Identification of SLC22A17 DNA methylation hotspot as a potential biomarker in cutaneous melanoma.

BACKGROUND: Cancer onset and progression are driven by genetic and epigenetic alterations leading to oncogene activation and the silencing of tumor suppressor genes. Among epigenetic mechanisms, DNA methylation (methDNA) is gaining growing interest in cancer. Promoter hypomethylation is associated with oncogene activation while intragenic methDNA can be involved in transcriptional elongation, alternative spicing, and the activation of cryptic start sites. Several genes involved in the modulation of the tumor microenvironment are regulated by methDNA, including the Solute Carrier Family 22 Member 17 (SLC22A17), which is involved in iron trafficking and extracellular matrix remodeling cooperating with the Gelatinase-Associated Lipocalin (NGAL) ligand. However, the exact role of intragenic methDNA in cancer has not been fully investigated. Therefore, the aim of the present study is to explore the role of methDNA in the regulation of SLC22A17 in cutaneous melanoma (CM), used as a tumor model. METHODS: Correlation and differential analyses between SLC22A17 expression and methDNA were performed using the data contained in The Cancer Genome Atlas and Gene Expression Omnibus databases. Functional studies on melanoma cell lines treated with 5-Azacytidine (5-Aza) were conducted to assess the correlation between methDNA and SLC22A17 expression. A validation study on the diagnostic potential of the in silico-identified SLC22A17 methDNA hotspot was finally performed by analyzing tissue samples obtained from CM patients and healthy controls. RESULTS: The computational analyses revealed that SLC22A17 was significantly downregulated in CM, and its expression was related to promoter hypomethylation and intragenic hypermethylation. Moreover, SLC22A17 overexpression and hypermethylation of two intragenic methDNA hotspots were associated with a better clinical outcome in CM patients. The correlation between SLC22A17 methDNA and expression was confirmed in 5-Aza-treated cells. In agreement with in silico analyses, the SLC22A17 promoter methylation hotspot showed higher methDNA levels in CM samples compared to nevi. In addition, the methDNA levels of this hotspot were positively correlated with advanced CM. CONCLUSIONS: The SLC22A17 methDNA hotspot could represent a promising biomarker for CM, highlighting the regulatory role of methDNA on SLC22A17 expression. These results pave the way for the identification of novel epigenetic biomarkers and therapeutic targets for the management of CM patients.

Signaling Role of Pericytes in Vascular Health and Tissue Homeostasis.

Pericytes are multipotent cells embedded within the vascular system, primarily surrounding capillaries and microvessels where they closely interact with endothelial cells. These cells are known for their intriguing properties due to their heterogeneity in tissue distribution, origin, and multifunctional capabilities. Specifically, pericytes are essential in regulating blood flow, promoting angiogenesis, and supporting tissue homeostasis and regeneration. These multifaceted roles draw on pericytes' remarkable ability to respond to biochemical cues, interact with neighboring cells, and adapt to changing environmental conditions. This review aims to summarize existing knowledge on pericytes, emphasizing their versatility and involvement in vascular integrity and tissue health. In particular, a comprehensive view of the major signaling pathways, such as PDGFß/ PDGFRß, TGF-ß, FOXO and VEGF, along with their downstream targets, which coordinate the behavior of pericytes in preserving vascular integrity and promoting tissue regeneration, will be discussed. In this light, a deeper understanding of the complex signaling networks defining the phenotype of pericytes in healthy tissues is crucial for the development of targeted therapies in vascular and degenerative diseases.

The potential of the nutraceutical berberine in the treatment of hepatocellular carcinoma and other liver diseases such as NAFLD and NASH.

Hepatocellular carcinoma (HCC) is a common cancer which unfortunately has poor outcomes. Common anti-cancer treatments such as chemotherapy and targeted therapy have not increased patient survival significantly. A common treatment for HCC patients is transplantation, however, it has limitations and complications. Novel approaches are necessary to more effectively treat HCC patients. Berberine (BBR) is a nutraceutical derived from various fruits and trees, which has been used for centuries in traditional medicine to treat various diseases such as diabetes and inflammation. More recently, the anti-proliferation effects of BBR have been investigated in the treatment of patients with various cancers, especially colorectal cancer, and in non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH). In this review, we will focus on studies with BBR in liver diseases.

