Spiradenoma and cylindroma are distinctive skin adnexal tumors with sweat gland differentiation and potential for malignant transformation and aggressive behaviour. We present the genomic analysis of 75 samples from 57 representative patients including 15 cylindromas, 17 spiradenomas, 2 cylindroma-spiradenoma hybrid tumors, and 24 low- and high-grade spiradenocarcinoma cases, together with morphologically benign precursor regions of these cancers. We reveal somatic or germline alterations of the CYLD gene in 15/15 cylindromas and 5/17 spiradenomas, yet only 2/24 spiradenocarcinomas. Notably, we find a recurrent missense mutation in the kinase domain of the ALPK1 gene in spiradenomas and spiradenocarcinomas, which is mutually exclusive from mutation of CYLD and can activate the NF-κB pathway in reporter assays. In addition, we show that high-grade spiradenocarcinomas carry loss-of-function TP53 mutations, while cylindromas may have disruptive mutations in DNMT3A. Thus, we reveal the genomic landscape of adnexal tumors and therapeutic targets.

Nanotechnology has been materialized as a proficient technology for the development of anticancer nanoparticles all the way through an environment-friendly approach. Conventionally, nanoparticles have been assembled by dissimilar methods, but regrettably rely on the negative impact on the natural environment. Amalgamation of nanoparticles by means of plant extract is alternate conservative methods. Scutellaria barbata species was used majorly as food or as medicines against various diseases, and extensive research was conducted for their therapeutic properties. The present research was mainly focused on the synthesis of gold nanoparticles from the Scutellaria barbata by green route method and evaluation of its anticancer activity against pancreatic cancer cell lines (PANC-1). The gold nanoparticles have been characterized by UV-visible spectroscopy, TEM, SAED, AFM, and FTIR analysis. The synthesized gold nanoparticles (AuNPs) possessed effective anticancer activity against pancreatic cancer cell lines (PANC-1). Hence, further research on this plant may lead to the development of novel anticancer drugs which can be used to combat pancreatic cancer.

Intratumoral heterogeneity, particularly the potential cancer stemness of single cancer cells, has not yet been fully elucidated in human esophageal cancer. Single‑cell transcriptome sequencing of two types of esophageal adenocarcinoma (EAC) and two types of esophageal squamous cell carcinoma (ESCC) tissues was performed, and the intratumoral cancer stemness of the types of esophageal cancer were characterized at the single‑cell level in the present study. By comparing the transcriptomic profiles of single cancer cells with high and low stemness in individual patients, it was revealed that the overexpression of cell cycle‑associated genes in EAC cells was highly correlated with stemness, whereas overexpression of genes involved in the signaling pathways of DNA replication and DNA damage repair was significantly correlated with stemness in ESCC. High expression of these stemness‑associated genes was correlated with poor prognosis of patients. Additionally, poly [ADP‑ribose] polymerase(PARP)4 was identified as a novel cancer stemness‑associated gene in ESCC and its association with survival was validated in a cohort of 121 patients with ESCC. These findings have profound potential implications for the use of cell cycle inhibitors in EAC and PARP inhibitors in ESCC, which may provide novel mechanistic insights into the plasticity of esophageal cancer.

BACKGROUND & AIMS: Barrett's esophagus (BE) is a precursor to esophageal adenocarcinoma (EAC). Progression from BE to cancer is associated with obesity, possibly due to increased abdominal pressure and gastroesophageal reflux disease, although this pathogenic mechanism has not been proven. We investigated whether environmental or dietary factors associated with obesity contribute to the progression of BE to EAC in mice.
METHODS: Tg(ED-L2-IL1RN/IL1B)#Tcw mice (a model of BE, called L2-IL1B mice) were fed a chow (control) or high-fat diet (HFD) or were crossbred with mice that express human interleukin (IL) 8 (L2-IL1B/IL8 mice). Esophageal tissues were collected and analyzed for gene expression profiles and by quantitative polymerase chain reaction, immunohistochemistry, and flow cytometry. Organoids were established from BE tissue of mice and cultured with serum from lean or obese individuals or with neutrophils from L2-IL1B mice. Feces from mice were analyzed by 16s ribosomal RNA sequencing and compared to 16s sequencing data from patients with dysplasia or BE. L2-IL1B were mice raised in germ-free conditions.
RESULTS: L2-IL1B mice fed an HFD developed esophageal dysplasia and tumors more rapidly than mice fed the control diet; the speed of tumor development was independent of body weight. The acceleration of dysplasia by the HFD in the L2-IL1B mice was associated with a shift in the gut microbiota and an increased ratio of neutrophils to natural killer cells in esophageal tissues compared with mice fed a control diet. We observed similar differences in the microbiomes from patients with BE that progressed to EAC vs patients with BE that did not develop into cancer. Tissues from dysplasias of L2-IL1B mice fed the HFD contained increased levels of cytokines that are produced in response to CXCL1 (the functional mouse homolog of IL8, also called KC). Serum from obese patients caused organoids from L2-IL1B/IL8 mice to produce IL8. BE tissues from L2-IL1B mice fed the HFD and from L2-IL1B/IL8 mice contained increased numbers of myeloid cells and cells expressing Cxcr2 and Lgr5 messenger RNAs (epithelial progenitors) compared with mice fed control diets. BE tissues from L2-IL1B mice raised in germ-free housing had fewer progenitor cells and developed less dysplasia than in L2-IL1 mice raised under standard conditions; exposure of fecal microbiota from L2-IL1B mice fed the HFD to L2-IL1B mice fed the control diet accelerated tumor development.
CONCLUSIONS: In a mouse model of BE, we found that an HFD promoted dysplasia by altering the esophageal microenvironment and gut microbiome, thereby inducing inflammation and stem cell expansion, independent of obesity.

