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The process of cancer initiation and development is a dynamic and complex mechanism involving multiple genetic and non-genetic variations. With the development of high throughput techniques like next-generation sequencing, the field of cancer biology extended beyond the protein-coding genes. It brought the functional role of noncoding RNAs into cancer-associated pathways. MicroRNAs (miRNAs) are one such class of noncoding RNAs regulating different cancer development aspects, including progression and metastasis. MicroRNA-1 (miR-1) is a highly conserved miRNA with a functional role in developing skeletal muscle precursor cells and cardiomyocytes, and acts as a consistent tumor suppressor gene. In humans, two discrete genes, MIR1–1 located on 20q13.333 and MIR1–2 located on 18q11.2 loci, encode for a single mature miR-1. Dysregulation or downregulation of miR-1 has been demonstrated in multiple cancers, including lung, breast, liver, prostate, colorectal, pancreatic, medulloblastoma, and gastric cancer. A vast number of studies have shown that miR-1 affects the multiple hallmarks of cancer like proliferation, invasion and metastasis, apoptosis, angiogenesis, chemosensitization, and immune modulation. The potential therapeutic applications of miR-1 in multiple cancer pathways provide a novel platform for developing anticancer therapies. This review focuses on the different antitumorigenic and therapeutic aspects of miR-1 including how it regulates tumor development and associated immunogenic functions.

Growth differentiation factor 15 or macrophage inhibitory cytokine-1 (GDF15/MIC-1) is a divergent member of the transforming growth factor β superfamily and has a diverse pathophysiological role in cancers, cardiometabolic disorder, and other diseases. GDF15 can control hematopoietic growth, energy homeostasis, adipose tissue metabolism, body growth, bone remodeling, and response to stress signals. The role of GDF15 in cancer development and progression is complicated and depends on the specific cancer type, stage, and tumor microenvironment. Recently, research on GDF15 and GDF15-associated signaling has accelerated due to the identification of the GDF15 receptor: glial cell line-derived neurotrophic factor (GDNF) family receptor α-like (GFRAL). Therapeutic interventions to target GDF15 and/or GFRAL revealed the mechanisms that drive its activity and might improve overall outcomes of patients with metabolic disorders or cancer. The present review highlights the structure and functions of GDF15 and its receptor, emphasizing the pleiotropic role of GDF15 in obesity, tumorigenesis, metastasis, immunomodulation, and cachexia.

Pancreatic cancer (PC) remains a major cause of cancer-related deaths primarily due to its inherent potential of therapy resistance. Checkpoint inhibitors have emerged as promising anti-cancer agents when used in combination with conventional anti-cancer therapies. Recent studies have highlighted a critical role of the Greatwall kinase (MASTL; Microtubule-associated serine/threonine-protein kinase-like) in promoting oncogenic malignancy and resistance to anti-cancer therapies; however, its role in PC remains unknown. Based on a comprehensive investigation involving PC patient samples, murine models of PC progression (Kras;PdxCre-KC and Kras;p53;PdxCre-KPC), and loss and gain of function studies, we report a previously undescribed critical role of MASTL in promoting cancer malignancy and therapy resistance. Mechanistically, MASTL promotes PC by modulating the epidermal growth factor receptor (EGFR) protein stability and, thereupon, kinase signaling. We further demonstrate that combinatorial therapy targeting MASTL promotes the efficacy of the cell-killing effects of Gemcitabine using both genetic and pharmacological inhibitions. Taken together, this study identifies a key role of MASTL in promoting PC progression and its utility as a novel target in promoting sensitivity to the anti-pancreatic cancer therapies.

BACKGROUND: Mucins are high molecular weight glycoproteins dysregulated in aggressive cancers. The role of mucins in disease progression, tumor proliferation, and chemotherapy resistance has been studied extensively. MAIN BODY: This article provides a comprehensive review of mucin’s function as a physical barrier and the implication of mucin overexpression in impeded drug delivery to solid tumors. Mucins regulate the epithelial to mesenchymal transition (EMT) of cancer cells via several canonical and non-canonical oncogenic signaling pathways. Furthermore, mucins play an extensive role in enriching and maintaining the cancer stem cell (CSC) population, thereby sustaining the self-renewing and chemoresistant cellular pool in the bulk tumor. It has recently been demonstrated that mucins regulate the metabolic reprogramming during oncogenesis and cancer progression, which account for tumor cell survival, proliferation, and drug-resistance. CONCLUSION: This review article focuses on delineating mucin’s role in oncogenic signaling and aberrant regulation of gene expressions, culminating in CSC maintenance, metabolic rewiring, and development of chemoresistance, tumor progression, and metastasis.

Secreted mucin 5AC (MUC5AC) is the most abundantly overexpressed member of the mucin family during early pancreatic intraepithelial neoplasia stage I (PanIN-I) of pancreatic cancer. To comprehend the contribution of Muc5ac in pancreatic cancer pathology, we genetically ablated it in an autochthonous murine model (KrasG12D; Pdx-1cre, KC), which mirrors the early stages of pancreatic cancer development. Neoplastic onset and the PanIN lesion progression were significantly delayed in Muc5ac knockout (KrasG12D; Pdx-1 cre; Muc5ac−/−, KCM) animals with a 50% reduction in PanIN-2 and 70% reduction in PanIN-3 lesions compared with KC at 50 weeks of age. High-throughput RNA-sequencing analysis from pancreatic tissues of KCM animals revealed a significant decrease in cancer stem cell (CSC) markers Aldh1a1, Klf4, EpCAM, and CD133. Furthermore, the silencing of MUC5AC in human pancreatic cancer cells reduced their tumorigenic propensity, as indicated by a significant decline in tumor formation frequency by limiting dilution assay upon subcutaneous administration. The contribution of MUC5AC in CSC maintenance was corroborated by a significant decrease in tumor burden upon orthotopic implantation of MUC5AC-depleted pancreatic cancer cells. Mechanistically, MUC5AC potentiated oncogenic signaling through integrin αvβ5, pSrc (Y416), and pSTAT3 (Y705). Phosphorylated STAT3, in turn, upregulated Klf4 expression, thereby enriching the self-renewing CSC population. A strong positive correlation of Muc5ac with Klf4 and pSTAT3 in the PanIN lesions of KC mouse pancreas reinforces the crucial involvement of MUC5AC in bolstering the CSC-associated tumorigenic properties of Kras-induced metaplastic cells, which leads to pancreatic cancer onset and progression. Significance: This study elucidates that de novo expression of MUC5AC promotes cancer cell stemness during Kras-driven pancreatic tumorigenesis and can be targeted for development of a novel therapeutic regimen.

Over the past three decades, monoclonal antibodies (mAbs) have revolutionized the landscape of cancer therapy. Still, this benefit remains restricted to a small proportion of patients due to moderate response rates and resistance emergence. The field has started to embrace better mAb-based formats with advancements in molecular and protein engineering technologies. The development of a therapeutic mAb with long-lasting clinical impact demands a prodigious understanding of target antigen, effective mechanism of action, gene engineering technologies, complex interplay between tumor and host immune system, and biomarkers for prediction of clinical response. This review discusses the various approaches used by mAbs for tumor targeting and mechanisms of therapeutic resistance that is not only caused by the heterogeneity of tumor antigen, but also the resistance imposed by tumor microenvironment (TME), including inefficient delivery to the tumor, alteration of effector functions in the TME, and Fc-gamma receptor expression diversity and polymorphism. Further, this article provides a perspective on potential strategies to overcome these barriers and how diagnostic and prognostic biomarkers are being used in predicting response to mAb-based therapies. Overall, understanding these interdependent parameters can improve the current mAb-based formulations and develop novel mAb-based therapeutics for achieving durable clinical outcomes in a large subset of patients.