Deciphering signaling pathways in hematopoietic stem cells: the molecular complexity of Myelodysplastic Syndromes (MDS) and leukemic progression.

Myelodysplastic Syndromes, a heterogeneous group of hematological disorders, are characterized by abnormalities in phosphoinositide-dependent signaling, epigenetic regulators, apoptosis, and cytokine interactions within the bone marrow microenvironment, contributing to disease pathogenesis and neoplastic growth. Comprehensive knowledge of these pathways is crucial for the development of innovative therapies that aim to restore normal apoptosis and improve patient outcomes.

Phospholipases in Gliomas: Current Knowledge and Future Perspectives from Bench to Bedside.

Phospholipases are essential intermediaries that work as hydrolyzing enzymes of phospholipids (PLs), which represent the most abundant species contributing to the biological membranes of nervous cells of the healthy human brain. They generate different lipid mediators, such as diacylglycerol, phosphatidic acid, lysophosphatidic acid, and arachidonic acid, representing key elements of intra- and inter-cellular signaling and being involved in the regulation of several cellular mechanisms that can promote tumor progression and aggressiveness. In this review, it is summarized the current knowledge about the role of phospholipases in brain tumor progression, focusing on low- and high-grade gliomas, representing promising prognostic or therapeutic targets in cancer therapies due to their influential roles in cell proliferation, migration, growth, and survival. A deeper understanding of the phospholipases-related signaling pathways could be necessary to pave the way for new targeted therapeutic strategies.

A miRNA screening identifies miR-192-5p as associated with response to azacitidine and lenalidomide therapy in myelodysplastic syndromes.

BACKGROUND: miRNAs are small non-coding RNAs that regulate gene expression and are linked to cancer development and progression. miRNA profiles are currently studied as new prognostic factors or therapeutic perspectives. Among hematological cancers, myelodysplastic syndromes at higher risk of evolution into acute myeloid leukemia are treated with hypomethylating agents, like azacitidine, alone or in combination with other drugs, such as lenalidomide. Recent data showed that, during azacitidine and lenalidomide therapy, the concurrent acquisition of specific point mutations affecting inositide signalling pathways is associated with lack or loss of response to therapy. As these molecules are implicated in epigenetic processes, possibly involving miRNA regulation, and in leukemic progression, through the regulation of proliferation, differentiation and apoptosis, here we performed a new miRNA expression analysis of 26 high-risk patients with myelodysplastic syndromes treated with azacitidine and lenalidomide at baseline and during therapy. miRNA array data were processed, and bioinformatic results were correlated with clinical outcome to investigate the translational relevance of selected miRNAs, while the relationship between selected miRNAs and specific molecules was experimentally tested and proven. RESULTS: Patients' overall response rate was 76.9% (20/26 cases): complete remission (5/26, 19.2%), partial remission (1/26, 3.8%), marrow complete remission (2/26, 7.7%), hematologic improvement (6/26, 23.1%), hematologic improvement with marrow complete remission (6/26, 23.1%), whereas 6/26 patients (23.1%) had a stable disease. miRNA paired analysis showed a statistically significant up-regulation of miR-192-5p after 4 cycles of therapy (vs baseline), that was confirmed by real-time PCR analyses, along with an involvement of BCL2, that was proven to be a miR-192-5p target in hematopoietic cells by luciferase assays. Furthermore, Kaplan-Meier analyses showed a significant correlation between high levels of miR-192-5p after 4 cycles of therapy and overall survival or leukemia-free survival, that was stronger in responders, as compared with patients early losing response and non-responders. CONCLUSIONS: This study shows that high levels of miR-192-5p are associated with higher overall survival and leukemia-free survival in myelodysplastic syndromes responding to azacitidine and lenalidomide. Moreover, miR-192-5p specifically targets and inhibits BCL2, possibly regulating proliferation and apoptosis and leading to the identification of new therapeutic targets.

