Analysis of head and neck carcinoma progression reveals novel and relevant stage-specific changes associated with immortalisation and malignancy.

We report changes in the genomic landscape in the development of head and neck squamous cell carcinomas HNSCC from potentially premalignant lesions (PPOLS) to malignancy and lymph node metastases. Likely pathological mutations predominantly involved a relatively small set of genes reported previously (TP53, KMT2D, CDKN2A, PIK3CA, NOTCH1 and FAT1) but also other predicted cancer drivers (MGA, PABPC3, NR4A2, NCOR1 and MACF1). Notably, all these mutations arise early and are present in PPOLs. The most frequent genetic changes, which follow acquisition of immortality and loss of senescence, are of consistent somatic copy number alterations (SCNAs) involving chromosomal regions enriched for genes in known and previously unreported cancer-related pathways. We mapped the evolution of SCNAs in HNSCC progression. One of the earliest SCNAs involved deletions of CSMD1 (8p23.2). CSMD1 deletions or promoter hypermethylation were present in all of the immortal PPOLs and occurred at high frequency in the immortal HNSCC cell lines. Modulation of CSMD1 in cell lines revealed significant suppression of proliferation and invasion by forced expression, and significant stimulation of invasion by knockdown of expression. Known cancer drivers NOTCH1, PPP6C, RAC1, EIF4G1, PIK3CA showed significant increase in frequency of SCNA in transition from PPOLs to HNSCC that correlated with their expression. In the later stages of progression, HNSCC with and without nodal metastases showed some clear differences including high copy number gains of CCND1, hsa-miR-548k and TP63 in the metastases group.

Depleted polymorphonuclear leukocytes in human metastatic liver reflect an altered immune microenvironment associated with recurrent metastasis.

BACKGROUND: Hepatic immunity, normally protective against neoplasia, is subverted in colorectal liver metastasis (CRLM). Here, we compare the inflammatory microenvironment of CRLM-bearing liver tissue to donor liver. METHODS: Twenty-five patients undergoing resection for CRLM were recruited, 13 of whom developed intrahepatic recurrence within 18 months. Biopsies were obtained from tumour and normal liver tissue adjacent to and distal from, the tumour. Donor liver biopsies were obtained during transplantation. Biopsies were cultured and conditioned media (CM) screened for 102 inflammatory mediators. Twelve of these were validated by Luminex assay. Transwell assays measured cancer cell chemotaxis. Polymorphonuclear leukocytes (PMN) and lymphocytes were quantified in H&E sections. RESULTS: Fewer periportal tissue-resident PMN were present in metastatic liver compared to donor liver. Patients with the fewest PMN in liver tissue distal to their tumour had a shorter time to intrahepatic recurrence (P < 0.001). IL-6, CXCL1, CXCL5, G-CSF, GM-CSF, VEGF, LIF, and CCL3 were higher in liver-bearing CRLM compared to donor tissue. Consequently, cancer cells migrated equally towards CM of all regions of metastatic liver but not towards donor liver CM. CONCLUSIONS: The local inflammatory environment may affect both immune cell infiltration and cancer cell migration contributing to recurrence following resection for CRLM.

Human Tumor-Infiltrating Myeloid Cells: Phenotypic and Functional Diversity.

Our current understanding of human tumor-resident myeloid cells is, for the most part, based on a large body of work in murine models or studies enumerating myeloid cells in patient tumor samples using immunohistochemistry (IHC). This has led to the establishment of the theory that, by and large, tumor-resident myeloid cells are either "protumor" M2 macrophages or myeloid-derived suppressor cells (MDSC). This concept has accelerated our understanding of myeloid cells in tumor progression and enabled the elucidation of many key regulatory mechanisms involved in cell recruitment, polarization, and activation. On the other hand, this paradigm does not embrace the complexity of the tumor-resident myeloid cell phenotype (IHC can only measure 1 or 2 markers per sample) and their possible divergent function in the hostile tumor microenvironment. Here, we examine the criteria that define human tumor-infiltrating myeloid cell subsets and provide a comprehensive and critical review of human myeloid cell nomenclature in cancer. We also highlight new evidence characterizing their contribution to cancer pathogenesis based on evidence derived from clinical studies drawing comparisons with murine studies where necessary. We then review the mechanisms in which myeloid cells are regulated by tumors in humans and how these are being targeted therapeutically.

Adaptation of a Cell-Based High Content Screening System for the In-Depth Analysis of Celiac Biopsy Tissue.

The IN Cell Analyzer 1000 possesses several distinguishing features that make it a valuable tool in research today. This fully automated high content screening (HCS) system introduced quantitative fluorescent microscopy with computerized image analysis for use in cell-based analysis. Previous studies have focused on live cell assays, where it has proven to be a powerful and robust method capable of providing reproducible, quantitative data. Using HCS as a tool to investigate antigen expression in duodenal biopsies, we developed a novel approach to tissue positioning and mapping. We adapted IN Cell Analyzer 1000's image acquisition and analysis software for the investigation of tissue transglutaminase (tTG) and smooth muscle alpha-actin (SM α-actin) staining in paraffin-embedded duodenal tissue sections from celiac patients and healthy controls. These innovations allowed a quantitative analysis of cellular structure and protein expression. The results from routine biopsy material indicated the intensity of protein expression was altered in celiac disease compared to normal biopsy material.