Pancreatic cancer (PC) is difficult to defeat due to mechanism (s) driving metastasis and drug resistance. Cancer stemness is a major challenging phenomenon associated with PC metastasis and limiting therapy efficacy. In this study, we evaluated the pre-clinical and clinical significance of eradicating pancreatic cancer stem cells (PCSC) and its components using a pan-EGFR inhibitor afatinib in combination with gemcitabine. Afatinib in combination with gemcitabine significantly reduced KrasG12D/+; Pdx-1 Cre (KC) (P

Purpose: The cytokine milieu in pancreatic ductal adenocarcinoma (PDAC) promotes tumor progression and immune suppression, contributing to the dismal prognosis of patients with PDAC. The roles of many of these cytokines, however, have not been thoroughly investigated in PDAC. Experimental Design: PDAC microarray and The Cancer Genome Atlas datasets were analyzed to identify cytokines and cognate receptors overexpressed in PDAC and associated with survival. Pathway and CIBERSORT analyses were used to elucidate potential mechanisms of altered patient survival. Comparative analysis of cytokine expression in KPC (K-rasG12D; TP53R172H; Pdx-1cre) and KC (K-rasG12D; Pdx-1cre) PDAC models and multicolor immunofluorescence (IF) staining of human PDAC–resected samples were used to validate these findings. Results: CXCL9 and CXCL10 were among the most highly overexpressed cytokines by bioinformatics analyses, while their receptor, CXCR3, was significantly overexpressed by IHC analysis. Higher CXCR3 ligand expression was associated with shorter overall survival, while high CXCR3 expression was associated with better survival. The CXCR3 ligands, CXCL4, 9, and 10, were overexpressed in KPC compared with KC mice. Pathway analysis of CXCR3- and CXCR3 ligand–associated genes showed that CXCR3 is a marker of antitumor immunity, while its ligands may promote immunosuppression. CIBERSORT and IF studies of PDAC tissues demonstrated that high CXCR3 expression was associated with increased CD8+ T-cell and naïve B-cell signatures and loss of plasma cell signatures. CXCR3 ligand expression was associated with increased CD8+ T-cell signatures and loss of natural killer–cell signatures. Conclusions: CXCR3 ligands are overexpressed in PDAC and are associated with poor survival likely related to alterations in tumor immune infiltrate/activity.

A dynamic mucosal layer shields the epithelial cells lining the body cavities and is made up of high molecular weight, heavily glycosylated, multidomain proteins called mucins. Mucins, broadly grouped into transmembrane and secreted mucins, are the first responders to any mechanical or chemical insult to the epithelia and help maintain tissue homeostasis. However, their intrinsic properties to protect and repair the epithelia are exploited during oncogenic processes, where mucins are metamorphosed to aid the tumor cells in their malignant journey. Diverse domains, like the variable number tandem repeats (VNTR), sea urchin sperm protein enterokinase and agrin (SEA), adhesion-associated domain (AMOP), nidogen-like domain (NIDO), epidermal growth factor-like domain (EGF), and von Willebrand factor type D domain (vWD) on mucins, including MUC1, MUC4, MUC5AC, MUC5B, and MUC16, have been shown to facilitate cell-to-cell and cell-to-matrix interactions, and cell-autonomous signaling to promote tumorigenesis and distant dissemination of tumor cells. Several obstacles have limited the study of mucins, including technical difficulties in working with these huge glycoproteins, the dearth of scientific tools, and lack of animal models; thus, the tissue-dependent and domain-specific roles of mucins during mucosal protection, chronic inflammation, tumorigenesis, and hematological dissemination of malignant cells are still unclear. Future studies should try to integrate information on the rheological, molecular, and biological characteristics of mucins to comprehensively delineate their pathophysiological role and evaluate their suitability as targets in future diagnostic and therapeutic strategies.

Many of the 20-member mucin family are evolutionarily conserved proteins that are often aberrantly expressed and glycosylated in various benign and malignant pathologies including oncogenic signaling leading to tumor invasion, metastasis, and immune evasion. Large size and extensive glycosylation present challenges to study mucin structure using traditional methods, including crystallography. We offer the hypothesis that the functional versatility of mucins may be attributed to the presence of intrinsically disordered regions (IDRs), which provide dynamism and flexibility; further, that these sites offer potential therapeutic targets. Herein, we examined the links between mucin structure and function based on IDRs, post-translational modifications (PTMs), and potential impact on their interactome. Using sequence-based bioinformatics tools, we observed that mucins are predicted to be moderately (20–40%) to highly (>40%) disordered and many conserved mucin domains could be disordered. Phosphorylation sites overlap with IDRs throughout the mucin sequences. Additionally, the majority of predicted O- and N- glycosylation sites in the tandem repeat regions occur within IDRs, and these IDRs contain a large number of functional motifs, i.e. molecular recognition features (MoRFs), which directly influence PPIs. This investigation provides a novel perspective and offers an insight into the complexity and dynamic nature of mucins.

Introduction Claudins are tight-junction proteins, which maintain an epithelial barrier in normal colon cells. Overexpression of Claudin-1 has been implicated for development of colon cancer. We postulated that Claudin-1 may be a useful target in near-infrared imaging and fluorescence-guided surgery. Methods We conjugated Claudin-1 antibody to LI-COR IR800DyeCW (Claudin-1-IRDye800CW). Western blotting of 9 human colon cancer cell lysates was performed. Animal imaging was performed with the LI-COR Pearl Trilogy Fluorescence Imaging System. A dose-response study was carried out with subcutaneous LS174T colon cancer cell line models. Increasing doses of Claudin-1-IRDye800CW via tail vein injection were administered to three groups of mice. Two groups of mice were used as controls (antibody alone, and dye alone). In vivo imaging was performed at 24, 48, and 72 h after administration of the conjugated dye. Orthotopic implantation of patient-derived tumors and cell lines was performed and peritoneal carcinomatosis models were created. After tumor growth, mice were administered Claudin-1-IRDye800CW and imaged in vivo 48 h later. The mice were euthanized and laparotomy was performed to assess internal organs and toxicity. Results Western blotting revealed that all colon cancer cell lysates expressed varying amounts of Claudin-1. All tumors demonstrated strong and specific fluorescence labeling at 800 nm, even with the lowest dose of 12.5 μg of Claudin-1-IRDye800CW. Conclusions Claudin-1 is a useful target for near-infrared antibody-based imaging for visualization of colorectal tumors for future use in fluorescence-guided surgery.

RNA polymerase II-associated factor 1 (PAF1)/pancreatic differentiation 2 (PD2) is a core subunit of the human PAF1 complex (PAF1C) that regulates the RNA polymerase II function during transcriptional elongation. PAF1/PD2 has also been linked to the oncogenesis of pancreatic ductal adenocarcinoma (PDAC). Here, we report that PAF1/PD2 undergoes posttranslational modification (PTM) through SUMOylation, enhancing the radiation resistance of PDAC cells. We identified that PAF1/PD2 is preferentially modified by small ubiquitin-related modifier 1 (SUMO 1), and mutating the residues (K)-150 and 154 by site-directed mutagenesis reduces the SUMOylation. Interestingly, PAF1/PD2 was found to directly interact with the promyelocytic leukemia (PML) protein in response to radiation, and inhibition of PAF1/PD2 SUMOylation at K-150/154 affects its interaction with PML. Our results demonstrate that SUMOylation of PAF1/PD2 increased in the radiated pancreatic cancer cells. Furthermore, inhibition of SUMOylation or PML reduces the cell growth and proliferation of PDAC cells after radiation treatment. These results suggest that SUMOylation of PAF1/PD2 interacts with PTM for PDAC cell survival. Furthermore, abolishing the SUMOylation in PDAC cells enhances the effectiveness of radiotherapy. Overall, our results demonstrate a novel PTM and PAF1/PD2 interaction through SUMOylation, and inhibiting the SUMOylation of PAF1/PD2 enhance the therapeutic efficacy for PDAC.

