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BACKGROUND An increase in thyroid cancers, predominantly papillary thyroid carcinoma (PTC), has been recently reported in children. METHODS The histopathology of 28 consecutive PTCs from the northeast United States was reviewed. None of the patients (ages 6-18 years; 20 females, 8 males) had significant exposure to radiation. Nucleic acid from tumors was tested for genetic abnormalities (n = 27). Negative results were reevaluated by targeted next-generation sequencing. RESULTS Seven of 27 PTCs (26%) had neurotrophic tyrosine kinase receptor (NTRK) fusion oncogenes (NTRK type 3/ets variant 6 [NTRK3/ETV6], n =5; NTRK3/unknown, n = 1; and NTRK type 1/translocated promoter region, nuclear basket protein [NTRK1/TPR], n = 1), including 5 tumors that measured >2 cm and 3 that diffusely involved the entire thyroid or lobe. All 7 tumors had lymphatic invasion, and 5 had vascular invasion. Six of 27 PTCs (22%) had ret proto-oncogene (RET) fusions (RET/PTC1, n = 5; RET/PTC3, n = 1); 2 tumors measured >2 cm and diffusely involved the thyroid, and 5 had lymphatic invasion, with vascular invasion in 2. Thirteen PTCs had the B-Raf proto-oncogene, serine/threonine kinase (BRAF) valine-to-glutamic acid mutation at position 600 (BRAFV600E) (13 of 27 tumors; 48%), 11 measured

Background: Clear cell renal cell carcinoma (ccRCC) is the most lethal form of kidney cancer. Small molecule VEGFR inhibitors are widely used but are not curative and various resistance mechanisms such as activation of the MET pathway have been described. Dual MET/VEGFR2 inhibitors have recently shown clinical benefit but limited preclinical data evaluates their effects in ccRCC. Methods: An interrogation of the Cancer Genome Atlas (TCGA) dataset was performed to evaluate oncogenic alterations in the MET/VEGFR2 pathway. We evaluated the in vitro effects of Cabozantinib, a dual MET/VEGFR2 inhibitor, using a panel of ccRCC cell lines. Drug effects of cell viability and proliferation, migration, cell scatter, anchorage independent growth, and downstream MET/VEGFR2 signaling pathways were assessed. Results: Twelve percent of TCGA cases had possible MET/HGF oncogenic alterations with co-occurrence noted (p

Our results showed that, at the same BMI level, Uygurs have greater WHR values, abdominal visceral fat content, and diabetes risks than Kazaks. In addition, values of HDL-C in Uygur subjects were lower than those in Kazak subjects, and values of creatinine, uric acid, diastolic blood pressure, blood glucose, and fructosamine in Uygur male subjects were lower than those in Kazak male subjects. In contrast, systolic blood pressure values in Uygur subjects were greater than those in Kazak subjects, and blood glucose values were greater in Uygur female subjects than in Kazak female subjects. Additionally, in Uygurs, visceral adipose tissue expression levels ofTBX1andTCF21were greater in obesity group than in normal and T2DM groups and lower in T2DM group than in normal group (P<0.01). The visceral adipose tissue expression levels ofAPNin normal group was greater than those in obesity and T2DM groups, and visceral adipose tissue expression levels ofTNF-αandMCP-1in normal group were lower than those in obesity and T2DM groups (P<0.01). In conclusion, T2DM in Uygurs was mainly associated with not only distribution of adipose tissue in body, but also change in metabolic activity and adipocytokines secretion of adipose tissue.

