Genomic characterization of Gli-activator targets in sonic hedgehog-mediated neural patterning

Sonic hedgehog (Shh) acts as a morphogen to mediate the specification of distinct cell identities in the ventral neural tube through a Gli-mediated (Gli1-3) transcriptional network. Identifying Gli targets in a systematic fashion is central to the understanding of the action of Shh. We examined this issue in differentiating neural progenitors in mouse. An epitope-tagged Gli-activator protein was used to directly isolate cis-regulatory sequences by chromatin immunoprecipitation (ChIP). ChIP products were then used to screen custom genomic tiling arrays of putative Hedgehog (Hh) targets predicted from transcriptional profiling studies, surveying 50-150 kb of non-transcribed sequence for each candidate. In addition to identifying expected Gli-target sites, the data predicted a number of unreported direct targets of Shh action. Transgenic analysis of binding regions in Nkx2.2, Nkx2.1 (Titf1) and Rab34 established these as direct Hh targets. These data also facilitated the generation of an algorithm that improved in silico predictions of Hh target genes. Together, these approaches provide significant new insights into both tissue-specific and general transcriptional targets in a crucial Shh-mediated patterning process

DNA repair biomarkers XPF and phospho-MAPKAP kinase 2 correlate with clinical outcome in advanced head and neck cancer.

BackgroundInduction chemotherapy is a common therapeutic option for patients with locoregionally-advanced head and neck cancer (HNC), but it remains unclear which patients will benefit. In this study, we searched for biomarkers predicting the response of patients with locoregionally-advanced HNC to induction chemotherapy by evaluating the expression pattern of DNA repair proteins.MethodsExpression of a panel of DNA-repair proteins in formalin-fixed paraffin embedded specimens from a cohort of 37 HNC patients undergoing platinum-based induction chemotherapy prior to definitive chemoradiation were analyzed using quantitative immunohistochemistry.ResultsWe found that XPF (an ERCC1 binding partner) and phospho-MAPKAP Kinase 2 (pMK2) are novel biomarkers for HNSCC patients undergoing platinum-based induction chemotherapy. Low XPF expression in HNSCC patients is associated with better response to induction chemoradiotherapy, while high XPF expression correlates with a worse response (p = 0.02). Furthermore, low pMK2 expression was found to correlate significantly with overall survival after induction plus chemoradiation therapy (p = 0.01), suggesting that pMK2 may relate to chemoradiation therapy.ConclusionsWe identified XPF and pMK2 as novel DNA-repair biomarkers for locoregionally-advanced HNC patients undergoing platinum-based induction chemotherapy prior to definitive chemoradiation. Our study provides insights for the use of DNA repair biomarkers in personalized diagnostics strategies. Further validation in a larger cohort is indicated

A new concurrent chemotherapy with vinorelbine and mitomycin C in combination with radiotherapy in patients with locally advanced squamous cell carcinoma of the head and neck

Objective: The purpose of this pilot study was to evaluate the feasibility and toxicity of concurrent chemotherapy with vinorelbine and mitomycin C in combination with accelerated radiotherapy (RT) in patients with locally advanced cancer of the head and neck. Patients and Methods: Between January 2003 and March 2004, 15 patients with T4/N2-3 squamous cell carcinoma (12/15) and with N3 cervical lymph node metastases of carcinoma of unknown primary (3/15) were treated with chemotherapy and simultaneous accelerated RT. Results: 11 patients completed therapy without interruption or dose reduction. Grade 3 - 4 acute mucosal toxicity was observed in 9/15 patients, grade 4 hematologic toxicity in 6/15 patients. At a median follow-up of 7.5 months, 2 patients have died of intercurrent disease, 2 patients have experienced local relapse; 5 patients are alive with no evidence of disease at the primary tumor site. Discussion: The described regimen is highly effective, but led to remarkable side effects

A self-organized model for cell-differentiation based on variations of molecular decay rates

