Search

NGS-Cancer Panel Gene list

Gene mutation summary.

Summary of copy number data.

Mutation summary of individual tissue samples (Mutations with a frequency {greater than or equal to}1%)

Sample ID and Tissue Information.

Fluorescence technology has many useful applications for both research and teaching, among them the detection of fluorescence in live transgenic organisms and of DNA in agarose gels. However, dedicated fluorescence imaging systems can be expensive and complex. We describe a simple apparatus for non-microscopic fluorescence imaging using affordable and readily available parts. We describe three activities of increasing complexity that utilize fluorescence illumination and teach principles of fluorescence. At the high school level, our more advanced activities can be used for lessons addressing NGSS performance expectations in HS-LS3 and HS-LS4.

Recent FDA approvals of immunotherapy for NSCLC provide patients new treatment options, and these approvals also highlight the importance of the immune response in cancer treatment. While immunotherapy provides patients a new treatment option, the therapy is effective in less than half of the treated patients. To attain greater insight into the tumor-immune microenvironment, NSCLC tumors were analyzed by IHC and RNA-seq. IHC was used to identify NSCLC tumors that contain low, moderate, or high levels of CD8+ positive cells as a manifestation of an active anti-tumor immune response. Gene expression analysis identified an emergent gene signature that is associated with high and moderate levels of CD8 in NSCLC. In addition, the NSCLC tumors also express a unique combination of genes that may indicate complex anti-tumor immune responses (INFG-related genes, STATs, CXCL9, OX40, PD-L1, PD-L2, IDO1, and CD47). Several NSCLC tumors also express the immune checkpoint PD-L1 and at least one additional immune inhibitory molecule (IDO1, PD-L2, or others), which may explain the lack of a therapeutic response to treatments that disrupt only one immune checkpoint pathway.

Inflammatory breast cancer (IBC) is a rare and aggressive form of breast cancer that remains poorly understood at the molecular level. Comprehensive tumor profiling was performed to understand clinically actionable alterations in IBC. Targeted next-generation sequencing (NGS) and IHC were performed to identify activated pathways in IBC tumor tissues. siRNA studies examined the impact of IBC genomic variants in cellular models. IBC tumor tissues were further characterized for immune infiltration and immune checkpoint expression by IHC. Genomic analysis identified recurrent alterations in core biologic pathways, including activating and targetable variants in HER/PI3K/mTOR signaling. High rates of activating HER3 point mutations were discovered in IBC tumors. Cell line studies confirmed a role for mutant HER3 in IBC cell proliferation. Immunologic analysis revealed a subset of IBC tumors associated with high CD8+/PD-L1+ lymphocyte infiltration. Immune infiltration positively correlated with an NGS-based estimate of neoantigen exposure derived from the somatic mutation rate and mutant allele frequency, iScore. Additionally, DNA mismatch repair alterations, which may contribute to higher iScores, occurred at greater frequency in tumors with higher immune infiltration. Our study identifies genomic alterations that mechanistically contribute to oncogenic signaling in IBC and provides a genetic basis for the selection of clinically relevant targeted and combination therapeutic strategies. Furthermore, an NGS-based estimate of neoantigen exposure developed in this study (iScore) may be a useful biomarker to predict immune infiltration in IBC and other cancers. The iScore may be associated with greater levels of response to immunotherapies, such as PD-L1/PD-1–targeted therapies. Mol Cancer Ther; 15(7); 1746–56. ©2016 AACR.

In their technical comment, Janz et al. take issue with our recent study examining the association between host breadth and diversification rates in the brush-footed butterflies (Lepidoptera: Nymphalidae) (Hamm and Fordyce 2015). Specifically, they are concerned that we misrepresent their "oscillation hypothesis" (OH) (Janz et al. 2006; Janz and Nylin 2008) and that one of our models was inadequate to test hypotheses regarding host breadth and diversification rate. Given our mutual interests in the macroevolutionary patterns of herbivorous insects, we appreciate the opportunity to respond to their concerns.

