Cytokines and inflammatory mediators: 25. Certolizumab Pegol has a Different Profile from the other Anti-TNFS, Including Golimumab, in a Variety of in Vitro Assays

Background: Activities of the anti-TNFs, certolizumab pegol (CZP), etanercept (ETA), infliximab (IFX) and adalimumab (ADA), have been compared in a range of in vitro assays. CZP is the only licensed PEGylated Fab' anti-TNF; ETA is a fusion protein with an IgG1 Fc, and IFX and ADA are both antibodies with an IgG1 Fc. Golimumab (GLM) is a monoclonal IgG1 TNF inhibitor recently approved for a number of indications; it is thus of interest to assess the in vitro activity of GLM. In vitro assays previously used were neutralisation of TNF in the L929 bioassay, inhibition of LPS-driven cytokine production by monocytes, induction of apoptosis in activated lymphocytes and monocytes, and induction of neutrophil necrosis. Methods: Neutralisation of human TNF was assessed in the L929 bioassay using a range of concentrations of the anti-TNFs and a fixed concentration of TNF (100 pg/mL). Activity of the anti-TNFs at inhibiting LPS-driven IL-1β secretion by monocytes was assessed by incubating peripheral blood monocytes with various concentrations of the anti-TNF for 1 hour (hr) and then washing the cells. LPS was added for 4 hrs, the supernatants collected and the IL-1β level measured by ELISA. To assess induction of apoptosis, peripheral blood lymphocytes were activated for 2 days with 2 μg/mL CD3/CD28 and monocytes with 300 U/mL IL-4 and GMCSF for 3 days. The effect of the anti-TNFs on apoptosis was assessed by Annexin V staining using flow cytometry 24 hrs later. The effect of the anti-TNFs on neutrophil necrosis was determined by measuring myeloperoxidase release after 12 hrs. An isotype-matched control was used in all assays except the L929 bioassay. Results: IC90 neutralisation activity of the anti-TNFs in the L929 bioassay was 0.3 ng/mL for ETA, 4 ng/mL for GLM, 15 ng/mL for ADA, and 20 ng/mL for IFX, compared with 2.5 ng/mL for CZP. CZP was the most potent inhibitor of LPS-driven IL-1β secretion (IC50 ∼0.1 ng/mL), followed by GLM (20 ng/mL) and IFX (50 ng/mL). GLM, ADA, IFX and ETA induced apoptosis of monocytes and lymphocytes to a similar degree reaching a level of 23% and ∼40% at 100 μg/mL, respectively. CZP caused no increase in apoptosis above the levels seen with the isotype-matched control. In the neutrophil necrosis assay, ADA,IFX and GLM caused ∼70% necrosis at 100 μg/mL, and ETA 48%. CZP did not increase the level of necrosis above the level of the control. Conclusions: Bioactivity of the IgG1 molecules GLM, IFX and ADA in neutralising human TNF was inferior to that of CZP and ETA. CZP, the only PEGylated anti-TNF, had a different profile to the other anti-TNFs as it was the most potent at inhibiting LPS-driven IL-1β production by monocytes, did not induce apoptosis of activated monocytes and lymphocytes, and did not cause neutrophil necrosis. The clinical relevance of these in vitro effects is unknown. Nevertheless, these assays show interesting in vitro differences between the anti-TNFs. Disclosure statement: G.F. and A.N. are employees of UC

Current patch test results with the European baseline series and extensions to it from the 'European Surveillance System on Contact Allergy' network, 2007-2008

BACKGROUND: The pattern of contact sensitization to the supposedly most important allergens assembled in the baseline series differs between countries, presumably at least partly because of exposure differences. Objectives. To describe the prevalence of contact sensitization to allergens tested in consecutive patients in the years 2007 and 2008, and to discuss possible differences. METHODS: Data from the 39 departments in 11 European countries comprising the European Surveillance System on Contact Allergy network (www.essca-dc.org) in this period have been pooled and analysed according to common standards. RESULTS: Patch test results with the European baseline series, and country-specific or department-specific additions to it, obtained in 25 181 patients, showed marked international variation. Metals and fragrances are still the most frequent allergens across Europe. Some allergens tested nationally may be useful future additions to the European baseline series, for example methylisothiazolinone, whereas a few long-term components of the European baseline series, namely primin and clioquinol, no longer warrant routine testing. CONCLUSIONS: The present analysis points to 'excess' prevalences of specific contact sensitization in some countries, although interpretation must be cautious if only few, and possibly specialized, centres are representing one country. A comparison as presented may help to target in-depth research into possible causes of 'excess' exposure, and/or consideration of methodological issues, including modifications to the baseline series

