Air Quality and Health Impacts of Onshore Oil and Gas Flaring and Venting Activities Estimated Using Refined Satellite-Based Emissions.

Emissions from flaring and venting (FV) in oil and gas (O&G) production are difficult to quantify due to their intermittent activities and lack of adequate monitoring and reporting. Given their potentially significant contribution to total emissions from the O&G sector in the United States, we estimate emissions from FV using Visible Infrared Imaging Radiometer Suite satellite observations and state/local reported data on flared gas volume. These refined estimates are higher than those reported in the National Emission Inventory: by up to 15 times for fine particulate matter (PM2.5), two times for sulfur dioxides, and 22% higher for nitrogen oxides (NOx). Annual average contributions of FV to ozone (O3), NO2, and PM2.5 in the conterminous U.S. (CONUS) are less than 0.15%, but significant contributions of up to 60% are found in O&G fields with FV. FV contributions are higher in winter than in summer months for O3 and PM2.5; an inverse behavior is found for NO2. Nitrate aerosol contributions to PM2.5 are highest in the Denver basin whereas in the Permian and Bakken basins, sulfate and elemental carbon aerosols are the major contributors. Over four simulated months in 2016 for the entire CONUS, FV contributes 210 additional instances of exceedances to the daily maximum 8-hr average O3 and has negligible contributions to exceedance of NO2 and PM2.5, given the current form of the national ambient air quality standards. FV emissions are found to cause over $7.4 billion in health damages, 710 premature deaths, and 73,000 asthma exacerbations among children annually.

Phase I LITESPARK-001 study of belzutifan for advanced solid tumors: Extended 41-month follow-up in the clear cell renal cell carcinoma cohort.

BACKGROUND: Accumulation of the HIF-2α transcription factor is an oncogenic event implicated in the tumorigenesis of clear cell renal cell carcinoma (ccRCC). In the phase I LITESPARK-001 study, the first-in-class HIF-2α inhibitor belzutifan demonstrated antitumor activity and an acceptable safety profile for pretreated patients with advanced ccRCC. Updated data with additional follow-up of > 40 months are presented. METHODS: LITESPARK-001 is an ongoing open-label study with a 3 + 3 dose-escalation design followed by an expansion phase. Patients with ccRCC enrolled at 7 sites received belzutifan 120 mg orally once daily until disease progression, unacceptable toxicity, or patient withdrawal. The data cutoff date was July 15, 2021. The primary end point was identifying the maximum tolerated dose and/or the recommended phase II dose. Secondary end points included objective response rate (ORR) and duration of response (DOR) per RECIST v1.1 by investigator assessment and safety. RESULTS: Median follow-up was 41.2 months (range, 38.2-47.7). Patients received a median of 3 (range, 1-9) prior systemic therapies. Of 55 patients, 14 (25 %) achieved an objective response. Median DOR was not reached (range, 3.1 + to 38.0 + months). Adverse events (AEs) attributed to study treatment by investigator assessment were reported in 53 patients (96 %). 22 patients (40 %) had grade 3 treatment-related AEs; the most common were anemia (n = 13; 24 %) and hypoxia (n = 7; 13 %). No grade 4 or 5 treatment-related AEs occurred. CONCLUSION: After a median follow-up of 41.2 months, belzutifan monotherapy demonstrated durable antitumor activity in patients with advanced ccRCC and acceptable safety. CLINICALTRIALS: gov. NCT02974738.

Unraveling the structural aspects of the G-quadruplex in SMO promoter and elucidating its contribution in transcriptional regulation.

