Antonios N. Gargalionis

Polycystin-1 and polycystin-2 are involved in the acquisition of aggressive phenotypes in colorectal cancer.

The polycystins PC1 and PC2 are emerging as major players in mechanotransduction, a process that influences all steps of the invasion/metastasis cascade. We hypothesized that PC1 and PC2 facilitate cancer aggressiveness. Immunoblotting, RT‐PCR, semi‐quantitative and quantitative real‐time PCR and FACS analyses were employed to investigate the effect of polycystin overexpression in colorectal cancer (CRC) cells. The impact of PC1 inhibition on cancer‐cell proliferation was evaluated through an MTT assay. In vitro data were analyzed by Students t‐test. HT29 human xenografts were treated with anti‐PC1 (extracellular domain) inhibitory antibody and analyzed via immunohistochemistry to determine the in vivo role of PC1 in CRC. Clinical significance was assessed by examining PC1 and PC2 protein expression in CRC patients (immunohistochemistry). In vivo and clinical data were analyzed by non‐parametric tests, Kaplan‐Meier curves, log‐rank test and Cox model. All statistical tests were two‐sided. PC1 overexpression promotes epithelial‐to‐mesenchymal transition (EMT) in HCT116 cells, while PC2 overexpression results in upregulation of the mTOR pathway in SW480 cells. PC1 inhibition causes reduced cell proliferation in CRC cells inducing tumor necrosis and suppressing EMT in HT29 tumor xenografts. In clinical study, PC1 and PC2 overexpression associates with adverse pathological parameters, including invasiveness and mucinous carcinomas. Moreover, PC1 overexpression appears as an independent prognostic factor of reduced recurrence‐free survival (HR = 1.016, p = 0.03) and lowers overall survival probability, while aberrant PC2 expression predicts poor overall survival (p = 0.0468). These results support, for the first time, a direct link between mechanosensing polycystins (PC1 and PC2) and CRC progression.

The molecular rationale of Src inhibition in colorectal carcinomas

Src has been one of the most studied proto‐oncogenes. The cellular Src (c‐Src) holds a critical role in several human malignancies and has emerged as a key factor that promotes tumor progression during the multistep process of colorectal cancer (CRC) pathogenesis. The robust activation of Src in CRC of aggressive phenotype and poor prognosis seems to be a subsequent event of a strong link between its deregulated activity and the tumors cell adhesion properties, invasiveness and metastatic potential. The rarely detected genetic defects drive interest in signaling networks that control Src kinase activity and integrate the association of Src with receptor tyrosine kinases (RTKs), such as the epidermal growth factor receptor (EGFR). Therefore, a dynamic crosstalk is being formed with oncogenic capacity and therapeutic applications, because Src inhibition seems to sensitize previously unresponsive cancer cells to chemotherapy and anti‐EGFR inhibitors. The present review explores the molecular basis behind Src inhibition in colorectal carcinomas. Furthermore, preclinical studies and clinical trials of Src inhibitors and combination regimens are discussed, providing new insights for further investigation and new therapeutic strategies.

Advanced glycation end products upregulate lysyl oxidase and endothelin-1 in human aortic endothelial cells via parallel activation of ERK1/2–NF-κB and JNK–AP-1 signaling pathways