Understanding the Roles of the Hedgehog Signaling Pathway during T-Cell Lymphopoiesis and in T-Cell Acute Lymphoblastic Leukemia (T-ALL).

The Hedgehog (HH) signaling network is one of the main regulators of invertebrate and vertebrate embryonic development. Along with other networks, such as NOTCH and WNT, HH signaling specifies both the early patterning and the polarity events as well as the subsequent organ formation via the temporal and spatial regulation of cell proliferation and differentiation. However, aberrant activation of HH signaling has been identified in a broad range of malignant disorders, where it positively influences proliferation, survival, and therapeutic resistance of neoplastic cells. Inhibitors targeting the HH pathway have been tested in preclinical cancer models. The HH pathway is also overactive in other blood malignancies, including T-cell acute lymphoblastic leukemia (T-ALL). This review is intended to summarize our knowledge of the biological roles and pathophysiology of the HH pathway during normal T-cell lymphopoiesis and in T-ALL. In addition, we will discuss potential therapeutic strategies that might expand the clinical usefulness of drugs targeting the HH pathway in T-ALL.

Advances in MDS/AML and inositide signalling.

Aberrant signaling pathways regulating proliferation and differentiation of hematopoietic stem cells (HSCs) can contribute to disease pathogenesis and neoplastic growth. Phosphoinositides (PIs) are inositol phospholipids that are implicated in the regulation of critical signaling pathways: aberrant regulation of Phospholipase C (PLC) beta1, PLCgamma1 and the PI3K/Akt/mTOR pathway play essential roles in the pathogenesis of Myelodysplastic Syndromes (MDS) and Acute Myeloid Leukemia (AML).

Effects of chloroquine and hydroxychloroquine on the sensitivity of pancreatic cancer cells to targeted therapies.

Approaches to improve pancreatic cancer therapy are essential as this disease has a very bleak outcome. Approximately 80% of pancreatic cancers are pancreatic ductal adenocarcinomas (PDAC). PDAC is a cancer which is difficult to effectively treat as it is often detected late in the disease process. Almost all PDACs (over 90%) have activating mutations in the GTPase gene KRAS. These mutations result in constitutive KRas activation and the mobilization of downstream pathways such as the Raf/MEK/ERK pathway. Small molecule inhibitors of key components of the KRas/Raf/MEK/ERK pathways as well as monoclonal antibodies (MoAbs) specific for upstream growth factor receptors such insulin like growth factor-1 receptor (IGF1-R) and epidermal growth factor receptors (EGFRs) have been developed and have been evaluated in clinical trials. An additional key regulatory gene frequently mutated (∼75%) in PDAC is the TP53 tumor suppressor gene which controls the transcription of multiple genes involved in cell cycle progression, apoptosis, metabolism, cancer progression and other growth regulatory processes. Small molecule mutant TP53 reactivators have been developed which alter the structure of mutant TP53 protein and restore some of its antiproliferative activities. Some mutant TP53 reactivators have been examined in clinical trials with patients with mutant TP53 genes. Inhibitors to the TP53 negative regulator Mouse Double Minute 2 (MDM2) have been developed and analyzed in clinical trials. Chloroquine and hydroxychloroquine are established anti-malarial and anti-inflammatory drugs that also prevent the induction of autophagy which can have effects on cancer survival. Chloroquine and hydroxychloroquine have also been examined in various clinical trials. Recent studies are suggesting effective treatment of PDAC patients may require chemotherapy as well as targeting multiple pathways and biochemical processes.

Bioinformatic analysis of the LCN2-SLC22A17-MMP9 network in cancer: The role of DNA methylation in the modulation of tumor microenvironment.