Associations of ErbB4 (ERBB4/HER4), the fourth member of the EGFR family, with cancer are variable, possibly as a result of structural diversity of this receptor. There are multiple structural isoforms of ERBB4 arising by alternative mRNA splicing, and a subset undergo proteolysis that releases membrane-anchored and soluble isoforms that associate with transcription factors and coregulators to modulate transcription. To compare the differential and common signaling activities of full-length (FL) and soluble intracellular isoforms of ERBB4, four JM-a isoforms (FL and soluble intracellular domain (ICD) CYT-1 and CYT-2) were expressed in isogenic MCF10A cells and their biologic activities were analyzed. Both FL and ICD CYT-2 promoted cell proliferation and invasion, and CYT-1 suppressed cell growth. Transcriptional profiling revealed several new and underexplored ERBB4-regulated transcripts, including: proteases/protease inhibitors (MMP3 and SERPINE2), the YAP/Hippo pathway (CTGF, CYR61, and SPARC), the mevalonate/cholesterol pathway (HMGCR, HMGCS1, LDLR, and DHCR7), and cytokines (IL8, CCL20, and CXCL1). Many of these transcripts were subsequently validated in a luminal breast cancer cell line that normally expresses ERBB4. Furthermore, ChIP-seq experiments identified ADAP1, APOE, SPARC, STMN1, and MXD1 as novel molecular targets of ERBB4. These findings clarify the diverse biologic activities of ERBB4 isoforms, and reveal new and divergent functions. Implications: ErbB4 as a regulator of Hippo and mevalonate pathways provides new insight into milk production and anabolic processes in normal mammary epithelia and cancer. Mol Cancer Res; 12(8); 1140–55. ©2014 AACR.

The JmjC domain-containing H3K4 histone demethylase jumonji AT-rich interactive domain 1B (JARID1B) (also known as KDM5B and PLU1) is overexpressed in breast cancer and is a potential target for breast cancer treatment. To investigate the in vivo function of JARID1B, we developed Jarid1b(-/-) mice and characterized their phenotypes in detail. Unlike previously reported Jarid1b(-/-) strains, the majority of these Jarid1b(-/-) mice were viable beyond embryonic and neonatal stages. This allowed us to further examine phenotypes associated with the loss of JARID1B in pubertal development and pregnancy. These Jarid1b(-/-) mice exhibited decreased body weight, premature mortality, decreased female fertility, and delayed mammary gland development. Related to these phenotypes, JARID1B loss decreased serum estrogen level and reduced mammary epithelial cell proliferation in early puberty. In mammary epithelial cells, JARID1B loss diminished the expression of key regulators for mammary morphogenesis and luminal lineage specification, including FOXA1 and estrogen receptor α. Mechanistically, JARID1B was required for GATA3 recruitment to the Foxa1 promoter to activate Foxa1 expression. These results indicate that JARID1B positively regulates mammary ductal development through both extrinsic and cell-autonomous mechanisms.

A fundamental goal in cancer biology is to identify the cells and signalling pathways that are keys to induce tumour regression. Here we use a spontaneously self-regressing tumour, cutaneous keratoacanthoma (KAs), to identify physiological mechanisms that drive tumour regression. By using a mouse model system that recapitulates the behaviour of human KAs, we show that self-regressing tumours shift their balance to a differentiation programme during regression. Furthermore, we demonstrate that developmental programs utilized for skin hair follicle regeneration, such as Wnt, are hijacked to sustain tumour growth and that the retinoic acid (RA) signalling pathway promotes tumour regression by inhibiting Wnt signalling. Finally, we find that RA signalling can induce regression of malignant tumours that do not normally spontaneously regress, such as squamous cell carcinomas. These findings provide new insights into the physiological mechanisms of tumour regression and suggest therapeutic strategies to induce tumour regression., Keratoacanthomas are skin tumours that spontaneously regress but the mechanisms leading to regression are unknown. Here, using a mouse chemical carcinogenesis model, the authors show that tumour regression is driven by activation of retinoic acid signalling that induces Wnt inhibition and tumour differentiation.

The development of high-throughput sequencing methods allows for the characterization of microbial communities in a wide range of environments on an unprecedented scale. However, insight into microbial community composition is limited by our ability to detect patterns in this flood of sequences. Here we compare the performance of 51 analysis techniques using real and simulated bacterial 16S rRNA pyrosequencing datasets containing either clustered samples or samples arrayed across environmental gradients. We find that many diversity patterns are evident with severely undersampled communities, and that methods vary widely in their ability to detect gradients and clusters. Chi-squared distances and Pearson correlation distances perform especially well for detecting gradients, while Gower and Canberra distances perform especially well for detecting clusters. These results also provide a basis for understanding tradeoffs between number of samples and depth of coverage, tradeoffs which are important to consider when designing studies to characterize microbial communities.