Endothelial dysfunction involves deregulation of the key extracellular matrix (ECM) enzyme lysyl oxidase (LOX) and the vasoconstrictor protein, endothelin-1 (ET-1), whose gene expression can be modulated by the transcriptional activators nuclear factor kappa B (NF-κB) and activator protein-1 (AP-1). Advanced glycation end products (AGEs) present an aggravating factor of endothelial dysfunction which upon engagement to their receptor RAGE induce upregulation of mitogen-activated protein kinases (MAPKs), leading to NF-κB and AP-1 potentiation. We hypothesized that AGEs could induce NF-κΒ- and AP-1-dependent regulation of LOX and ET-1 expression via the AGE/RAGE/MAPK signaling axis. Western blot, real-time qRT-PCR, FACS analysis and electrophoretic mobility-shift assays were employed in human aortic endothelial cells (HAECs) following treatment with AGE-bovine serum albumin (AGE-BSA) to investigate the signaling pathway towards this hypothesis. Furthermore, immunohistochemical analysis of AGEs, RAGE, LOX and ET-1 expression was conducted in aortic endothelium of a rat experimental model exposed to high- or low-AGE content diet. HAECs exposed to AGE-BSA for various time points exhibited upregulation of LOX and ET-1 mRNA levels in a dose- and time-dependent manner. Exposure of HAECs to AGE-BSA also showed specific elevation of phospho(p)-ERK1/2 and p-JNK levels in a dose- and time-dependent fashion. AGE administration significantly increased NF-κΒ- and AP-1-binding activity to both LOX and ET-1 cognate promoter regions. Moreover, LOX and ET-1 overexpression in rat aortic endothelium upon high-AGE content diet confirmed the functional interrelation of these molecules. Our findings demonstrate that AGEs trigger NF-κΒ- and AP-1-mediated upregulation of LOX and ET-1 via the AGE/RAGE/MAPK signaling cascade in human endothelial cells, thus contributing to distorted endothelial homeostasis by impairing endothelial barrier function, altering ECM biomechanical properties and cell proliferation.

Deciphering signaling networks in osteosarcoma pathobiology

Osteosarcoma is the most frequent type of primary bone tumors among children and adolescents. During the past years, little progress has been made regarding prognosis of osteosarcoma patients, especially for those with metastatic disease. Genomic instability and gene alterations are common, but current data do not reveal a consistent and repeatable pattern of osteosarcoma development, thus paralleling the tumors high heterogeneity. Critical signal transduction pathways have been implicated in osteosarcoma pathobiology and are being evaluated as therapeutic targets, including receptor activator for nuclear factor-κB (RANK), Wnt, Notch, phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin, and mechanotransduction pathways. Herein, we recapitulate and discuss recent advances in the context of molecular mechanisms and signaling networks that contribute to osteosarcoma progression and metastasis, towards patient-tailored and novel-targeted treatments.

Recent Advances in Mechanobiology of Osteosarcoma.

Mechanotransduction is a key process by which cells perceive extracellular mechanical cues/intercellular physical interactions and transform them into intracellular biochemical signals. This physiological process is crucial during bone development and bone remodeling throughout childhood and adult life, whereas several aberrations during this process have emerged as a distinct pathogenic molecular entity in bone maladies and tumor formation. The present review focuses on recent advances regarding the mechanobiology of osteosarcoma, the most common type of bone cancer. Special emphasis is given on the mechano‐responsive signal transduction pathways underlying osteosarcoma pathology and on specific mechanosensitive molecules engaged in osteosarcoma development. J. Cell. Biochem. 118: 232–236, 2017.

Polycystins: Mechanosensors with Diagnostic and Prognostic Potential in Cancer

Polycystins are key regulators of cell-to-cell and cell-to-extracellular matrix (ECM) physical interactions that have been recently implicated in oncogenic mechanisms. Polycystins may be emerging as potential diagnostic and prognostic markers in specific types of malignancy, offering a promising research area for the development of novel biomarkers in solid tumors.

The Role of Platelets in Cardiovascular Disease: Molecular Mechanisms

The role of platelets in atherosclerotic process and subsequently in the pathophysiology of cardiovascular disease is essential as platelets in addition to their contribution to thrombosis and hemostasis modulating inflammatory reactions and immune response. Platelets after adhesion on the injured vascular endothelium and activation release a wide range of molecules stored in platelets granules such as chemokines, proinflammatory molecules and other biological response modulators accelerating interaction among platelets, endothelial cells and leukocytes. These interactions establish a localized inflammatory response that promotes the atherosclerotic process. Moreover, activated platelets give rise to microparticles another active participant within the blood stream. The purpose of this review is to present the role of platelets in the above mechanisms giving an emphasis on the nature of the platelet derived- molecules and their contribution to the atherosclerotic process.