Several features of cancer cells such as proliferation, invasion, metastatic spreading, and drug resistance are affected by their interaction with several tumor microenvironment (TME) components, including neutrophil gelatinase-associated lipocalin (NGAL), solute carrier family 22 member 17 (SLC22A17), and matrix metallopeptidase 9 (MMP9). These molecules play a key role in tumor growth, invasion, and iron-dependent metabolism of cancer cells. However, the precise epigenetic mechanisms underlying the gene regulation of Lipocalin 2 (LCN2), SLC22A17, and MMP9 in cancer still remain unclear. To this purpose, computational analysis was performed on TCGA and GTEx datasets to evaluate the expression and DNA methylation status of LCN2, SLC22A17, and MMP9 genes in different tumor types. Correlation analysis between gene/isoforms expression and DNA methylation levels of LCN2, SLC22A17, and MMP9 was performed to investigate the role of DNA methylation in the modulation of these genes. Protein network analysis was carried out using reverse phase protein arrays (RPPA) data to identify protein-protein interactions of the LCN2-SLC22A17-MMP9 network. Furthermore, survival analysis was performed according to gene expression and DNA methylation levels. Our results demonstrated that LCN2 and MMP9 were mainly upregulated in most tumor types, whereas SLC22A17 was largely downregulated, representing a specific hallmark signature for all gastrointestinal tumors. Notably, the expression of LCN2, SLC22A17, and MMP9 genes was negatively affected by promoter methylation. Conversely, intragenic hypermethylation was associated with the overexpression of SLC22A17 and MMP9 genes. Protein network analysis highlighted the role of the LCN2-SLC22A17-MMP9 network in TME by the interaction with fibronectin 1 and claudin 7, especially in rectal tumors. Moreover, the impact of expression and methylation status of LCN2, SLC22A17, and MMP9 on overall survival and progression free interval was tumor type-dependent. Overall, our analyses provide a detailed overview of the expression and methylation status of LCN2, SLC22A17, and MMP9 in all TCGA tumors, indicating that the LCN2-SLC22A17-MMP9 network was strictly regulated by DNA methylation within TME. Our findings pave the way for the identification of novel DNA methylation hotspots with diagnostic and prognostic values and suitable for epi-drug targeting.

Effects of TP53 Mutations and miRs on Immune Responses in the Tumor Microenvironment Important in Pancreatic Cancer Progression.

Approximately 90% of pancreatic cancers are pancreatic ductal adenocarcinomas (PDAC). PDAC is the fourth leading cause of cancer death world-wide. Therapies for PDAC are largely ineffective due to the dense desmoplastic tumor microenvironment which prevents chemotherapeutic drugs and small molecule inhibitors from exerting effective anti-cancer effects. In this review, we will discuss the roles of TP53 and miRs on the PDAC tumor microenvironment and how loss of the normal functions of TP53 promote tumor progression. The TP53 gene is mutated in approximately 50% of pancreatic cancers. Often, these TP53 mutations are point mutations which confer additional functions for the TP53 proteins. These are called gain of function (GOF) mutations (mut). Another class of TP53 mutations are deletions which result in loss of the TP53 protein; these are referred to TP53-null mutations. We have organized this review into various components/properties of the PDAC microenvironment and how they may be altered in the presence of mutant TP53 and loss of certain miR expression.

Pathobiology and Therapeutic Relevance of GSK-3 in Chronic Hematological Malignancies.

Glycogen synthase kinase-3 (GSK-3) is an evolutionarily conserved, ubiquitously expressed, multifunctional serine/threonine protein kinase involved in the regulation of a variety of physiological processes. GSK-3 comprises two isoforms (α and ß) which were originally discovered in 1980 as enzymes involved in glucose metabolism via inhibitory phosphorylation of glycogen synthase. Differently from other proteins kinases, GSK-3 isoforms are constitutively active in resting cells, and their modulation mainly involves inhibition through upstream regulatory networks. In the early 1990s, GSK-3 isoforms were implicated as key players in cancer cell pathobiology. Active GSK-3 facilitates the destruction of multiple oncogenic proteins which include ß-catenin and Master regulator of cell cycle entry and proliferative metabolism (c-Myc). Therefore, GSK-3 was initially considered to be a tumor suppressor. Consistently, GSK-3 is often inactivated in cancer cells through dysregulated upstream signaling pathways. However, over the past 10-15 years, a growing number of studies highlighted that in some cancer settings GSK-3 isoforms inhibit tumor suppressing pathways and therefore act as tumor promoters. In this article, we will discuss the multiple and often enigmatic roles played by GSK-3 isoforms in some chronic hematological malignancies (chronic myelogenous leukemia, chronic lymphocytic leukemia, multiple myeloma, and B-cell non-Hodgkin's lymphomas) which are among the most common blood cancer cell types. We will also summarize possible novel strategies targeting GSK-3 for innovative therapies of these disorders.