Development of novel methods is needed for the synthesis of nanoparticles. Attention on such particles has elevated disquiet about the eco-friendly manner of their fabrication methods. In the present study, we equipped and synthesized gold nanoparticles (AuNPs) from Panax notoginseng; investigated as an environmentally friendly and non-toxic substrate. Amalgamation of AuNPs was distinguished by numerous studies such as UV-absorbance and it shows peak values in the range of 520-550 nm. Nanoparticles sizes are confirmed by dynamic light scattering analysis and it shows 100 nm. Moreover, transmission electron microscopy (TEM) and energy dispersive X-ray analysis (EDX) confirmed the shape of the Au particles present in the synthesized materials. FTIR analysis studies showed the active molecules positioned in the plane of the synthesized particles. The anticancer potential of AuNPs was evaluated in PANC-1 cells. Additionally, AuNPs efficiently induced cytotoxicity, ROS and apoptosis by intonating intrinsic apoptotic gene expressions in PANC-1 cells. Finally, our study confirmed the synthesis of AuNPs from Panax notoginseng, which showed anticancer effects in an environmentally friendly manner.

Cervical cancer is the third most common highest mortality in women worldwide. The use of standard chemotherapeutic drugs against cervical cancer patients received several side effects. Therefore, we focused phytoconsituents-mediated synthesis of gold nanoparticles (AuNPs) considered as greatest attention in the treatment of cervical cancer. In this present study, we reported that green synthesis of AuNPs by using with Alternanthera Sessilis aqueous extract. Synthesis of AuNPs were characterized by UV visible spectroscopy, energy dispersive X-ray (EDX), selected area diffraction pattern (SAED), Fourier transform infrared spectroscopy (FTIR), high-resolution transmission electron microscopy (HR-TEM) and atomic force microscope. Synthesized AuNPs confirmed by the UV absorption maximum at 535 and crystal structure of gold AuNPs was further confirmed by EDX and SAED. TEM and atomic force microscopy images show the size and morphological distribution of nanoparticles. FTIR analysis was confirmed the hydroxyl groups, amine and alkaline groups of biomolecules are present in the AuNPs. Moreover, AuNPs induce cytotoxicity in cervical cancer cells and also induce apoptosis through modulating intrinsic apoptotic mechanisms in cervical cancer cells. This green synthesis of AuNPs from Alternanthera sessilis approach was easy, large scaled up and eco-friendly.

Short interfering RNA (siRNA) showed to be a viable alternative to a better prognosis in cancer therapy. Nevertheless, the successful application of this strategy still depends on the development of nanocarriers for the safe delivery of siRNA into the diseased tissue, which mostly occurs by passive accumulation. When an external magnetic field is applied, magnetic nanoparticles biodistribution is partially modulated to favor accumulation in a target tissue. In this work we designed a novel magnetic responsive siRNA nanocarrier. The new delivery system is composed of superparamagnetic iron oxide nanoparticles (SPIONs) coated with calcium phosphate (CaP) and PEG-polyanion block copolymers, which are known to be biocompatible. The nanoparticles presented rounded shape with small size and narrow distribution suitable for biomedical applications. TEM images showed dark spheres in the core surrounded by a lower electron density material in the corona. The X-ray photoelectron spectra (XPS) confirmed CaP-polymer coating of the magnetic core. In addition, the coating procedure did not affect the superparamagnetic property as showed using a vibrating sample magnetometer (VSM). With a high loading efficiency (80%), the nanoparticles enhanced vascular endothelium growth factor (VEGF) silencing in breast cancer cells in vitro, at gene and protein levels (~60% and 40%, respectively), without associated toxicity. Iron and siRNA quantification showed that the novel nanoparticles move towards a magnetic source carrying siRNA molecules. Therefore, these novel nanoparticles are a promising tool for cancer therapy based on RNAi effect, added by a magnetic capability to further modulate siRNA accumulation in the target tissue.