Many areas of research in ecology and evolutionary biology depend on the quantification of dietary niche width. For herbivorous insects, diet breadth has most often been measured as simply the number and type of host plant taxa attacked. We propose an index of host range (which we refer to as "ordinated diet breadth") based on observed associations between plants and insects, and the calculation of multivariate distances among plants in ordination space. Similarities and distances are calculated based on host association and, in this context, potentially encompass multiple properties of plants, including phytochemistry, phenology, and other plant traits. This approach can distinguish between herbivores that utilize suites of hosts that are commonly used together and herbivores that attack unusual host combinations, and thus have a relatively broad diet breadth. For illustration, we use a data set of nymphalid butterfly host records, and compare taxonomic and ordinated host range. For a large number of butterfly taxa, we find that host use is clustered in multivariate space with respect to associations observed across all of the butterfly taxa. Applications are discussed, including a hypothesis test of nonrandom host association, and prediction of shifts and expansions of diet breadth.

Herbivorous insects represent one of the most successful animal radiations known. They occupy a wide range of niches, feed on a great variety of plants, and are species rich; yet the factors that influence their diversification are poorly understood. Host breadth is often cited as a major factor influencing diversification, and, according to the Oscillation Hypothesis, shifts from generalist to specialist feeding states increase the diversification rate for a clade. We explored the relationship between host breadth and diversification within the Nymphalidae (Lepidoptera) and explicitly tested predictions of the Oscillation Hypothesis. We found strong evidence of diversification rate heterogeneity, but no difference in host breadth between clades with a higher diversification rate compared to their sisters. We also found some clades exhibited phylogenetic nonindependence in host breadth and these clades had lower host plant turnover than expected by chance, suggesting host breadth is evolutionarily constrained. Finally, we found that transitions among host breadth categories varied, but the likelihood of reductions in host breadth was greater than that of increases. Our results indicate host breadth is decoupled from diversification rate within the Nymphalidae, and that constraints on diet breadth might play an important role in the evolution of herbivorous insects.

The Mitchell's satyr butterfly, Neonympha mitchellii mitchellii French 1889 is a federally-listed endangered species found in parts of the eastern United States of America. Because of its endangered status, considerable research efforts have been devoted to understanding its biology, ecology, and its conservation. Despite these efforts, information about N. m. mitchellii has not been summarized for more than a decade. Here we summarize and expand upon the work conducted by governmental and not-for-profit agencies that have produced reports that are not easily accessible to researchers or interested lepidopterists. In addition to summarizing the literature, we present data from feeding trials and also demonstrate that microclimates exist that may be exploited by larvae. We conclude by identifying key areas of needed research and describe steps imperative to the recovery the Mitchell's satyr.

Estimates of animal abundance are essential to conservation biology and are sorely lacking for many endangered species in the United States of America. This lack of knowledge may disproportionately affect butterflies in the USA, which form the largest group of federally protected insects (20 of 62 species). The Mitchell's satyr butterfly, Neonympha mitchellii mitchellii, is a federally endangered species found at 18 highly isolated sites in the Eastern USA. Currently, surveys are conducted by meandering through habitat and recording the number of butterflies observed. These surveys are efficient in terms of staffing and time, but the data from these surveys cannot be used to estimate abundance. Mark release recapture surveys generate estimates of demographic parameters and have been conducted, albeit infrequently, and require high staffing levels and weeks of fieldwork to generate estimates with reasonable error. I employed hierarchical distance sampling along line transects to estimate N. m. mitchellii abundance at one site in lower Michigan, USA. This method requires one observer to traverse a series of transects at a walking pace and record the number of butterflies observed and their perpendicular distance to the transect line. My results suggest that this method is as cost efficient as meander surveys, but generates reasonable estimates of butterfly abundance.

Atypical teratoid rhabdoid tumors (ATRTs) are rare, highly malignant central nervous system tumors that occur during infancy and early childhood. Their poor outcome and resistance to conventional chemotherapies and radiotherapy, urges the development of new therapies. Recent studies have evaluated the effects of histone deacetylase inhibitors (HDACi) as a new potential treatment for ATRTs. However, most HDACi act unselectively against all, or at least several, histone deacetylase (HDAC) family members. We hypothesized that specific HDAC family members are deregulated in ATRT and therefore a more selective class of HDACi would be beneficial to patients with ATRT.To test our hypothesis, we evaluated the expression level of different HDAC family members in ATRTs. Eight ATRTs were compared to six medulloblastoma samples in regards to the level of expression of the 18 HDAC family members as determined by microarray gene expression profiling.HDAC1 was the only member of the HDAC family to be significantly differentially expressed in ATRTs (FC = 4.728; p value = 0.00003).A class of HDACi specifically targeting HDAC1 may allow for the desired therapeutic benefits with fewer side effects for children with ATRT.

