Experimentally induced liver tumors in mice harbor activating mutations in either Catnb (␤-catenin) or Ha-ras, according to the carcinogenic treatment. We have now investigated by microarray analysis the gene expression profiles in tumors of the two genotypes. In total, 364 genes or expressed sequences with aberrant expression relative to normal liver were identified, but only 30 of these demonstrated unidirectional changes in both tumor types. Several functional clusters were identified that involve changes in amino acid utilization and ammonia disposition in Catnb-mutated tumors as opposed to alterations in lipid and cholesterol metabolism in Ha-ras-mutated tumors. Moreover, several genes coding for inhibitory molecules within the Wnt-signaling pathway were upregulated in Catnb-mutated tumors, suggesting induction of a negative feedback loop, whereas Ha-ras-mutated tumors showed alterations in the expression of several genes functional in monomeric G-protein signaling. We conclude that mouse hepatoma cells adopt different evolutionary strategies that allow for their selective outgrowth under variable environmental conditions. Human hepatocellular cancers (HCC) lack RAS mutations but are frequently mutated in CTNNB1, the human Catnb ortholog. The set of genes aberrantly expressed in Catnb-mutated mouse tumors was used to screen, by expression profiling, for dysregulation of orthologous genes within a panel of 25 HCCs, of which 10 were CTNNB1-mutated. HCCs with activated ␤-catenin displayed a gene expression profile that was similar to Catnb-mutated mouse tumors but distinct from the other human HCCs. In conclusion, expression fingerprints may be used for diagnostic purposes and potential new therapeutic intervention strategies. If, however, DEN treatment is combined with subsequent chronic administration of the liver tumor promoter phenobarbital (PB) according to a classical initiation-promotion protocol, tumors predominate that lack ras mutations but show activating mutations in the Catnb (␤-catenin) proto-oncogene instead. 3 On histological examination, liver tumors generated in the absence or presence of the tumor promoter PB demonstrate considerable differences in hematoxylin-eosin-stained sections: the former are often basophilic and are generally composed of comparatively small cells, whereas the latter are often eosinophilic and contain larger cells with enlarged nuclei. 4,5 Several additional differences have been described if individual markers were used for discrimination of tumor types including glutamine synthetase (GS), which is strongly increased in expression in Catnb-mutated but undetectable in ras-mutated mouse hepatocytes. 6 This suggests that mutation in either of the two genes produces divergent phenotypes; however, comparative genome-

Endocrine disruption is a specific form of toxicity, where natural and/or anthropogenic chemicals, known as “endocrine disruptors” (EDs), trigger adverse health effects by disrupting the endogenous hormone system. There is need to harmonize guidance on the regulation of EDs, but this has been hampered by what appeared as a lack of consensus among scientists. This publication provides summary information about a consensus reached by a group of world-leading scientists that can serve as the basis for the development of ED criteria in relevant EU legislation. Twenty-three international scientists from different disciplines discussed principles and open questions on ED identification as outlined in a draft consensus paper at an expert meeting hosted by the German Federal Institute for Risk Assessment (BfR) in Berlin, Germany on 11–12 April 2016. Participants reached a consensus regarding scientific principles for the identification of EDs. The paper discusses the consensus reached on background, definition of an ED and related concepts, sources of uncertainty, scientific principles important for ED identification, and research needs. It highlights the difficulty in retrospectively reconstructing ED exposure, insufficient range of validated test systems for EDs, and some issues impacting on the evaluation of the risk from EDs, such as non-monotonic dose–response and thresholds, modes of action, and exposure assessment. This report provides the consensus statement on EDs agreed among all participating scientists. The meeting facilitated a productive debate and reduced a number of differences in views. It is expected that the consensus reached will serve as an important basis for the development of regulatory ED criteria.

All cells and organisms exhibit stress-coping mechanisms to ensure survival. Cytoplasmic protein-RNA assemblies termed stress granules are increasingly recognized to promote cellular survival under stress. Thus, they might represent tumor vulnerabilities that are currently poorly explored. The translation-inhibitory eIF2α kinases are established as main drivers of stress granule assembly. Using a systems approach, we identify the translation enhancers PI3K and MAPK/p38 as pro-stress-granule-kinases. They act through the metabolic master regulator mammalian target of rapamycin complex 1 (mTORC1) to promote stress granule assembly. When highly active, PI3K is the main driver of stress granules; however, the impact of p38 becomes apparent as PI3K activity declines. PI3K and p38 thus act in a hierarchical manner to drive mTORC1 activity and stress granule assembly. Of note, this signaling hierarchy is also present in human breast cancer tissue. Importantly, only the recognition of the PI3K-p38 hierarchy under stress enabled the discovery of p38’s role in stress granule formation. In summary, we assign a new pro-survival function to the key oncogenic kinases PI3K and p38, as they hierarchically promote stress granule formation.

The toxicological relevance of effects observed at molecular stage, which occur at dose levels well below classical no-observed adverse effect levels is currently subject to controversial scientific debate. While the importance of molecular effects for the identification of a mode of action or an adverse outcome pathway is undisputed, their impact for other regulatory purposes remains uncertain. Here, we report the results of a 28-day rat-feeding study including three widely used hepatotoxic (tri)azole fungicides (cyproconazole, epoxiconazole and prochloraz) administered individually at five dose levels, ranging from slightly above the reference values to a clear toxic effect dose. Parameters analysed included pathology, histopathology, clinical chemistry and particularly effects on the molecular level. Since azole fungicides are considered to cause liver toxicity by a mechanism involving the constitutive androstane receptor (CAR), a known CAR activator (phenobarbital, PB) was administered to investigate potential similarities between triazoles and PB-mediated liver toxicity by pathway-focused gene expression analysis. Our results show an increase in liver weights and additionally histopathological changes (hepatocellular hypertrophy) for all substances at the top dose levels. The effects on liver weight were most pronounced for cyproconazole by which also the animals receiving the next lower dose were affected. In addition, vacuolisation of hepatocytes was observed at the top dose level. No such findings were obtained with any substance at lower doses to which consumers and operators might be exposed to. In contrast, the expression of sensitive marker genes (like some cytochrome-P-450 isoforms) was significantly affected also at the lower dose levels. While some of these changes, like the induction of genes related to fatty acid and phospholipid metabolism (e.g. Fasn, Fat/Cd36, Ppargc1a) or xenobiotic metabolism (Cyp1a1, Cyp2b1, Cyp3a2), could be associated with high dose effects like hepatocellular vacuolisation or hypertrophy, a histopathological correlate was lacking for others.

No abstract