Low Percentage of KRAS Mutations Revealed by Locked Nucleic Acid Polymerase Chain Reaction: Implications for Treatment of Metastatic Colorectal Cancer

Metastatic colorectal cancer (mCRC) is frequently characterized by the presence of mutations of the KRAS oncogene, which are generally associated with a poor response to treatment with anti-epidermal growth factor receptor (anti-EGFR) monoclonal antibodies. With the methods currently used, a case is classified as KRAS-mutated when approximately 20% of the cells bear an activating KRAS mutation. These considerations raise the question of whether cells with a mutated KRAS can be found in mCRC cases classified as KRAS wild-type when more sensitive methods are used. In addition, the issue arises of whether these mCRC cases with low proportion of KRAS-mutated cells could account at least in part for the therapeutic failure of anti-EGFR therapies that occur in 40–60% of cases classified as KRAS wild type. In this study, we compared the classical assays with a very sensitive test, a locked nucleic acid (LNA) polymerase chain reaction (PCR), capable of detecting KRAS-mutated alleles at extremely low frequency (detection sensitivity limit 0.25% mutated DNA/wild-type DNA). By analyzing a cohort of 213 mCRC patients for KRAS mutations, we found a 20.6% discordance between the sequencing/TheraScreen methods and the LNA-PCR. Indeed, 44 mCRC patients initially considered KRAS wild type were reclassified as KRAS mutated by using the LNA-PCR test. These patients were more numerous among individuals displaying a clinical failure to anti-EGFR therapies. Failure to respond to these biological treatments occurred even in the absence of mutations in other EGFR pathway components such as BRAF.     

Lipid-transfer proteins in biosynthetic pathways

Compartmentalization is a defining feature of eukaryotic cells that allows the spatial segregation of different functions, such as protein and lipid synthesis, and ensures their fidelity and efficiency. This imposes the need for an intense flux of metabolic intermediates between segregated enzymatic activities, as seen for the sequential transport of neosynthesized proteins through the segments of the secretory pathway during their post-translational modification. For lipid synthesis, the identification of proteins that transfer lipids between membranes has revealed an additional mechanism for this intercompartment exchange. The intense interest elicited by the lipid-transfer proteins over the last few years has led to the definition of their central role in key processes, such as lipid metabolism, membrane trafficking, and signaling.     

Synthesis, chaperoning, and metabolism of proteins are regulated by NT-3/TrkC signaling in the medulloblastoma cell line DAOY

The human medulloblastoma cell line DAOY was transfected with Tropomyosin receptor kinase (TrkC), a marker for good prognostic outcome. Following TrkC-activation by its ligand neurotrophin-3, protein extracts from DAOY cells were run on 2DE with subsequent MALDI-TOF-TOF analysis and quantification in order to detect downstream effectors. Protein levels of translational, splicing, processing, chaperone, protein handling, and metabolism machineries were shown to depend on neurotrophin-3-induced TrkC activation probably representing pharmacological targets.     

Hematology and serum chemistry values in captive and wild pichis, Zaedyus pichiy (Mammalia, Dasypodidae)

As part of an ongoing study on the health status of pichis, Zaedyus pichiy (Mammalia, Dasypodidae), blood was collected under manual restraint from 72 free-ranging pichis captured in Mendoza Province, Argentina, between November 2001 and December 2006, and from 22 captive-kept pichis in January 2007. Reference values were established for hematology and serum chemistry. Pichis had lower leukocyte counts and higher mean corpuscular volumes than most other mammals. Blood values were similar for captive and wild pichis, and only a few significant differences were found among genders or age classes.     

The role of the social sciences and economics in understanding and informing tree biosecurity policy and planning: a global summary and synthesis

Increased global biosecurity threats to trees, woods and forests have been strongly linked to the upsurge in worldwide trade and the expansion of tourism. A whole range of social, economic and political actors are implicated and affected by the movement of pests and diseases along these international pathways. A number of factors affect the actions of stakeholders, and wider public, including their values and motivations, how risks are perceived and acted upon, their ability to act, as well as the existing regulatory and economic environment. Understanding these factors is key to any future attempts to improve biosecurity policy and practice, and we present available evidence on six key dimension: (1) the role of different stakeholders and the broader public within tree health; (2) levels of knowledge and awareness of tree pests and diseases amongst the variety of end-user ‘stakeholder’ groups, and influences on their attitudes and practices; (3) social acceptability of management approaches; (4) the impact of formal and informal governance arrangements; (5) risk communication; (6) economic analyses on the impact of tree pests. We conclude by identifying evidence gaps and emphasising the need for better integration within the social sciences and between the social and natural sciences to promote effective interdisciplinary and policy-relevant contributions to tree health.     