Background Genotyping platforms such as single nucleotide polymorphism (SNP) arrays are powerful tools to study genomic aberrations in cancer samples. Allele specific information from SNP arrays provides valuable information for interpreting copy number variation (CNV) and allelic imbalance including loss-of-heterozygosity (LOH) beyond that obtained from the total DNA signal available from array comparative genomic hybridization (aCGH) platforms. Several algorithms based on hidden Markov models (HMMs) have been designed to detect copy number changes and copy-neutral LOH making use of the allele information on SNP arrays. However heterogeneity in clinical samples, due to stromal contamination and somatic alterations, complicates analysis and interpretation of these data. Methods We have developed MixHMM, a novel hidden Markov model using hidden states based on chromosomal structural aberrations. MixHMM allows CNV detection for copy numbers up to 7 and allows more complete and accurate description of other forms of allelic imbalance, such as increased copy number LOH or imbalanced amplifications. MixHMM also incorporates a novel sample mixing model that allows detection of tumor CNV events in heterogeneous tumor samples, where cancer cells are mixed with a proportion of stromal cells. Conclusions We validate MixHMM and demonstrate its advantages with simulated samples, clinical tumor samples and a dilution series of mixed samples. We have shown that the CNVs of cancer cells in a tumor sample contaminated with up to 80% of stromal cells can be detected accurately using Illumina BeadChip and MixHMM. Availability The MixHMM is available as a Python package provided with some other useful tools at http://genecube.med.yale.edu:8080/MixHMM.

Airborne microorganisms have been studied for centuries, but the majority of this research has relied on cultivation-dependent surveys that may not capture all of the microbial diversity in the atmosphere. As a result, our understanding of airborne microbial ecology is limited despite the relevance of airborne microbes to human health, various ecosystem functions, and environmental quality. Cultivation-independent surveys of small-subunit rRNA genes were conducted in order to identify the types of airborne bacteria and fungi found at a single site (Boulder, CO) and the temporal variability in the microbial assemblages over an 8-day period. We found that the air samples were dominated by ascomycete fungi of the Hypocreales order and a diverse array of bacteria, including members of the proteobacterial and Cytophaga-Flavobacterium-Bacteroides groups that are commonly found in comparable culture-independent surveys of airborne bacteria. Bacterium/fungus ratios varied by 2 orders of magnitude over the sampling period, and we observed large shifts in the phylogenetic diversity of bacteria present in the air samples collected on different dates, shifts that were not likely to be related to local meteorological conditions. We observed more phylogenetic similarity between bacteria collected from geographically distant sites than between bacteria collected from the same site on different days. These results suggest that outdoor air may harbor similar types of bacteria regardless of location and that the short-term temporal variability in airborne bacterial assemblages can be very large.

The vertebrate gut harbors a vast community of bacterial mutualists, the composition of which is modulated by the host immune system. Many gastrointestinal (GI) diseases are expected to be associated with disruptions of host-bacterial interactions, but relatively few comprehensive studies have been reported. We have used the rhesus macaque model to investigate forces shaping GI bacterial communities. We used DNA bar coding and pyrosequencing to characterize 141,000 sequences of 16S rRNA genes obtained from 100 uncultured GI bacterial samples, allowing quantitative analysis of community composition in health and disease. Microbial communities of macaques were distinct from those of mice and humans in both abundance and types of taxa present. The macaque communities differed among samples from intestinal mucosa, colonic contents, and stool, paralleling studies of humans. Communities also differed among animals, over time within individual animals, and between males and females. To investigate changes associated with disease, samples of colonic contents taken at necropsy were compared between healthy animals and animals with colitis and undergoing antibiotic therapy. Communities from diseased and healthy animals also differed significantly in composition. This work provides comprehensive data and improved methods for studying the role of commensal microbiota in macaque models of GI diseases and provides a model for the large-scale screening of the human gut microbiome., Author Summary Bacterial mutualists within the gastrointestinal tract aid digestion, promote development of the gut immune system, and provide competitive barriers to pathogen invasion. The host, in return, provides bacteria with safe housing and food during lean times. The composition of the gut microbiota is controlled in part by the host immune system. In a variety of disease states, immune function can be altered, and gut morbidity is often associated, leading to the hypothesis that alterations in the GI microbiota may contribute to disease. In this study, the gut microbiota was characterized in 100 samples from rhesus macaques using pyrosequencing, which allowed 141,000 sequences from 16S rRNA genes to be generated and analyzed. Healthy animals were compared to animals with gut disorders, induced, for example by advanced simian AIDS. Many factors contributed to changes in the microbiota, including the sex of the animal of origin. Animals with chronic colitis showed differences in composition of the GI microbiota compared to healthy animals, providing an association between altered microbiota and disease.