Systemic properties of living cells are the result of molecular dynamics governed by so-called genetic regulatory networks (GRN). These networks capture all possible features of cells and are responsible for the immense levels of adaptation characteristic to living systems. At any point in time only small subsets of these networks are active. Any active subset of the GRN leads to the expression of particular sets of molecules (expression modes). The subsets of active networks change over time, leading to the observed complex dynamics of expression patterns. Understanding of this dynamics becomes increasingly important in systems biology and medicine. While the importance of transcription rates and catalytic interactions has been widely recognized in modeling genetic regulatory systems, the understanding of the role of degradation of biochemical agents (mRNA, protein) in regulatory dynamics remains limited. Recent experimental data suggests that there exists a functional relation between mRNA and protein decay rates and expression modes. In this paper we propose a model for the dynamics of successions of sequences of active subnetworks of the GRN. The model is able to reproduce key characteristics of molecular dynamics, including homeostasis, multi-stability, periodic dynamics, alternating activity, differentiability, and self-organized critical dynamics. Moreover the model allows to naturally understand the mechanism behind the relation between decay rates and expression modes. The model explains recent experimental observations that decay-rates (or turnovers) vary between differentiated tissue-classes at a general systemic level and highlights the role of intracellular decay rate control mechanisms in cell differentiation.Comment: 16 pages, 5 figure

Reductive glutamine metabolism is a function of the α-ketoglutarate to citrate ratio in cells

Reductively metabolized glutamine is a major cellular carbon source for fatty acid synthesis during hypoxia or when mitochondrial respiration is impaired. Yet, a mechanistic understanding of what determines reductive metabolism is missing. Here we identify several cellular conditions where the α-ketoglutarate/citrate ratio is changed due to an altered acetyl-CoA to citrate conversion, and demonstrate that reductive glutamine metabolism is initiated in response to perturbations that result in an increase in the α-ketoglutarate/citrate ratio. Thus, targeting reductive glutamine conversion for a therapeutic benefit might require distinct modulations of metabolite concentrations rather than targeting the upstream signalling, which only indirectly affects the process.German Science Foundation (Grant FE1185)National Institutes of Health (U.S.) (Ruth L. Kirschstein National Research Service Award Postdoctoral Fellowship F32 CA132358)National Institutes of Health (U.S.) (Grant 5-P30-CA14051-39)Damon Runyon Cancer Research FoundationBurroughs Wellcome FundSmith Family FoundationNational Institutes of Health (U.S.) (Grant 1R01CA160458-01A1

Evaluation of a novel rash scale and a serum proteomic predictor in a randomized phase II trial of sequential or concurrent cetuximab and pemetrexed in previously treated non-small cell lung cancer

BACKGROUND: Candidate predictive biomarkers for epidermal growth factor receptor inhibitors (EGFRi), skin rash and serum proteomic assays, require further qualification to improve EGFRi therapy in non-small cell lung cancer (NSCLC). In a phase II trial that was closed to accrual because of changes in clinical practice we examined the relationships among candidate biomarkers, quantitative changes in tumor size, progression-free and overall survival. METHODS: 55 patients with progressive NSCLC after platinum therapy were randomized to receive (Arm A) cetuximab, followed by pemetrexed at progression, or (Arm B) concurrent cetuximab and pemetrexed. All received cetuximab monotherapy for the first 14 days. Pre-treatment serum and weekly rash assessments by standard and EGFRi-induced rash (EIR) scales were collected. RESULTS: 43 patients (20-Arm A, 23-Arm B) completed the 14-day run-in. Median survival was 9.1 months. Arm B had better median overall (Arm B = 10.3 [95% CI 7.5, 16.8]; Arm A = 3.5 [2.8, 11.7] months P = 0.046) and progression-free survival (Arm B = 2.3 [1.6, 3.1]; Arm A = 1.6 [0.9, 1.9] months P = 0.11). The EIR scale distributed ratings among 6 rather than 3 categories but ordinal scale rash severity did not predict outcomes. The serum proteomic classifier and absence of rash after 21 days of cetuximab did. CONCLUSIONS: Absence of rash after 21 days of cetuximab therapy and the serum proteomic classifier, but not ordinal rash severity, were associated with NSCLC outcomes. Although in a small study, these observations were consistent with results from larger retrospective analyses. TRIAL REGISTRATION: Clinicaltrials.gov Identifier NCT0020393