We located a 25 nt G-rich sequence in the promoter region of SMO oncogene. We performed an array of biophysical and biochemical assays and confirmed the formation of a parallel G quadruplex (SMO1-GQ) by the identified sequence. SMO1-GQ is highly conserved in primates. For a comprehensive characterization of the SMO quadruplex structure, we have performed spectroscopic and in silico analysis with established GQ binder small molecules TMPyP4 and BRACO-19. We observed comparatively higher stable interaction of BRACO-19 with SMO1-GQ. Structure-based, rational drug design against SMO1-GQ to target SMO oncogene requires a detailed molecular anatomy of the G-quadruplex. We structurally characterised the SMO1-GQ using DMS footprinting assay and molecular modelling, docking, and MD simulation to identify the probable atomic regions that interact with either of the small molecules. We further investigated SMO1-GQ in vivo by performing chromatin immunoprecipitation (ChIP) assay. ChIP data revealed that this gene element functions as a scaffold for a number of transcription factors: specificity protein (Sp1), nucleolin (NCL), non-metastatic cell 2 (NM23-H2), cellular nucleic acid binding protein (CNBP), and heterogeneous nuclear ribonucleoprotein K (hnRNPK) which reflects the SMO1-P1 G-quadruplex to be the master regulator of SMO1 transcriptional activity. The strong binding interaction detected between SMO1-GQ and BRACO-19 contemplates the potential of the G quadruplex as a promising anti-cancer druggable target to downregulate SMO1 oncogene driven cancers.Communicated by Ramaswamy H. Sarma.

Using evolutionary constraint to define novel candidate driver genes in medulloblastoma.

Current knowledge of cancer genomics remains biased against noncoding mutations. To systematically search for regulatory noncoding mutations, we assessed mutations in conserved positions in the genome under the assumption that these are more likely to be functional than mutations in positions with low conservation. To this end, we use whole-genome sequencing data from the International Cancer Genome Consortium and combined it with evolutionary constraint inferred from 240 mammals, to identify genes enriched in noncoding constraint mutations (NCCMs), mutations likely to be regulatory in nature. We compare medulloblastoma (MB), which is malignant, to pilocytic astrocytoma (PA), a primarily benign tumor, and find highly different NCCM frequencies between the two, in agreement with the fact that malignant cancers tend to have more mutations. In PA, a high NCCM frequency only affects the BRAF locus, which is the most commonly mutated gene in PA. In contrast, in MB, >500 genes have high levels of NCCMs. Intriguingly, several loci with NCCMs in MB are associated with different ages of onset, such as the HOXB cluster in young MB patients. In adult patients, NCCMs occurred in, e.g., the WASF-2/AHDC1/FGR locus. One of these NCCMs led to increased expression of the SRC kinase FGR and augmented responsiveness of MB cells to dasatinib, a SRC kinase inhibitor. Our analysis thus points to different molecular pathways in different patient groups. These newly identified putative candidate driver mutations may aid in patient stratification in MB and could be valuable for future selection of personalized treatment options.

Noncoding RNA as an influential epigenetic modulator with promising roles in cancer therapeutics.

The epigenetic landscape has an important role in cellular homeostasis and its deregulation leads to cancer. Noncoding (nc)RNA networks function as major regulators of cellular epigenetic hallmarks via regulation of vital processes, such as histone modification and DNA methylation. They are integral intracellular components affecting multiple oncogenic pathways. Thus, it is important to elucidate the effects of ncRNA networks on epigenetic programming that lead to the initiation and progression of cancer. In this review, we summarize the effects of epigenetic modification influenced by ncRNA networks and crosstalk between diverse classes of ncRNA, which could aid the development of patient-specific cancer therapeutics targeting ncRNAs, thereby altering cellular epigenetics.

Belzutifan plus cabozantinib for patients with advanced clear cell renal cell carcinoma previously treated with immunotherapy: an open-label, single-arm, phase 2 study.