Lung adenocarcinoma (LUAD) is a heterogeneous and deadly subtype of non-small cell lung cancer that metastasizes rapidly to distant organs. Recent transcriptional profiling of primary LUADs by TCGA identified three molecular subgroups with unique clinical and biological features. Amongst the most aggressive LUADs are tumors classified in the proximal inflammatory (PI) subtype. However, the molecular mechanisms that dictate metastatic competence of this tumor type remain undefined. Using a bioinformatic approach, we found that PI tumors are significantly enriched for the expression of extracellular matrix genes and their corresponding cell surface receptors compared to tumors of other subtypes. Among these receptors, hyaluronan mediated motility receptor (HMMR) was found to be significantly upregulated. Knockdown of HMMR in LUAD cells decreased their potential to invade, migrate, and form colonies in vitro. In an experimental mouse model of metastasis, suppression of HMMR decreased metastatic incidence and burden. Interestingly, using an orthotopic model of LUAD, suppression of HMMR attenuated tumor initiating capacity of LUAD cells within an inflammatory microenvironment without affecting tumor progression. In distant organs, HMMR is similarly essential for tumor re-initiation and expansion of micrometastases to macrometastases. Our findings suggest that the aggressive behavior of PI LUADs requires the interaction between HMMR and the tumor microenvironment. Citation Format: Laura E. Stevens, Minghui Zhao, Zongzhi Liu, Don Nguyen. A novel molecular subset of metastatic lung adenocarcinoma is defined by the function of the proteoglycan receptor HMMR. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2269. doi:10.1158/1538-7445.AM2015-2269

The focus of this paper is to utilize the acidity of hexafluoroalcohol (HFA) in addressing performance deficiencies associated with current 193nm methacrylate resist materials. In this study, we have designed and developed a variety of HFA pendant methacrylate monomers and the corresponding imaging polymers for ArF lithography. It was shown that typical swelling behavior observed in methacrylate resists can be substantially reduced or eliminated by replacing commonly used multicylcic lactone polar functionalities with acidic HFA side chains. The incorporation of aliphatic spacers between HFA and polymer backbone were found to be more effective than cyclic hindered moieties, in achieving linear dissolution characteristics. The typical poor etch stability associated with fluorine atoms in HFA can be substantially minimized by designing side chains with a combination of appropriate cyclic and aliphatic moieties and fine-tuning the corresponding polymer compositions. PEB sensitivity of high activation energy protecting group (e.g., methyladamentyl group) based methacrylate resists can be substantially improved through the incorporation of acidic HFA side chains (6nm/C to

The genomic sequence of a rice (Oryza sativa L.) glycine-rich protein (GRP) gene, designated Osgrp-2, has been previously determined (GenBank U40708). Primer extension analysis indicated that transcription starts 47 bp upstream of the translation start codon. To gain an insight into the transcriptional regulation of this gene, the 2,401-bp promoter sequence and a series of its 5' deletions were transcriptionally fused to the beta-glucuronidase (GUS) gene. GUS activity was subsequently assayed in a transient expression system of tobacco ( Nicotiana tabacum L.) protoplasts, which revealed the presence of a positive regulatory region (-2290 to -1406) and two negative regulatory regions (-2401 to -2291 and -1405 to -1022) in the Osgrp-2 promoter for the promoter activity. The positive regulatory region displayed an enhancer-like activity when fused to the cauliflower mosaic virus (CaMV) 35S minimal promoter (-89 to +6) to drive GUS expression and assayed on tobacco leaves by the Agrobacterium-mediated transient expression technique (agroinfiltration). Histochemical staining for GUS activity on transgenic tobacco plants has further indicated a preferential expression in vascular tissues of stems and leaves conferred by the positive regulatory region. A 1,023-bp fragment of the Osgrp-2 promoter (-1021 to +2) fused with GUS was transformed into tobacco and proved to be capable of conferring vascular-specific expression. Further 5' and 3' deletion analysis of the 1,023-bp promoter revealed that a 99-bp fragment located from -497 to -399 contained cis-elements responsible for vascular-specific expression.

