer, we generated a transgenic (TG) mouse which overexpresses the human LH-R (hLH-R) in the female reproductive tract, under the control of the oviduct-specific glycoprotein (OGP) mouse promoter (mogp-1). The transgene was highly expressed in the uterus, ovary and liver, but only in the uterus morphological and molecular alterations (increased proliferation and trans-differentiation in the endometrial layer) were detected. A transcriptomic analysis on the uteri of young TG mice showed an up regulation of genes involved in cell cycle control and a down regulation of genes related to the immune system and the metabolism of xenobiotics. Aged TG females developed tumor masses in the uteri, which resembled an Endometrial Cancer (EC). Microarray and immunohistochemistry data indicated the deregulation of signaling pathways which are known to be altered in human ECs. The analysis of a cohort of 126 human ECs showed that LH-R overexpression is associated with early-stage tumors. Overall, our data led support to conclude that LH-R overexpression may directly contribute to trigger the neoplastic transformation of the endometrium.

Genome wide CRISPR screening reveals a role for sialylation in the tumorigenesis and chemoresistance of acute myeloid leukemia cells.

Aberrant activation of cytokine and growth factor signal transduction pathways confers enhanced survival and proliferation properties to acute myeloid leukemia (AML) cells. However, the mechanisms underlying the deregulation of signaling pathways in leukemia cells are unclear. To identify genes capable of independently supporting cytokine-independent growth, we employed a genome-wide CRISPR/Cas9-mediated loss-of-function screen in GM-CSF-dependent human AML TF-1cells. More than 182 genes (p<0.01) were found to suppress the cytokine-independent growth of TF-1cells. Among the top hits, genes encoding key factors involved in sialylation biosynthesis were identified; these included CMAS, SLC35A1, NANS, and GNE. Knockout of either CMAS or SLC35A1 enabled cytokine-independent proliferation and survival of AML cells. Furthermore, NSG (NOD/SCID/IL2Rγ-/-) mice injected with CMAS or SLC35A1-knockout TF-1cells exhibited a shorter survival than mice injected with wild-type cells. Mechanistically, abrogation of sialylation biosynthesis in TF-1cells induced a strong activation of ERK signaling, which sensitized cells to MEK inhibitors but conferred resistance to JAK inhibitors. Further, the surface level of α2,3-linked sialic acids was negatively correlated with the sensitivity of AML cell lines to MEK/ERK inhibitors. We also found that sialylation modulated the expression and stability of the CSF2 receptor. Together, these results demonstrate a novel role of sialylation in regulating oncogenic transformation and drug resistance development in leukemia. We propose that altered sialylation could serve as a biomarker for targeted anti-leukemic therapy.

Penile Cancer-Derived Cells Molecularly Characterized as Models to Guide Targeted Therapies.

Penile cancer (PeCa) is a common disease in poor and developing countries, showing high morbidity rates. Despite the recent progress in understanding the molecular events involved in PeCa, the lack of well-characterized in vitro models precludes new advances in anticancer drug development. Here we describe the establishment of five human primary penile cancer-derived cell cultures, including two epithelial and three cancer-associated fibroblast (CAF) cells. Using high-throughput genomic approaches, we found that the epithelial PeCa derived- cells recapitulate the molecular alterations of their primary tumors and present the same deregulated signaling pathways. The differentially expressed genes and proteins identified are components of key oncogenic pathways, including EGFR and PI3K/AKT/mTOR. We showed that epithelial PeCa derived cells presented a good response to cisplatin, a common therapeutic approach used in PeCa patients. The growth of a PeCa-derived cell overexpressing EGFR was inhibited by EGFR inhibitors (cetuximab, gefitinib, and erlotinib). We also identified CAF signature markers in three PeCa-derived cells with fibroblast-like morphology, indicating that those cells are suitable models for PeCa microenvironment studies. We thus demonstrate the utility of PeCa cell models to dissect mechanisms that promote penile carcinogenesis, which are useful models to evaluate therapeutic approaches for the disease.

