Abstract
Background: Previous studies have found that particular pathologic features are more common in breast cancers arising in BRCA mutation carriers. However, the biologic and molecular bases for ...the morphologic associations are not clear. This study is conducted to analyze pathologic and molecular features in tumors stratified by BRCA1 or BRCA2 mutation status using the breast cancer samples that have comprehensive molecular portraits characterized by the Cancer Genome Atlas (TCGA).
Methods: The digital slides of breast cancer samples submitted for comprehensive molecular profiling to the TCGA were reviewed by expert breast pathologists, who were unaware of the BRCA status or other molecular signatures. Each tumor was evaluated and scored for histologic type, nuclear pleomorphism, tubule formation, mitosis, stromal inflammation, and necrosis. 562 cases had both pathology and tumor exome sequencing data available and constituted the current study population. We determined the association of somatic BRCA1 and BRCA2 mutation status with pathologic features and molecular characteristics (mutation of PIK3CA and TP53, and molecular subtypes defined by PAM50 mRNA data) using the Fisher exact test for categorical variables and the Wilcoxon test for ordinal variables.
Results: Of the 562 tumors, 514 had no BRCA1 or BRCA2 mutation, while 48 (8.5%) of tumors were found to harbor a BRCA1 mutation (n = 16, 3%), BRCA2 mutation (n = 30, 5%), or mutation in both (n = 2, 0.3%). BRCA1 and BRCA2 mutational status showed no significant association with lobular features, tubule formation, nuclear pleomorphism, or stromal inflammation (all p > 0.05), although there was a trend for increased nuclear pleomorphism in BRCA2 mutant cases (p = 0.07). The lack of significant association of BRCA1/2 mutational status with these features may be due to our study's relatively small number of BRCA1/2 mutant cases. Both BRCA1 and BRCA2 mutations were associated with a higher mitotic count (p = 0.03 and 0.04, respectively). BRCA2 mutation showed no association with necrosis (p = 1), while BRCA1 mutation status was associated with increased necrosis (OR = 2.7, p = 0.04). BRCA2 mutation status showed no significant association with PAM50 subtype (p = 0.37), while BRCA1 mutation status was significantly associated with PAM50 molecular subtype (p = 0.005), with the greatest enrichment among Basal-like (7/70 Basal-like with BRCA1 mutation, 10%) and depletion among Luminal-B (0/79 Luminal-B with BRCA1 mutation, 0%). Neither BRCA1 nor BRCA2 mutations were significantly association with PIK3CA mutations (p = 0.39, 0.08, respectively). BRCA2 mutation status was not associated with TP53 mutations (p = 0.65), while BRCA1 mutation status was associated with increased TP53 mutations (OR = 4.0, p = 0.005).
Conclusion: Tumors with BRCA1 and BRCA2 alterations are associated with specific pathologic and molecular features. However, there is molecular and morphologic heterogeneity within these cancers. These factors need to be considered when designing algorithms for BRCA testing and targeted therapy in BRCA-related cancers.
Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-05-15.
Inhibition of NO synthesis promotes P-selectin expression on endothelial cells; however, the precise mechanism is unclear. Because NO has been shown to inhibit protein kinase C (PKC) activity, we ...examined the hypothesis that the NO synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME) stimulates P-selectin expression on platelets via PKC activation. Ten-minute incubation with either phorbol 12-myristate 13-acetate (PMA), thrombin, or L-NAME significantly increased P-selectin expression on platelets (as assessed by flow-cytometric analysis) and PKC activity of platelet membranes. Increased P-selectin expression induced by either PMA, thrombin, or L-NAME was significantly attenuated by the selective PKC inhibitor UCN-01 (7-hydroxystaurosporine). Furthermore, L-NAME-induced P-selectin expression was significantly attenuated by either L-arginine, 8-bromo-cGMP, or sodium nitroprusside (SNP). Interestingly, L-NAME further potentiated P-selectin upregulation by thrombin. L-NAME, thrombin, and PMA also significantly increased polymorphonuclear leukocyte adherence to the coronary artery endothelium, an effect that was significantly attenuated by the anti-P-selectin monoclonal antibody PB1.3 or by UCN-01, L-arginine, 8-bromo-cGMP or SNP but not by D-arginine or the nonblocking anti-P-selectin monoclonal antibody NBP1.6. These results indicate that inhibition of NO synthesis induces rapid P-selectin expression, which appears to be at least partially mediated by PKC activation in platelets. Similar effects and mechanisms of L-NAME on P-selectin function were also observed in endothelial cells, another site of P-selectin expression. Thus, PKC activation may play an important role in cell-to-cell interaction when NO production is compromised. (Arterioscler Thromb Vasc Biol. 1995;15:2068-2075.).