BACKGROUND: Barrett's esophagus (BE) is most commonly seen as the condition in which the normal squamous epithelium lining of the esophagus is replaced by goblet cells. Many studies show that BE is a predisposing factor for the development of esophageal adenocarcinoma (EAC), a particularly lethal cancer. The use of single nucleotide polymorphisms (SNPs) to map BE/EAC genes has previously provided insufficient genetic information to fully characterize the heterogeneous nature of the disease. We therefore hypothesize that rigorous interrogation of other types of genomic changes, e.g. tracts of homozygosity (TOH), repetitive elements, and insertion/deletions, may provide a comprehensive understanding of the development of BE/EAC.
RESULTS: First, we used a case-control framework to identify TOHs by using SNPs and tested for association with BE/EAC. Second, we used a case only approach on a validation series of eight samples subjected to exome sequencing to identify repeat elements and insertion/deletions. Third, insertion/deletions and repeat elements identified in the exomes were then mapped onto genes in the significant TOH regions. Overall, 24 TOH regions were significantly differentially represented among cases, as compared to controls (adjusted-P = 0.002-0.039). Interestingly, four BE/EAC-associated genes within the TOH regions consistently showed insertions and deletions that overlapped across eight exomes. Predictive functional analysis identified NOTCH, WNT, and G-protein inflammation pathways that affect BE and EAC.
CONCLUSIONS: The integration of common TOHs (cTOHs) with repetitive elements, insertions, and deletions within exomes can help functionally prioritize factors contributing to low to moderate penetrance predisposition to BE/EAC.

Ovarian cancer remains the most lethal type of cancer among all gynecological malignancies. The majority of patients are diagnosed with ovarian cancer at the late stages of the disease. Therefore, there exists an imperative need for the development of early ovarian cancer diagnostic techniques. Exosomes, secreted by various cell types, play pivotal roles in intercellular communication, which emerge as promising diagnostic and prognostic biomarkers for ovarian cancer. In this study, we present for the first time, at least to the best of our knowledge, the proteomics profiling of exosomes derived from the plasma of patients with ovarian cancer via liquid chromatography tandem mass spectrometry (LC‑MS/MS) with tandem mass tagging (TMT). The exosomes enriched from patient plasma samples were characterized by nanoparticle tracking analysis (NTA), dynamic light scattering (DLS), transmission electron microscopy (TEM) and western blot analysis. The size of the plasma exosomes fell into the range of 30 to 100 nm in diameter. The exosomal marker proteins, CD81 and TSG101, were clearly stained in the exosome samples; however, there was no staining for the endoplasmic reticulum protein, calnexin. A total of 294 proteins were identified with all exosome samples. Among these, 225 proteins were detected in both the cancerous and non‑cancerous samples. Apart from universal exosomal proteins, exosomes derived from ovarian cancer patient plasma also contained tumor‑specific proteins relevant to tumorigenesis and metastasis, particularly in epithelial ovarian carcinoma (EOC). Patients with EOC often suffer from coagulation dysfunction. The function of exosomes in coagulation was also examined. Several genes relevant to the coagulation cascade were screened out as promising diagnostic and prognostic factors that may play important roles in ovarian cancer progression and metastasis. On the whole, in this study, we successfully isolated and purified exosomes from plasma of patients with EOC, and identified a potential role of these exosomes in the coagulation cascade, as well as in the diagnosis and prognosis of patients.

The aim of the present investigation is to explore the innovative platform for the synthesis of plant-based nanoparticles, which contain biocompatible and biodegradable carrier of chitosan loaded with phloretin hydrophobic phytochemical applied as a stable anticancer agent. Treatment of cancer uses chemotherapeutic drugs as the cells are resistant to other drugs. However, the usage of therapeutic drug is limited by its poor solubility and low bioavailability. To overcome this problem, we fabricated the phloretin loaded chitosan nanoparticles (PhCsNPs) and physicochemical properties of PhCsNPs were characterized by FTIR, XRD, DLS, SEM and TEM. The findings indicated that the synthesized PhCsNPs were spherical and homogeneous in shape with the size distribution of 80-100 nm and exhibited stability in ultimate drug releasing profile. Further, we substantiated the anticancer efficiency of PhCsNPs through bio-assessment, such as cytotoxicity measurement, intracellular ROS, mitochondrial dysfunction, lipid peroxidation measurement, antioxidants status, apoptotic associated gene expression profile and cell cycle analysis in human oral cancer cell lines. The findings suggested that PhCsNPs augmented the mitochondrial-mediated apoptotic mechanism through the stimulation of oxidative stress, depletion of cellular antioxidants and cell cycle arrest. Our data suggested that PhCsNPs could be used as an efficient therapeutic agent for the treatment of oral cancer.