BACKGROUND: Few treatment options are available for patients with advanced renal cell carcinoma who have received previous anti-PD-1-based or anti-PD-L1-based immunotherapy. Combining belzutifan, an HIF-2α inhibitor, with cabozantinib, a multitargeted tyrosine-kinase inhibitor of VEGFR, c-MET, and AXL, might provide more antitumoural effects than either agent alone. We aimed to investigate the antitumour activity and safety of belzutifan plus cabozantinib in patients with advanced clear cell renal cell carcinoma that was previously treated with immunotherapy. METHODS: This open-label, single-arm, phase 2 study was conducted at ten hospitals and cancer centres in the USA. Patients were enrolled into two cohorts. Patients in cohort 1 had treatment-naive disease (results will be reported separately). In cohort 2, eligible patients were aged 18 years or older with locally advanced or metastatic clear cell renal cell carcinoma, measurable disease according to Response Evaluation Criteria in Solid Tumours version 1.1, an Eastern Cooperative Oncology Group performance status score of 0 or 1, and had previously received immunotherapy and up to two systemic treatment regimens. Patients were given belzutifan 120 mg orally once daily and cabozantinib 60 mg orally once daily until disease progression, unacceptable toxicity, or patient withdrawal. The primary endpoint was confirmed objective response assessed by the investigator. Antitumour activity and safety were assessed in all patients who received at least one dose of study treatment. This trial is registered with ClinicalTrials.gov, NCT03634540, and is ongoing. FINDINGS: Between Sept 27, 2018, and July 14, 2020, 117 patients were screened for eligibility, 52 (44%) of whom were enrolled in cohort 2 and received at least one dose of study treatment. Median age was 63·0 years (IQR 57·5-68·5), 38 (73%) of 52 patients were male, 14 (27%) were female, 48 (92%) were White, two (4%) were Black or African American, and two were Asian (4%). As of data cutoff (Feb 1, 2022), median follow-up was 24·6 months (IQR 22·1-32·2). 16 (30·8% [95% CI 18·7-45·1]) of 52 patients had a confirmed objective response, including one (2%) who had a complete response and 15 (29%) who had partial responses. The most common grade 3-4 treatment-related adverse event was hypertension (14 [27%] of 52 patients). Serious treatment-related adverse events occurred in 15 (29%) patients. One death was considered treatment related by the investigator (respiratory failure). INTERPRETATION: Belzutifan plus cabozantinib has promising antitumour activity in patients with pretreated clear cell renal cell carcinoma and our findings provide rationale for further randomised trials with belzutifan in combination with a VEGFR tyrosine-kinase inhibitor. FUNDING: Merck Sharp & Dohme (a subsidiary of Merck & Co) and the National Cancer Institute.

Leveraging base-pair mammalian constraint to understand genetic variation and human disease.

Thousands of genomic regions have been associated with heritable human diseases, but attempts to elucidate biological mechanisms are impeded by an inability to discern which genomic positions are functionally important. Evolutionary constraint is a powerful predictor of function, agnostic to cell type or disease mechanism. Single-base phyloP scores from 240 mammals identified 3.3% of the human genome as significantly constrained and likely functional. We compared phyloP scores to genome annotation, association studies, copy-number variation, clinical genetics findings, and cancer data. Constrained positions are enriched for variants that explain common disease heritability more than other functional annotations. Our results improve variant annotation but also highlight that the regulatory landscape of the human genome still needs to be further explored and linked to disease.

Leveraging Base Pair Mammalian Constraint to Understand Genetic Variation and Human Disease.

Although thousands of genomic regions have been associated with heritable human diseases, attempts to elucidate biological mechanisms are impeded by a general inability to discern which genomic positions are functionally important. Evolutionary constraint is a powerful predictor of function that is agnostic to cell type or disease mechanism. Here, single base phyloP scores from the whole genome alignment of 240 placental mammals identified 3.5% of the human genome as significantly constrained, and likely functional. We compared these scores to large-scale genome annotation, genome-wide association studies (GWAS), copy number variation, clinical genetics findings, and cancer data sets. Evolutionarily constrained positions are enriched for variants explaining common disease heritability (more than any other functional annotation). Our results improve variant annotation but also highlight that the regulatory landscape of the human genome still needs to be further explored and linked to disease.

Identification and characterization of a flexile G-quadruplex in the distal promoter region of stemness gene REX1.