Background: The receptor tyrosine kinase HER2 is overexpressed in 18-20% of breast cancers and is the target of the highly effective therapy, trastuzumab. Coamplicons are frequent in HER2 amplified tumors, yet their role in response to T is not defined. To identify amplicons which might predict response to T in patients we performed gene expression and copy number analysis in tumor tissue from a preoperative trial with brief exposure to T, followed by T+C. Methods: Fresh tumor core biopsies were taken before at a 2 week timepoint after a single dose of T (8mg/m2) from 80 HER2-overpressing, early breast cancer patients enrolled on a clinical trial of T>T+C. Nucleic acids were extracted using Qiagen AllPrep and were analyzed with Illumina Ref8 and Illumina QUAD 610 SNP arrays. Eight commonly amplified regions (11q13, 14q32, 17q12, 20q13, 21q22, 8p22, 8q24) were identified by FISH probe coordinates and copy number calculated using the GPHMM algorithm. Amplification was defined as 4 or more copies of the amplicon. Gene expression was analyzed in Bioconductor using LIMMA analysis. Genomic biomarkers were compared pathologic complete response (pCR) vs objective response (CR+PR) and non-response (SD+PD) by RECIST criteria at the surgical timepoint. Results: There were 50 tumor pairs with adequate RNA for gene expression and 40 baseline tumors for copy number analysis with 28 paired samples. In the 40 baseline biopsies, the 11q13 amplicon was present in 14/40 (35%) of tumors but never seen in tumors with a pCR (t-test; p=0.004975). Of note, in the tumors with amplification of 11q13 there was no evidence of change in copy number after exposure to T, however numbers were small (7 pairs). Gene expression of the 11q13 region showed no significant change in common amplicon genes after one dose of T. The 17q12 amplicon, which harbors the HER2 locus, was amplified in 35/40 (87.5%) of cases - copy number was not associated with response, however, HER2 and GRB7 gene expression levels significantly declined in pCR tumors compared with Conclusions: Integrated genomic analysis after brief exposure to T may provide useful predictors for response to T+C in early stage, HER2 positive breast cancer. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD07-08.