Tobacco-smoke exposure indicators and urinary mutagenicity

In this study, the correlation of indicators of external (i.e. mean daily intake of condensate, nicotine, tobacco and tobacco proteins, and daily number of cigarettes smoked) and of internal tobacco-smoke exposure (i.e. urinary 1-pyrenol, nicotine and its metabolites and trans,trans-muconic acid) with urinary mutagenicity, detected on YG1024 Salmonella typhimurium strain with S9, were examined in 118 smokers.An increase in urinary mutagenicity was clearly significantly correlated with each external and internal indicators of exposure to tobacco smoke (correlation coefficient (r) ranging between 0.22 and 0.54, P<0.01), with a greater extent in the case of indicators of internal dose. In multiple regression analysis, among the indicators of external exposure, daily tobacco intake was the only variable significantly associated with urinary mutagenicity (t=2.47, P=0.015, with partial contribution to r²=5.15%). Instead, when all indicators of exposure (external and internal) were considered in the analysis, the influence of urinary 1-pyrenol on urinary mutagenicity was predominant, followed by those of urinary trans,trans-muconic acid and nicotine plus metabolites (t=4.63, 2.73 and 2.08, P<0.001, P=0.002 and 0.04, with partial contribution to r²=17.0, 6.66 and 3.96%, respectively), with no influence at all of external tobacco-smoke exposure indicators.In conclusion, our results show that indicators of internal dose are better correlated with formation of mutagens in urine of smokers. Among these, the best indicator was urinary 1-pyrenol and this result designates the combustion processes of tobacco as the determining step for the formation of urinary mutagens. However, as these biomarkers cannot be analysed the amount of daily tobacco intake represent the best valuable index of external (presumptive) exposure to tobacco-smoke genotoxins.     

hERG1 Potassium Channel Expression in Colorectal Adenomas: Comparison with Other Preneoplastic Lesions of the Gastrointestinal Tract

Preneoplastic lesions represent a useful target for early diagnosis and follow-up of gastrointestinal malignancies. hERG1 channel expression was tested by immunohistochemistry (IHC) in a cohort of colorectal adenoma samples belonging to Italian subjects. Overall, hERG1 was expressed in 56.5% of cases with both high staining intensity and a high percentage of positive cells. Moreover, hERG1 was expressed in a higher percentage of dysplastic adenomas with respect to hyperplastic lesions, and the proportion of positive samples further increased in patients with high-grade dysplasia. Comparing hERG1 expression in other preneoplastic lesions of the GI tract (gastric dysplasia and Barrett’s esophagus), it emerged that in all the conditions, hERG1 was expressed with a diffused pattern, throughout the cell, with variable staining intensity within the samples. The highest expression was detected in gastric dysplasia samples and the lowest in Barrett’s esophagus at similar levels observed in colorectal adenomas. Our results show that hERG1 is aberrantly expressed in human preneoplastic lesions of the gastrointestinal tract and has a different pattern of expression and role in the different sites. Overall, the detection of hERG1 expression in preneoplastic lesions could represent a novel diagnostic or prognostic marker of progression in the gastrointestinal tract.     

Enhancing effect of lithium and potassium ions on lectin-induced lymphocyte proliferation in aging and down's syndrome subjects

The effect of different concentrations of LiCl or KCl (0.6–20 meq/liter) on PHA-stimulated lymphocytes from young, old, and Down's syndrome subjects was studied. LiCl showed a dramatic enhancing effect on [³H]thymidine incorporation induced by a suboptimal dose of PHA in old subjects and Down's syndrome patients. An increase of [³H]thymidine incorporation in human lymphocytes stimulated by a suboptimal dose of PHA was also observed with KCl. This effect was higher in old subjects than that observed in young and Down's subjects. LiCl and KCl can modulate and partially restore the derangement in early events of mitogen stimulation which seems to be present in lymphocytes from both old and Down's syndrome subjects.