We have identified a parallel G-quadruplex (R1WT) in the distal promoter region (-821 base-pairs upstream of the TSS) of the pluripotent gene REX1. Through biophysical and biochemical approach, we have characterized the G-quadruplex (GQ) as a potential molecular switch that may control REX1 promoter activity to determine the transcriptional fate. Small- molecule interactive study of the monomeric form of R1WT (characterized as R1mut2) with TMPyP4 and BRACO-19 revealed GQ destabilization upon interaction with TMPyP4 and stabilization upon interaction with BRACO-19. This distinctive drug interactivity suggests the in cellulo R1WT to be a promising drug target. The endogenous existence of R1WT was confirmed by BG4 antibody derived chromatin immunoprecipitation experiment. Here in, we also report the endogenous interaction of GQ specific transcription factors (TFs) with R1WT region in the human chromatin of cancer cell. The wild-type G-quadruplex was found to interact with four important transcription factors, (i) specificity protein (Sp1) (ii) non-metastatic cell 2 (NM23-H2): a diphosphatase (iii) cellular nucleic acid binding protein (CNBP) and (iv) heterogenous nuclear ribonucleoprotein K (hnRNPK) in the REX1 promoter. In contrast, nucleolin protein (NCL) binding was found to be low to the said G-quadruplex. The flexibility of R1WT between folded and unfolded states, obtained from experimental and computational analysis strongly suggests R1WT to be an important gene regulatory element in the genome. It controls promoter DNA relaxation with the coordinated interaction of transcription factors, the deregulation of which seeds stemness characteristic in cancer cells for further metastatic progression.

Curcumin arrests G-quadruplex in the nuclear hyper-sensitive III1 element of c-MYC oncogene leading to apoptosis in metastatic breast cancer cells.

c-MYC is deregulated in triple negative breast cancer (TNBC) pointing to be a promising biomarker for breast cancer treatment. Precise level of MYC expression is important in the control of cellular growth and proliferation. Designing of c-MYC-targeted antidotes to restore its basal level of cellular expression holds an optimistic approach towards anti-cancer treatment. MYC transcription is dominantly controlled by Nuclear Hypersensitive Element III-1 (NHEIII1) upstream of the promoter region possessing G-Quadruplex silencer element (Pu-27). We have investigated the selective binding-interaction profile of a natural phytophenolic compound Curcumin with native MYC G-quadruplex by conducting an array of biophysical experiments and in silico based Molecular Docking and Molecular Dynamic (MDs) simulation studies. Curcumin possesses immense anti-cancerous properties. We have observed significantly increased stability of MYC-G Quadruplex and thermodynamic spontaneity of Curcumin-MYC GQ binding with negative ΔG value. Transcription of MYC is tightly regulated by a complex mechanism involving promoters, enhancers and multiple transcription factors. We have used Curcumin as a model drug to understand the innate mechanism of controlling deregulated MYC back to its basal expression level. We have checked MYC-expression at transcriptional and translational level and proceeded for Chromatin Immuno-Precipitation assay (ChIP) to study the occupancy level of SP1, Heterogeneous nuclear ribonucleoprotein K (hnRNPK), Nucleoside Diphosphate Kinase 2 (NM23-H2) and Nucleolin at NHEIII1 upon Curcumin treatment of MDA-MB-231 cells. We have concluded that Curcumin binding tends to drive the equilibrium towards stable G-quadruplex formation repressing MYC back to its threshold-level. On retrospection of the synergistic effect of upregulated c-MYC and BCL-2 in cancer, we have also reported a new pathway [MYC-E2F-1-BCL-2-axis] through which Curcumin trigger apoptosis in cancer cells.Communicated by Ramaswamy H. Sarma.

Three-year review of a capacity building pilot for a sustainable regional network on food, nutrition and health systems education in India.