Climbing plants provide efficient pathways for ants to access patchy arboreal resources. However, plant stems vary greatly in physical characteristics that are likely to influence ant locomotion. We collected, measured and identified ants foraging on 671 stems of climbing plants at the La Selva Biological Station, Costa Rica. We applied tuna baits to 70% of the observed stems to attract ants to a broad range of stem sizes. We used these data to examine relationships between relative stem roughness, growth form (herbaceous or woody), stem diameter and the body length of foraging ants representing 58 species. The size of the largest ants found on stems generally increased with stem size up to 3.2 mm diameter, whereas the size of the smallest ants present on stems did not vary with stem diameter. The largest ants in the forest (Paraponera clavata) used small stems (

The future of drug design and the development of new therapeutics will rely on our ability to unravel the complexities of the epigenome in normal and disease states. Proper epigenetic regulation is essential for normal differentiation in embryogenesis and development. Conversely, abnormal epigenetic regulation is a feature of complex diseases, including cancer, diabetes, heart disease and other pathologies. Epigenetic therapies hold promise for a wide range of biological applications, from cancer treatment to the establishment of induced pluripotent stem cells. The creation of more specific and effective epigenetic therapies, however, requires a more complete understanding of epigenomic landscapes. Here, we give a historical overview of the epigenomics field and how epigenetic modifications can affect embryo development and disease etiology. We also discuss the impact of current and future epigenetic drugs.

The ant Tapinoma sessile (Say) 1836 is one of the most widely distributed ants in North America; yet, it has received very little attention from biologists apart from its ability to infest houses. The original species description does not adequately account for the phenotypic variation present in this species of typically darkly concolored ant. In California, a bicolored morph was discovered that superficially resembles T. sessile; hence, its species status was questioned. Comparative morphometric analysis indicates that the bicolored ants are consistently distinguishable across a range of characters, and allometric techniques conclusively demonstrate that these two morphs are divergent. Based on the analysis of morphological data it was concluded that the bicolored ant is not T. sessile and is described as a new species, Tapinoma schreiberi. A neotype of T. sessile also is designated in this article, and measurement data are presented to assist taxonomists and delimit the morphological boundaries of T. sessile.

Genetic sampling of endangered species can inform conservation management and potentially aid the long-term survival of a species. However, when dealing with very small populations of rare species, the sacrifice of whole animals may not be desirable or permitted. We set out to develop a demonstrably non-lethal method of obtaining DNA from the federally-endangered Mitchell’s satyr butterfly, Neonympha mitchellii mitchellii. Because of its endangered status we developed our methods on related species. In greenhouse and fields trials, we demonstrate that removal of small amounts of hind wing (2–3 mm2) has no significant impact on the behavior or survival of Vanessa cardui and Satyrodes eurydice. Based on these studies we were successful in obtaining a permit from the US Fish and Wildlife Service to sample DNA from N. m. mitchellii populations. We suggest that our results can be extended to the sampling of other rare butterfly species.

In their technical comment, Janz et al. take issue with our recent study examining the association between host breadth and diversification rates in the brush-footed butterflies (Lepidoptera: Nymphalidae) (Hamm and Fordyce 2015). Specifically, they are concerned that we misrepresent their "oscillation hypothesis" (OH) (Janz et al. 2006; Janz and Nylin 2008) and that one of our models was inadequate to test hypotheses regarding host breadth and diversification rate. Given our mutual interests in the macroevolutionary patterns of herbivorous insects, we appreciate the opportunity to respond to their concerns.

Members of the plant genus Selaginella (de Beauvois 1805) have few known insect herbivores even though they are considered by some to be ‘living fossils’, with extant taxa virtually indistinguishable from 300 Mya fossils. Butterflies are well-known herbivores, and the satyrs are among the most speciose of them despite having radiated ∼35 Mya ago. Nearly all satyrs feed on grass or sedges, but members of the Neotropical genus Euptychia Hubner 1818 feed on Selaginella ; little is known about the degree to which this butterfly favors this ancient plant over those that its close relatives utilize. To advance our knowledge of Euptychia natural history, we conducted a series of experiments to examine oviposition preference and growth rates across a series of potential host plants on a Euptychia westwoodi population in Costa Rica. We found that Euptychia westwoodi Butler 1867 exhibit a strong preference to oviposit on Selaginella eurynota over the sympatric Selaginella arthritica , though they perform equally well as larvae on both plants. We did not observe oviposition on a sympatric grass that is commonly consumed by close relatives of E. westwoodi , and when larvae were offered the grass they refused to eat. These results suggest that E. westwoodi in Costa Rica exhibit a strong preference for Selaginella and may have lost the ability to feed on a locally abundant grass commonly used by other Satyrinae.

