Prevalence of germ-line mutations in p16, p19ARF, and CDK4 in familial melanoma:analysis of a clinic-based population

Five to ten percent of individuals with melanoma have another affected family member, suggesting familial predisposition. Germ-line mutations in the cyclin-dependent kinase (CDK) inhibitor p16 have been reported in a subset of melanoma pedigrees, but their prevalence is unknown in more common cases of familial melanoma that do not involve large families with multiple affected members. We screened for germ-line mutations in p16 and in two other candidate melanoma genes, p19ARF and CDK4, in 33 consecutive patients treated for melanoma; these patients had at least one affected first or second degree relative (28 independent families). Five independent, definitive p16 mutations were detected (18%, 95% confidence interval: 6%, 37%), including one nonsense, one disease-associated missense, and three small deletions. No mutations were detected in CDK4. Disease-associated mutations in p19ARF, whose transcript is derived in part from an alternative codon reading frame of p16, were only detected in patients who also had mutations inactivating p16. We conclude that germ-line p16 mutations are present in a significant fraction of individuals who have melanoma and a positive family history.

Characterisation of Bioresorbable Composite Performance for Validation of New On-Line Process Monitoring Technology

There is an increasing interest in the biomedical field to create implantable medical devices to provide a temporary mechanical function for use inside the human body. In many of these applications bioresorbable polymer composites using PLLA with β-TCP , are increasingly being used due to their biocompatability, biodegradability and mechanical strength.1,3 These medical devices can be manufactured using conventional plastics processing methods such as injection moulding and extrusion, however there is great need to understand and control the process due to a lack of knowledge on the influence of processing on material properties. With the addition of biocompatible additives there is also a requirement to be able to predict the quality and level of dispersion within the polymer matrix. On-line UV-Vis spectroscopy has been shown to monitor the quality of fillers in polymers. This can eliminate time consuming and costly post-process evaluation of additive dispersion. The aim of this work was to identify process and performance relationships of PLLA/β-TCP composites with respect to melt-extrusion conditions. This is part of a wider study into on-line process monitoring of bioresorbable polymers as used in the medical industry.
These results show that final properties of the PLLA/ β-TCP composite are highly influenced by the particle size and loading. UV-Vis spectroscopy can be used on-line to monitor the final product and this can be utilised as a valuable tool for quality control in an application where consistent performance is of paramount importance.

Activated protein C inhibits lipopolysaccharide-induced nuclear translocation of nuclear factor kappaB (NF-kappaB) and tumour necrosis factor alpha (TNF-alpha) production in the THP-1 monocytic cell line

Activated protein C (APC) protects against sepsis in animal models and inhibits the lipopolysacharide (LPS)-induced elaboration of proinflammatory cytokines from monocytes. The molecular mechanism responsible for this property is unknown. We assessed the effect of APC on LPS-induced tumour necrosis factor alpha (TNF-alpha) production and on the activation of the central proinflammatory transcription factor nuclear factor-kappaB (NF-kappaB) in a THP-1 cell line. Cells were preincubated with varying concentrations of APC (200 microg/ml, 100 microg/ml and 20 microg/ml) before addition of LPS (100 ng/ml and 10 microg/ml). APC inhibited LPS-induced production of TNF-alpha both in the presence and absence of fetal calf serum (FCS), although the effect was less marked with 10% FCS. APC also inhibited LPS-induced activation of NF-kappaB, with APC (200 microg/ml) abolishing the effect of LPS (100 ng/ml). The ability of APC to inhibit LPS-induced translocation of NF-kappaB is likely to be a significant event given the critical role of the latter in the host inflammatory response.

Balancing the (carbon) budget: Using linear inverse models to estimate carbon flows and mass-balance 13C:15N labelling experiments in low oxygen sediments.

Over 1 million km² of seafloor experience permanent low-oxygen conditions within oxygen minimum zones (OMZs). OMZs are predicted to grow as a consequence of climate change, potentially affecting oceanic biogeochemical cycles. The Arabian Sea OMZ impinges upon the western Indian continental margin at bathyal depths (150 - 1500 m) producing a strong depth dependent oxygen gradient at the sea floor. The influence of the OMZ upon the short term processing of organic matter by sediment ecosystems was investigated using in situ stable isotope pulse chase experiments. These deployed doses of ¹³C:¹⁵N labeled organic matter onto the sediment surface at four stations from across the OMZ (water depth 540 - 1100 m; [O₂] = 0.35 - 15 μM). In order to prevent experimentally anoxia, the mesocosms were not sealed. ¹³C and ¹⁵N labels were traced into sediment, bacteria, fauna and ¹³C into sediment porewater DIC and DOC. However, the DIC and DOC flux to the water column could not be measured, limiting our capacity to obtain mass-balance for C in each experimental mesocosm. Linear Inverse Modeling (LIM) provides a method to obtain a mass-balanced model of carbon flow that integrates stable-isotope tracer data with community biomass and biogeochemical flux data from a range of sources. Here we present an adaptation of the LIM methodology used to investigate how ecosystem structure influenced carbon flow across the Indian margin OMZ. We demonstrate how oxygen conditions affect food-web complexity, affecting the linkages between the bacteria, foraminifera and metazoan fauna, and their contributions to benthic respiration. The food-web models demonstrate how changes in ecosystem complexity are associated with oxygen availability across the OMZ and allow us to obtain a complete carbon budget for the stationa where stable-isotope labelling experiments were conducted.