BACKGROUND: In Kolkata (India), there are high rates of malnourished children (45.9%) under the age of three, impacting growth, organ development, function, and cognition. Mothers have a major role to play during this crucial development stage, with research showing nutrition knowledge, attitudes and practices (KAP) of mothers are important determinants of childhood malnutrition. AIMS: To document 3 years of capacity building towards a sustainable nutrition education network in Kolkata, India, while assessing the ability to perform data collection in the form of needs assessments, impact assessments and capacity reviews. METHODS: Descriptive review and analysis of engagement and impact from 3 years of work by the NNEdPro Global Centre for Nutrition and Health, initiating locally led nutrition education interventions. Mapping to the Indian National Nutrition Strategy was also performed to review adherence to nationwide priorities surrounding nutrition and determine the wider application potential of the network. RESULTS: Two simultaneous projects were taken forward by a team of local healthcare professionals and student champions. Project 1-medical college workshops for medical student nutrition education with added focus on underserved populations, Project 2-preparation for a 'Mobile Teaching Kitchen' (MTK) in marginalised communities to empower local women as nutrition educators.Data collection methods used for analysing markers of impact and sustainability were semi-structured interviews of the community members, and KAP questionnaires to assess response to educational sessions. CONCLUSION: With local support it is possible to create and sustain fieldwork for an extended period with meaningful outputs and impact. This initiative demonstrates that it is possible to use healthcare professionals, students and volunteers with low-intensity training and a low-cost approach to produce action research with considerable impact and results in rapid, reliable and robust manner.

Long noncoding RNAs in cancer immunity: a new avenue in drug discovery.

The central role of the nonprotein-coding portion of the genome, such as long noncoding (lnc)RNAs is emerging as a hidden player manipulating the immune system in cancer. lncRNAs, in association with their interacting partners, regulate the expression of various immune system genes, which are perturbed during cancer. The tissue-specific expression of lncRNAs and their importance in cellular proliferation, the tumor microenvironment (TME), epithelial-mesenchymal transition (EMT), and modulation of the cells of the innate and adaptive immune system have novel therapeutic implications in establishing lncRNAs as biomarkers and targets to overcome cancer-associated immunosuppression. In this review, we establish and strengthen the link between lncRNAs and cancer immunity.

Nuclear Receptor Binding Protein 2 Is Downregulated in Medulloblastoma, and Reduces Tumor Cell Survival upon Overexpression.

Pseudokinases, comprising 10% of the human kinome, are emerging as regulators of canonical kinases and their functions are starting to be defined. We previously identified the pseudokinase Nuclear Receptor Binding Protein 2 (NRBP2) in a screen for genes regulated during neural differentiation. During mouse brain development, NRBP2 is expressed in the cerebellum, and in the adult brain, mainly confined to specific neuronal populations. To study the role of NRBP2 in brain tumors, we stained a brain tumor tissue array for NRPB2, and find its expression to be low, or absent, in a majority of the tumors. This includes medulloblastoma (MB), a pediatric tumor of the cerebellum. Using database mining of published MB data sets, we also find that NRBP2 is expressed at a lower level in MB than in the normal cerebellum. Recent studies indicate that MB exhibits frequent epigenetic alternations and we therefore treated MB cell lines with drugs inhibiting DNA methylation or histone deacetylation, which leads to an upregulation of NRBP2 mRNA expression, showing that it is under epigenetic regulation in cultured MB cells. Furthermore, forced overexpression of NRBP2 in MB cell lines causes a dramatic decrease in cell numbers, increased cell death, impaired cell migration and inhibited cell invasion in vitro. Taken together, our data indicate that downregulation of NRBP2 may be a feature by which MB cells escape growth regulation.

Whole-genome sequencing of glioblastoma reveals enrichment of non-coding constraint mutations in known and novel genes.