Lantana camaraL. (Verbenaceae) is a shrub of Neotropical origin that has spread to at least 60 countries (Dayet al. 2003), and its ability to rapidly occupy disturbed habitat has led to it being named one of the ten worst weeds on the planet (Cronk & Fuller 1995, Sharmaet al. 2005). A number of traits may have contributed to its success as an invasive species. For example,L. camaraincreases the available soil nitrogen in otherwise nitrogen-poor ecosystems (Sharma & Raghubanshi 2009), it is extremely resilient to disturbance (Gentle & Guggin 1997a, Hiremath & Sundram 2005), it is allelopathic to native plants (Achhireddy & Singh 1984, Gentle & Duggin 1997b) and its fruits are dispersed by a variety of birds (Mandon-Dalgeret al. 2004, Swarbricket al. 1998). A potentially unappreciated aspect ofL. camarabiology that may also contribute to its invasiveness relates to pollination ecology.

Members of the plant genus Selaginella are often considered living fossils, as extant taxa are virtually indistinguishable from 300 Ma fossils. In contrast, the brush-footed butterflies are a relatively recent radiation, having diversified primarily within the last 60 Ma. Satyrs are among the most derived of these butterflies, likely radiated ~35 Ma and are known primarily for their high diversity and propensity to feed on grasses and sedges. In contrast to its close relatives, the Neotropical satyr genus Euptychia also feeds on Selaginella, which is thought to be nutrient poor. Using no choice feeding experiments, I compared growth rates Costa Rican E. westwoodi that were offered two species of Selaginella, to those that were offered Lasiascis rusifolia, a grass commonly fed upon by close relatives. I discovered E. westwoodi larvae fed on two species of Selaginella and there was no difference in mass gained between the two species. However, larvae refused to feed on L. ruscifolia and lost mass over the course of the trial and expired unless they were transferred to Selaginella. To the best of my knowledge, these are the first data to report larval performance of the butterfly genus Euptychia or any Selaginella feeding insect. . Though far from conclusive, these results support the proposition that Euptychia have lost the ability to feed on other host plants and are now specialized on grasses.

Background: Inflammatory breast cancer (IBC) is the most aggressive and lethal form of primary breast cancer. Inflammatory signaling pathways are active in IBC, but the function of the immune response remains elusive. ErbB receptors play a role in IBC. ERBB2 is amplified in 50% of IBCs, and ERBB3 is also mutated in IBC. Currently, Lapatinib, a dual ErbB inhibitor, is used in IBC patients with ERBB2 amplification. Although immune cell inflammatory signaling may promote IBC tumor growth and metastasis, the presence of cytotoxic tumor-associated lymphocytes has also been associated with a more favorable breast cancer prognosis. The contradictory nature of the immune data highlights the need to elucidate the relationship between the immune response and subsequent treatments. Given the active IBC immune component and the recent clinical benefit of immune checkpoint inhibitors in cancer treatment, IBC’s may represent a clinically unique breast cancer population that may benefit from immunomodulating agents. Interestingly, Myc is a downstream effector of ErbB signaling, and Myc is thought to regulate the expression of immune checkpoint proteins CD47 and PD-L1. The presence of PD-L1-positive IBC immune infiltrate suggests that IBC’s may benefit from therapies that disrupt PD-L1 signaling together with ErbB inhibitors. METHODS: IHC was used to examine immune checkpoint signaling and characterize the tumor-immune infiltrate. Tumor tissues were also characterized using an RNA-seq panel that examined the expression of 377 immune-related genes. Cell lines (BT474, SKR3, AU565, and SUM225) were treated with Lapatinib and Neratinib to examine the relationship between growth factor receptors and downstream immune signaling pathways. RESULTS: RNA-seq revealed the expression of specific immunosuppressive signaling pathways in tumors. Treatment of breast cancer cells with ErbB inhibitors resulted in a decrease in the levels of PD-L1. Treatment with Lapatinib and Neratinib diminished PD-L1 in all cell lines. Levels of Phospho-Stat3 decreased in BT474 and Sum225 but not in SKBR3 and AU565, implying that PD-L1 is regulated by another mechanism (ERK-MYC). CONCLUSION: Our results provide mechanistic insight into ErbB receptor activation and the expression of downstream signaling molecules (Stat3, Myc, PD-L1, and others). In addition, our unique RNA-seq immune signature reveals the expression of several genes that may serve as biomarkers of inhibitory immune signaling pathways. Our laboratory is currently examining the correlation between PD-L1 expression and the activation of ErbB2 in model systems and clinical trials using ErbB inhibitors in combination with PD-L1 inhibitors. Our immune panel gene signature may serve as a useful diagnostic test that, in conjunction with traditional ErbB testing, can identify patients that will benefit from combination therapy of an ErbB inhibitor and an immune checkpoint inhibitor. Note: This abstract was not presented at the meeting. Citation Format: Christopher A. Hamm, Sumin Zhao, Cesar A. Santa-Maria, Massimo Cristofanilli, Neil L. Spector, Sarah Bacus. Targeted therapies to ERBB receptors downregulate expression of PD-L1: implications in combination therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2636. doi:10.1158/1538-7445.AM2017-2636