Wild type and gain of function mutant TP53 can regulate the sensitivity of pancreatic cancer cells to chemotherapeutic drugs, EGFR/Ras/Raf/MEK, and PI3K/mTORC1/GSK-3 pathway inhibitors, nutraceuticals and alter metabolic properties.

TP53 is a master regulator of many signaling and apoptotic pathways involved in: aging, cell cycle progression, gene regulation, growth, apoptosis, cellular senescence, DNA repair, drug resistance, malignant transformation, metastasis, and metabolism. Most pancreatic cancers are classified as pancreatic ductal adenocarcinomas (PDAC). The tumor suppressor gene TP53 is mutated frequently (50-75%) in PDAC. Different types of TP53 mutations have been observed including gain of function (GOF) point mutations and various deletions of the TP53 gene resulting in lack of the protein expression. Most PDACs have point mutations at the KRAS gene which result in constitutive activation of KRas and multiple downstream signaling pathways. It has been difficult to develop specific KRas inhibitors and/or methods that result in recovery of functional TP53 activity. To further elucidate the roles of TP53 in drug-resistance of pancreatic cancer cells, we introduced wild-type (WT) TP53 or a control vector into two different PDAC cell lines. Introduction of WT-TP53 increased the sensitivity of the cells to multiple chemotherapeutic drugs, signal transduction inhibitors, drugs and nutraceuticals and influenced key metabolic properties of the cells. Therefore, TP53 is a key molecule which is critical in drug sensitivity and metabolism of PDAC.

Phytol and Heptacosane Are Possible Tools to Overcome Multidrug Resistance in an In Vitro Model of Acute Myeloid Leukemia.

Drug resistance is the ability of cancer cells to gain resistance to both conventional and novel chemotherapy agents, and remains a major problem in cancer therapy. Resistance mechanisms are multifactorial and involve more strictly pharmacological factors, such as P-glycoprotein (P-gp) and biological factors such as inhibitor of apoptosis proteins (IAPs) and the nuclear factor-kappa B (NF-κB) pathway. Possible therapeutic strategies for the treatment of acute myeloid leukemia (AML) have increased in recent years; however, drug resistance remains a problem for most pa-tients. Phytol and heptacosane are the major compounds of Euphorbia intisy essential oil (EO) which were demonstrated to inhibit P-gp in a multidrug resistant in vitro model of AML. This study investigated the mechanism by which phytol and heptacosane improve P-gp-mediated drug transport. Phytol suppresses the P-gp expression via NF-κB inhibition and does not seem to act on the efflux system. Heptacosane acts as a substrate and potent P-gp inhibitor, demonstrating the ability to retain the substrate doxorubicin inside the cell and enhancing its cytotoxic effects. Our results suggest that these compounds act as non-toxic modulators of P-gp through different mechanisms and are able to revert P-gp-mediated drug resistance in tumor cells.

Effects of the Mutant TP53 Reactivator APR-246 on Therapeutic Sensitivity of Pancreatic Cancer Cells in the Presence and Absence of WT-TP53.

