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A group of European FOCIS Centers of Excellence adapted panels of the Human Immunophenotyping Consortium (HIPC) for whole blood analysis. Using four core panels [T/regulatory T cell/B/natural killer (T/Treg/B/NK) and myeloid cells] the main leukocyte populations were analyzed in a clinical-diagnostic setting in a harmonized manner across different platforms. As a first step, the consortium presents here the absolute and relative frequencies of the leukocyte subpopulations in the peripheral blood of more than 300 healthy volunteers across six different European centers.

The Basophil Activation Test (BAT) is a valuable allergy diagnostic tool but is time-consuming and requires skilled personnel, which has limited its clinical use. We therefore developed and clinically tested a microfluidic immunoaffinity BAT (miBAT) technique where we captured basophils directly from whole blood followed by in vitro activation and quantification of activation markers. For the first time basophils captured from whole blood, from both allergic patients and healthy donors, have been activated using allergens., QC 20210330

Tissue-resident memory T (T-RM) cells are CD8(+) T lymphocytes that reside in the tissues, including tumours. This T cell subset possesses a magnitude of cytotoxicity, but its epigenetic regulation has not been studied. Here, we investigate the impact of perforin DNA methylation in T-RM cells and correlate it with their functional potential. Fifty-three urothelial urinary bladder cancer (UBC) patients were recruited prospectively. The DNA methylation status of the perforin gene (PRF1) locus in T-RM cells was investigated by pyrosequencing. Flow cytometry with ViSNE analysis and in-vitro stimulation were used to evaluate T-RM cell phenotypes. We discovered that tumour T-RM cells have low DNA methylation in the PRF1 locus (329% methylation), which corresponds to increased numbers of perforin-expressing T-RM cells. Surprisingly, programmed cell death 1 (PD-1) expression is high in tumour T-RM cells, suggesting exhaustion. Following interleukin-15 and T cell receptor stimulation, perforin and T-bet expressions are enhanced, indicating that T-RM cells from tumours are not terminally exhausted. Moreover, a high number of T-RM cells infiltrating the tumours corresponds to lower tumour stage in patients. In conclusion, T-RM cells from UBC tumours are epigenetically cytotoxic with signs of exhaustion. This finding identifies T-RM cells as potential new targets for cancer immunotherapy.

Patients with advanced urothelial urinary bladder cancer (UBC) have a pessimistic prognosis, although modern urology can offer preoperative chemotherapy followed by radical surgery. Attempts to find additional, other than adjuvant cytotoxic treatment options need to be explored. The field of immune therapy may be of particular value in UBC since immunotherapy of superficial UBC using an unspecific immunotherapy modality, namely BCG, has been so successful. In addition, UBC is one of the most genetically unstable cancers carrying many neo-antigens as potential targets for T lymphocyte recognition. A tempting option aiming at highly individualized treatment could be autologous cell therapy based on the concepts of sentinel node detection. The sentinel nodes, representing the foremost arena for the struggle between the immune system of the patient and the tumor assault, can offer high quantities of highly specialized T lymphocytes. These T cells have evolved in response to a number of specific tumor antigens reaching the tumor draining sentinel nodes en route. Adoptive immunotherapy within this framework entails possibilities of harvesting tumor specific T cells from the sentinel nodes and allowing them to expand in vitro. Expanded and strengthened T cells can then be reinfused as a safe and non-toxic adjuvant immunotherapy. The first pilot trials have been published utilizing this method in two solid cancers, colon and UBC. Results demonstrate objective responses in some patients without any negative side effects. The future challenge is to increase the translational research in this specific field, to further improve immunobiological techniques, to optimize patient selection and to perform larger randomized controlled clinical trials.

The sentinel node is the first lymph node to receive lymphatic drainage from a tumour and is usually the first site of metastases. Today, the sentinel node is used for tumour staging. Here, we focus on its immunological role and investigate lymphocytic function in sentinel nodes, identified intraoperatively by peritumoural dye injection, from 15 patients with colon cancer. Tumour infiltrating lymphocytes, sentinel and nonsentinel lymph node cells and peripheral blood leukocytes were studied by flow cytometry, proliferation assays and interferon-gamma secretion after activation with autologous tumour homogenate. Whereas tumour-infiltrating lymphocytes were nonresponsive in the proliferation assays, lymphocytes from sentinel nodes proliferated dose dependently and secreted interferon-gamma upon stimulation with tumour homogenate. The responses were of varying magnitude and tended to be weaker in metastatic sentinel nodes. Sentinel node lymphocytes represents an enriched source of tumour reactive lymphocytes, and may be useful in future trials of adoptive immunotherapy., Marits, P Karlsson, M Dahl, K Larsson, P Wanders, A Thorn, M Winqvist, O eng Research Support, Non-U.S. Gov't England 2006/04/28 09:00 Br J Cancer. 2006 May 22;94(10):1478-84.