BACKGROUND: Glioblastoma (GBM) has one of the worst 5-year survival rates of all cancers. While genomic studies of the disease have been performed, alterations in the non-coding regulatory regions of GBM have largely remained unexplored. We apply whole-genome sequencing (WGS) to identify non-coding mutations, with regulatory potential in GBM, under the hypothesis that regions of evolutionary constraint are likely to be functional, and somatic mutations are likely more damaging than in unconstrained regions. RESULTS: We validate our GBM cohort, finding similar copy number aberrations and mutated genes based on coding mutations as previous studies. Performing analysis on non-coding constraint mutations and their position relative to nearby genes, we find a significant enrichment of non-coding constraint mutations in the neighborhood of 78 genes that have previously been implicated in GBM. Among them, SEMA3C and DYNC1I1 show the highest frequencies of alterations, with multiple mutations overlapping transcription factor binding sites. We find that a non-coding constraint mutation in the SEMA3C promoter reduces the DNA binding capacity of the region. We also identify 1776 other genes enriched for non-coding constraint mutations with likely regulatory potential, providing additional candidate GBM genes. The mutations in the top four genes, DLX5, DLX6, FOXA1, and ISL1, are distributed over promoters, UTRs, and multiple transcription factor binding sites. CONCLUSIONS: These results suggest that non-coding constraint mutations could play an essential role in GBM, underscoring the need to connect non-coding genomic variation to biological function and disease pathology.

Pembrolizumab alone or with chemotherapy versus cetuximab with chemotherapy for recurrent or metastatic squamous cell carcinoma of the head and neck (KEYNOTE-048): a randomised, open-label, phase 3 study.

BACKGROUND: Pembrolizumab is active in head and neck squamous cell carcinoma (HNSCC), with programmed cell death ligand 1 (PD-L1) expression associated with improved response. METHODS: KEYNOTE-048 was a randomised, phase 3 study of participants with untreated locally incurable recurrent or metastatic HNSCC done at 200 sites in 37 countries. Participants were stratified by PD-L1 expression, p16 status, and performance status and randomly allocated (1:1:1) to pembrolizumab alone, pembrolizumab plus a platinum and 5-fluorouracil (pembrolizumab with chemotherapy), or cetuximab plus a platinum and 5-fluorouracil (cetuximab with chemotherapy). Investigators and participants were aware of treatment assignment. Investigators, participants, and representatives of the sponsor were masked to the PD-L1 combined positive score (CPS) results; PD-L1 positivity was not required for study entry. The primary endpoints were overall survival (time from randomisation to death from any cause) and progression-free survival (time from randomisation to radiographically confirmed disease progression or death from any cause, whichever came first) in the intention-to-treat population (all participants randomly allocated to a treatment group). There were 14 primary hypotheses: superiority of pembrolizumab alone and of pembrolizumab with chemotherapy versus cetuximab with chemotherapy for overall survival and progression-free survival in the PD-L1 CPS of 20 or more, CPS of 1 or more, and total populations and non-inferiority (non-inferiority margin: 1·2) of pembrolizumab alone and pembrolizumab with chemotherapy versus cetuximab with chemotherapy for overall survival in the total population. The definitive findings for each hypothesis were obtained when statistical testing was completed for that hypothesis; this occurred at the second interim analysis for 11 hypotheses and at final analysis for three hypotheses. Safety was assessed in the as-treated population (all participants who received at least one dose of allocated treatment). This study is registered at ClinicalTrials.gov, number NCT02358031. FINDINGS: Between April 20, 2015, and Jan 17, 2017, 882 participants were allocated to receive pembrolizumab alone (n=301), pembrolizumab with chemotherapy (n=281), or cetuximab with chemotherapy (n=300); of these, 754 (85%) had CPS of 1 or more and 381 (43%) had CPS of 20 or more. At the second interim analysis, pembrolizumab alone improved overall survival versus cetuximab with chemotherapy in the CPS of 20 or more population (median 14·9 months vs 10·7 months, hazard ratio [HR] 0·61 [95% CI 0·45-0·83], p=0·0007) and CPS of 1 or more population (12·3 vs 10·3, 0·78 [0·64-0·96], p=0·0086) and was non-inferior in the total population (11·6 vs 10·7, 0·85 [0·71-1·03]). Pembrolizumab with chemotherapy improved overall survival versus cetuximab with chemotherapy in the total population (13·0 months vs 10·7 months, HR 0·77 [95% CI 0·63-0·93], p=0·0034) at the second interim analysis and in the CPS of 20 or more population (14·7 vs 11·0, 0·60 [0·45-0·82], p=0·0004) and CPS of 1 or more population (13·6 vs 10·4, 0·65 [0·53-0·80], p<0·0001) at final analysis. Neither pembrolizumab alone nor pembrolizumab with chemotherapy improved progression-free survival at the second interim analysis. At final analysis, grade 3 or worse all-cause adverse events occurred in 164 (55%) of 300 treated participants in the pembrolizumab alone group, 235 (85%) of 276 in the pembrolizumab with chemotherapy group, and 239 (83%) of 287 in the cetuximab with chemotherapy group. Adverse events led to death in 25 (8%) participants in the pembrolizumab alone group, 32 (12%) in the pembrolizumab with chemotherapy group, and 28 (10%) in the cetuximab with chemotherapy group. INTERPRETATION: Based on the observed efficacy and safety, pembrolizumab plus platinum and 5-fluorouracil is an appropriate first-line treatment for recurrent or metastatic HNSCC and pembrolizumab monotherapy is an appropriate first-line treatment for PD-L1-positive recurrent or metastatic HNSCC. FUNDING: Merck Sharp & Dohme.