© 2017 Bell et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Temporal isolation remains an understudied, and potentially under-appreciated, mechanism of reproductive isolation. Phenological differences have been discovered in populations of the pine white butterfly (Neophasia menapia), a typically univoltine species found throughout western North America. At two locations in the Coast Range of California there are two periods of adult emergence per year, one in early summer (July) and one in late summer/autumn (September/October). Differences in flight time are accompanied by differences in wing shape and pigmentation. Here we use a combination of population genomics and morphological analyses to assess the extent to which temporal isolation is able to limit gene flow between sympatric early and late flights. Not only did we detect both genetic and morphological differences between early and late flights at the two sites, we also found that the patterns of differentiation between the two flights were different at each location, suggesting an independent origin for the two sympatric flights. Additionally, we found no evidence that these sympatric flights originated via colonization from any of the other sampled localities. We discuss several potential hypotheses about the origin of these temporally isolated sympatric flights.

Herbivorous insects represent one of the most successful animal radiations known. They occupy a wide range of niches, feed on a great variety of plants, and are species rich; yet the factors that influence their diversification are poorly understood. Host breadth is often cited as a major factor influencing diversification, and, according to the Oscillation Hypothesis, shifts from generalist to specialist feeding states increase the diversification rate for a clade. We explored the relationship between host breadth and diversification within the Nymphalidae (Lepidoptera) and explicitly tested predictions of the Oscillation Hypothesis. We found strong evidence of diversification rate heterogeneity, but no difference in host breadth between clades with a higher diversification rate compared to their sisters. We also found some clades exhibited phylogenetic nonindependence in host breadth and these clades had lower host plant turnover than expected by chance, suggesting host breadth is evolutionarily constrained. Finally, we found that transitions among host breadth categories varied, but the likelihood of reductions in host breadth was greater than that of increases. Our results indicate host breadth is decoupled from diversification rate within the Nymphalidae, and that constraints on diet breadth might play an important role in the evolution of herbivorous insects.

Temperature-related studies were conducted on Drosophila suzukii Matsumura (Diptera: Drosophilidae: Drosophilini). From 10-28°C, temperature had a significant impact on blueberries, Vaccinium corymbosum L. (Ericales: Ericaceae), and cherries, Prunus avium (L.) L. 1755 (Rosales: Rosaceae), important commercial hosts of D. suzukii. Temperature had a significant influence on D. suzukii developmental period, survival, and fecundity, with decreasing developmental periods as temperatures increased to 28°C. At 30°C, the highest temperature tested, development periods increased, indicating that above this temperature the developmental extremes for the species were approached. D. suzukii reared on blueberries had lower fecundity than reared on cherries at all temperatures where reproduction occurred. The highest net reproductive rate (R(o)) and intrinsic rate of population increase (r(m)) were recorded on cherries at 22°C and was 195.1 and 0.22, respectively. Estimations using linear and nonlinear fit for the minimum, optimal, and maximum temperatures where development can take place were respectively, 7.2, 28.1, and 42.1°C. The r(m) values were minimal, optimal, and maximal at 13.4, 21.0, and 29.3°C, respectively. Our laboratory cultures of D. suzukii displayed high rates of infection for Wolbachia spp. (Rickettsiales: Rickettsiaceae), and this infection may have impacted fecundity found in this study. A temperature-dependent matrix population estimation model using fecundity and survival data were run to determine whether these data could predict D. suzukii pressure based on environmental conditions. The model was applied to compare the 2011 and 2012 crop seasons in an important cherry production region. Population estimates using the model explained different risk levels during the key cherry harvest period between these seasons.