Proteomic analysis identified LBP and CD14 as key proteins in blood/biphasic calcium phosphate microparticle interactions

Immediately upon implantation, scaffolds for bone repair are exposed to the patient's blood. Blood proteins adhere to the biomaterial surface and the protein layer affects both blood cell functions and biomaterial bioactivity. Previously, we reported that 80-200 µm biphasic calcium phosphate (BCP) microparticles embedded in a blood clot, induce ectopic woven bone formation in mice, when 200-500 µm BCP particles induce mainly fibrous tissue. Here, in a LC-MS/MS proteomic study we compared the differentially expressed blood proteins (plasma and blood cell proteins) and the deregulated signaling pathways of these osteogenic and fibrogenic blood composites. We showed that blood/BCP-induced osteogenesis is associated with a higher expression of fibrinogen (FGN) and an upregulation of the Myd88- and NF-κB-dependent TLR4 signaling cascade. We also highlighted the key role of the LBP/CD14 proteins in the TLR4 activation of blood cells by BCP particles. As FGN is an endogenous ligand of TLR4, able to modulate blood composite stiffness, we propose that different FGN concentrations modify the blood clot mechanical properties, which in turn modulate BCP/blood composite osteoactivity through TLR4 signaling. The present findings provide an insight at the protein level, into the mechanisms leading to an efficient bone reconstruction by blood/BCP composites. Statement of significanceUpon implantation, scaffolds for bone repair are exposed to the patient's blood. Blood proteins adhere to bone substitute surface and this protein layer affects both biomaterial bioactivity and bone healing. Therefore, for the best outcome for patients, it is crucial to understand the molecular interactions between blood and bone scaffolds. Biphasic calcium phosphate (BCP) ceramics are considered as the gold standard in bone reconstruction surgery. Here, using proteomic analyses we showed that the osteogenic properties of 80-200 µm BCP particles embedded in a blood clot is associated with a higher expression of fibrinogen. Fibrinogen upregulates the Myd88- and NF-κB-dependent TLR4 pathway in blood cells and, BCP-induced TLR4 activation is mediated by the LBP and CD14 proteins.

Pathway-Directed Therapy in Multiple Myeloma.

Simple SummaryMultiple Myeloma is a cancer of plasma cells in the bone marrow. While effective treatments are available and many patients can now live years with the disease, most patients ultimately run out of treatment options. Pathway-directed therapy looks at genetic aberrations in the tumor cells and tries to blockade the tumor’s Achilles heel. Since myeloma cells show changes in several pathways, which can vary between different patients, agents targeting these pathways often only show activity in few patients. Pathway-directed therapy in myeloma is therefore often combined with personalized medicine, which aims to identify drugs that might interfere with the most important pathways in a particular patient. There are several pathways that can be targeted in myeloma, and, in combination with personalized medicine, some have shown promising results. However, there is still a challenge in identifying suitable patients and preventing resistance to single drugs, most likely caused by other pathways assuming the function of the blockaded one. Further research is therefore required to improve pathway-directed therapy.Multiple Myeloma (MM) is a malignant plasma cell disorder with an unmet medical need, in particular for relapsed and refractory patients. Molecules within deregulated signaling pathways, including the RAS/RAF/MEK/ERK, but also the PI3K/AKT-pathway belong to the most promising evolving therapeutic targets. Rationally derived compounds hold great therapeutic promise to target tumor-specific abnormalities rather than general MM-associated vulnerabilities. This paradigm is probably best depicted by targeting mutated BRAF: while well-tolerated, remarkable responses have been achieved in selected patients by inhibition of BRAFV600E alone or in combination with MEK. Targeting of AKT has also shown promising results in a subset of patients as monotherapy or to resensitize MM-cells to conventional treatment. Approaches to target transcription factors, convergence points of signaling cascades such as p53 or c-MYC, are emerging as yet another exciting strategy for pathway-directed therapy. Informed by our increasing knowledge on the impact of signaling pathways in MM pathophysiology, rationally derived Precision-Medicine trials are ongoing. Their results are likely to once more fundamentally change treatment strategies in MM.