BACKGROUND: An altered Wnt-signaling activation has been reported during Barrett's esophagus progression, but with rarely detected mutations in APC and β-catenin (CTNNB1) genes.
METHODS: In this study, a robust in-depth expression pattern analysis of frizzled receptors, co-receptors, the Wnt-ligands Wnt3a and Wnt5a, the Wnt-signaling downstream targets Axin2, and CyclinD1, as well as the activation of the intracellular signaling kinases Akt and GSK3β was performed in an in vitro cell culture model of Barrett's esophagus. Representing the Barrett's sequence, we used normal esophageal squamous epithelium (EPC-1, EPC-2), metaplasia (CP-A) and dysplasia (CP-B) to esophageal adenocarcinoma (EAC) cell lines (OE33, OE19) and primary specimens of squamous epithelium, metaplasia and EAC.
RESULTS: A loss of Wnt3a expression was observed beginning from the metaplastic cell line CP-A towards dysplasia (CP-B) and EAC (OE33 and OE19), confirmed by a lower staining index of WNT3A in Barrett's metaplasia and EAC, than in squamous epithelium specimens. Frizzled 1-10 expression analysis revealed a distinct expression pattern, showing the highest expression for Fzd2, Fzd3, Fzd4, Fzd5, Fzd7, and the co-receptor LRP5/6 in EAC cells, while Fzd3 and Fzd7 were rarely expressed in primary specimens from squamous epithelium.
CONCLUSION: Despite the absence of an in-depth characterization of Wnt-signaling-associated receptors in Barrett's esophagus, by showing variations of the Fzd- and co-receptor profiles, we provide evidence to have a significant role during Barrett's progression and the underlying pathological mechanisms.

Squamous cell carcinoma antigen-1 (SCCA1) overexpression is associated with poor prognosis and chemoresistance in several tumor types, however, the underlying mechanisms remain elusive. Here, we report SCCA1 in relation to the immune and peritumoral adipose tissue microenvironment in early and advanced esophageal adenocarcinoma (EAC). In our series of patients with EAC, free SCCA1 serum levels were associated with significantly worse overall survival, and SCCA1-IgM serum levels showed a trend to a worse overall survival. Serum SCCA1 and intratumoral SCCA1 were inversely correlated with immune activation markers. In agreement with these findings, SCCA1 induced the expression of the immune checkpoint molecule programmed death ligand-1 on monocytes and a direct correlation of these 2 molecules was observed in sequential tumor sections. Furthermore, SCCA1 mRNA expression within the tumor was inversely correlated with stem cell marker expression both within the tumor and in the peritumoral adipose tissue. In vitro, in EAC cell lines treated with different chemotherapeutic drugs, cell viability was significantly modified by SCCA1 presence, as cells overexpressing SCCA1 were significantly more resistant to cell death. In conclusion, poor prognosis in EAC overexpressing SCCA1 is due to reduced tumor chemosensitivity as well as intratumoral immunity impairment, likely induced by this molecule.

Our aim was to characterize and validate that the location and age of onset of the tumor are both important criteria to classify colorectal cancer (CRC). We analyzed clinical and molecular characteristics of early-onset CRC (EOCRC) and late-onset CRC (LOCRC), and we compared each tumor location between both ages-of-onset. In right-sided colon tumors, early-onset cases showed extensive Lynch syndrome (LS) features, with a relatively low frequency of chromosomal instability (CIN), but a high CpG island methylation phenotype. Nevertheless, late-onset cases showed predominantly sporadic features and microsatellite instability cases due to

Marie Unna hereditary hypotrichosis (MUHH) and multiple familial trichoepithelioma (MFT) are both autosomal dominant disorders. Recently, certain genes (HR and EPS8L3) have been found to be responsible for MUHH, while CYLD has been demonstrated to be the main pathogenic gene in MFT patients. However, there exist a number of CYLD mutation-negative MFT cases, for which the causative gene has been unknown. Here, we identified a large, five-generation Han Chinese family with several patients presenting with MUHH and MFT. Sanger sequencing of three genes in 13 family members was performed. We found that the c.1A>G mutation in an inhibitory upstream open-reading frame of HR (U2HR) was present in all MUHH patients, while no pathogenic variants were found in the 3'- or 5'-untranslated regions, exons or flanking intronic sequences of EPS8L3 or CYLD in any family members. Subsequently, whole-genome sequencing was performed for five affected and one unaffected family member. We found no CYLD variants but identified an FABP12 variant (rs536105592 G>A) in the patients with both MUHH and MFT. These results suggest that the U2HR mutation was responsible for MUHH and the FABP12 variant may be coincidental in the accompanying MFT in this unique pedigree. This report deepens our understanding of the genetic basis of hair follicle diseases.

All-trans-retinoic acid (ATRA) is a potent inducer of cellular differentiation, growth arrest, and apoptosis as well as a front-line therapy for acute promyelocytic leukemia (APL). The present study provides evidence that induction of autophagy is required for ATRA to induce differentiation of APL (NB4) cells into granulocytes. ATRA treatment causes ~12-fold increase in the number of acidic vesicular organelles and induces marked up-regulation of LC3-II, autophagy-related 5 (ATG5), and Beclin-1. Transmission electron microscopy (TEM) revealed a decrease in mitochondria and ATRA-induced differentiation. To determine the role of autophagy in the differentiation of APL, we knocked down ATG5 in NB4 cells to find that ATRA-induced differentiation is significantly inhibited during ATG5 knock down in cells, indicating the role of autophagy in differentiation of APL. Further experiments revealed restriction of autophagy during ATRA-induced differentiation and inhibition of tissue transglutaminase 2 (TG2) and phospho-focal adhesion kinase (p-FAK), which are known to have roles in differentiation and cell attachment. We examined expression of Beclin-1 and B-cell lymphoma-2 (Bcl-2) and levels of mechanistic target of rapamycin (mTOR) after ATRA treatment. ATRA inhibits Bcl-2, up-regulates Beclin-1 expression, and reduces induction of mTOR activation/phosphorylation in NB4 cells. Our results reveal that autophagy has roles in regulation of differentiation, mitochondria elimination, and cell attachment during ATRA-induced APL differentiation.