Mast cells modulate proliferation, migration and stemness of glioma cells through downregulation of GSK3ß expression and inhibition of STAT3 activation.

Glioblastoma (GBM) heterogeneity is the main obstacle to efficient treatment due to the existence of subpopulation of cells with increased tumorigenicity and network of tumor associated parenchymal cells in the tumor microenvironment. We previously demonstrated that mast cells (MCs) infiltrate mouse and human gliomas in response to variety of signals in a glioma grade-dependent manner. However, the role of MCs in glioma development and the mechanisms behind MCs-glioma cells interaction remain unidentified. In the present study, we show that MCs upon activation by glioma cells produce soluble factors including IL-6, which are documented to be involved in cancer-related activities. We observe 'tumor educated' MCs decrease glioma cell proliferation and migration, reduce self-renewal capacity and expression of stemness markers but in turn promote glioma cell differentiation. 'Tumor educated' MC derived mediators exert these effects via inactivation of STAT3 signaling pathway through GSK3ß down-regulation. We identified 'tumor educated' MC derived IL-6 as one of the contributors among the complex mixture of MCs mediators, to be partially involved in the observed MC induced biological effect on glioma cells. Thus, MC mediated abolition of STAT3 signaling hampers glioma cell proliferation and migration by suppressing their stemness and inducing differentiation via down-regulation of GSK3ß expression. Targeting newly identified inflammatory MC-STAT3 axis could contribute to patient tailored therapy and unveil potential future therapeutic opportunities for patients.

Mast Cell Infiltration in Human Brain Metastases Modulates the Microenvironment and Contributes to the Metastatic Potential.

Metastatic brain tumors continue to be a clinical problem, despite new therapeutic advances in cancer treatment. Brain metastases (BMs) are among the most common mass lesions in the brain that are resistant to chemotherapies, have a very poor prognosis, and currently lack any efficient diagnostic tests. Predictions estimate that about 40% of lung and breast cancer patients will develop BM. Despite this, very little is known about the immunological and genetic aberrations that drive tumorigenesis in BM. In this study, we demonstrate the infiltration of mast cells (MCs) in a large cohort of human BM samples with different tissues of origin for primary cancer. We applied patient-derived BM cell models to the study of BM cell-MC interactions. BM cells when cocultured with MCs demonstrate enhanced growth and self-renewal capacity. Gene set enrichment analyses indicate increased expression of signal transduction and transmembrane proteins related genes in the cocultured BM cells. MCs exert their effect by release of mediators such as IL-8, IL-10, matrix metalloprotease 2, and vascular endothelial growth factor, thereby permitting metastasis. In conclusion, we provide evidence for a role of MCs in BM. Our findings indicate MCs' capability of modulating gene expression in BM cells and suggest that MCs can serve as a new target for drug development against metastases in the brain.