Storage proteins are deposited into protein storage vacuoles (PSVs) during plant seed development and maturation and stably accumulate to high levels; subsequently, during germination the storage proteins are rapidly degraded to provide nutrients for use by the embryo. Here, we show that a PSV has within it a membrane-bound compartment containing crystals of phytic acid and proteins that are characteristic of a lytic vacuole. This compound organization, a vacuole within a vacuole whereby storage functions are separated from lytic functions, has not been described previously for organelles within the secretory pathway of eukaryotic cells. The partitioning of storage and lytic functions within the same vacuole may reflect the need to keep the functions separate during seed development and maturation and yet provide a ready source of digestive enzymes to initiate degradative processes early in germination.

The modern delineation of taxonomic groups is often aided by analyses of molecular data, which can also help inform conservation biology. Two subspecies of the butterfly Neonympha mitchellii are classified as federally endangered in the United States: Neonympha mitchellii mitchellii, the Mitchell’s satyr, and Neonympha mitchellii francisi, the Saint Francis’s satyr. The recent discovery of additional disjunct populations of N. mitchellii in the southeastern US could have important implications for both legal protection and management decisions. We elucidated the relationships among 48 individuals representing 5 N. mitchellii populations using 6 molecular markers (5 nuclear and 1 mitochondrial) under a variety of analytical frameworks. Phylogenetic analysis resulted in moderately supported clades that corresponded with the geographic region where samples originated. Clustering analyses identified 3 groups, wherein the 2 named subspecies formed separate clusters. Coalescent analyses indicated evolutionary divergence between N. m. mitchellii and all other populations but weakly supported divergence among N. m. francisi and the recently discovered populations. Hence, the 2 currently accepted subspecies were clearly different from one another, but the recently discovered populations could not be completely distinguished from N. m. francisi or each other. We propose that N. m. mitchellii and N. m. francisi continue to be managed as separate endangered species.

Drosophila suzukii Matsumura (spotted wing drosophila) has recently become a serious pest of a wide variety of fruit crops in the United States as well as in Europe, leading to substantial yearly crop losses. To enable basic and applied research of this important pest, we sequenced the D. suzukii genome to obtain a high-quality reference sequence. Here, we discuss the basic properties of the genome and transcriptome and describe patterns of genome evolution in D. suzukii and its close relatives. Our analyses and genome annotations are presented in a web portal, SpottedWingFlyBase, to facilitate public access.

Malignant rhabdoid tumors (MRT) can occur in a variety of anatomical sites. The most frequent locations are the brain, where they are named atypical teratoid/rhabdoid tumors (AT/RT), and the kidney, where they are named rhabdoid tumors of the kidney (RTK). MRTs at all sites are recognized as the same entity due to their similar morphology, aggressive behavior, and a common genetic abnormality, an inactivating mutation of the SMARCB1/INI-1/hSNF5/BAF47 gene. We aim to investigate potential molecular differences between AT/RT and RTK.We analyzed the microRNA (miRNA) and gene expression (GE) profiles of 10 RTK, 13 AT/RT, and 2 human MRT cell lines (G401-RTK and MON-AT/RT). Illumina V2 MicroRNA Chips (Illumina, Inc., CA, USA) were used for miRNA analysis, and Illumina HT-12 whole genome expression arrays were used for GE analysis.The distribution of p values from GE showed a significant difference between RTK and AT/RT, with 20 % of the genes having p values ≤0.05 and the principal component analysis of the GE data showed separation between RTK and AT/RT. However, the miRNA expression failed to identify the different tumor groups. Among the 122 genes significantly differentially expressed between AT/RT and RTK, we found both genes related to brain development (i.e., FABP7, 22-fold increase in AT/RT) and genes related to kidney development (i.e., TCF21, sixfold increase in RTK).Based on our results, we hypothesized that although MRT are indeed the same tumor, independent of the site of origin, the GE differences reflect the influence of microenvironment over tumor development.