Aberrant activation of Wnt/β-catenin signaling plays a central role in the pathogenesis of a wide variety of malignancies and is typically caused by mutations in core Wnt pathway components driving constitutive, ligand-independent signaling. In multiple myelomas (MMs), however, these pathway intrinsic mutations are rare despite the fact that most tumors display aberrant Wnt pathway activity. Recent studies indicate that this activation is caused by genetic and epigenetic lesions of Wnt regulatory components, sensitizing MM cells to autocrine Wnt ligands and paracrine Wnts emanating from the bone marrow niche. These include deletion of the tumor suppressor CYLD, promotor methylation of the Wnt antagonists WIF1, DKK1, DKK3, and sFRP1, sFRP2, sFRP4, sFRP5, as well as overexpression of the co-transcriptional activator BCL9 and the R-spondin receptor LGR4. Furthermore, Wnt activity in MM is strongly promoted by interaction of both Wnts and R-spondins with syndecan-1 (CD138) on the MM cell-surface. Functionally, aberrant canonical Wnt signaling plays a dual role in the pathogenesis of MM: (I) it mediates proliferation, migration, and drug resistance of MM cells; (II) MM cells secrete Wnt antagonists that contribute to the development of osteolytic lesions by impairing osteoblast differentiation. As discussed in this review, these insights into the causes and consequences of aberrant Wnt signaling in MM will help to guide the development of targeting strategies. Importantly, since Wnt signaling in MM cells is largely ligand dependent, it can be targeted by drugs/antibodies that act upstream in the pathway, interfering with Wnt secretion, sequestering Wnts, or blocking Wnt (co)receptors.

BACKGROUND & AIMS: Esophageal adenocarcinoma (EAC) is resistant to standard chemoradiation treatments, and few targeted therapies are available. We used large-scale tissue profiling and pharmacogenetic analyses to identify deregulated signaling pathways in EAC tissues that might be targeted to slow tumor growth or progression.
METHODS: We collected 397 biopsy specimens from patients with EAC and nonmalignant Barrett's esophagus (BE), with or without dysplasia. We performed RNA-sequencing analyses and used systems biology approaches to identify pathways that are differentially activated in EAC vs nonmalignant dysplastic tissues; pathway activities were confirmed with immunohistochemistry and quantitative real-time polymerase chain reaction analyses of signaling components in patient tissue samples. Human EAC (FLO-1 and EsoAd1), dysplastic BE (CP-B, CP-C, CP-D), and nondysplastic BE (CP-A) cells were incubated with pharmacologic inhibitors or transfected with small interfering RNAs. We measured effects on proliferation, colony formation, migration, and/or growth of xenograft tumors in nude mice.
RESULTS: Comparisons of EAC vs nondysplastic BE tissues showed hyperactivation of transforming growth factor-β (TGFB) and/or Jun N-terminal kinase (JNK) signaling pathways in more than 80% of EAC samples. Immunohistochemical analyses showed increased nuclear localization of phosphorylated JUN and SMAD proteins in EAC tumor tissues compared with nonmalignant tissues. Genes regulated by the TGFB and JNK pathway were overexpressed specifically in EAC and dysplastic BE. Pharmacologic inhibition or knockdown of TGFB or JNK signaling components in EAC cells (FLO-1 or EsoAd1) significantly reduced cell proliferation, colony formation, cell migration, and/or growth of xenograft tumors in mice in a SMAD4-independent manner. Inhibition of the TGFB pathway in BE cell lines reduced the proliferation of dysplastic, but not nondysplastic, cells.
CONCLUSIONS: In a transcriptome analysis of EAC and nondysplastic BE tissues, we found the TGFB and JNK signaling pathways to be hyperactivated in EACs and the genes regulated by these pathways to be overexpressed in EAC and dysplastic BE. Inhibiting these pathways in EAC cells reduces their proliferation, migration, and formation of xenograft tumors. Strategies to block the TGFB and JNK signaling pathways might be developed for treatment of EAC.

Autophagy, a dynamic pathway in which intracellular membrane structures sequester portions of the cytosol for degradation, plays multiple roles in physiological and pathological processes. Autophagy may have suppressive and promotive roles in the formation and progression of cancer. A growing number of methods to identify, quantify, and manipulate autophagy have been developed. Because most of these methods are semiquantitative and have significant limitations, it is important to emphasize that a combination of these assays is recommended for the analysis of autophagy. Here, I briefly discuss the autophagic process, its role in disease, and I summarize some of the best-known and most widely used methods to study autophagy in vitro in the context of cancer, including transmission electron microscopy (TEM), detection and quantification of the autophagy protein LC3 by western blot, and the use of GFP-LC3 to quantify puncta by fluorescence microscopy and tandem labeled RFP/mCherry-GFP-LC3 fluorescence microscopy to measure autophagic flux.