BackgroundMedulloblastoma (MB), the most frequent malignant pediatric brain tumor, is subdivided into four primary subgroups, wingless-type (WNT), sonic hedgehog (SHH), group 3, and group 4. Haploinsufficiency of chromosome 17p13.3 and c-Myc amplification distinguish high-risk group 3 tumors associated with rapid metastasis, recurrence and early mortality. We sought to identify the role of miR-212-3p, which resides on chromosome 17p13.3, in the pathophysiology of group 3 MB.MethodsWe first determined miR-212-3p expression in group 3 MB using several publicly-available datasets with confirmatory studies in vitro. We then identified epigenetic regulation by studying methylation and HDAC modifications along the promoter region. We used two systems for expression restoration, i.e. transient transfection or stable induction, to delineate miR-212-3p’s tumor suppressive and biochemical properties via assays assessing cancer proliferation, migration, invasion, colony formation, along with cell cycle and apoptosis analyses. We then compared MB and miR target databases to isolate a putative target whose biochemical and oncogenic properties were similarly elucidated using either transient silencing of target expression or stable induction of miR-212-3p.ResultsRNA expression analyses revealed dramatically reduced miR-212-3p levels in group 3 tumors and cell lines mainly through epigenetic silencing via histone modifications. Restoring miR-212-3p expression reduced in vitro cancer cell proliferation, migration, colony formation, and wound healing. Elevated miR-212-3p levels shifted c-Myc phosphorylation (from serine-62 to threonine-58), triggering destabilization and degradation; concurrently, its pro-apoptotic binding partners, i.e., Bin-1 and P19ARF, were upregulated with subsequent elevated apoptotic signals. Using a combination of transcriptomic data and dual luciferase assay, we isolated an oncogenic target of miR-212-3p, i.e. NFIB, a nuclear transcription factor implicated in metastasis and recurrence in various cancers. Increased expression of NFIB was confirmed in group 3 tumors, with poor survival shown in high-expressing patients. Transient NFIB silencing in vitro reduced cancer cell proliferation, colony formation, migration, and invasion. Concurrently, in group 3 MB cells, reduced medullosphere formation along with decreased expression of stem cell markers (Nanog, Oct4, Sox2, CD133) were noted.ConclusionThese results substantiate the tumor-suppressive role of miR-212-3p in group 3 MB and provide a potential therapeutic oncogenic target implicated in metastasis and tumor recurrence.

An imbalance in the crosstalk between the host and gut microbiota affects the intestinal barrier function, which results in inflammatory diseases and colorectal cancer. The colon epithelium protects itself from a harsh environment and various pathogenic organisms by forming a double mucus layer, primarily comprising mucins. Recent studies are focusing on how dietary patterns alter the gut microbiota composition, which in turn regulates mucin expression and maintains the intestinal layers. In addition, modulation of gut microbiota by microbiotic therapy (involving fecal microbiota transplantation) has emerged as a significant factor in the pathologies associated with dysbiosis. Therefore, proper communication between host and gut microbiota via different dietary patterns (prebiotics and probiotics) is needed to maintain mucus composition, mucin synthesis, and regulation. Here, we review how the interactions between diet and gut microbiota and bacterial metabolites (postbiotics) regulate mucus layer functionalities and mucin expression in human health and disease.

Simple Summary The current obstacles for discovering new drugs for cancer therapy have necessitated the development of the alternative strategy of drug repurposing, the identification of new uses for approved or investigational drugs for new therapeutic purposes. Niclosamide (Nic) is a Food and Drug Administration (FDA)-approved anti-helminthic drug, reported to have anti-cancer effects, and is being assessed in various clinical trials. In the current study, we assessed the therapeutic efficacy of Nic on pancreatic cancer (PC) in vitro. Our results revealed mitochondrial stress and mTORC1-dependent autophagy as the predominant players of Nic-induced PC cell death. This study provided a novel mechanistic insight for anti-cancer efficacy of Nic by increasing p-Gsk3β that modulates molecular signaling(s), including inhibition of hedgehog (Hh) signaling-mediated cellular proliferation and increased apoptosis through mTORC1-dependent autophagy may prove helpful for the development of novel PC therapies. Abstract Niclosamide (Nic), an FDA-approved anthelmintic drug, is reported to have anti-cancer efficacy and is being assessed in clinical trials for various solid tumors. Based on its ability to target multiple signaling pathways, in the present study, we evaluated the therapeutic efficacy of Nic on pancreatic cancer (PC) in vitro. We observed an anti-cancerous effect of this drug as shown by the G0/G1 phase cell cycle arrest, inhibition of PC cell viability, colony formation, and migration. Our results revealed the involvement of mitochondrial stress and mTORC1-dependent autophagy as the predominant players of Nic-induced PC cell death. Significant reduction of Nic-induced reactive oxygen species (ROS) and cell death in the presence of a selective autophagy inhibitor spautin-1 demonstrated autophagy as a major contributor to Nic-mediated cell death. Mechanistically, Nic inhibited the interaction between BCL2 and Beclin-1 that supported the crosstalk of autophagy and apoptosis. Further, Nic treatment resulted in Gsk3β inactivation by phosphorylating its Ser-9 residue leading to upregulation of Sufu and Gli3, thereby negatively impacting hedgehog signaling and cell survival. Nic induced autophagic cell death, and p-Gsk3b mediated Sufu/Gli3 cascade was further confirmed by Gsk3β activator, LY-294002, by rescuing inactivation of Hh signaling upon Nic treatment. These results suggested the involvement of a non-canonical mechanism of Hh signaling, where p-Gsk3β acts as a negative regulator of Hh/Gli1 cascade and a positive regulator of autophagy-mediated cell death. Overall, this study established the therapeutic efficacy of Nic for PC by targeting p-Gsk3β mediated non-canonical Hh signaling and promoting mTORC1-dependent autophagy and cell death.

Aberrant protein glycosylation has been shown to have a significant contribution in aggressive cancer, including pancreatic cancer (PC). Emerging evidence has implicated the involvement of cancer stem cells (CSCs) in PC aggressiveness; however, the contribution of glycosylation on self-renewal properties and maintenance of CSC is understudied. Here, using several in vitro and in vivo models lacking C1GALT1 expression, we identified the role of aberrant O-glycosylation in stemness properties and aggressive PC metastasis. A loss in C1GALT1 was found to result in the truncation of O-glycosylation on several glycoproteins with an enrichment of Tn carbohydrate antigen. Mapping of Tn-bearing glycoproteins in C1GALT1 KO cells identified significant Tn enrichment on CSC glycoprotein CD44. Notably, a loss of C1GALT1 in PC cells was found to enhance CSC features (side population-SP, ALDH1+, and tumorspheres) and self-renewal markers NANOG, SOX9, and KLF4. Furthermore, a loss of CD44 in existing C1GALT1 KO cells decreased NANOG expression and CSC features. We determined that O-glycosylation of CD44 activates ERK/NF-kB signaling, which results in increased NANOG expression in PC cells that facilitated the alteration of CSC features, suggesting that NANOG is essential for PC stemness. Finally, we identified that loss of C1GALT1 expression was found to augment tumorigenic and metastatic potential, while an additional loss of CD44 in these cells reversed the effects. Overall, our results identified that truncation of O-glycans on CD44 increases NANOG activation that mediates increased CSC activation.

Purpose: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy demonstrating aberrant and progressive expression of mucins. The contribution of individual mucins has been extensively investigated in PDAC; however, comprehensive mucin profiling including splice variants in PDAC tumors has not been reported. Experimental Design: Using publicly available RNA sequencing (RNA-seq) datasets, we assess the expression of mucin family members and their splice variants (SV) in PDAC tumor samples for the first time. Mucin SVs that are correlated with PDAC patient survival are validated in a cohort of patient tumor samples. Further, we use computational methods to derive novel pancreatic tumor subtypes using mucin expression signatures and their associated activated pathways. Results: Principal component analysis identified four novel mucin-based PDAC subtypes. Pathway analysis implicated specific biological signatures for each subtype, labeled (i) immune activated, (ii) progressive, (iii) pancreatitis-initiated, and (iv) anti-inflammatory/PanIN-initiated. Assessing mucin SVs, significantly longer survival is observed with higher expression of 4 MUC1 and 1 MUC13 SVs, whereas patients expressing 2 MUC4 and 1 MUC16 SVs had shorter survival. Using a whole-transcriptome correlation, a three-gene panel, including ESRP2, PTK6, and MAGEH1, is designated to assess PDAC tumor sample cellularity by PCR. One MUC4 SV and one MUC13 SV are quantified in a separate PDAC patient cohort, and their effects on survival are experimentally validated. Conclusions: Altogether, we demonstrate the unique expression pattern of mucins, four mucin-based PDAC subtypes, and the contribution of MUC1, MUC4, and MUC16 SVs in PDAC patient survival.