Serglycin as a potential biomarker for glioma: association of serglycin expression, extent of mast cell recruitment and glioblastoma progression.

Serglycin is an intracellular proteoglycan with a unique ability to adopt highly divergent structures by glycosylation with variable types of glycosaminoglycans (GAGs) when expressed by different cell types. Serglycin is overexpressed in aggressive cancers suggesting its protumorigenic role. In this study, we explored the expression of serglycin in human glioma and its correlation with survival and immune cell infiltration. We demonstrate that serglycin is expressed in glioma and that increased expression predicts poor survival of patients. Analysis of serglycin expression in a large cohort of low- and high-grade human glioma samples reveals that its expression is grade dependent and is positively correlated with mast cell (MC) infiltration. Moreover, serglycin expression in patient-derived glioma cells is significantly increased upon MC co-culture. This is also accompanied by increased expression of CXCL12, CXCL10, as well as markers of cancer progression, including CD44, ZEB1 and vimentin.In conclusion, these findings indicate the importance of infiltrating MCs in glioma by modulating signaling cascades involving serglycin, CD44 and ZEB1. The present investigation reveals serglycin as a potential prognostic marker for glioma and demonstrates an association with the extent of MC recruitment and glioma progression, uncovering potential future therapeutic opportunities for patients.

Serglycin proteoglycans limit enteropathy in Trichinella spiralis-infected mice.

BACKGROUND: Serglycin proteoglycans are essential for maturation of secretory granules and for the correct granular storage of cationic proteases in hematopoietic cells, e.g. mast cells. However, little is known about the in vivo functions of serglycin proteoglycans during infection. Here we investigated the potential role of serglycin proteoglycans in host defense after infection with the nematode Trichinella spiralis. RESULTS: Twelve days post infection lack of serglycin proteoglycans caused significantly increased enteropathy. The serglycin-deficient mice showed significantly increased intestinal worm burden, reduced recruitment of mast cells to the intestinal crypts, decreased levels of the mast cell proteases MCPT5 and MCPT6 in intestinal tissue, decreased serum levels of TNF-α, IL-1ß, IL-10 and IL-13, increased levels of IL-4 and total IgE in serum, and increased intestinal levels of the neutrophil markers myeloperoxidase and elastase, as compared to wild type mice. At five weeks post infection, increased larvae burden and inflammation were seen in the muscle tissue of the serglycin-deficient mice. CONCLUSIONS: Our results demonstrate that the serglycin-deficient mice were more susceptible to T. spiralis infection and displayed an unbalanced immune response compared to wild type mice. These findings point to an essential regulatory role of serglycin proteoglycans in immunity.

Targeting Serglycin Prevents Metastasis in Murine Mammary Carcinoma.

In hematopoietic cells, serglycin proteoglycans mainly contribute to proper storage and secretion of inflammatory mediators via their negatively charged glycosaminoglycans. Serglycin proteoglycans are also expressed in cancer cells where increased expression has been linked to poor prognosis. However, the serglycin-dependent mediators promoting cancer progression remain to be determined. In the present study we report that genetic ablation of serglycin proteoglycan completely blocks lung metastasis in the MMTV-PyMT-driven mouse breast cancer model, while serglycin-deficiency did not affect primary tumour growth or number of mammary tumours. Although E-cadherin expression was higher in the serglycin-deficient primary tumour tissue, indicating reduced invasiveness, serglycin-deficient tumour cells were still detected in the circulation. These data suggest that serglycin proteoglycans play a role in extravasation as well as colonization and growth of metastatic cells. A microarray expression analysis and functional annotation of differentially expressed genes identified several biological pathways where serglycin may be important. Our results suggest that serglycin and serglycin-dependent mediators are potential drug targets to prevent metastatic disease/dissemination of cancer.