Somatic hypermutation and class switch recombination of immunoglobulin (Ig) genes occur in germinal center (GC) B cells and are initiated through deamination of cytidine to uracil by activation induced cytidine deaminase (AID). Resulting uracil-guanine (U-G) mismatches are processed by UNG-dependent base-excision repair (BER) and MSH2-dependent mismatch repair (MMR) pathways to yield mutations and DNA strand lesions. Although off-target AID activity also contributes to oncogenic point mutations and chromosome translocations associated with B cell lymphomas, the role of downstream AID-associated DNA repair pathways in lymphomagenesis is not defined. Through deregulated expression of BCL6, IµHABcl6 mice develop an AID-dependent GC-derived lymphoma that resembles human diffuse large B cell lymphoma (DLBCL). We have previously demonstrated that IµHABcl6 Ung-/-Msh2-/- mice have a similar incidence (35% vs 27%) but a 2.5-fold shorter median time to development of B220+ IgM+ PNAhi CD138- DLBCL compared with IµHABcl6 mice (6.5 months vs. 16.2 months; P = 0.0003). This suggests that AID-associated DNA repair pathways serve to protect the GC B cell and delay BCL6-driven lymphomagenesis. To investigate the individual contribution of BER and MMR in the pathogenesis of GC-derived lymphoma, we have now generated IµHABcl6 Ung-/- and IµHABcl6 Msh2-/- single-deficient mice. The majority of IµHABcl6 Ung-/- mice remained healthy beyond 20 months with only 3 of 22 (13.6%) mice becoming sick starting at ∼16 months. Sick mice were found to have splenic lymphomas comprised of mature B220+ IgM+ PNAlow CD138- B cells. Histological examination revealed expanded follicles with a population of small lymphocytes, consistent with a follicular B cell lymphoma which has been shown to arise in Ung-/- mice. In contrast, 18 of 22 (81.8%) IµHABcl6 Msh2-/- mice rapidly succumbed to malignancy starting at ∼3 months and had a median survival of 6 months. Of 15 tumors available for analysis, there was 1 histiocytic sarcoma, 1 squamous cell carcinoma, 4 T cell lymphomas, and 9 B220+ IgM- PNA- CD138- pre-B cell lymphomas (determined by histology, immunophenotyping and gene expression profiling). None of the IµHABcl6 Ung-/- or IµHABcl6 Msh2-/- mice developed DLBCL. Since lack of UNG is strongly protective when MSH2 is present, we conclude that in the setting of deregulated BCL6, UNG promotes the development of DLBCL. In contrast, MSH2 is protective against the development of tumors in general and does not facilitate DLBCL in the absence of UNG. Combined with the observation that IµHABcl6 Ung-/-Msh2-/- mice develop DLBCL with a significantly shorter latency than IµHABcl6 mice, this data indicates that a complex interplay between AID-associated BER and MMR produces a net protective effect against lymphomagenesis. In the absence of UNG and MSH2, AID-generated U-G mismatches are not processed into strand lesions and are simply replicated, yielding C/G to T/A transition mutations. Thus, to assess how combined lack of UNG and MSH2 might promote the accelerated development of BCL6-driven lymphoma, we carried out spectral karyotyping and sequence analysis of AID target genes (IgJH4, cMyc, Pim1, RhoH, Cd79a, CD79b, H2afx, Pax5, and Cd83) in lymphomas from the different genotypes. IµHABcl6 DLBCLs (3/3) harbored various complex chromosome abnormalities, consistent with previous findings. Numerous clonal and sub-clonal chromosome abnormalities including translocations, duplications, deletions, and aneuploidies were also detected in IµHABcl6 Ung-/-Msh2-/- (4/4) and IµHABcl6 Ung-/- (2/2) lymphomas. Pre-B cell tumors from IµHABcl6 Msh2-/- mice could not be stimulated to produce metaphase chromosomes. Clonal and non-clonal mutations of the IgJH4 intronic region were identified in lymphomas from IµHABcl6 (2/3), IµHABcl6 Ung-/-Msh2-/- (4/4), and IµHABcl6 Ung-/- (2/3) mice, consistent with ongoing AID activity. No mutations were detected in 3 pre-B cell lymphomas, consistent with their pre-GC origin. Six clonal mutations within AID hotspots (all C/G to T/A) were identified in Pim1, RhoH, and Pax5 in 2 of 4 IµHABcl6 Ung-/-Msh2-/- DLBCLs. None of the other genotypes carried any clonal mutations of non-Ig genes. Thus, chromosome abnormalities in GC B cell lymphomas can arise through mechanisms independent of BER and MMR but may be due to off-target effects of AID on other genes that regulate cell cycle, apoptosis, or genomic stability. Disclosures: No relevant conflicts of interest to declare.