Cyclic nucleotide phosphodiesterases (PDEs) are superfamily of enzymes that regulate the spatial and temporal relationship of second messenger signaling in the cellular system. Among the 11 different families of PDEs, phosphodiesterase 1 (PDE1) sub-family of enzymes hydrolyze both 3’,5’-cyclic adenosine monophosphate (cAMP) and 3’,5’-cyclic guanosine monophosphate (cGMP) in a mutually competitive manner. The catalytic activity of PDE1 is stimulated by their binding to Ca(2+)/calmodulin (CaM), resulting in the integration of Ca(2+) and cyclic nucleotide-mediated signaling in various diseases. The PDE1 family includes three subtypes, PDE1A, PDE1B and PDE1C, which differ for their relative affinities for cAMP and cGMP. These isoforms are differentially expressed throughout the body, including the cardiovascular, central nervous system and other organs. Thus, PDE1 enzymes play a critical role in the pathophysiology of diseases through the fundamental regulation of cAMP and cGMP signaling. This comprehensive review provides the current research on PDE1 and its potential utility as a therapeutic target in diseases including the cardiovascular, pulmonary, metabolic, neurocognitive, renal, cancers and possibly others.

Farnesyl diphosphate synthase (FDPS), a mevalonate pathway enzyme, is highly expressed in several cancers, including prostate cancer (PCa). To date, the mechanistic, functional, and clinical significance of FDPS in cancer remains unexplored. We evaluated the FDPS expression and its cancer-associated phenotypes using in vitro and in vivo methods in PTEN-deficient and sufficient human and mouse PCa cells and tumors. Interestingly, FDPS overexpression synergizes with PTEN deficiency in PTEN conditionally knockout mice (P

The Ras family of GTPases is involved in cell proliferation, cell survival, and angiogenesis. It is upregulated in several cancers, including pancreatic cancer (PC) and leads to uncontrolled growth and aggressiveness. PC is well known to be a lethal disease with poor prognosis, plagued by limited therapeutic modalities. MicroRNAs (miRNAs), which are short non-coding RNA molecules, have recently emerged as regulators of signaling networks and have shown potential to target pathway components for therapeutic use in several malignancies. K-Ras mutations are widespread in PC cases (90%), with mutations detectable as early as pancreatic intraepithelial neoplasias and in later metastatic stages alike; therefore, these mutations in K-Ras are obvious drivers and potential targets for PC therapy. Several K-Ras targeting miRNAs have lately been discovered, and many of them have shown promise in combating pancreatic tumor growth in vitro and in mouse models. However, the field of miRNA therapy is still in its infancy, and miRNA mimics or anti-miRNA oligonucleotides that target Ras pathway have thus far not been evaluated in PC patients. In this review, we summarize the role of several miRNAs that regulate oncogenic K-Ras signaling in PC, with their prospective roles as therapeutic agents for targeting K-Ras pathway.

Background Pancreatic cancer (PC) is among foremost causes of cancer related deaths worldwide due to generic symptoms, lack of effective screening strategies and resistance to chemo- and radiotherapies. The risk factors associated with PC include several metabolic disorders such as obesity, insulin resistance and type 2 diabetes mellitus (T2DM). Studies have shown that obesity and T2DM are associated with PC pathogenesis; however, their role in PC initiation and development remains obscure. Main body Several biochemical and physiological factors associated with obesity and/or T2DM including adipokines, inflammatory mediators, and altered microbiome are involved in PC progression and metastasis albeit by different molecular mechanisms. Deep understanding of these factors and causal relationship between factors and altered signaling pathways will facilitate deconvolution of disease complexity as well as lead to development of novel therapies. In the present review, we focuses on the interplay between adipocytokines, gut microbiota, adrenomedullin, hyaluronan, vanin and matrix metalloproteinase affected by metabolic alteration and pancreatic tumor progression. Conclusions Metabolic diseases, such as obesity and T2DM, contribute PC development through altered metabolic pathways. Delineating key players in oncogenic development in pancreas due to metabolic disorder could be a beneficial strategy to combat cancers associated with metabolic diseases in particular, PC.

BACKGROUND & AIMS: Pancreatic ductal adenocarcinomas (PDACs) produce higher levels of truncated O-glycan structures (such as Tn and sTn) than normal pancreata. Dysregulated activity of core 1 synthase, glycoprotein-N-acetylgalactosamine 3-beta-galactosyltransferase 1 (C1GALT1) leads to increased expression of these truncated O-glycans. We investigated whether and how truncated O-glycans contributes to development and progression of PDAC using mice with disruption of C1galt1. METHODS: We crossed C1galt1 floxed mice (C1galt1(loxP/loxP)) with Kras(G12D/+); Trp53(R172H/+); Pdx1- Cre mice (KPC mice) to create KPCC mice. Growth and progression of pancreatic tumors were compared between KPC and KPCC mice; pancreatic tissues were collected and analyzed by immunohistochemistry; immunofluorescence; and Sirius red, alcian blue, and lectin staining. We used the CRISPR/Cas9 system to disrupt C1GALT1 in human PDAC cells (T3M4 and CD18/HPAF) and levels of O-glycans were analyzed by lectin blotting, mass spectrometry and lectin-pull down assay. Orthotopic studies and RNA sequencing analyses are performed with control and C1GALT1 knockout PDAC cells. C1GALT1 expression was analyzed in well differentiated (n=36) and poorly differentiated (n=23) PDAC samples by immunohistochemistry. RESULTS: KPCC mice had significantly shorter survival times (median, 102 days) than KPC mice (median, 200 days), and developed early pancreatic intraepithelial neoplasias at 3 weeks, PDAC at 5 weeks, and metastases at 10 weeks compared to KPC. Pancreatic tumors that developed in KPCC mice were more aggressive than those of KPC mice (more invasive and metastases), had a decreased amount of stroma, and had increased production of Tn. Poorly differentiated PDAC specimens had significantly lower levels of C1GALT1 than welldifferentiated PDACs. Human PDAC cells with knockout of C1GALT1 had aberrant glycosylation of MUC16 compared with control cells, and increased expression of genes that regulate tumorigenesis and metastasis. CONCLUSIONS: In studies of KPC mice with disruption of C1galt1, we found that loss of C1galt1 promotes development of aggressive PDACs and increased metastasis. Knockout of C1GALT1 leads to increased tumorigenicity and truncation of O-glycosylation on MUC16, which could contribute to increased aggressiveness.

The receptor tyrosine kinase MET is frequently involved in malignant transformation and inhibiting its activity in MET-dependent cancers is associated with improved clinical outcomes. Emerging evidence also suggests that mitochondria play an essential role in tumorigenesis and Dynamin Related Protein (DRP1), a key component of the mitochondrial fission machinery, has emerged as an attractive therapeutic target. Here, we report that inhibiting MET activity with the tyrosine kinase inhibitor MGCD516 attenuates viability, migration, and invasion of non-small cell lung cancer (NSCLC) and malignant pleural mesothelioma (MPM) cell lines in vitro, and significantly retards tumor growth in vivo. Interestingly, MGCD516 treatment also results in altered mitochondrial morphology in these cell lines. Furthermore, inhibiting MET pharmacologically or knocking down its expression using siRNA, decreases DRP1 activity alluding to possible crosstalk between them in these two cancers. Consistently, a combination of MGCD516 and mdivi-1, a quinazolinone reported to inhibit mitochondrial fission, is more effective in attenuating proliferation of NSCLC and MPM cell lines than either drug alone. Considered together, the present study has uncovered a novel mechanism underlying mitochondrial regulation by MET that involves crosstalk with DRP1, and suggests that a combination therapy targeting both MET and DRP1 could be a novel strategy for NSCLC and MPM.

FACT ( fa cilitates c hromatin t ranscription), an essential and evolutionarily conserved heterodimer from yeast to humans, controls transcription and is found to be upregulated in various cancers. However, the basis for such upregulation is not clearly understood. Our recent results deciphering a new ubiquitin-proteasome system regulation of the FACT subunit SPT16 in orchestrating transcription in yeast hint at the involvement of the proteasome in controlling FACT in humans, with a link to cancer. To test this, we carried out experiments in human embryonic kidney (HEK293) cells, which revealed that human SPT16 undergoes ubiquitylation and that its abundance is increased following inhibition of the proteolytic activity of the proteasome, thus implying proteasomal regulation of human SPT16. Furthermore, we find that the increased abundance/expression of SPT16 in HEK293 cells alters the transcription of genes, including ones associated with cancer, and that the proteasomal degradation of SPT16 is impaired in kidney cancer (Caki-2) cells to upregulate SPT16. Like human SPT16, murine SPT16 in C2C12 cells also undergoes ubiquitylation and proteasomal degradation to regulate transcription. Collectively, our results reveal a proteasomal regulation of mammalian SPT16, with physiological relevance in controlling transcription, and implicate such proteasomal control in the upregulation of SPT16 in cancer.