Esophageal adenocarcinoma (EAC) is a poor-prognosis cancer type with rapidly rising incidence. Understanding of the genetic events driving EAC development is limited, and there are few molecular biomarkers for prognostication or therapeutics. Using a cohort of 551 genomically characterized EACs with matched RNA sequencing data, we discovered 77 EAC driver genes and 21 noncoding driver elements. We identified a mean of 4.4 driver events per tumor, which were derived more commonly from mutations than copy number alterations, and compared the prevelence of these mutations to the exome-wide mutational excess calculated using non-synonymous to synonymous mutation ratios (dN/dS). We observed mutual exclusivity or co-occurrence of events within and between several dysregulated EAC pathways, a result suggestive of strong functional relationships. Indicators of poor prognosis (SMAD4 and GATA4) were verified in independent cohorts with significant predictive value. Over 50% of EACs contained sensitizing events for CDK4 and CDK6 inhibitors, which were highly correlated with clinically relevant sensitivity in a panel of EAC cell lines and organoids.

Gene therapy is a promising alternative that ensures effective treatment and cure for cancer. Here, we report graphene-reinforced chitosan (CS) construct based non-viral vector for tumor-targeted gene therapy. The therapeutic gene, pDNA-TNF-α, was loaded on to chitosan-carboxylated graphene oxide (CS-CGO) construct via electrostatic interaction. The pDNA-TNF-α-CS-CGO thus obtained was further passivated with 4,7,10-trioxa-1,13-tridecanediamine for protecting the vector from the mononuclear phagocyte system that contributes to the prolongation of circulation half-life. The surface passivated carrier (PEG-pDNA-TNF-α-CS-CGO) then festooned with the folic acid derived carbon dots (C-dots) for targeting folate receptors that are overexpressed in most of the cancer cells. The results of TEM images and zeta potential values ensured the occurrence of desired changes in each stage of C-dot-PEG-pDNA-TNF-α-CS-CGO formulation. After 14 days of incubation, the anti-angiogenesis effect was observed for final formulation in the chorioallantoic membrane. The results of in vitro gene expression study in cancer cell line show a comparatively higher transfection efficacy of the developed system (C-dot-PEG-pDNA-TNF-α-CS-CGO) than pDNA-TNF-α. The efficiency of the developed gene delivery system was further confirmed using a developed and validated artificial tumor cell apparatus.

The molecular mechanisms underlying the antineoplastic properties of metformin combined with nelfinavir remain elusive. To explore this question, transmission electron microscopy (TEM) was used to observe the combinatorial effect of inducing autophagosome formation in human cervical cancer cells. Western blotting respectively assayed protein expression of LC3I, LC3II, Beclin-1, Autophagy-related protein 7 (Atg7), Autophagy-related protein 3 (Atg3), NAD-dependent deacetylase sirtuin-3 (SIRT3) and major histocompatibility complex class I chain-related gene A (MICA). Lactate dehydrogenase (LDH) cytotoxicity assay evaluated natural killer (NK) cell cytotoxicity in the presence of metformin and nelfinavir in combination or each drug alone. Using tumor xenografts in a nude mouse model, antitumor efficacy of the drug combination was assessed. We found that the drug combination could induce autophagosome formation in human cervical cancer cells. The biomarker proteins of autophagy, including Beclin-1, Atg7 and Atg3, decreased, but the ratios of LC3I/II increased. We also found that this drug combination sensitizes human cervical cancer cells to NK cell-mediated lysis by increasing the protein of SIRT3 and MICA. Moreover, this drug combination markedly induced autophagy of SiHa xenografts in nude mice. Therefore, it can be concluded that metformin, in combination with nelfinavir, can induce SIRT3/mROS-dependent autophagy and sensitize NK cell-mediated lysis in human cervical cancer cells and cervical cancer cell xenografts in nude mice. Thus, our findings have revealed the detailed molecular mechanisms underlying the antitumor effects of metformin in combination with nelfinavir in cervical cancer.

Brooke-Spiegler syndrome (BSS; a.k.a. tuban tumor syndrome) is an autosomal dominant inherited skin disorder caused by germline mutations in the CYLD tumor suppressor gene. BSS is characterized by multiple skin adnexal tumors, mainly cylindromas and spiradenomas on the head and neck. The tumors are often severely disfiguring and require repeated surgical interventions. Here, we describe a four-generation BSS-family with a novel germline c.1613_1614delGC CYLD mutation that introduces a premature STOP codon predicted to result in a truncated, inactivated CYLD protein. In addition, we present a pilot study describing establishment of the first patient-derived xenografts (PDXs) from cutaneous CYLD-defective cylindromas. Fresh tumor tissues from cylindromas were transplanted into immunocompromised mice to generate PDXs. One xenograft showed progressive tumor growth after 3 months whereas the others remained unchanged in size during the 6 months study period. Histopathological and immunohistochemical analyses of the PDXs revealed that they recapitulate the histological and molecular features of their respective primary tumors, including expression of NTRK3 and the oncogenic driver MYB. In summary, we present the first preclinical BSS-model that morphologically and genetically recapitulates human CYLD-defective cylindromas. This model will be useful for preclinical therapeutic drug testing and for further studies of the molecular pathogenesis of inherited cylindromas.