Understanding the Hedgehog Signaling Pathway in Acute Myeloid Leukemia Stem Cells: A Necessary Step toward a Cure

Simple SummaryThe Hedgehog signaling pathway is related to the cell cycle. In particular, it is considered to play a fundamental role in the quiescence of leukemic stem cell (i.e., a temporary resting state without cell replication). Leukemic stem cells are the cells supposed to give rise to the relapses of the leukemia. Therefore, the Hedgehog pathway must be understood to improve the current treatments against acute myeloid leukemia and avoid the relapse of the disease. In this review, we gather the present knowledge about the physiological Hedgehog pathway function, the aberrant activation of Hedgehog in leukemia, and highlight the lack of evidence regarding some aspects of this important pathway. Finally, we summarize the acute myeloid leukemia treatments targeting this signaling pathway.A better understanding of how signaling pathways govern cell fate is fundamental to advances in cancer development and treatment. The initialization of different tumors and their maintenance are caused by the deregulation of different signaling pathways and cancer stem cell maintenance. Quiescent stem cells are resistant to conventional chemotherapeutic treatments and, consequently, are responsible for disease relapse. In this review we focus on the conserved Hedgehog (Hh) signaling pathway which is involved in regulating the cell cycle of hematopoietic and leukemic stem cells. Thus, we examine the role of the Hh signaling pathway in normal and leukemic stem cells and dissect its role in acute myeloid leukemia. We explain not only the connection between illness and the signaling pathway but also evaluate innovative therapeutic approaches that could affect the outcome of patients with acute myeloid leukemia. We found that many aspects of the Hedgehog signaling pathway remain unknown. The role of Hh has only been proven in embryo and hematopoietic stem cell development. Further research is needed to elucidate the role of GLI transcription factors for therapeutic targeting. Glasdegib, an SMO inhibitor, has shown clinical activity in acute myeloid leukemia; however, its mechanism of action is not clear.

MiR-320a/SP1 negative reciprocal interaction contributes to cell growth and invasion in colorectal cancer

BackgroundTranscription factors (TFs) may be engaged in reciprocal regulatory circuits with certain miRNAs to maintain cellular homeostasis. Disequilibrium of the reciprocities by certain tumor-related stimuli may give rise to deregulation of downstream cellular signaling pathways, thus promoting malignant tumor phenotypes. Specificity Protein 1 (SP1) is the most representative member of the tumor-related transcription factors. Previous studies disclosed that SP1 can transcriptionally regulate miRNAs and coding genes to facilitate tumor progression. In our study, we used bioinformatic analysis to predict several SP1-binding sites within the miR-320a promoter and found that SP1 is a predicted target gene of miR-320a. Therefore, we hypothesize a reciprocal regulatory link between SP1 and miR-320a that participates in colorectal cancer (CRC) developmentMethodsWe performed bioinformatic analysis, quantitative polymerase chain reaction (qPCR), immunoblotting, dual-luciferase reporter assays, and a series of in vitro and in vivo functional assays to describe a novel SP1/miR-320a reciprocal interaction in CRCResultsFirst, we found that miR-320a was significantly downregulated in CRC tissues and cell lines. Consistent with findings in other cancers, miR-320a exhibited inhibitory effects on cell growth and invasion of CRC in vitro and in vivo. Moreover, we identified SP1 as a target gene of miR-320a, and ectopic SP1 expression partly abolished miR-320a-induced inhibitory effects. Conversely, we confirmed that SP1 interacts with the miR-320a promoter, leading to depression of miR-320a. This illustrates a double-negative feedback loop between miR-320a and SP1. Additionally, based on the fact that SP1 promotes MACC1 transcription, we determined via immunoblotting that the oncogenic MACC1/MET signaling pathway was inactivated in the context of miR-320a-induced SP1 downregulationConclusionTaken together, our study is the first to describe a miR-320a/SP1 negative reciprocal interaction, which contributes to cell growth and invasion in CRC through modulation of the MACC1/MET signaling pathway.

The Clinicopathological Significance of YAP/TAZ Expression in Hepatocellular Carcinoma with Relation to Hypoxia and Stemness.

