Abstract
Monitoring immune responses to solid cancers may be a better prognostic tool than conventional staging criteria, and it can also serve as an important criterion for the selection of individualized therapy. Multiparametric phenotyping by mass cytometry extended possibilities for immunoprofiling. However, careful optimization of each step of such method is necessary for obtaining reliable results. Also, with respect to procedure length and costs, sample preparation, staining, and storage should be optimized. Here, we designed a panel of 31 antibodies which allows for identification of several subpopulations of lymphoid and myeloid cells in a solid tumor and peripheral blood simultaneously. For sample preparation, disaggregation of tumor tissue with two different collagenases combined with DNase I was compared, and removal of dead or tumor cells by magnetic separation was evaluated. Two possible procedures of barcoding for single-tube staining of several samples were examined. While the palladium-based barcoding affected the stability of several antigens, the staining with two differently labeled CD45 antibodies was suitable for cells isolated from a patient's blood and tumor. The storage of samples in the intercalation solution for up to two weeks did not influence results of the analysis, which allowed the measurement of samples collected within this interval on the same day. This procedure optimized on samples from patients with head and neck squamous cell carcinoma enabled identification of various immune cells including rare subpopulations.
Highlights
Cancer generation and progression are critically affected by the host immune system
Cancer immunotherapy was revitalized in recent years, and its clinical use is progressively increasing, especially after the US Food and Drug Administration (FDA) approval of the monoclonal antibodies ipilimumab, in 2011, and nivolumab and pembrolizumab, in 2014, targeting the immune checkpoints cytotoxic T lymphocyte-associated antigen 4 (CTLA-4; CD152) and programmed cell death protein 1 (PD-1; CD279), respectively
The development of cancer immunotherapy is associated with the detection of immune reactions, cells, and markers that enables the monitoring of the effect of therapy but is important for prognosis and prediction of treatment success because only a minority of patients is responsive to immunotherapy
Summary
Cancer immunotherapy was revitalized in recent years, and its clinical use is progressively increasing, especially after the US Food and Drug Administration (FDA) approval of the monoclonal antibodies ipilimumab, in 2011, and nivolumab and pembrolizumab, in 2014, targeting the immune checkpoints cytotoxic T lymphocyte-associated antigen 4 (CTLA-4; CD152) and programmed cell death protein 1 (PD-1; CD279), respectively. Besides these antibodies, other promising immunotherapeutic approaches against malignant diseases—adoptive transfer of modified T cells, cancer vaccines, and chimeric monoclonal antibodies called bispecific T cell engager (BiTE)—are available [1, 2].
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