BACKGROUND: Esophageal adenocarcinoma (EAC) is an aggressive disease with high mortality and an overall 5-year survival rate of less than 20%. Barrett's esophagus (BE) is the only known precursor of EAC, and patients with BE have a persistent and excessive risk of EAC over time. Individuals with BE are up to 30-125 times more likely to develop EAC than the general population. Thus, early detection of EAC and BE could significantly improve the 5-year survival rate of EAC. Due to the limitations of endoscopic surveillance and the lack of clinical risk stratification strategies, molecular biomarkers should be considered and thoroughly investigated.
AIM: To explore the transcriptome changes in the progression from normal esophagus (NE) to BE and EAC.
METHODS: Two datasets from the Gene Expression Omnibus (GEO) in NCBI Database (https://www.ncbi.nlm.nih.gov/geo/) were retrieved and used as a training and a test dataset separately, since NE, BE, and EAC samples were included and the sample sizes were adequate. This study identified differentially expressed genes (DEGs) using the R/Bioconductor project and constructed trans-regulatory networks based on the Transcriptional Regulatory Element Database and Cytoscape software. Enrichment of Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) terms was identified using the Database for Annotation, Visualization, and Integrated Discovery (DAVID) Bioinformatics Resources. The diagnostic potential of certain DEGs was assessed in both datasets.
RESULTS: In the GSE1420 dataset, the number of up-regulated DEGs was larger than that of down-regulated DEGs when comparing EAC
CONCLUSION: After the construction and analyses of the trans-regulatory networks in EAC and BE, the results indicate that COL1A1 and MMP1 could be potential biomarkers for EAC and BE, respectively.

Esophageal adenocarcinoma (EAC) has a dismal prognosis, and survival benefits of recent multimodality treatments remain small. Cancer-associated fibroblasts (CAFs) are known to contribute to poor outcome by conferring therapy resistance to various cancer types, but this has not been explored in EAC. Importantly, a targeted strategy to circumvent CAF-induced resistance has yet to be identified. By using EAC patient-derived CAFs, organoid cultures, and xenograft models we identified IL-6 as the stromal driver of therapy resistance in EAC. IL-6 activated epithelial-to-mesenchymal transition in cancer cells, which was accompanied by enhanced treatment resistance, migratory capacity, and clonogenicity. Inhibition of IL-6 restored drug sensitivity in patient-derived organoid cultures and cell lines. Analysis of patient gene expression profiles identified ADAM12 as a noninflammation-related serum-borne marker for IL-6-producing CAFs, and serum levels of this marker predicted unfavorable responses to neoadjuvant chemoradiation in EAC patients. These results demonstrate a stromal contribution to therapy resistance in EAC. This signaling can be targeted to resensitize EAC to therapy, and its activity can be measured using serum-borne markers.

BACKGROUND: The incidence of esophageal adenocarcinoma (EAC) is rising rapidly in the US and Western countries. The development of Barrett's esophagus (BE) and its progression to EAC have been linked to chronic gastroesophageal reflux disease (GERD). Exposure of BE and EAC cells to acidic bile salts (ABS) in GERD conditions induces high levels of oxidative stress and DNA damage. In this study, we investigated the role of insulin-like growth factor binding protein 2 (IGFBP2) in regulating ABS-induced DNA double-strand breaks.
METHODS: Real-time RT-PCR, western blot, immunohistochemistry, immunofluorescence, co-immunoprecipitation, flow cytometry, and cycloheximide (CHX) chase assays were used in this study. To mimic GERD conditions, a cocktail of acidic bile salts (pH 4) was used in 2D and 3D organotypic culture models. Overexpression and knockdown of IGFBP2 in EAC cells were established to examine the functional and mechanistic roles of IGFBP2 in ABS-induced DNA damage.
RESULTS: Our results demonstrated high levels of IGFBP2 mRNA and protein in EAC cell lines as compared to precancerous Barrett's cell lines, and IGFBP2 is frequently overexpressed in EACs (31/57). Treatment of EAC cells with ABS, to mimic GERD conditions, induced high levels of IGFBP2 expression. Knocking down endogenous IGFBP2 in FLO1 cells (with constitutive high levels of IGFBP2) led to a significant increase in DNA double-strand breaks and apoptosis, following transient exposure to ABS. On the other hand, overexpression of exogenous IGFBP2 in OE33 cells (with low endogenous levels of IGFBP2) had a protective effect against ABS-induced double-strand breaks and apoptosis. We found that IGFBP2 is required for ABS-induced nuclear accumulation and phosphorylation of EGFR and DNA-PKcs, which are necessary for DNA damage repair activity. Using co-immunoprecipitation assay, we detected co-localization of IGFBP2 with EGFR and DNA-PKcs, following acidic bile salts treatment. We further demonstrated, using cycloheximide chase assay, that IGFBP2 promotes EGFR protein stability in response to ABS exposure.
CONCLUSIONS: IGFBP2 protects EAC cells against ABS-induced DNA damage and apoptosis through stabilization and activation of EGFR - DNA-PKcs signaling axis.