Wind tunnel studies provide a valuable experimental approach that can be used to investigate the influence of specific environmental parameters and to make generalizations about insect behavior. In ...this study, we designed an experiment to test the sensitivity of butterflies to isolated environmental parameters in the context of understanding edge responses. We tested the behavior of 21 different butterfly species in response to certain stimuli, including food source, feeder color, temperature, and UV light. Certain butterfly species (e.g. Heliconius melpomene and Papilio polytes) were particularly active in the wind tunnel setup. All butterfly species tested preferred blue feeders over white, yellow or pink. Investigation of the UV content of the different feeders and the butterflies' preferred nectar plant showed a similar wavelength response, which could indicate a UV preference in butterflies. We also observed species-dependent temperature preferences. Papilio lowii had a significant preference for the warm side (36.0°C) of the wind tunnel, whereas Papilio polytes showed a significant preference for the cold side (25.3°C).
Leishmania donovani is an obligatory intracellular parasite which cycles between the midgut of sand flies (extracellular promastigote) and the phagolysosomes of mammalian macrophages (intracellular ...amastigote). Promastigotes have been readily cultured, whereas axenic cultures of amastigotes have only recently been developed. A new method for in vitro differentiation of
L. donovani promastigotes into amastigotes is presented, in which promastigotes are exposed to environmental changes that mimic the in vivo process. First, promastigotes are subjected to 37°C+5% CO
2 for 24 h, and then are shifted to pH 5.5. Under these conditions, differentiation is completed within 120 h. In the reverse process, amastigotes are induced to differentiate back to promastigotes by transferring them to promastigote growth conditions (medium 199 at pH 7.4 and 26°C). Axenic amastigotes closely resemble animal-derived amastigotes. They manifest all seven proteins of the amastigote-specific A2 gene family. They down-regulate lipophosphoglycan (LPG) synthesis and do not express it on their surface. LPG is up-regulated 2 h after inducing amastigotes to differentiate to promastigotes. Within 6 h, parasites resume the promastigote level of this molecule, although differentiation is completed only after 48 h. Axenic amastigotes also express amastigote-like metabolic activities of proline uptake, as well as thymidine and proline incorporation. In conclusion, the results indicate that the method developed for in vitro differentiation of
L. donovani promastigotes to amastigotes is efficient and yields organisms resembling animal-derived amastigotes. Being able to induce in vitro differentiation of
L. donovani provides us with an excellent tool to study
Leishmania development and differentiation.
Following DNA damage, human cells arrest primarily in the G(1) and G(2) phases of the cell cycle. Here, we show that after irradiation, human cancer cells with targeted deletion of PTEN or naturally ...occurring PTEN mutations can exert G(1) and G(2) arrests but are unable to arrest in size. Pharmacological inhibition of phosphoinositol-3-kinase or mTOR in PTEN(-/-) cells restored the size arrest, whereas siRNA-mediated depletion of TSC2 in PTEN(+/+) cells attenuated the size arrest. Radiation treatment potentiated Akt activation in PTEN(-/-) but not PTEN(+/+) cells. Finally, abrogation of the size arrest via PTEN deletion conferred radiosensitivity both in vitro and in vivo. These results identify a new tumor suppressor gene-regulated, DNA damage-inducible arrest that occurs simultaneously with the G(1) and G(2) arrests but is genetically separable from them. We suggest that aberrant regulation of cell size during cell cycle arrest may be important in human cancer pathogenesis.
Abstract
The latent structure of schizotypy and psychosis-spectrum symptoms remains poorly understood. Furthermore, molecular genetic substrates are poorly defined, largely due to the substantial ...resources required to collect rich phenotypic data across diverse populations. Sample sizes of phenotypic studies are often insufficient for advanced structural equation modeling approaches. In the last 50 years, efforts in both psychiatry and psychological science have moved toward (1) a dimensional model of psychopathology (eg, the current Hierarchical Taxonomy of Psychopathology HiTOP initiative), (2) an integration of methods and measures across traits and units of analysis (eg, the RDoC initiative), and (3) powerful, impactful study designs maximizing sample size to detect subtle genomic variation relating to complex traits (the Psychiatric Genomics Consortium PGC). These movements are important to the future study of the psychosis spectrum, and to resolving heterogeneity with respect to instrument and population. The International Consortium of Schizotypy Research is composed of over 40 laboratories in 12 countries, and to date, members have compiled a body of schizotypy- and psychosis-related phenotype data from more than 30000 individuals. It has become apparent that compiling data into a protected, relational database and crowdsourcing analytic and data science expertise will result in significant enhancement of current research on the structure and biological substrates of the psychosis spectrum. The authors present a data-sharing infrastructure similar to that of the PGC, and a resource-sharing infrastructure similar to that of HiTOP. This report details the rationale and benefits of the phenotypic data collective and presents an open invitation for participation.