Background: Receptor tyrosine kinases (RTKs) modulate many oncogenic signaling pathways. RTKs can be activated as a result of feedback loops initiated from inhibition of downstream signaling components and contribute to resistance to small molecule targeted therapies. Target-specific inhibitors have provided many advances in cancer treatment, but it is becoming increasingly evident that single-agent treatment approaches are suboptimal. To address this, we performed single-agent compound screens against a panel of melanoma cell lines to select candidate agents for testing in combination agent screens (Cancer Discov 2013 3:52-67). One of the top single-agents effective in mutant BRAF melanomas was dovitinib (TKI258, CHIR258), a multi-RTK inhibitor shown to inhibit numerous type III, IV, and V RTKs. We present data examining the target profile of dovitinib in short-term cultured melanoma cell lines and describe effective partner agents for further testing as combination therapies. Materials and Methods: Twenty-five melanoma cell lines were screened against 150 agents. The most efficacious forty agents from the single-agent screen were collated and three consensus concentrations were determined for each compound to perform pair-wise combinations (Cancer Discov 2013 3:52-67). Growth inhibition was evaluated by ATP luminescent detection by CellTiterGlo®. Colony formation experiments were performed to confirm positive drug interactions. Phosphoproteomic approaches were used to further evaluate targets of dovitinib. Results: Dovitinib was one of the top hits from this single-agent screen that was effective in mutant BRAF melanoma cell lines. Dovitinib does not inhibit mutant BRAF directly, but inhibits RTKs. We found that multiple RTK targets of dovitinib are expressed and active in mutant BRAF melanoma cell lines. Furthermore, we found that dovitinib combines well with vemurafenib and MEK and AKT inhibitors to inhibit colony formation ability and cell proliferation. This was seen in both vemurafenib-sensitive and resistant cell lines. Conclusions: These findings further demonstrate the utility of using both single- and combination agent screening to determine genotype specific compound effects. Furthermore, these results support the rationale for using dovitinib as a partner agent with several small molecule inhibitors for melanoma combination therapy, especially in the vemurafenib-resistant setting. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B275. Citation Format: Casey G. Langdon, Matthew A. Held, Andrew B. Koo, Michael Klein, Zongzhi Liu, James T. Platt, David F. Stern. Dovitinib as a single-agent or in combination with other small molecule inhibitors for mutant BRAF melanomas. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B275.

Lung cancer is the leading cause of cancer-related deaths worldwide. This poor prognosis is due to the rapid metastatic progression of this disease, yet the mechanisms that govern lung cancer dissemination and colonization are unclear. Through an integrated approach, we identified an alveolar cell-selective gene signature that stratifies lung adenocarcinoma (ADC) into molecular subtypes of distinct prognosis and differentiation states. This transcriptional program is regulated in part by the cell fate transcription factors, GATA6 and HOPX, which are normally required for proper alveolar differentiation. In an experimental model, suppression of GATA6 and HOPX cooperatively enhances metastatic competence of lung ADC cells. Whole genome RNA sequencing reveals that this transcriptional network can modulate specific target genes and pathways involved in airway epithelial differentiation and invasion. Our findings identify a novel cell lineage molecular program whose perturbation enhances lung cancer metastasis. Citation Format: William K.C. Cheung, Minghui Zhao, Zongzhi Liu, Paul D. Cao, Laura E. Stevens, Thomas F. Westbrook, Don X. Nguyen. A lineage-selective transcriptional pathway linking lung differentiation and cancer metastasis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3872. doi:10.1158/1538-7445.AM2013-3872

Lung cancer is the leading cause of cancer-related deaths worldwide. This poor prognosis is due to the rapid metastatic progression of this disease, yet the mechanisms that govern lung cancer dissemination and colonization are unclear. Through an integrated approach, we identified an alveolar cell-selective gene signature that stratifies lung adenocarcinoma (ADC) into molecular subtypes of distinct prognosis and differentiation states. This transcriptional program is regulated in part by the cell fate transcription factors, GATA6 and HOPX, which are normally required for proper alveolar differentiation. In an experimental model, suppression of GATA6 and HOPX cooperatively enhances metastatic competence of lung ADC cells. Whole genome RNA sequencing reveals that this transcriptional network can modulate specific target genes and pathways involved in airway epithelial differentiation and invasion. Our findings identify a novel cell lineage molecular program whose perturbation enhances lung cancer metastasis. Citation Format: William K.C. Cheung, Minghui Zhao, Zongzhi Liu, Paul D. Cao, Laura E. Stevens, Thomas F. Westbrook, Don X. Nguyen. A lineage-selective transcriptional pathway linking lung differentiation and cancer metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr B56.