Background Chondrosarcomas are malignant cartilage tumors that do not respond to traditional chemotherapy or radiation. The 5-year survival rate of histologic grade III chondrosarcoma is less than 30%. An animal model of chondrosarcoma has been established - namely, the Swarm Rat Chondrosarcoma (SRC) - and shown to resemble the human disease. Previous studies with this model revealed that tumor microenvironment could significantly influence chondrosarcoma malignancy. Methods To examine the effect of the microenvironment, SRC tumors were initiated at different transplantation sites. Pyrosequencing assays were utilized to assess the DNA methylation of the tumors, and SAGE libraries were constructed and sequenced to determine the gene expression profiles of the tumors. Based on the gene expression analysis, subsequent functional assays were designed to determine the relevancy of the specific genes in the development and progression of the SRC. Results The site of transplantation had a significant impact on the epigenetic and gene expression profiles of SRC tumors. Our analyses revealed that SRC tumors were hypomethylated compared to control tissue, and that tumors at each transplantation site had a unique expression profile. Subsequent functional analysis of differentially expressed genes, albeit preliminary, provided some insight into the role that thymosin-β4, c-fos, and CTGF may play in chondrosarcoma development and progression. Conclusion This report describes the first global molecular characterization of the SRC model, and it demonstrates that the tumor microenvironment can induce epigenetic alterations and changes in gene expression in the SRC tumors. We documented changes in gene expression that accompany changes in tumor phenotype, and these gene expression changes provide insight into the pathways that may play a role in the development and progression of chondrosarcoma. Furthermore, specific functional analysis indicates that thymosin-β4 may have a role in chondrosarcoma metastasis.

The aim of this study is to search for new therapeutic targets for atypical teratoid-rhabdoid tumors (ATRT).To achieve this, we compared the expression of 365 microRNAs among ATRT, medulloblastomas, and normal brain.MiR-221 and miR-222 were within the top differentially expressed microRNAs. The deregulated expression of miR221/222 was demonstrated to inhibit the expression of the tumor suppressor and inhibitor of cell cycle p27(Kip1). Here, we demonstrated the negative regulation of p27(Kip1) by miR-221/222 in ATRT using microarray, real-time reverse transcriptase polymerase chain reaction, and immunohistochemistry.As anti-miR therapy was recently proposed as an alternative treatment for cancer, these findings suggest that anti-miR-221/222 therapy might have therapeutic potential in ATRT.

Background Inflammatory breast Cancer (IBC) is a rare aggressive breast cancer in which cancer cells block the lymph vessels in the skin of the breast. IBC tumors have a high rate of ERBB2 positivity and response to ERBB2 targeted therapies. Due to the rarity of this cancer, the molecular etiology of this disease is poorly understood. Materials & Methods A 208 gene next-generation sequencing (NGS) panel was used to analyze 20 IBC patient tumor and matched normal samples. Tissues were stained for cell signaling proteins and immune-markers. Cell line studies were performed to understand the impact of genomic variants on therapeutic selection. Results Common pathway alterations reoccur among IBC samples that influence genome stability, PI3K signaling, and ERBB signaling. NGS revealed alterations in both the ERBB/PI3K pathways, including: ERBB2 amplification, ERBB3 mutations, and activating PI3K mutations. Immunohistochemistry (IHC) staining for pS6 and pERBB3 identified ERBB/PI3K pathway activity in IBC samples. Cell line studies using siRNA and neutralizing antibodies demonstrate that mutant ERBB3 signaling contributes to IBC proliferation. IBC tumors with high levels of CD8+ immune infiltrate have a significantly higher somatic mutation rate than other IBC tumors. A proposed score (iScore) based on the somatic mutation rate and the average mutant allele frequency, showed greater correlation with the level of CD8+ immune infiltration. Furthermore, tumors with high CD8+ infiltrating lymphocytes were associated with a higher frequency of alterations in DNA mismatch repair (MMR) genes. IHC revealed high levels of the immune checkpoint signaling molecule PD-L1 in the inflammatory infiltrate of IBC tumors. Conclusions This study identifies a higher level of ERBB3 mutations than reported in other cancers and an important role for ERBB3 mutation in IBC. ERBB3 targeted therapies in combination with ERBB/PI3K drugs may be important for IBC treatment. Furthermore, high somatic mutations rates in a subset of IBC tumors harboring MMR mutations lead to greater levels of PD-L1+ immune infiltrates, which suggests a possible benefit from immunotherapies such as anti-PD-L1 antibodies. iScore, a more predictive value of immune infiltration in tumors, may be indicative of the level of neoantigen exposure to the immune system. The utility of the newly proposed iScore is currently being investigated as a method to predict immune cell infiltrates and immunotherapy response in other tumor types. Citation Format: Diarmuid Moran, Christopher A. Hamm, Kakuturu Rao, Patricia Bacon-Trusk, Karen Pry, Victor Velculescu, Massimo Cristofanilli, Sarah S. Bacus. Genomic analysis identifies drug targetable pathways and predicts immune infiltration in inflammatory breast cancer tumors. [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 3887. doi:10.1158/1538-7445.AM2015-3887