Lung cancer (LC) is a heterogeneous disease consisting mainly of two subtypes, non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC), and remains the leading cause of death worldwide. Despite recent advances in therapies, the overall 5-year survival rate of LC remains less than 20%. The efficacy of current therapeutic approaches is compromised by inherent or acquired drug-resistance and severe off-target effects. Therefore, the identification and development of innovative and effective therapeutic approaches are critically desired for LC. The development of RNA-mediated gene inhibition technologies was a turning point in the field of RNA biology. The critical regulatory role of different RNAs in multiple cancer pathways makes them a rich source of targets and innovative tools for developing anticancer therapies. The identification of antisense sequences, short interfering RNAs (siRNAs), microRNAs (miRNAs or miRs), anti-miRs, and mRNA-based platforms holds great promise in preclinical and early clinical evaluation against LC. In the last decade, RNA-based therapies have substantially expanded and tested in clinical trials for multiple malignancies, including LC. This article describes the current understanding of various aspects of RNA-based therapeutics, including modern platforms, modifications, and combinations with chemo-/immunotherapies that have translational potential for LC therapies.

The Plexins family of proteins are well-characterized transmembrane receptors of semaphorins, axon guidance cue molecules, that mediate the cell attraction or repelling effects for such cues. Plexins and their ligands are involved in numerous cellular activities, such as motility, invasion, and adhesion to the basement membrane. The detachment of cells and the gain in motility and invasion are hallmarks of the cancer metastasis cascade, thus generating interest in exploring the role of plexins in cancer metastasis. Semaphorin–plexin complexes can act as tumor promoters or suppressors, depending upon the cancer type, and are under investigation for therapeutic purposes. Our group has identified Semaphorin-5A (SEMA5A)/Plexin-B3 as an attractive targetable complex for pancreatic cancer (PC) metastasis. However, our understanding of the Plexin-B3 function and pathological expression in PC is limited, and our present study delineates the role of Plexin-B3 in PC malignancy. We examined the pathological expression of Plexin-B3 in PC tumors and metastasis using a human tissue microarray, disease progression model of PDX-Cre-Kras(G12D) (KC) mice, and different metastatic sites obtained from the KrasG12D, Trp53R172H, Pdx1-Cre (KPC) mice model. We observed a higher Plexin-B3 expression in PC tumor cores than the normal pancreas, and different metastatic sites were positive for Plexin-B3 expression. However, in the KC mice model, the Plexin-B3 expression increased initially and then decreased with the disease progression. Next, to evaluate the functional role of Plexin-B3, we utilized T3M-4- and CD18/HPAF-Control and -Plexin B3 knockdown cells for different in vivo and in vitro studies. The knockdown of Plexin-B3 enhanced the in vitro cellular migration, invasiveness, and impaired colony formation in three-dimensional culture, along with an increase in cellular spread and remodeling of the actin filaments. We also observed a higher metastasis in nude mice injected with T3M-4- and CD18/HPAF-shPlexin-B3 cells compared to their respective control cells. Furthermore, we observed a lower number of proliferating Ki-67-positive cells and higher ALDH1-A1-positive cells in the tumors formed by Plexin-B3 knockdown cells compared to tumors formed by the control cells. Together, our data suggest that the loss of Plexin-B3 is associated with the interference of cell division machinery and the induction of stem cell-like characteristics in PC cells.

An improved understanding of stem cell niches, organogenesis, and disease models has paved the way for developing a three-dimensional (3D) organoid culture system. Organoid cultures can be derived from primary tissues (single cells or tissue subunits), adult stem cells (ASCs), induced pluripotent stem cells (iPSCs), or embryonic stem cells (ESCs). As a significant technological breakthrough, 3D organoid models offer a promising approach for understanding the complexities of human diseases ranging from the mechanistic investigation of disease pathogenesis to therapy. Here, we discuss the recent applications, advantages, and limitations of organoids as in vitro models for studying metabolomics, drug development, infectious diseases, and the gut microbiome. We further discuss the use of organoids in cancer modeling using high throughput sequencing approaches.

Background & Aims Tumor-microenvironment factors and cancer stem cells (CSCs) play a critical role in the aggressiveness of pancreatic cancer (PC). However, the degree to which tumor-microenvironment factors promote stemness remains unexplored. Here, we examined whether cancer-associated fibroblasts (CAFs) promote CSC features in PC. Methods PC cells were treated long-term (30, 60, and 90 days) with conditioned media (CM)-derived from normal human fibroblasts (NFs) and CAFs. The stemness features of tumorsphere formation and stemness populations, along with CSCs markers, were analyzed using 2-dimensional and 3-dimensional sodium alginate bead-based co-culture models. Immunohistochemistry and immunofluorescence staining were performed for CSCs and fibroblast markers in autochthonous KrasG12D/+; Trp53R172H/+; Pdx1-Cre mice and human pancreatic tumors. Polymerase chain reaction array and gene knockdown were performed to identify the mechanism of stemness enrichment. Results Long-term treatment of PC cells with CAF-CM enriched stemness, as indicated by significantly higher CD44+, ALDH+, and AF+ populations in PC cells. Increased tumorsphere formation and elevated CSC, self-renewal, and drug-resistance markers in CAF-CM–treated PC cells were observed. In addition, CAFs co-cultured with PC cells in the 3-dimensional model showed a substantial increase in stemness features. CD44 and α–smooth muscle actin were positively correlated and their expressions progressively increased from the early to late stages of KrasG12D/+; Trp53R172H/+; Pdx1-Cre mouse and human pancreatic tumors. Osteopontin/secreted phosphoprotein 1 was identified as the top differentially overexpressed gene in CAF-CM–treated PC cells and knockdown of osteopontin/secreted phosphoprotein 1 significantly reduced stemness characteristics in CAF-CM–treated PC cells. Conclusions Our data uncovered novel insight into the interplay between CAF and enrichment of stemness population through the osteopontin/secreted phosphoprotein 1–CD44 axis in PC.

Purpose: Metabolic reprogramming and cancer stem cells drive the aggressiveness of pancreatic ductal adenocarcinoma (PDAC). However, the metabolic and stemness programs of pancreatic precursor lesions (PPL), considered early PDAC development events, have not been thoroughly explored. Experimental Design: Meta-analyses using gene expression profile data from NCBI Gene Expression Omnibus and IHC on tissue microarrays (TMA) were performed. The following animal and cellular models were used: cerulean-induced KrasG12D; Pdx1 Cre (KC) acinar-to-ductal metaplasia (ADM) mice, KrasG12D; Smad4Loss; Pdx-1 Cre (KCSmad4−) intraductal papillary mucinous neoplasm (IPMN) mice, LGKC1 cell line derived from the doxycycline-inducible Gnas IPMN model, and human IPMN organoids. Flow cytometry, Seahorse extracellular flux analyzer, qRT-PCR, and sphere assay were used to analyze metabolic and stemness features. SR18292 was used to inhibit PGC1α, and short hairpin RNA was used to knockdown (KD) PGC1α. Results: The meta-analysis revealed a significant upregulation of specific stemness genes in ADM-mediated pancreatic intraepithelial neoplasms (PanIN) and IPMN. Meta- and TMA analyses followed by in vitro and in vivo validation revealed that ADM/PanIN exhibit increased PGC1α and oxidative phosphorylation (OXPhos) but reduced CPT1A. IPMN showed elevated PGC1α, fatty acid β-oxidation (FAO) gene expression, and FAO-OXPhos. PGC1α was co-overexpressed with its coactivator NRF1 in ADM/PanINs and with PPARγ in IPMN. PGC1α KD or SR18292 inhibited the specific metabolic and stemness features of PPLs and repressed IPMN organoid growth. Conclusions: ADM/PanINs and IPMNs show specific stemness signatures with unique metabolisms. Inhibition of PGC1α using SR18292 diminishes the specific stemness by targeting FAO-independent and FAO-dependent OXPhos of ADM/PanINs and IPMNs, respectively.