The TP53 tumor suppressor is mutated in ~75% of pancreatic cancers. The mutant TP53 protein in pancreatic ductal adenocarcinomas (PDAC) promotes tumor growth and metastasis. Attempts have been made to develop molecules that restore at least some of the properties of wild-type (WT) TP53. APR-246 is one such molecule, and it is referred to as a mutant TP53 reactivator. To understand the potential of APR-246 to sensitize PDAC cells to chemotherapy, we introduced a vector encoding WT-TP53 into two PDAC cell lines, one lacking the expression of TP53 (PANC-28) and one with a gain-of-function (GOF) mutant TP53 (MIA-PaCa-2). APR-246 increased drug sensitivity in the cells containing either a WT or mutant TP53 protein with GOF activity, but not in cells that lacked TP53. The introduction of WT-T53 into PANC-28 cells increased their sensitivity to the TP53 reactivator, chemotherapeutic drugs, and signal transduction inhibitors. The addition of WT-TP53 to PDAC cells with GOF TP53 also increased their sensitivity to the drugs and therapeutics, indicating that APR-246 could function in cells with WT-TP53 and GOF TP53. These results highlight the importance of knowledge of the type of TP53 mutation that is present in cancer patients before the administration of drugs which function through the reactivation of TP53.

Effects of the MDM2 inhibitor Nutlin-3a on sensitivity of pancreatic cancer cells to berberine and modified berberines in the presence and absence of WT-TP53.

Approaches to improve pancreatic cancer therapy are essential as this disease has a very bleak outcome. Approximately 80% of pancreatic cancers are pancreatic ductal adenocarcinomas (PDAC). A key regulatory gene frequently mutated (∼75%) in PDAC is the TP53 tumor suppressor gene which controls the transcription of multiple genes involved in cell cycle progression, apoptosis, cancer progression and other growth regulatory processes. The mouse double minute 2 homolog (MDM2) gene product is a nuclear-localized E3 ubiquitin ligase and negatively regulates the TP53 protein which results in its proteasomal degradation. Various MDM2 inhibitors have been isolated and examined in clinical trials, especially in patients with hematological malignancies. Nutlin-3a is one of the first MDM2 inhibitors isolated. Berberine (BBR) is a natural product found in many fruits and berries and used in traditional medicine for centuries. It has many biological effects, and some are anti-proliferative in nature. BBR may activate the expression of TP53 and inhibit cell cycle progression as well as other events important in cell growth. To understand more about the potential of compounds like BBR and chemical modified BBRs (NAX compounds) to sensitize PDAC cells to MDM2 inhibitors, we introduced either WT-TP53 or the pLXSN empty vector control into two PDAC cell lines, one lacking expression of TP53 (PANC-28) and one with gain-of-function mutant TP53 on both alleles (MIA-PaCa-2). Our results indicate that nutlin-3a was able to increase the sensitivity to BBR and certain NAX compounds. The effects of nutlin-3a were usually more substantial in those cells containing an introduced WT TP53 gene. These results highlight the importance of knowledge of the type of TP53 mutation that is present in cancer patients before the administration of drugs which function by stabilization of the TP53 protein.

Role of PLCγ1 in the modulation of cell migration and cell invasion in glioblastoma.

Phosphoinositide-specific phospholipases C (PLCs) are a class of enzymes involved in several cell activities, such as cell cycle regulation, proliferation, differentiation and cytoskeletal dynamics. Among these enzymes, PLCγ1 is one of the most expressed PLCs in the brain, contributing to a complex network in the developing nervous system. Several studies have shown that PLCγ1 signaling imbalance is linked to several brain disorders, including glioblastoma, the most aggressive brain tumor in adults. Indeed, it has been demonstrated a link between PLCγ1 inhibition and the arrest of glioma cell motility of fetal rat brain aggregates and the impairment of cell invasion abilities following its down-regulation. This study aims to determine the pathological influence of PLCγ1 in glioblastoma, through a translational study which combines in silico data, data from glioblastoma patients' samples and data on engineered cell lines. We found out that PLCγ1 gene expression correlates with the pathological grade of gliomas, and it is higher in fifty patients' glioblastoma tissue samples compared to twenty healthy controls. Moreover, it was demonstrated that PLCγ1 silencing in U87-MG leads to a reduction in cell migration and invasion abilities. The opposite trend was observed following PLCγ1 overexpression, suggesting an interesting possible involvement of PLCγ1 in gliomas' aggressiveness.