Densities of internally mixed organic-inorganic particles from mobility diameter measurements of aerodynamically classified aerosols

Accurate knowledge of particle density is essential for many aspects of aerosol science. Yet, density is often characterized poorly and incompletely for internally mixed particles, particularly for dry particles, with previous studies focused primarily on deliquescent (aqueous) droplets. Instead, densities for dry internally mixed particles are often inferred from mass composition measurements in combination with predictive models assuming ideal mixing, with the accuracy of such models not estimated. We determined particle densities from mobility diameter measurements (using a Scanning Mobility Particle Sizer, SMPS) for dried particles classified by their aerodynamic size (using an Aerosol Aerodynamic Classifier, AAC) for a range of two-component organic-inorganic particles containing known proportions of organic and inorganic species. We examined all permutations of mixing between four different organic (water soluble nigrosin dye, citric acid, polyethylene glycol-400, and ascorbic acid) and three different inorganic (sodium chloride, ammonium sulfate, and sodium nitrate) species. The accuracy and precision in our measured particle densities were ∼5% and ∼1%, respectively, for nonvolatile particles. Substantial deviations in particle density from ideal mixing (up to 20%) were observed. We tested descriptions of the non-ideal mixing for our systems by representing the volume change of mixing using Redlich-Kister (RK) polynomials in terms of mass fraction or in terms of mole fraction, with both approaches performing similarly.</p

Densities of internally mixed organic-inorganic particles from mobility diameter measurements of aerodynamically classified aerosols

Accurate knowledge of particle density is essential for many aspects of aerosol science. Yet, density is often characterized poorly and incompletely for internally mixed particles, particularly for dry particles, with previous studies focused primarily on deliquescent (aqueous) droplets. Instead, densities for dry internally mixed particles are often inferred from mass composition measurements in combination with predictive models assuming ideal mixing, with the accuracy of such models not estimated. We determined particle densities from mobility diameter measurements (using a Scanning Mobility Particle Sizer, SMPS) for dried particles classified by their aerodynamic size (using an Aerosol Aerodynamic Classifier, AAC) for a range of two-component organic-inorganic particles containing known proportions of organic and inorganic species. We examined all permutations of mixing between four different organic (water soluble nigrosin dye, citric acid, polyethylene glycol-400, and ascorbic acid) and three different inorganic (sodium chloride, ammonium sulfate, and sodium nitrate) species. The accuracy and precision in our measured particle densities were ∼5% and ∼1%, respectively, for nonvolatile particles. Substantial deviations in particle density from ideal mixing (up to 20%) were observed. We tested descriptions of the non-ideal mixing for our systems by representing the volume change of mixing using Redlich-Kister (RK) polynomials in terms of mass fraction or in terms of mole fraction, with both approaches performing similarly.</p

Phosphoglycerate dehydrogenase diverts glycolytic flux and contributes to oncogenesis

Most tumors exhibit increased glucose metabolism to lactate, however, the extent to which glucose-derived metabolic fluxes are used for alternative processes is poorly understood [1, 2]. Using a metabolomics approach with isotope labeling, we found that in some cancer cells a relatively large amount of glycolytic carbon is diverted into serine and glycine metabolism through phosphoglycerate dehydrogenase (PHGDH). An analysis of human cancers showed that PHGDH is recurrently amplified in a genomic region of focal copy number gain most commonly found in melanoma. Decreasing PHGDH expression impaired proliferation in amplified cell lines. Increased expression was also associated with breast cancer subtypes, and ectopic expression of PHGDH in mammary epithelial cells disrupted acinar morphogenesis and induced other phenotypic alterations that may predispose cells to transformation. Our findings show that the diversion of glycolytic flux into a specific alternate pathway can be selected during tumor development and may contribute to the pathogenesis of human cancer.National Institutes of Health (U.S.)National Cancer Institute (U.S.)Smith Family FoundationDamon Runyon Cancer Research FoundationBurroughs Wellcome Fun