I isolated and characterized 8 polymorphic anonymous nuclear loci from the federally endangered Mitchell’s satyr butterfly, Neonympha mitchellii mitchellii (Lepidoptera: Nymphalidae). All loci reported here are polymorphic and will serve as important tools in the conservation of this rare butterfly.

Ependymoma is a common pediatric central nervous system (CNS) tumor that is believed to originate from ependymal cells located in the lining of ventricular surfaces in the brain. Ependymoma presents great challenges in treatment despite the advances in neurosurgical techniques and adjuvant therapy. To uncover protein interaction networks that are consistently compromised in pediatric ependymomas, we conducted a computational analysis of the array CGH data derived from 203 primary ependymomas from the St. Jude Children's Research Hospital cohort, published by Johnson and colleagues [Nature, 466(7306):632-6, 2010]. The analysis (Bischof and Soares, manuscript in preparation) revealed 15 regions of copy number variation (CNVs) that occurred in at least 50% of the tumors. Peak genes from these CNVs were selected for a confirmatory study using DNA digital PCR assays with an independent ependymoma cohort from the Ann & Robert H Lurie Children's Hospital of Chicago. In this study, all ependymomas displayed copy number alterations in at least one of the 10 genes tested by digital PCR. The digital PCR data sheds light into the biology of ependymomas as the 10 candidate genes have a wide range of cellular functions; from neural stem cell development, to tumor suppression, to the pathogenesis of glioblastoma. In conclusion, we have confirmed that the occurrence of DNA copy number variation in these 10 genes prevail in ependymomas. Indeed, all tumors exhibited at least one of these CNVs. These alterations provide insights into the protein interaction networks underlying development of ependymomas while uncovering targets that may be exploited for therapeutic intervention. Citation Format: Christopher A. Hamm, Fabricio F. Costa, Jared M. Bischof, Elio F. Vanin, Maria de Fatima Bonaldo, Steve Iannaccone, Veena Rajaram, David George, Tadanori Tomita, Stewart Goldman, Lawrence J. Jennings, Richard J. Gilbertson, Marcelo B. Soares. Identification of genes that frequently exhibit copy number alterations in pediatric ependymomas. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3111. doi:10.1158/1538-7445.AM2014-3111

Abnormal patterns of DNA methylation are observed in several types of human cancer. While localized DNA methylation of CpG islands has been associated with gene silencing, the effect that genome-wide loss of methylation has on tumorigenesis is not completely known. To examine its effect on tumorigenesis, we induced DNA demethylation in a rat model of human chondrosarcoma using 5-aza-2-deoxycytidine. Rat specific pyrosequencing assays were utilized to assess the methylation levels in both LINEs and satellite DNA sequences following 5-aza-2-deoxycytidine treatment. Loss of DNA methylation was accompanied by an increase in invasiveness of the rat chondrosarcoma cells, in vitro, as well as by an increase in tumor growth in vivo. Subsequent microarray analysis provided insight into the gene expression changes that result from 5-aza-2-deoxycytidine induced DNA demethylation. In particular, two genes that may function in tumorigenesis, sox-2 and midkine, were expressed at low levels in control cells but upon 5-aza-2-deoxycytidine treatment these genes became overexpressed. Promoter region DNA analysis revealed that these genes were methylated in control cells but became demethylated following 5-aza-2-deoxycytidine treatment. Following withdrawal of 5-aza-2-deoxycytidine, the rat chondrosarcoma cells reestablished global DNA methylation levels that were comparable to that of control cells. Concurrently, invasiveness of the rat chondrosarcoma cells, in vitro, decreased to a level indistinguishable to that of control cells. Taken together these experiments demonstrate that global DNA hypomethylation induced by 5-aza-2-deoxycytidine may promote specific aspects of tumorigenesis in rat chondrosarcoma cells.