Interleukin-8 is associated with proliferation, migration, angiogenesis and chemosensitivity in vitro and in vivo in colon cancer cell line models

Interleukin-8 (IL-8), a chemokine with a defining CXC amino acid motif, is known to possess tumorigenic and proangiogenic properties. Overexpression of IL-8 has been detected in many human tumors, including colorectal cancer (CRC), and is associated with poor prognosis. The goal of our study was to determine the role of IL-8 overexpression in CRC cells in vitro and in vivo. We stably transfected the IL-8 cDNA into two human colon cancer cell lines, HCT116 and Caco2, and selected IL-8-secreting transfectants. Real-time RT-PCR confirmed that IL-8 mRNA was overexpressed in IL-8 transfectants with 45- to 85-fold higher than parental cells. The IL-8-transfected clones secreted 19- to 28-fold more IL-8 protein than control and parental cells as detected by ELISA. The IL-8 transfectants demonstrated increased cellular proliferation, cell migration and invasion based on functional assays. Growth inhibition studies showed that IL-8 overexpression lead to a significant resistance to oxaliplatin (p < 0.0001). Inhibition of IL-8 overexpression with small interfering RNA reversed the observed increases in tumorigenic functions and oxaliplatin resistance, suggesting that IL-8 not only provides a proliferative advantage but also promotes the metastatic potential of colon cancer cells. Using a tumor xenograft model, IL-8-expressing cells formed significantly larger tumors than the control cells with increased microvessel density. Together, these findings indicate that overexpression of IL-8 promotes tumor growth, metastasis, chemoresistance and angiogenesis, implying IL-8 to be an important therapeutic target in CRC.

Selecting the best targeted agent in first-line treatment of unresectable liver metastases from colorectal cancer:does the bench have the answers?

For physicians facing patients with organ-limited metastases from colorectal cancer, tumor shrinkage and sterilization of micrometastatic disease is the main goal, giving the opportunity for secondary surgical resection. At the same time, for the majority of patients who will not achieve a sufficient tumor response, disease control remains the predominant objective. Since FOLFOX or FOLFIRI have similar efficacies, the challenge is to define which could be the most effective targeted agent (anti-EGFR or anti-VEGF) to reach these goals. Therefore, a priori molecular identification of patients that could benefit from anti-EGFR or anti-VEGF monoclonal antibodies (i.e. the currently approved targeted therapies for metastatic colorectal cancer) is of critical importance. In this setting, the KRAS mutation status was the first identified predictive marker of response to anti-EGFR therapy. Since it has been demonstrated that tumors with KRAS mutation do not respond to anti-EGFR therapy, KRAS status must be determined prior to treatment. Thus, for KRAS wild-type patients, the choices that remain are either anti-VEGF or anti-EGFR. In this review, we present the most updated data from translational research programs dealing with the identification of biomarkers for response to targeted therapies.

Pharmacogenetic angiogenesis profiling for first-line Bevacizumab plus oxaliplatin-based chemotherapy in patients with metastatic colorectal cancer

PURPOSE: There is substantial germline genetic variability within angiogenesis pathway genes, thereby causing interindividual differences in angiogenic capacity and resistance to antiangiogenesis therapy. We investigated germline polymorphisms in genes involved in VEGF-dependent and -independent angiogenesis pathways to predict clinical outcome and tumor response in metastatic colorectal cancer (mCRC) patients treated with bevacizumab and oxaliplatin-based chemotherapy.

EXPERIMENTAL DESIGN: A total of 132 patients treated with first-line bevacizumab and FOLFOX or XELOX were included in this study. Genomic DNA was isolated from whole-blood samples by PCR-RFLP or direct DNA sequencing. The endpoints of the study were progression-free survival (PFS), overall survival (OS), and response rate (RR).

RESULTS: The minor alleles of EGF rs444903 A>G and IGF-1 rs6220 A>G were associated with increased OS and remained significant in multivariate Cox regression analysis (HR: 0.52; 95% CI: 0.31-0.87; adjusted P = 0.012 and HR: 0.60; 95% CI: 0.36-0.99; adjusted P = 0.046, respectively). The minor allele of HIF1α rs11549465 C>T was significantly associated with increased PFS but lost its significance in multivariate analysis. CXCR1 rs2234671 G>C, CXCR2 rs2230054 T>C, EGFR rs2227983 G>A, and VEGFR-2 rs2305948 C>T predicted tumor response, with CXCR1 rs2234671 G>C remaining significant in multiple testing (P(act) = 0.003).

CONCLUSION: In this study, we identified common germline variants in VEGF-dependent and -independent angiogenesis genes predicting clinical outcome and tumor response in patients with mCRC receiving first-line bevacizumab and oxaliplatin-based chemotherapy.