Pancreatic cancer (PC) is acquired postnatally; to mimic this scenario, we developed an inducible KrasG12D; Ptf1a-CreER™ (iKC) mouse model, in which Kras is activated postnatally at week 16 upon tamoxifen (TAM) administration. Upon TAM treatment, iKC mice develop pancreatic intraepithelial neoplasia (PanIN) lesions and PC with metastasis at the fourth and fortieth weeks, respectively, and exhibited acinar-to-ductal metaplasia (ADM) and transdifferentiation. Kras activation upregulated the transcription factors Ncoa3, p-cJun and FoxM1, which in turn upregulated expression of transmembrane mucins (Muc1, Muc4 and Muc16) and secretory mucin (Muc5Ac). Interestingly, knockdown of KrasG12D in multiple PC cell lines resulted in downregulation of MUC1, MUC4, MUC5AC and MUC16. In addition, iKC mice exhibited ADM and transdifferentiation. Our results show that the iKC mouse more closely mimics human PC development and can be used to investigate pancreatic ductal adenocarcinoma (PDAC) biomarkers, early onset of PDAC, and ADM. The iKC model can also be used for preclinical strategies such as targeting mucin axis alone or in combination with neo-adjuvant, immunotherapeutic approaches and to monitor chemotherapy response., Summary: In this study, we developed a tamoxifen-inducible transgenic mouse model that closely mimics the age-related onset of pancreatic cancer (PC). We showed that activation of Kras (iKC) specifically regulates transcription factors that promote mucin expression and acinar to ductal metaplasia, whereas, loss of KrasG12D allele in PC cells resulted in the reduction of mucin proteins such as MUC1, MUC4, MUC5AC and MUC16.

Pancreatic ductal adenocarcinoma (PDAC) metastasizes to distant organs, which is the primary cause of mortality; however, specific features mediating organ-specific metastasis remain unexplored. Emerging evidence demonstrates that cancer stem cells (CSCs) and cellular metabolism play a pivotal role in metastasis. Here we investigated the role of distinct subtypes of pancreatic CSCs and their metabolomic signatures in organ-specific metastatic colonization. We found that PDAC consists of ALDH+/CD133+ and drug-resistant (MDR1+) subtypes of CSCs with specific metabolic and stemness signatures. Human PDAC tissues with gemcitabine treatment, autochthonous mouse tumors from KrasG12D; Pdx1-Cre (KC) and KrasG12D; Trp53R172H; Pdx-1 Cre (KPC) mice, and KPC- Liver/Lung metastatic cells were used to evaluate the CSC, EMT (epithelial-to-mesenchymal transition), and metabolic profiles. A strong association was observed between distinct CSC subtypes and organ-specific colonization. The liver metastasis showed drug-resistant CSC- and EMT-like phenotype with aerobic glycolysis and fatty acid β-oxidation-mediated oxidative (glyco-oxidative) metabolism. On the contrary, lung metastasis displayed ALDH+/CD133+ and MET-like phenotype with oxidative metabolism. These results were obtained by evaluating FACS-based side population (SP), autofluorescence (AF+) and Alde-red assays for CSCs, and Seahorse-based oxygen consumption rate (OCR), extracellular acidification rate (ECAR), and fatty acid β-oxidation (FAO)-mediated OCR assays for metabolic features along with specific gene signatures. Further, we developed in vitro human liver and lung PDAC metastasis models by using a combination of liver or lung decellularized scaffolds, a co-culture, and a sphere culture methods. PDAC cells grown in the liver-mimicking model showed the enrichment of MDR1+ and CPT1A+ populations, whereas the PDAC cells grown in the lung-mimicking environment showed the enrichment of ALDH+/CD133+ populations. In addition, we observed significantly elevated expression of ALDH1 in lung metastasis and MDR1/LDH-A expression in liver metastasis compared to human primary PDAC tumors. Our studies elucidate that distinct CSCs adapt unique metabolic signatures for organotropic metastasis, which will pave the way for the development of targeted therapy for PDAC metastasis.

Purpose: Docetaxel plays an indispensable role in the management of advanced prostate cancer. However, more than half of patients do not respond to docetaxel, and those good responders frequently experience significant cumulative toxicity, which limits its dose duration and intensity. Hence, a second agent that could increase the initial efficacy of docetaxel and maintain tolerability at biologically effective doses may improve outcomes for patients. Experimental Design: We determined phosphodiesterase 5 (PDE5) expression levels in human and genetically engineered mouse (GEM) prostate tissues and tumor-derived cell lines. Furthermore, we investigated the therapeutic benefits and underlying mechanism of PDE5 inhibitor sildenafil in combination with docetaxel using in vitro, Pten conditional knockout (cKO), derived tumoroid and xenograft prostate cancer models. Results: PDE5 expression was higher in both human and mouse prostate tumors and cancer cell lines compared with normal tissues/cells. In GEM prostate-derived cell lines, PDE5 expression increased from normal prostate (wild-type) epithelial cells to androgen-dependent and castrated prostate-derived cell lines. The addition of physiologically achievable concentrations of sildenafil enhanced docetaxel-induced prostate cancer cell growth inhibition and apoptosis in vitro, reduced murine 3D tumoroid growth, and in vivo tumorigenicity as compared with docetaxel alone. Furthermore, sildenafil enhanced docetaxel-induced NO and cGMP levels thereby augmenting antitumor activity. Conclusions: Our results demonstrate that sildenafil's addition could sensitize docetaxel chemotherapy in prostate cancer cells at much lesser concentration than needed for inducing cell death. Thus, the combinatorial treatment of sildenafil and docetaxel may improve anticancer efficacy and reduce chemotherapy-induced side-effects among patients with advanced prostate cancer.

BackgroundDifferential expression of mucins has been associated with several cancers including colorectal cancer (CRC). In normal physiological conditions, secretory mucin MUC5AC is not expressed in the colonic mucosa, whereas its aberrant expression is observed during development of colon cancer and its precursor lesions. To date, the molecular mechanism of MUC5AC in CRC progression and drug resistance remains obscure.MethodsMUC5AC expression was determined in colon tissue microarray by immunohistochemistry. A RNA interference and CRISPR/Cas9-mediated system was used to knockdown/knockout the MUC5AC in CRC cell lines to delineate its role in CRC tumorigenesis using in vitro functional assays and in vivo (sub-cutaneous and colon orthotopic) mouse models. Finally, CRC cell lines and xenograft models were used to identify the mechanism of action of MUC5AC.ResultsOverexpression of MUC5AC is observed in CRC patient tissues and cell lines. MUC5AC expression resulted in enhanced cell invasion and migration, and decreased apoptosis of CRC cells. MUC5AC interacted with CD44 physically, which was accompanied by the activation of Src signaling. Further, the presence of MUC5AC resulted in enhanced tumorigenesis and appearance of metastatic lesions in orthotopic mouse model. Additionally, up-regulation of MUC5AC resulted in resistance to 5-fluorouracil (5-FU) and oxaliplatin, and its knockout increased sensitivity to these drugs. Finally, we observed that up-regulation of MUC5AC conferred resistance to 5-FU through down-regulation of p53 and its target genep21and up-regulation of β-catenin and its target genesCD44andLgr5.ConclusionOur findings suggest that differential expression of secretory mucin MUC5AC results in enhanced tumorigenesis and also confers chemoresistance via CD44/β-catenin/p53/p21 signaling.

Brain metastasis (BM) predominantly occurs in triple-negative (TN) and epidermal growth factor 2 (HER2)-positive breast cancer (BC) patients, and currently, there is an unmet need for the treatment of these patients. BM is a complex process that is regulated by the formation of a metastatic niche. A better understanding of the brain metastatic processes and the crosstalk between cancer cells and brain microenvironment is essential for designing a novel therapeutic approach. In this context, the aberrant expression of miRNA has been shown to be associated with BM. These non-coding RNAs/miRNAs regulate metastasis through modulating the formation of a metastatic niche and metabolic reprogramming via regulation of their target genes. However, the role of miRNA in breast cancer brain metastasis (BCBM) is poorly explored. Thus, identification and understanding of miRNAs in the pathobiology of BCBM may identify a novel candidate miRNA for the early diagnosis and prevention of this devastating process. In this review, we focus on understanding the role of candidate miRNAs in the regulation of BC brain metastatic processes as well as designing novel miRNA-based therapeutic strategies for BCBM.

