Higher Number of EBI3 Cells in Mucosal Chronic Hyperplastic Candidiasis May Serve to Regulate IL-17-Producing Cells.

Previous research into the inflammatory cell infiltrate of chronic hyperplastic candidosis (CHC) determined that the immune response is primarily composed of T cells, the majority of which are T helper (CD4+) cells. This present investigation used immunohistochemistry to further delineate the inflammatory cell infiltrate in CHC. Cells profiled were those expressing IL-17A cytokine, EBI3 and IL-12A subunits of the IL-35 cytokine, and FoxP3+ cells. Squamous cell papilloma (with Candida infection) and oral lichen planus tissues served as comparative controls to understand the local immune responses to Candida infection. The results demonstrated that Candida-induced inflammation and immune regulation co-exist in the oral mucosa of CHC and that high prevalence of cells expressing the EBI3 cytokine subunit may play an important role in this regulation. This balance between inflammation and immune tolerance toward invading Candida in the oral mucosa may be critical in determining progress of infection.

Immunohistochemical Expression Patterns of Inflammatory Cells Involved in Chronic Hyperplastic Candidosis.

The profile of the inflammatory cell infiltrate in chronic hyperplastic candidosis (CHC) was determined in oral mucosal biopsies by immunohistochemistry. One tonsillar tissue section was included as an immunohistochemistry control, whilst squamous papilloma (n = 4) with secondary Candida infection was used as Candida controls. Oral lichen planus tissues (n = 10) provided negative controls for Candida presence, as well as positive controls for inflammation. Immunohistochemistry employed antibodies specific for CD3+ (T lymphocytes), CD4+ (T helper cells), CD8+ (cytotoxic T cells), and CD20+ (B lymphocytes). Manual counting of stained cells from digitised images determined the proportion of each cell type relative to the total number of cells, and these were assessed in the mucosa, the epithelium, and the lamina propria. The mean proportion of CD3+ cells was significantly higher than CD20+ cells in all tissue types. For CHC, the mean proportion of CD3+ cells in entire tissues was 15.6%, with the highest proportion in the lamina propria (32.6%) compared with the epithelium (3.9%). CD20+ cells were in much lower proportions (1.8%) in CHC, with the highest proportion (3.6%) in the lamina propria. T lymphocytes were predominately CD4+ cells (9.0%) compared with CD8+ cells (4.4%). CD4+ cells were most prevalent in the lamina propria (23.1%) compared with the epithelium (mean = 3.2%). From these results, it was concluded that the immune response invoked by Candida in CHC is primarily driven by the T helper cells.

Pre-trial inter-laboratory analytical validation of the FOCUS4 personalised therapy trial.

INTRODUCTION: Molecular characterisation of tumours is increasing personalisation of cancer therapy, tailored to an individual and their cancer. FOCUS4 is a molecularly stratified clinical trial for patients with advanced colorectal cancer. During an initial 16-week period of standard first-line chemotherapy, tumour tissue will undergo several molecular assays, with the results used for cohort allocation, then randomisation. Laboratories in Leeds and Cardiff will perform the molecular testing. The results of a rigorous pre-trial inter-laboratory analytical validation are presented and discussed. METHODS: Wales Cancer Bank supplied FFPE tumour blocks from 97 mCRC patients with consent for use in further research. Both laboratories processed each sample according to an agreed definitive FOCUS4 laboratory protocol, reporting results directly to the MRC Trial Management Group for independent cross-referencing. RESULTS: Pyrosequencing analysis of mutation status at KRAS codons12/13/61/146, NRAS codons12/13/61, BRAF codon600 and PIK3CA codons542/545/546/1047, generated highly concordant results. Two samples gave discrepant results; in one a PIK3CA mutation was detected only in Leeds, and in the other, a PIK3CA mutation was only detected in Cardiff. pTEN and mismatch repair (MMR) protein expression was assessed by immunohistochemistry (IHC) resulting in 6/97 discordant results for pTEN and 5/388 for MMR, resolved upon joint review. Tumour heterogeneity was likely responsible for pyrosequencing discrepancies. The presence of signet-ring cells, necrosis, mucin, edge-effects and over-counterstaining influenced IHC discrepancies. CONCLUSIONS: Pre-trial assay analytical validation is essential to ensure appropriate selection of patients for targeted therapies. This is feasible for both mutation testing and immunohistochemical assays and must be built into the workup of such trials. TRIAL REGISTRATION NUMBER: ISRCTN90061564.