0000000000198956
AUTHOR
Sjoerd H. Van Der Burg
Hierarchical modeling for rare event detection and cell subset alignment across flow cytometry samples.
Flow cytometry is the prototypical assay for multi-parameter single cell analysis, and is essential in vaccine and biomarker research for the enumeration of antigen-specific lymphocytes that are often found in extremely low frequencies (0.1% or less). Standard analysis of flow cytometry data relies on visual identification of cell subsets by experts, a process that is subjective and often difficult to reproduce. An alternative and more objective approach is the use of statistical models to identify cell subsets of interest in an automated fashion. Two specific challenges for automated analysis are to detect extremely low frequency event subsets without biasing the estimate by pre-processing…
Managing Multi-center Flow Cytometry Data for Immune Monitoring.
With the recent results of promising cancer vaccines and immunotherapy 1 – 5 , immune monitoring has become increasingly relevant for measuring treatment-induced effects on T cells, and an essential tool for shedding light on the mechanisms responsible for a successful treatment. Flow cytometry is the canonical multi-parameter assay for the fine characterization of single cells in solution, and is ubiquitously used in pre-clinical tumor immunology and in cancer immunotherapy trials. Current state-of-the-art polychromatic flow cytometry involves multi-step, multi-reagent assays followed by sample acquisition on sophisticated instruments capable of capturing up to 20 parameters per cell at a…
Consensus nomenclature for CD8(+) T cell phenotypes in cancer
International audience; Whereas preclinical investigations and clinical studies have established that CD8+ T cells can profoundly affect cancer progression, the underlying mechanisms are still elusive. Challenging the prevalent view that the beneficial effect of CD8+ T cells in cancer is solely attributable to their cytotoxic activity, several reports have indicated that the ability of CD8+ T cells to promote tumor regression is dependent on their cytokine secretion profile and their ability to self-renew. Evidence has also shown that the tumor microenvironment can disarm CD8+ T cell immunity, leading to the emergence of dysfunctional CD8+ T cells. The existence of different types of CD8+ T…
Flow Cytometry in Cancer Immunotherapy: Applications, Quality Assurance, and Future
Cancer immunotherapy seeks to elicit or augment the antitumor immune response in a patient in order to enlist the help of the patient’s own immune system for tumor control. In this context, immune monitoring provides evidence of immunogenicity, guides the choice and dosage of antigens, assesses the effects of immune modulators and therapy combinations, and has the potential to reveal early biomarkers of clinical efficacy. In view of their role in the anticancer immune response, the quantity and quality of tumor antigen-specific effector CD4+ and CD8+ T cells are of particular interest, and characterization of regulatory T cells and myeloid-derived suppressor cells is increasingly relevant. …
Classification of current anticancer immunotherapies.
© 2014. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Development of an RNA-based kit for easy generation of TCR-engineered lymphocytes to control T-cell assay performance.
Cell-based assays to monitor antigen-specific T-cell responses are characterized by their high complexity and should be conducted under controlled conditions to lower multiple possible sources of assay variation. However, the lack of standard reagents makes it difficult to directly compare results generated in one lab over time and across institutions. Therefore TCR-engineered reference samples (TERS) that contain a defined number of antigen-specific T cells and continuously deliver stable results are urgently needed. We successfully established a simple and robust TERS technology that constitutes a useful tool to overcome this issue for commonly used T-cell immuno-assays. To enable users t…
Cryopreservation of MHC Multimers: Recommendations for Quality Assurance in Detection of Antigen Specific T Cells
Fluorescence-labeled peptide-MHC class I multimers serve as ideal tools for the detection of antigen-specific T cells by flow cytometry, enabling functional and phenotypical characterization of specific T cells at the single cell level. While this technique offers a number of unique advantages, MHC multimer reagents can be difficult to handle in terms of stability and quality assurance. The stability of a given fluorescence-labeled MHC multimer complex depends on both the stability of the peptide-MHC complex itself and the stability of the fluorochrome. Consequently, stability is difficult to predict and long-term storage is generally not recommended. We investigated here the possibility of…
Generation of TCR-Engineered T Cells and Their Use To Control the Performance of T Cell Assays
Abstract The systematic assessment of the human immune system bears huge potential to guide rational development of novel immunotherapies and clinical decision making. Multiple assays to monitor the quantity, phenotype, and function of Ag-specific T cells are commonly used to unravel patients’ immune signatures in various disease settings and during therapeutic interventions. When compared with tests measuring soluble analytes, cellular immune assays have a higher variation, which is a major technical factor limiting their broad adoption in clinical immunology. The key solution may arise from continuous control of assay performance using TCR-engineered reference samples. We developed a simp…
Immunoguiding, the Final Frontier in the Immunotherapy of Cancer
T cells play an important role in cancer. This notion is strongly supported by an enormous number of trials on the clinical impact of tumor-infiltrating T cells and studies consequently showing that therapeutic interventions which are based on transfer, activation and expansion, or de-blocking of tumor-specific T cells, which have met with clinical success. To optimally profit from the flow of newly developed immune-based therapeutics aiming to reinforce the systemic and local tumor-specific T-cell response, it will be required to identify biomarkers that provide the rationale to use a particular therapy, that measure the effect of intervention, and that can predict the outcome of a therape…
Recommendations from the iSBTc-SITC/FDA/NCI Workshop on Immunotherapy Biomarkers
Abstract Purpose: To facilitate development of innovative immunotherapy approaches, especially for treatment concepts exploiting the potential benefits of personalized therapy, there is a need to develop and validate tools to identify patients who can benefit from immunotherapy. Despite substantial effort, we do not yet know which parameters of antitumor immunity to measure and which assays are optimal for those measurements. Experimental Design: The iSBTc-SITC (International Society for Biological Therapy of Cancer-Society for Immunotherapy of Cancer), FDA (Food and Drug Administration), and NCI (National Cancer Institute) partnered to address these issues for immunotherapy of cancer. Here…
Harmonization of Immune Biomarker Assays for Clinical Studies
Assays that measure a patient's immune response play an increasingly important role in the development of immunotherapies. The inherent complexity of these assays and independent protocol development between laboratories result in high data variability and poor reproducibility. Quality control through harmonization--based on integration of laboratory-specific protocols with standard operating procedures and assay performance benchmarks--is one way to overcome these limitations. Harmonization guidelines can be widely implemented to address assay performance variables. This process enables objective interpretation and comparison of data across clinical trial sites and also facilitates the ide…