Glycosylation is the most commonly occurring post-translational modifications, and is believed to modify over 50% of all proteins. The process of glycan modification is directed by different glycosyltransferases, depending on the cell in which it is expressed. These small carbohydrate molecules consist of multiple glycan families that facilitate cell–cell interactions, protein interactions, and downstream signaling. An alteration of several types of O-glycan core structures have been implicated in multiple cancers, largely due to differential glycosyltransferase expression or activity. Consequently, aberrant O-linked glycosylation has been extensively demonstrated to affect biological function and protein integrity that directly result in cancer growth and progression of several diseases. Herein, we provide a comprehensive review of several initiating enzymes involved in the synthesis of O-linked glycosylation that significantly contribute to a number of different cancers.

Summary Tumor heterogeneity and cisplatin resistance are major causes of tumor relapse and poor survival. Here, we show that in lung cancer, interaction between paxillin (PXN) and integrin β4 (ITGB4), components of the focal adhesion (FA) complex, contributes to cisplatin resistance. Knocking down PXN and ITGB4 attenuated cell growth and improved cisplatin sensitivity, both in 2D and 3D cultures. PXN and ITGB4 independently regulated expression of several genes. In addition, they also regulated expression of common genes including USP1 and VDAC1, which are required for maintaining genomic stability and mitochondrial function, respectively. Mathematical modeling suggested that bistability could lead to stochastic phenotypic switching between cisplatin-sensitive and resistant states in these cells. Consistently, purified subpopulations of sensitive and resistant cells re-created the mixed parental population when cultured separately. Altogether, these data point to an unexpected role of the FA complex in cisplatin resistance and highlight a novel non-genetic mechanism., Graphical Abstract, Highlights • In lung cancer, interaction between PXN and ITGB4 contributes to cisplatin resistance. • Knocking down PXN and ITGB4 attenuated cell growth and improved cisplatin sensitivity. • PXN and ITGB4 regulate expression of USP1 and VDAC1, respectively. • The data underscore a novel non-genetic mechanism for chemoresistance in lung cancer., Cell Biology; Mathematical Biosciences; Cancer

Aberrant glycosylation plays a critical role in tumor aggressiveness, progression, and metastasis. Emerging evidence associates cancer initiation and metastasis to the enrichment of cancer stem cells (CSCs). Several universal markers have been identified for CSCs characterization; however, a specific marker has not yet been identified for different cancer types. Specific glycosylation variation plays a major role in the progression and metastasis of different cancers. Interestingly, many of the CSC markers are glycoproteins and undergo differential glycosylation. Given the importance of CSCs and altered glycosylation in tumorigenesis, the present review will discuss current knowledge of altered glycosylation of CSCs and its application in cancer research.

Background Ovarian cancer is a complicated malady associated with cancer stem cells (CSCs) contributing to 238,700 estimated new cases and 151,900 deaths per year, worldwide. CSCs comprise a tiny fraction of tumor-bulk responsible for cancer recurrence and eventual mortality. CSCs or tumor initiating cells are responsible for self-renewal, differentiation and proliferative potential, tumor initiation capability, its progression, drug resistance and metastatic spread. Although several biomarkers are implicated in these processes, their distribution within the ovary and association with single cell type has neither been established nor demonstrated across ovarian tumor developmental stages. Therefore, precise identification, thorough characterization and effective targeted destruction of dormant and highly proliferating potent CSC populations is an immediate need. Results In view of this, distribution of various CSC (ALDH1/2, C-KIT, CD133, CD24 and CD44) and cell proliferation (KI67) specific markers in the ovarian surface epithelium (OSE) and cortex regions in normal ovary, and benign, borderline and high grade metastatic ovarian tumors by immuno-histochemistry and confocal microscopy was studied. We further confirmed their expression by RT-PCR analysis. Co-expression analysis of stem cell (OCT4, SSEA4) and CSC (ALDH1/2, CD44 and LGR5) markers with proliferation marker (KI67) in HG tumors revealed dual positive proliferating stem and CSCs, few non-proliferating stem/CSC (SSEA4+/KI67− and ALDH1/2+/KI67−) and only KI67+ cells in cortex, signifying dynamic populations and interesting cellular hierarchy in cortex region. Smaller spherical (≤ 5 μm) and larger spindle/elliptical shaped (~ 10 μm) cell populations with high nucleo-cytoplasmic ratio were detected across all samples (including normal ovaries) but with variable distribution and characteristic stage-wise marker expression across different tumor stages. Conclusions Diverse stem and CSC populations expressing characteristic markers revealing distinct phenotypes (spherical ≤5 μm and spindle/elliptical ~ 10 μm) were distributed within different tumor stages studied signifying dynamic and probable functional hierarchy within these cell types. Involvement of extra-ovarian sites of origin of stem and CSCs requires rigorous evaluation. Quantitative analysis of potent CSC populations, their mechanisms and pathways for self-renewal, chemo-resistance, metastatic spread etc. with respect to various markers studied, will provide better insights and targets for developing effective therapeutics to prevent metastasis and eventually help improve patient mortality. Electronic supplementary material The online version of this article (10.1186/s13048-018-0439-3) contains supplementary material, which is available to authorized users.

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most challenging malignancies. Desmoplasia and tumor-supporting inflammation are hallmarks of PDAC. The tumor microenvironment contributes significantly to tumor progression and spread. Cancer-associated fibroblasts (CAFs) facilitate therapy resistance and metastasis. Recent reports emphasized the concurrence of multiple subtypes of CAFs with diverse roles, fibrogenic, and secretory. C-X-C motif chemokine receptor 2 (CXCR2) is a chemokine receptor known for its role during inflammation and its adverse role in PDAC. Oncogenic Kras upregulates CXCR2 and its ligands and, thus, contribute to tumor proliferation and immunosuppression. CXCR2 deletion in a PDAC syngeneic mouse model produced increased fibrosis revealing a potential undescribed role of CXCR2 in CAFs. In this study, we demonstrate that the oncogenic Kras-CXCR2 axis regulates the CAFs function in PDAC and contributes to CAFs heterogeneity. We observed that oncogenic Kras and CXCR2 signaling alter CAFs, producing a secretory CAF phenotype with low fibrogenic features; and increased secretion of pro-tumor cytokines and CXCR2 ligands, utilizing the NF-κB activity. Finally, using syngeneic mouse models, we demonstrate that oncogenic Kras is associated with secretory CAFs and that CXCR2 inhibition promotes activation of fibrotic cells (myofibroblasts) and impact tumors in a mutation-dependent manner.

Three-dimensional (3D) cultures use the property of some cells to self-organize in matrices and generate structures that can be programmed to represent an organ or a pathology. Organoid cultures are the 3D cultivation of source tissue (ranging from cells to tissue fragments) in a support matrix and specialized media that nearly resembles the physiological environment. Depending on the source tissue, growth factors, and inhibitors provided, organoids can be programmed to recapitulate the biology of a system and progression of pathology. Organoids are genetically stable, and genetically amenable, making them very suitable tools to study tissue homeostasis and cancer. In this Review, we focus on providing recent technical advances from published literature to efficiently use organoids as a tool for disease modeling and therapeutics. Also, we discuss stem cell biology principles used to generate multiple organoids and their characteristics, with a brief description of methodology. A major theme of this review is to expand organoid applications to the study disease progression and drug response in different cancers. We also discuss shortcomings, limitations, and advantages of developed 3D cultures, with the rationale behind the methodology.