Targeting DNA Damage Response and Replication Stress in Pancreatic Cancer

Background and aims: Continuing recalcitrance to therapy cements pancreatic cancer (PC) as the most lethal malignancy, which is set to become the second leading cause of cancer death in our society. The study aim was to investigate the association between DNA damage response (DDR), replication stress and novel therapeutic response in PC to develop a biomarker driven therapeutic strategy targeting DDR and replication stress in PC. Methods: We interrogated the transcriptome, genome, proteome and functional characteristics of 61 novel PC patient-derived cell lines to define novel therapeutic strategies targeting DDR and replication stress. Validation was done in patient derived xenografts and human PC organoids. Results: Patient-derived cell lines faithfully recapitulate the epithelial component of pancreatic tumors including previously described molecular subtypes. Biomarkers of DDR deficiency, including a novel signature of homologous recombination deficiency, co-segregates with response to platinum (P < 0.001) and PARP inhibitor therapy (P < 0.001) in vitro and in vivo. We generated a novel signature of replication stress with which predicts response to ATR (P < 0.018) and WEE1 inhibitor (P < 0.029) treatment in both cell lines and human PC organoids. Replication stress was enriched in the squamous subtype of PC (P < 0.001) but not associated with DDR deficiency. Conclusions: Replication stress and DDR deficiency are independent of each other, creating opportunities for therapy in DDR proficient PC, and post-platinum therapy

"This is the first time I've talked about this": considerations when conducting qualitative research interviews with adolescents and young adults with cancer

ualitative research is a useful and important way to describe and analyze the experiences of adolescents and young adults (AYAs) with cancer. However, undertaking qualitative research with AYAs living with, or beyond, a cancer diagnosis requires careful planning and a well-informed approach for participants with a high level of vulnerability and who are at a crucial stage developmentally. This article reflects on the challenges of including AYAs with cancer between the ages of 16 and 25 years in qualitative research. By drawing on each author's own experiences of engaging in qualitative research interviews with AYAs with cancer, the article provides suggestions regarding how best to manage challenges and yield valuable data describing their experiences. Insights that are shared between qualitative researchers in this field may assist in preparing for the challenges posed by conducting qualitative research with this group and may help researchers to manage this activity successfully

Genomic analyses identify molecular subtypes of pancreatic cancer

© 2016 Macmillan Publishers Limited. All rights reserved.Integrated genomic analysis of 456 pancreatic ductal adenocarcinomas identified 32 recurrently mutated genes that aggregate into 10 pathways: KRAS, TGF-β, WNT, NOTCH, ROBO/SLIT signalling, G1/S transition, SWI-SNF, chromatin modification, DNA repair and RNA processing. Expression analysis defined 4 subtypes: (1) squamous; (2) pancreatic progenitor; (3) immunogenic; and (4) aberrantly differentiated endocrine exocrine (ADEX) that correlate with histopathological characteristics. Squamous tumours are enriched for TP53 and KDM6A mutations, upregulation of the TP63ΔN transcriptional network, hypermethylation of pancreatic endodermal cell-fate determining genes and have a poor prognosis. Pancreatic progenitor tumours preferentially express genes involved in early pancreatic development (FOXA2/3, PDX1 and MNX1). ADEX tumours displayed upregulation of genes that regulate networks involved in KRAS activation, exocrine (NR5A2 and RBPJL), and endocrine differentiation (NEUROD1 and NKX2-2). Immunogenic tumours contained upregulated immune networks including pathways involved in acquired immune suppression. These data infer differences in the molecular evolution of pancreatic cancer subtypes and identify opportunities for therapeutic development