Mechanosensor polycystin-1 potentiates differentiation of human osteoblastic cells by upregulating Runx2 expression via induction of JAK2/STAT3 signaling axis

Polycystin-1 (PC1) has been proposed as a chief mechanosensing molecule implicated in skeletogenesis and bone remodeling. Mechanotransduction via PC1 involves proteolytic cleavage of its cytoplasmic tail (CT) and interaction with intracellular pathways and transcription factors to regulate cell function. Here we demonstrate the interaction of PC1-CT with JAK2/STAT3 signaling axis in mechanically stimulated human osteoblastic cells, leading to transcriptional induction of Runx2 gene, a master regulator of osteoblastic differentiation. Primary osteoblast-like PC1-expressing cells subjected to mechanical-stretching exhibited a PC1-dependent increase of the phosphorylated(p)/active form of JAK2. Specific interaction of PC1-CT with pJAK2 was observed after stretching while pre-treatment of cells with PC1 (anti-IgPKD1) and JAK2 inhibitors abolished JAK2 activation. Consistently, mechanostimulation triggered PC1-mediated phosphorylation and nuclear translocation of STAT3. The nuclear phosphorylated(p)/DNA-binding competent pSTAT3 levels were augmented after stretching followed by elevated DNA-binding activity. Pre-treatment with a STAT3 inhibitor either alone or in combination with anti-IgPKD1 abrogated this effect. Moreover, PC1-mediated mechanostimulation induced elevation of Runx2 mRNA levels. ChIP assays revealed direct regulation of Runx2 promoter activity by STAT3/Runx2 after mechanical-stretching that was PC1-dependent. Our findings show that mechanical load upregulates expression of Runx2 gene via potentiation of PC1–JAK2/STAT3 signaling axis, culminating to possibly control osteoblastic differentiation and ultimately bone formation.

MicroRNAs in colorectal neoplasia: from pathobiology to clinical applications.

MicroRNAs (miRNAs) are small, non-coding RNAs that can post-transcriptionally regulate gene expression via messenger RNA (mRNA) targeting. During the past few years several miRNA groups emerged as critical components of developmental and pathological processes, among them being cancer. In colorectal cancer (CRC) specifically, numerous miRNA molecules have been identified up- or downregulated functioning as tumor-specific markers with oncogenic and tumor-suppressive properties. Their dysregulation impacts impaired cellular processes such as cell proliferation, apoptosis, angiogenesis, invasion and metastasis. The detection of extracellular miRNAs in plasma and fecal samples of CRC patients tends to provide novel, non-invasive biomarkers in favor of CRC diagnosis and, at the same time, data from in vivo and in vitro CRC models reveal promising therapeutic applications through miRNA inhibition and miRNA delivery.

Cancer mechanobiology: Effects and therapeutic perspectives: Cancer mechanobiology