Metastasis and drug resistance are two major challenges in cancer therapies. The occurrence of these two events requires cancer cells to gain new mutations or change their expression profiling, or both, adapt to new environment (i.e. distal organs or drug treatments). These changes likely follow Darwin's evolutionary theory: heterogeneous of cancer cells are generated randomly and more aggressive cells are selected by the tissue environment. Stable inheritance of genome and epigenetic states could delay metastasis and drug resistance. Trimethylated lysine 4 in histone H3 (H3K4me3) is the epigenetic marker for transcriptionally active promoters and the binding site for transcriptional activators. Removing the methyl groups from trimethylated H3K4 on gene promoter regions is a common epigenetic strategy for gene silencing. The JARID1 family members are to date the only known proteins responsible for catalyzing demethylation of trimethylated H3K4. Therefore they are essential in cell differentiation and development. Even though JARID1A (RBP2) was implicated in tumorigenesis of certain cancer types, its exact roles are still poorly understood. I hypothesized that RBP2 contributes to changes of epigenetic states during the occurrence of metastasis and drug resistance. Consistent with my hypothesis, I found that RBP2 is highly expressed in metastatic breast cancer cells and Herceptin tolerant population. Loss of RBP2 decreased lung metastatic gene signature and increase drug sensitivity. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-389. doi:1538-7445.AM2012-LB-389

The recent introduction of massively parallel pyrosequencers allows rapid, inexpensive analysis of microbial community composition using 16S ribosomal RNA (rRNA) sequences. However, a major challenge is to design a workflow so that taxonomic information can be accurately and rapidly assigned to each read, so that the composition of each community can be linked back to likely ecological roles played by members of each species, genus, family or phylum. Here, we use three large 16S rRNA datasets to test whether taxonomic information based on the full-length sequences can be recaptured by short reads that simulate the pyrosequencer outputs. We find that different taxonomic assignment methods vary radically in their ability to recapture the taxonomic information in full-length 16S rRNA sequences: most methods are sensitive to the region of the 16S rRNA gene that is targeted for sequencing, but many combinations of methods and rRNA regions produce consistent and accurate results. To process large datasets of partial 16S rRNA sequences obtained from surveys of various microbial communities, including those from human body habitats, we recommend the use of Greengenes or RDP classifier with fragments of at least 250 bases, starting from one of the primers R357, R534, R798, F343 or F517.

Pyrosequencing technology allows us to characterize microbial communities using 16S ribosomal RNA (rRNA) sequences orders of magnitude faster and more cheaply than has previously been possible. However, results from different studies using pyrosequencing and traditional sequencing are often difficult to compare, because amplicons covering different regions of the rRNA might yield different conclusions. We used sequences from over 200 globally dispersed environments to test whether studies that used similar primers clustered together mistakenly, without regard to environment. We then tested whether primer choice affects sequence-based community analyses using UniFrac, our recently-developed method for comparing microbial communities. We performed three tests of primer effects. We tested whether different simulated amplicons generated the same UniFrac clustering results as near-full-length sequences for three recent large-scale studies of microbial communities in the mouse and human gut, and the Guerrero Negro microbial mat. We then repeated this analysis for short sequences (100-, 150-, 200- and 250-base reads) resembling those produced by pyrosequencing. The results show that sequencing effort is best focused on gathering more short sequences rather than fewer longer ones, provided that the primers are chosen wisely, and that community comparison methods such as UniFrac are surprisingly robust to variation in the region sequenced. © 2007 The Author(s).

Glycine-rich proteins (GRPs) belong to a kind of important structural proteins of plant cell walls. The expression of GRP genes is regulated spatially and developmentally as well as by various environmental stresses, thus providing a good model for the study of plant gene expression. We obtained the genomic sequence of a new GRP gene (Osgrp- 2) from a rice genomic library. The transcription start site of Osgrp- 2 was determined by 5'-rapid amplification of cDNA ends (RACE) and a 2.4-kb promoter sequence was thus delimited. The spatial and developmental expression pattern as well as the wound-inducible character of Osgrp- 2 in rice plants was analyzed in detail. Furthermore, the gene was mapped onto rice chromosome 10 by analysis of restriction fragment length polymorphism (RFLP).