GSK3 as a Regulator of Cytoskeleton Architecture: Consequences for Health and Disease.

Glycogen synthase kinase 3 (GSK3) was initially isolated as a critical protein in energy metabolism. However, subsequent studies indicate that GSK-3 is a multi-tasking kinase that links numerous signaling pathways in a cell and plays a vital role in the regulation of many aspects of cellular physiology. As a regulator of actin and tubulin cytoskeleton, GSK3 influences processes of cell polarization, interaction with the extracellular matrix, and directional migration of cells and their organelles during the growth and development of an animal organism. In this review, the roles of GSK3-cytoskeleton interactions in brain development and pathology, migration of healthy and cancer cells, and in cellular trafficking of mitochondria will be discussed.

The NUPR1/p73 axis contributes to sorafenib resistance in hepatocellular carcinoma.

The multikinase inhibitor sorafenib was the first drug approved by the FDA for treating patients with advanced hepatocellular carcinoma (HCC). However, sorafenib resistance remains a major challenge for improving the effectiveness of HCC treatment. Previously, we identified several genes modulated after sorafenib treatment of human HCC cells, including the stress-inducible nuclear protein 1 (NUPR1) gene. Multiple studies have shown that NUPR1 regulates autophagy, apoptosis, and chemoresistance. Here, we demonstrate that treatment of HCC cells with sorafenib resulted in the activation of autophagic flux. NUPR1 knock-down (KD) in HCC cells was associated with increased p62 expression, suggesting an impairment of autophagic flux, and with a significant increase of cell sensitivity to sorafenib. In NUPR1 KD cells, reduced levels of NUPR1 were associated with the increased expression of p73 as well as its downstream transcription targets PUMA, NOXA, and p21. Simultaneous silencing of p73 and NUPR1 in HCC cells resulted in increased resistance to sorafenib, as compared to the single KD of either gene. Conversely, pharmacological activation of p73, via the novel p73 small molecule activator NSC59984, determined synergistic anti-tumor effects in sorafenib-treated HCC cells. The combination of NSC59984 and sorafenib, when compared to either treatment alone, synergistically suppressed tumor growth of HCC cells in vivo. Our data suggest that the activation of the p73 pathway achieved by NUPR1 KD potentiates sorafenib-induced anti-tumor effects in HCC cells. Moreover, combined pharmacological therapy with the p73 activator NSC59984 and sorafenib could represent a novel approach for HCC treatment.

GSK-3: a multifaceted player in acute leukemias.

Glycogen synthase kinase 3 (GSK-3) consists of two isoforms (α and ß) that were originally linked to glucose metabolism regulation. However, GSK-3 is also involved in several signaling pathways controlling many different key functions in healthy cells. GSK-3 is a unique kinase in that its isoforms are constitutively active, while they are inactivated mainly through phosphorylation at Ser residues by a variety of upstream kinases. In the early 1990s, GSK-3 emerged as a key player in cancer cell pathophysiology. Since active GSK-3 promotes destruction of multiple oncogenic proteins (e.g., ß-catenin, c-Myc, Mcl-1) it was considered to be a tumor suppressor. Accordingly, GSK-3 is frequently inactivated in human cancer via aberrant regulation of upstream signaling pathways. More recently, however, it has emerged that GSK-3 isoforms display also oncogenic properties, as they up-regulate pathways critical for neoplastic cell proliferation, survival, and drug-resistance. The regulatory roles of GSK-3 isoforms in cell cycle, apoptosis, DNA repair, tumor metabolism, invasion, and metastasis reflect the therapeutic relevance of these kinases and provide the rationale for combining GSK-3 inhibitors with other targeted drugs. Here, we discuss the multiple and often conflicting roles of GSK-3 isoforms in acute leukemias. We also review the current status of GSK-3 inhibitor development for innovative leukemia therapy.