Heightened Expression of CD39 by Regulatory T Lymphocytes Is Associated with Therapeutic Remission in Inflammatory Bowel Disease.

BACKGROUND: To evaluate whether changes in expression of CD39 by regulatory T lymphocytes (Treg) impact treatment response in inflammatory bowel disease. To then define the biological role of expression of CD39 on Treg in an animal model of colitis. METHODS: A prospective study of consecutive patients commencing anti-tumor necrosis factor therapy with infliximab (IFX) or adalimumab (ADA), who were then followed for 12 months. Treatment responses were defined both symptomatically and by endoscopy showing mucosal healing. Peripheral blood Tregs were quantified by flow cytometry. Functional importance of CD39 expression by Treg was determined in an adoptive T-cell transfer model of colitis. RESULTS: Forty-seven patients (ulcerative colitis, n = 22; Crohn's disease, n = 25) were recruited; 16 patients were complete responders and 13 nonresponders to anti-tumor necrosis factor. CD39 expression by Treg was lower in active inflammatory bowel disease and increased significantly after treatment in responders (CD39Treg/total Treg; 8% at baseline to 22.5% at late time point, P < 0.001). Responders were more likely to have therapeutic drug levels and in multivariate analysis therapeutic drug levels were associated with higher expression of CD39 by FoxP3 Treg and lower frequencies of interleukin 17A expressing cells. Tregs with genetic deletion of CD39 exhibit decrements in potential to suppress intestinal inflammation in a murine (CD45RB) T-cell transfer model of colitis in vivo, when compared with wild-type Treg. CONCLUSIONS: Increased expression of CD39 by peripheral blood Treg is observed in the setting of clinical and endoscopic remission in inflammatory bowel disease. Deficiency of CD39 expression by Treg can be linked to inability to suppress experimental colitis.

Targeting EGFR in metastatic colorectal cancer beyond the limitations of KRAS status: alternative biomarkers and therapeutic strategies.

Patients with metastatic colorectal cancer have a very poor prognosis. Incorporation of targeted molecular therapies, such as the anti-EGFR receptor monoclonal antibodies cetuximab and panitumumab, into treatment regimens has improved outcomes for patients with wild-type RAS tumors. Yet, response rates remain low and overall survival times are short. Increased understanding of oncogenic signaling pathways within the tumor, and how these are regulated by the inflammatory tumor microenvironment, is a priority to facilitate the development of biomarkers to better guide the use of existing therapies and to develop new ones. Here, we review recent preclinical and clinical progress in the development of biomarkers for predicting response to anti-EGFR therapy in metastatic colorectal cancer.

Complication of etanercept treatment for rheumatoid arthritis--purulent pericarditis caused by a commensal organism.

The patient presented with increasing fatigue and dyspnoea. The patient had medical history of rheumatoid arthritis for which she had been taking methotrexate for the past 15 years and etanercept for the past 6 years. Initial diagnosis was cardiac failure but further investigation by echocardiogram revealed a large pericardial effusion. Empirical piperacillin-tazobactam was started due to moderately raised inflammatory markers. Four hundred millilitre of frank pus was aspirated from the pericardial sac and antimicrobial treatment was changed to meropenem. Gram positive cocci were seen in the initial Gram stain, but conventional cultures remained negative. However, 16S ribosomal RNA gene sequencing of the pus sample detected the presence of Parvimonas micra genome. Reaccumulation of the effusion required further drainage where again P micra was detected by 16S ribosomal RNA gene sequencing. Two weeks of meropenem was completed followed by treatment with benzylpenicillin and metronidazole.

Randomized controlled trial of a prompt list to help advanced cancer patients and their caregivers to ask questions about prognosis and end-of-life care.

PURPOSE: To determine whether provision of a question prompt list (QPL) influences advanced cancer patients'/caregivers' questions and discussion of topics relevant to end-of-life care during consultations with a palliative care (PC) physician. PATIENTS AND METHODS: This randomized controlled trial included patients randomly assigned to standard consultation or provision of QPL before consultation, with endorsement of the QPL by the physician during the consultation. Consecutive eligible patients with advanced cancer referred to 15 PC physicians from nine Australian PC services were invited to participate. Consultations were audiotaped, transcribed, and analyzed by blinded coders; patients completed questionnaires before, within 24 hours, and 3 weeks after the consultation. RESULTS: A total of 174 patients participated (92 QPL, 82 control). Compared with controls, QPL patients and caregivers asked twice as many questions (for patients, ratio, 2.3; 95% CI, 1.7 to 3.2; P < .0001), and patients discussed 23% more issues covered by the QPL (95% CI, 11% to 37%; P < .0001). QPL patients asked more prognostic questions (ratio, 2.3; 95% CI, 1.3 to 4.0; P = .004) and discussed more prognostic (ratio, 1.43; 95% CI, 1.1 to 1.8, P = .003) and end-of-life issues (30% v 10%; P = .001). Fewer QPL patients had unmet information needs about the future (21 = 4.14; P = .04), which was the area of greatest unmet information need. QPL consultations (average, 38 minutes) were longer (P = .002) than controls (average, 31 minutes). No differences between groups were observed in anxiety or patient/physician satisfaction. CONCLUSION: Providing a QPL and physician endorsement of its use assists terminally ill cancer patients and their caregivers to ask questions and promotes discussion about prognosis and end-of-life issues, without creating patient anxiety or impairing satisfaction.