Background/Aims: Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) activation has been implicated in hepatocarcinogenesis and hepatic progenitor cell differentiation, and hypoxia has been shown to induce nuclear translocation of YAP in cancer cells. Here, we aimed to investigate the relationship between hypoxia, YAP and TAZ expression and stemness-related marker expression in human hepatocellular carcinomas (HCCs) and its clinical implications. Methods: Immunohistochemical stains were performed on tissue microarrays from 305 surgically resected HCCs, and the expression status of YAP and TAZ were correlated with CAIX, stemness markers (K19, EpCAM) and epithelial-mesenchymal transition (EMT)-related markers (uPAR, ezrin). The clinicopathological significance of YAP/TAZ expression was analyzed with relation to CAIX expression status. Results: YAP and TAZ expression were seen in 13.4 and 4.3% of HCCs, respectively. YAP/TAZ-positive HCCs frequently demonstrated higher serum AFP levels, microvascular invasion, advanced tumor stage, increased proliferative activity and expression of stemness- and EMT-related markers, CAIX, p53 and Smad2/3 (p < 0.05, all). Interestingly, YAP/TAZ-positivity was associated with microvascular invasion, higher serum AFP levels, stemness and EMT-related marker expression only in tumors expressing CAIX (p < 0.05, all), while these associations were not seen in CAIX-negative HCCs. Conclusions: YAP/TAZ expression is associated with vascular invasion, stemness and EMT in HCCs with hypoxia marker expression. The effect of Hippo signaling pathway deregulation in HCC may depend on the presence or absence of a hypoxic microenvironment, and hypoxia marker expression status should be taken into account when considering the use of YAP/TAZ as markers of aggressive biologic behavior in HCC.

Environmental Exposure to Heavy Metals Contributes to Diseases Via Deregulated Wnt Signaling Pathways.

Wnt signaling plays a critical role during embryogenesis and is responsible for regulating the homeostasis of the adult stem cells and cells fate via a multitude of signaling pathways and associated transcription factors, receptors, effectors, and inhibitors. For this review, published articles were searched from PubMed Central, Embase, Medline, and Google Scholar. The search terms were Wnt, canonical, noncanonical, signaling pathway, β-catenin, environment, and heavy metals. Published articles on Wnt signaling pathways and heavy metals as contributing factors for causing diseases via influencing Wnt signaling pathways were included. Wnt canonical or noncanonical signaling pathways are the key regulators of stem cell homeostasis that control many mechanisms. There is an adequate balance between β-catenin dependent and independent Wnt signaling pathways and remain highly conserved throughout different development stages. Environmental heavy metal exposure may cause either inhibition or overexpression of any component of Wnt signaling pathways such as Wnt protein, transcription factors, receptors, ligands, or transducers to impede normal cellular function via negatively affecting Wnt signaling pathways. Environmental exposure to heavy metals potentially contributes to diseases via deregulated Wnt signaling pathways.

Hodgkin Lymphoma in Nigeria: Prevalence, Mechanisms of Tumorigenesis and Diagnosis

Hodgkin lymphoma (HL) is an uncommon neoplasm where tumor stems from a certain kind of leukocytes known as B-cell. This lymphoid malignancy involves outlying lymph gland which may extend to tissues. Research estimates 40% of patients are diagnosed from constitutive signs while early detection is a challenge. The Hodgkin Reed-Stenberg cells is established as the hallmark of Hodgkin lymphoma but multiple underlying mechanisms are involved. Hodgkin lymphoma spreads across different regions with limitations of effective control, hence there is a need to fully understand its tumorigenesis and diagnosis. We reviewed the multiphase deregulated signalling pathways and several interactions amongst cells, these characteristics can proffer new breakthrough to target therapeutics either through direct inhibition of affected signalling pathways or Hodgkin Reed-Stenberg cells associated with diverse cellular infiltrates within the lymph cell. HL incidence in Nigeria is similar to the incidence in other regions but there is a higher death rate in Nigeria. Nigeria bears the second highest burden of HIV globally which contributes to incidence of HL. Also, Epstein-Barr virus increases the risk of the disease, which is also prevalent in Nigeria. Lymphomas are associated with higher cancer-related deaths especially in HIV-infected persons with 2.5times lower 2-year survival chance compared to HIV-uninfected persons and late stage of lymphoma was also a contributing factor to the survival outcomes. Hence, to reduce the burden of the disease, an understanding of the aetiology, epidemiology, and biochemical mechanisms of the lymphoma is needed which are herein reviewed.