A growing body of evidence provides new insights regarding the effect of aberrant mechanosensing of the extracellular matrix (ECM) and corresponding devious mechanotransduction of cancer cells and their surrounding stromal cells in tumorigenesis. Among others, distorted tissue mechanics has been associated with tumor invasion and metastasis. Recently, Glentis et al. highlighted the central role that mechanosensing plays during these processes. Cancer-associated fibroblasts (CAFs) are the most profuse type of tumor-surrounding stromal cells. Data show that CAFs interact physically and remodel basement membrane using mechanical forces. They cooperate with cancer cells to shape an invasive carcinoma acting independently from matrix metalloproteinases (MMPs). Therefore, inhibition of CAFs-mediated mechanical forces represents a putative therapeutic strategy for invasive tumors. However, effects of cancer mechanics seem to be not only restricted during dissemination of cancer cells and neoangiogenesis, but they also emerge as a feature with genetic and epigenetic consequences. Respective studies reveal an association between DNA methylation and histone modifications with matrix mechanosensing, whereas genomic alterations with cancer parallel tissue stiffness. Stiffer tissues (bone, muscle, skin) present with increased genomic alterations (copy number variations and somatic mutations) compared to tumors that develop in softer tissues (brain, marrow). A potential hypothesis is that this may be due to aberrant mechanoregulation of DNA repair pathways. Data suggest that constricted migration through tight structures leads to compressed nucleus and reduced density of DNA repair-associated proteins in U2OS osteosarcoma cells. DNA rapture, DNA breaks, cytoplasmic mislocalization of DNA repair-associated proteins in U2OS, mesenchymal stem cells (MSCs) and A549 cells have been also observed during migration through constricted areas, causing increased DNA damage and reduced DNA repair factors, chromosome copy-number variations, epithelial-to-mesenchymal transition-like phenotypic changes and clonal heterogeneity. The emerging impact of cancer mechanobiology on cancer hallmarks offers potential new therapeutic avenues thus marking a new field termed mechanopharmacology, which aims at exploiting tumor–microenvironment physical associations. This requires integration of biophysics and bioengineering, along with tools such as traction force microscopy, elastic micropillar assays, micropipette aspiration, atomic force microscopy and multicellular three-dimensional systems. More specifically, new strategies tend to exploit the mechanical properties of the metastatic niche. They involve generation of a mechanoresponsive cell system of MSCs that is modulated by the biomechanical cues of the tumor microenvironment to target metastases. In this system, activation of the mechano-induced transcriptional coactivators Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding motif (TAZ) is triggered by sensing of tissue stiffness. This activation is able to induce MSC-based expression of antitumor agents [cytosine deaminase (CD) converts the inactive prodrug 5-fluorocytosine to the active drug 5-fluorouracil] and kill breast cancer cells in vitro in response to tissue stiffening. In a mouse metastatic model of breast cancer data show that MSCs respond to certain mechanical stimuli and deliver CD selectively to the metastatic niche that experiences alterations of tissue mechanics. Because a prodrug was delivered there were limited off-target effects. YAP/TAZ have been also associated with mechanisms of drug resistance. YAP activation confers resistance to mitogen-activated protein kinase kinase (MAPKK, MEK) and RAF inhibitors in BRAF-mutant tumor types and the level of activation parallels the level of resistance. YAP/TAZ activation is also associated with resistance to BRAF mutant melanoma cells. In corroboration, the human epidermal growth factor receptor-2 (HER-2) targeting drug lapatinib in HER-2 amplified breast cancer cells displayed differential antiproliferative effects that were correlated inversely with the elastic properties of the surrounding medium. This modulus-dependent resistance was mediated through YAP and TAZ activation, which when silenced abrogated such resistance in vitro and in vivo. The aforementioned studies imply that abolishment of YAP/TAZ-mediated resistance to targeted agents could be achieved through inhibition of upstream kinases of the adhesive complex that integrate their function in YAP/TAZ [Src, focal adhesion kinase (FAK), Rho, Rho-associated protein kinase (ROCK), integrin]. This inhibition could take place along with simultaneous targeting of oncogenic mutants. Alternatively, direct YAP/TAZ or activator protein-1 (AP-1) targeting could be employed abrogating the effect of YAP/TAZ. Thorough investigation of cancer biomechanics offers novel mechanoregulated molecules as promising therapeutic targets and/or biomarkers. Mechanosensitive polycystins form complexes at the cell surface in order to regulate Conflict of interest: The authors declare that they have no potential conflicts of interest. DOI: 10.1002/ijc.31221 History: Online 15 Dec 2017 Correspondence to: Athanasios G. Papavassiliou, Department of Biological Chemistry, Medical School, National and Kapodistrian University of Athens, 75 M. Asias Street, 11527 Athens, Greece, Tel.: 1[30-210-746-2508], Fax: 1[30-210-746-2703], E-mail: papavas@med.uoa.gr E di to ri al