BACKGROUND: HER2 (ERBB2 or HER2/neu) is a tyrosine-kinase increasing cell proliferation. Overexpression/amplification of HER2 is correlated with worse prognosis in solid malignancies. Consequently, HER2 targeting is established in breast and upper gastrointestinal tract cancer. There are conflicting data concerning the impact of HER2 overexpression on esophageal adenocarcinoma (EAC), as most studies do not differ between cancers of the esophagus/gastroesophageal junction and the stomach. The aim of this study was to analyze the expression/amplification of HER2 in EAC in correlation to clinicopathological data to verify its prognostic impact.
METHODS: We analyzed 428 EAC patients that underwent transthoracic thoraco-abdominal esophagectomy between 1997 and 2014. We performed HER2 immunohistochemistry (IHC) according to the guidelines and fluorescence-in-situ-hybridization (FISH) for IHC score2+, using tissue micro arrays (TMA) with up to eight biopsies from the surface and infiltration area of a single tumor for evaluating HER2-heterogeneity and single-spot TMA. The HER2-status was correlated with clinicopathological data.
RESULTS: HER2-positivity was found in up to 14.9% in our cohort (IHC score 3+ or IHC score 2+ with gene amplification) and demonstrated a significantly better overall survival (OS) in correlation to HER2-negative tumors (median OS 70.1 vs. 24.6 months, p = 0.006). HER2-overexpression was more frequently seen in lower tumor stages (pT1/pT2, p = 0.038), in the absence of lymphatic metastases (pN0/pN+, p = 0.020), and was significantly associated with better histological grading (G1/G2) (p = 0.041).
CONCLUSION: We demonstrated a positive prognostic impact of HER2 overexpression in a large cohort of EAC, contrary to other solid malignancies including gastric cancer and breast cancer, but consistent to the results of a large study on EAC from 2012.

Human papillomavirus (HPV) is a necessary but insufficient cause of a subset of oral squamous cell carcinomas (OSCCs) that is increasing markedly in frequency. To identify contributory, secondary genetic alterations in these cancers, we used comprehensive genomics methods to compare 149 HPV-positive and 335 HPV-negative OSCC tumor/normal pairs. Different behavioral risk factors underlying the two OSCC types were reflected in distinctive genomic mutational signatures. In HPV-positive OSCCs, the signatures of APOBEC cytosine deaminase editing, associated with anti-viral immunity, were strongly linked to overall mutational burden. In contrast, in HPV-negative OSCCs, T>C substitutions in the sequence context 5'-ATN-3' correlated with tobacco exposure. Universal expression of HPV

Gastrointestinal stromal tumors (GISTs) are the most common type of mesenchymal tumor of the digestive tract. MicroRNAs (miRNAs) are short non-coding RNAs, which control gene expression at a post-transcriptional level. Dysregulated miRNAs are involved in various types of human disease, including cancer. In the present study, it was revealed that miRNA-182 (miR-182) expression was significantly upregulated in human GISTs compared with adjacent normal tissues. Overexpression of miR-182 enhanced GIST-T1 cell growth, with increased proliferation and decreased apoptosis. miR-182 upregulation also promoted colony formation and migration of GIST-T1 cells. In addition, cylindromatosis (CYLD) was identified as a direct target of miR-182. Overexpression of miR-182 suppressed CYLD expression and enhanced downstream nuclear factor (NF)-κB activation. It was also determined that the expression of CYLD was downregulated in association with upregulated miR-182 in human GISTs. In conclusion, these results demonstrated that miR-182 promoted GIST cell growth by negatively regulating CYLD expression. These findings indicated that miR-182 antagonist may be a promising therapeutic strategy for the treatment of human GIST.

A lectin with a molecular mass of 12 ± 1 kDa was isolated for the first time from Geodorum densiflorum (Lam.) rhizome (GDL). The lectin exhibited hemagglutination activity both in mice and human erythrocytes which was inhibited by 4-nitrophenyl-β-D-glucopyranoside among the tested 26 sugars. The lectin was heat stable and showed its full activity in the pH range from 5.0 to 9.0. The lectin did not lose its activity in the presence of urea but the activity lost significantly when treated with EDTA. Divalent cation Ca

Recent studies have shown that sex and sex steroids influence the composition of the gut microbiome. These studies also indicate that steroid regulation of the gut microbiome may play a role in pathological situations of hormonal excess, such as PCOS. Indeed, studies demonstrated that PCOS is associated with decreased alpha diversity and changes in specific Bacteroidetes and Firmicutes, previously associated with metabolic dysregulation. These studies suggest that androgens may regulate the gut microbiome in females and that hyperandrogenism may be linked with a gut 'dysbiosis' in PCOS. Future mechanistic studies will be required to elucidate how sex steroids regulate the composition and function of the gut microbial community and what the consequences of this regulation are for the host.