In humans, guanylyl cyclase C (GCC) is expressed by mucosal cells lining the intestine, from the duodenum to the rectum, but
not by extraintestinal tissues. Expression of GCC persists after mucosal ...cells undergo neoplastic transformation, and this
protein has been identified in all primary and metastatic colorectal tumors examined to date, suggesting that GCC may be a
highly specific biomarker for colorectal cancer. The utility of GCC as a diagnostic biomarker and therapeutic target is predicated,
in part, on defining the variability of its expression in colorectal cancer cells. Similarly, the utility of this biomarker
to define tumor burden in diagnosing, staging, and postoperative surveillance of patients is predicated on quantifying GCC
expression in cancer cells in tissues and blood. The present studies examined the heterogeneity of GCC expression in eight
human colorectal carcinoma cell lines in vitro representing the full spectrum of cytological differentiation. Quantification
of GCC expression by ligand binding and stimulation of cGMP accumulation demonstrated that functional GCC expression is heterogeneous
in different colorectal cancer cell lines. Qualitative reverse transcription (RT)-PCR demonstrated that all colorectal cancer
cells examined expressed GCC mRNA. However, GCC expression varied 100-fold in different colorectal cancer cell lines, determined
by a novel quantitative RT-PCR assay. Functional and molecular expressions of GCC were unrelated to the differentiation state
of cancer cells. These studies suggest that GCC is heterogeneously expressed by colorectal cancer cells in vitro and suggest
a role for quantitative RT-PCR analysis in the development of diagnostic tests using GCC as a biomarker for metastatic colorectal
cancer.
Escherichia coli heat-stable enterotoxins (ST) are small peptides of 18 or 19 amino acids that bind to specific cell surface receptors located on the intestinal brush border and activate guanylate ...cyclase, resulting in an increase in the intracellular cyclic guanosine 3',5'-monophosphate content of the cell. The present study examined whether receptors for ST are expressed by primary and metastatic human colonic tumors in vivo.
Plasma membranes prepared from surgical tissue samples from normal colon, liver and lung, primary colonic adenocarcinomas, and colon carcinomas metastatic to lung and liver were analyzed for the structural and functional characteristics of constituent ST receptors.
All primary and metastatic colonic tumors examined bound ST, showing receptors of high (pmol/L) and low (nmol/L) affinity with densities that were similar to those in normal colon. Also, affinity cross-linking of labeled ST to membranes showed similar binding proteins in primary and metastatic tumors and normal colon. ST binding and affinity-labeled proteins were not detected in normal extraintestinal tissues. Guanylate cyclase was activated by ST in membranes from all colonic tumors studied, with efficacies and potencies that were similar to those in normal colon. ST did not activate this enzyme in normal extraintestinal tissues.
Receptors for ST are expressed by primary and metastatic human colonic tumors in vivo, with structural and functional characteristics that are similar to those in normal human colon.
Epitope tagging is a powerful and commonly used approach for studying the physical properties of proteins and their functions and localization in eukaryotic cells. In the case of Saccharomyces ...cerevisiae, it has been possible to exploit the high efficiency of homologous recombination to tag proteins by modifying their endogenous genes, making it possible to tag virtually every endogenous gene and perform genome-wide proteomics experiments. However, due to the relative inefficiency of homologous recombination in cultured human cells, epitope-tagging approaches have been limited to ectopically expressed transgenes, with the attendant limitations of their nonphysiological transcriptional regulation and levels of expression. To overcome this limitation, a modification and extension of adeno-associated virus-mediated human somatic cell gene targeting technology is described that makes it possible to simply and easily create an endogenous epitope tag in the same way that it is possible to knock out a gene. Using this approach, we have created and validated human cell lines with epitope-tagged alleles of two cancer-related genes in a variety of untransformed and transformed human cell lines. This straightforward approach makes it possible to study the physical and biological properties of endogenous proteins in human cells without the need for specialized antibodies for individual proteins of interest.
Antibodies against alloantigens on donor platelets are a major cause of refractoriness to platelet transfusions in patients with thrombocytopenia. These antibodies usually arise in response to HLA ...class I antigens on leukocytes and platelets, and they may result from maternal–fetal incompatibility or repeated blood transfusions. Resources to supply HLA-compatible platelets to patients who have refractoriness to platelet transfusions from random donors are often limited. Moreover, up to 60 percent of transfusions of presumably HLA-compatible platelets fail to increase the platelet count in patients whose thrombocytopenia is refractory to transfusions.
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Previous studies of ways to prevent alloimmunization in patients with thrombocytopenia . . .
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