Of the four primary subgroups of medulloblastoma, the most frequent cytogenetic abnormality, i17q, distinguishes Groups 3 and 4 which carry the highest mortality; haploinsufficiency of 17p13.3 is a marker for particularly poor prognosis. At the terminal end of this locus lies miR‐1253, a brain‐enriched microRNA that regulates bone morphogenic proteins during cerebellar development. We hypothesized miR‐1253 confers novel tumor‐suppressive properties in medulloblastoma. Using two different cohorts of medulloblastoma samples, we first studied the expression and methylation profiles of miR‐1253. We then explored the anti‐tumorigenic properties of miR‐1253, in parallel with a biochemical analysis of apoptosis and proliferation, and isolated oncogenic targets using high‐throughput screening. Deregulation of miR‐1253 expression was noted, both in medulloblastoma clinical samples and cell lines, by epigenetic silencing via hypermethylation; specific de‐methylation of miR‐1253 not only resulted in rapid recovery of expression but also a sharp decline in tumor cell proliferation and target gene expression. Expression restoration also led to a reduction in tumor cell virulence, concomitant with activation of apoptotic pathways, cell cycle arrest and reduction of markers of proliferation. We identified two oncogenic targets of miR‐1253, CDK6 and CD276, whose silencing replicated the negative trophic effects of miR‐1253. These data reveal novel tumor‐suppressive properties for miR‐1253, i.e., (i) loss of expression via epigenetic silencing; (ii) negative trophic effects on tumor aggressiveness; and (iii) downregulation of oncogenic targets.

Cancer remains a leading cause of death in the USA and around the world. Although the current synthetic inhibitors used in targeted therapies have improved patient prognosis, toxicity and development of resistance to these agents remain a challenge. Plant-derived natural products and their derivatives have historically been used to treat various diseases, including cancer. Several leading chemotherapeutic agents are directly or indirectly based on botanical natural products. Beyond these important drugs, however, a number of crude herbal or botanical preparations have also shown promising utility for cancer and other disorders. One such natural resource is derived from certain plants of the family Annonaceae, which are widely distributed in tropical and subtropical regions. Among the best known of these is Annona muricata, also known as soursop, graviola or guanabana. Extracts from the fruit, bark, seeds, roots and leaves of graviola, along with several other Annonaceous species, have been extensively investigated for anticancer, anti-inflammatory and antioxidant properties. Phytochemical studies have identified the acetogenins, a class of bioactive polyketide-derived constituents, from the extracts of Annonaceous species, and dozens of these compounds are present in different parts of graviola. This review summarizes current literature on the therapeutic potential and molecular mechanism of these constituents from A.muricata against cancer and many non-malignant diseases. Based on available data, there is good evidence that these long-used plants could have both chemopreventive and therapeutic potential. Appropriate attention to safety studies will be important to assess their effectiveness on various diseases caused or promoted by inflammation.

Heavily glycosylated secreted mucin MUC5AC, by the virtue of its cysteine-rich repeats, can form inter- and intramolecular disulfide linkages resulting in complex polymers, which in turn craft the framework of the polymeric mucus gel on epithelial cell surfaces. MUC5AC is a molecule with versatile functional implications including barrier functions to epithelial cells, host–pathogen interaction, immune cell attraction to sites of premalignant or malignant lesions and tumor progression in a context-dependent manner. Differential expression, glycosylation and localization of MUC5AC have been associated with a plethora of benign and malignant pathologies. In this era of robust technologies, overexpression strategies and genetically engineered mouse models, MUC5AC is emerging as a potential diagnostic, prognostic and therapeutic target for various malignancies. Considering the clinical relevance of MUC5AC, this review holistically encompasses its genomic organization, domain structure, glycosylation patterns, regulation, functional and molecular connotation from benign to malignant pathologies. Furthermore, we have here explored the incipient and significant experimental tools that are being developed to study this structurally complex and evolutionary conserved gel-forming mucin.

Pancreatic differentiation 2 (PD2)/RNA polymerase II–associated factor 1 (PAF1) is the core subunit of the human PAF1 complex (PAF1C) that regulates the promoter-proximal pausing of RNA polymerase II as well as transcription elongation and mRNA processing and coordinates events in mRNA stability and quality control. As an integral part of its transcription-regulatory function, PD2/PAF1 plays a role in posttranslational histone covalent modifications as well as regulates expression of critical genes of the cell-cycle machinery. PD2/PAF1 alone, and as a part of PAF1C, provides distinct roles in the maintenance of self-renewal of embryonic stem cells and cancer stem cells, and in lineage differentiation. Thus, PD2/PAF1 malfunction or its altered abundance is likely to affect normal cellular functions, leading to disease states. Indeed, PD2/PAF1 is found to be upregulated in poorly differentiated pancreatic cancer cells and has the capacity for neoplastic transformation when ectopically expressed in mouse fibroblast cells. Likewise, PD2/PAF1 is upregulated in pancreatic and ovarian cancer stem cells. Here, we concisely describe multifaceted roles of PD2/PAF1 associated with oncogenic transformation and implicate PD2/PAF1 as an attractive target for therapeutic development to combat malignancy. Cancer Res; 78(2); 313–9. ©2018 AACR.

Ovarian cancer is most lethal among gynecological cancers with often fatal consequences due to lack of effective biomarkers and relapse, which propels ovarian cancer research into unique directions to establish solid targeted therapeutics. “Ovarian stem cells” expressing germline pluripotent markers serve as novel paradigm with potential to address infertility, menopause, and probably influence tumor initiation. Cancer stem cells (CSCs) pose vital role in tumor recurrence and hence it is extremely important to study them with respect to ovarian stem cells across various cancer stages and normal ovaries. Pluripotent (OCT4, NANOG, SOX2, SSEA1, and SSEA4), germline (IFITM3, VASA/DDX4), and cancer stem (CD44, LGR5) cell specific markers were characterized for protein and mRNA expression in tumor tissues to understand their distribution in the surface epithelium and ovarian cortex in benign, borderline, and high-grade malignant stages. To elucidate whether pluripotent ovarian germline stem cells and CSCs are common subset of stem cells in tumor tissues, VASA was colocalized with known pluripotent stem (OCT4, SSEA1, SSEA4) and CSC (CD44, LGR5) specific markers by confocal microscopy. Single, smaller spherical (≤5 μm), and larger elliptical fibroblast like (≥10 μm) cells (also in clusters or multiples) were detected implying probable functional behavioral significance of cells in tumor initiation and metastasis across various cancer stages. Cells revealed characteristic staining pattern in ovarian surface epithelium (OSE) and cortex regions exclusive for each marker. Co-expression studies revealed specific subpopulations existing simultaneously in OSE and cortex and that a dynamic hierarchy of (cancer) stem cells with germline properties prevails in normal ovaries and cancer stages. Novel insights into CSC biology with respect to ovarian and germline stem cell perspective were obtained. Understanding molecular signatures and distribution within ovarian tissue may enable identification of precise tumor-initiating CSC populations and signaling pathways thus improving their efficient targeting and strategies to prevent their dissemination causing fatal relapse.

Semaphorin-5A (SEMA5A) has differential cell surface expression between normal and cancer cells and represents an attractive target for therapeutic intervention in pancreatic cancer (PC). In this study, we delineated the pathological expression and significance of SEMA5A during PC progression and metastasis. We utilized human tissue microarrays and different PC mouse models (Pdx1-cre; LSL- Kras(G12D), Pdx1-Cre; LSL-Kras(G12D); LSL-p53(R172H) and RIP1-Tag2) to analyze SEMA5A expression during PC progression. Using human patients and different mouse models, we demonstrated that SEMA5A expression was highest in liver metastases, followed by primary pancreatic tumors, and the lowest expression was found in the normal pancreas. SEMA5A expression was localized on tumor cells with no staining in the surrounding stroma. To understand the functional significance of SEMA5A, we treated PC cell lines with recombinant SEMA5A. We observed an increase in migration, chemotaxis, and scattering of PC cells. To delineate the signaling axis of SEMA5A, we generated SEMA5A receptor-Plexin-B3 knockdown in T3M-4 and CD18/HPAF PC cell lines and observed that the effect of SEMA5A treatment was absent in the Plexin-B3 knockdown counterparts of T3M-4 and CD18/HPAF cells. SEMA5A treatment leads to phosphorylation of cMET in Plexin-B3 dependent manner. Our data demonstrate that there is an increase in SEMA5A expression during PC progression and the elevation of this expression takes place at metastatic sites especially the liver in both exocrine and endocrine tumors. SEMA5A can elicit a migratory response in cells by activating cMET through the Plexin-B3 receptor. In conclusion, SEMA5A signaling represents a potential molecule for targeting metastasis in pancreatic cancer.