TBNK cells: What are They and What Can They Tell Us?

Only about 1% of the blood is made up white blood cells, or leukocytes, yet they have critical functions as part of the immune system and can provide a wealth of information in research studies. Amongst these many leukocyte cell types, lymphocytes – which includes T cells, B cells and NK cells – all originate from a common lymphoid progenitor (CLP) in the hematopoietic stem cell lineage. Despite sharing this common origin, these precious cells have specialized functions working together as parts of the innate and adaptive immune responses. 

The immune system has been implicated to have some kind of role in almost every disease. Identifying and characterizing the many subtypes of TBNK cells and other immune factors can identify key biomarkers related to a specific disease that can be then used in clinical development programs as secondary or exploratory endpoints to evaluate immune-related adverse events or efficacy of a certain drug. Understanding the kinetics of immune cell subtypes and various cytokines within natural disease progression can also help reveal potential targets for immunomodulation in therapeutic development programs.(1) Additionally, individuals with the same disease can show differences in certain activated cell markers and cytokines indicative of distinct disease phenotypes, enabling patient stratifications in trials and more personalized therapeutics.  

Overall, activation of the immune response begins with the innate immune system recognizing a foreign invader or cancerous cell, followed by cytokine release, complement activation and acute inflammation. This speedy reaction triggers a more targeted response from the adaptive immune system which relies on antigen-presenting cells to activate specific T helper cells. These activated T cells coordinate an antigen-specific immune response involving humoral immunity from B cells and cell-mediated immunity from other T cells. Although this is a largely simplified overview, many other branches of cell types and factors act in synergy to drive elaborate signalling cascades for proper immune function. 

Development and function of T cells 

T cells start differentiating in the bone marrow and migrate to the thymus gland to mature into thymocytes. As a whole, T cells attack foreign cells, cancer cells and viral-infected cells by essentially acting as detectives to scope out pathogens through a unique T cell receptor that develops during maturation. T cell progenitor cells can differentiate into three main T cell subtypes: T helper 1 lymphocytes (Th1 cells), T helper 2 lymphocytes (Th2 cells) and cytotoxic T (Tc) cells. TH1 cells mediate inflammatory reactions and immunity to intracellular microbes. Th2 cells primarily help B lymphocytes produce antibodies, but also act to downregulate the inflammatory activities of Th1 cells. Cytotoxic T cells clear virus-infected cells and tumor cells typically by inducing apoptosis. Many different subsets of T cells respond to infection and this response further drives the formation of memory T cells that recognize specific antigens in preparation for subsequent infection. T cells release various cytokines, such as TNF-α, TNF-ß, interferon-γ, IL-2, IL-4, IL-6 and IL-8, that trigger downstream activation of other immune cells and responses. Cytokines, like IL-2 for example, are upregulated in T cells upon antigenic or mitogenic stimulation, which can lead to clonal expansion of T cells. Many of these cytokines also function to activate and recruit other cell types such as B cells, NK cells, macrophages and neutrophils.(2) Antigen-presenting cells, such as B cells, macrophages and dendritic cells, as well as natural killer cells can also further activate T cell functions, further demonstrating the cross-talk amongst these various cell types. The intricacies of both the innate and adaptive immune responses demonstrate just how complex these pathways are and also how many of these factors have redundant functions, acting as fail-safe mechanisms to ultimately maintain proper immune regulation. By characterizing these relationships in both normal and disease states, targets for drug development can be identified to harness or repair the functions of these various cell types to fight disease. 

Development and function of B cells

B cell progenitor cells in the bone marrow migrate to the lymph nodes to differentiate into immature B cells. They make antibodies that help your body fight infections and play a critical role in establishing the adaptive immune response to specific antigens. Exposure to an antigen differentiates naïve B cells into either plasma or memory cells. Plasma cells produce and release specific antibodies into the blood that recognize a specific antigen. These free-floating antibodies will bind to cell or pathogen based antigens and trigger a signalling cascade that recruits many different cell types to attack that cell and similarly infected cells. Memory cells are long-lived lymphocytes that carry the code to produce the particular antibody, but do not directly produce antibodies, unless activated to differentiate into plasma cells. Activation of these memory cells typically occurs when an antigen is reintroduced, long after the initial exposure to it. This is much faster than the differentiation process from naïve B cells to plasma/memory cells and is a way for the immune system to remember previous pathogenic encounters without producing too many unnecessary antibodies in circulation. Among the cytokines secreted by B cells is IL-6, the primary inducer of fever, hormones, and T and B cell expansion upon injury and infection.(2) Over time, plasma cells and the antibodies they produce decrease in quantity, leaving the memory cells to take over antibody production if the antigen reappears.  

Development and function of NK cells

Natural killer (NK) cells produce substances that kill tumor cells or viral-infected cells as part of the innate immune system. They are like poison darts targeting infected cells which can also recruit other factors to finish the job. They comprise a minor fraction of the total lymphocyte population, yet they reside throughout lymphoid and non-lymphoid tissues.(3). Unlike most other immune cells, NK cells can function without requiring major histocompatibility complex class I  (MHC-I) antigens or antibodies to perform their cell-killing roles, which allows them to respond much faster against viral infection or cancerous cells. NK cells primarily rely on markers that indicate cells in ‘distress’, but they can also detect antibody-coated target cells and function in the antibody-dependent cell cytotoxicity pathway as well. Among NK cells secreted cytokines is IFN-γ, which primarily functions in the antiviral response, the activation of macrophages and in clearing intracellular mycobacteria.(2) IFN-γ, which is also secreted by activated Th1 T cells, can stimulate macrophages, increase antigen processing and expression of MHC molecules, promote Ig class switching, and control proliferation of transformed cells.   

How are they used in research

Profiling of TBNK cells forms the bedrock for immune-related research, providing valuable information on patient response to therapies and vaccines. Flow cytometry represents a workhorse method for classifying and sorting specific cell subtypes using fluorescently labeled antibodies to extracellular and intracellular markers.(4) Measuring cytokine levels through various assays informs on immune function and activation. For example, TNF-α represents a proinflammatory biomarker that can be tracked in studies of autoimmune disease progression and therapy evaluation.(2) 

In vaccine development, characterizing the neutralizing or binding activity of antibodies and the various B and T cell subsets that drive cell-mediated immunity informs on the strength and durability of the immune response after immunization. This is critical, particularly in early clinical development stages, to inform the dose selection, vaccination schedule and potential for efficacy against disease that will need to be evaluated later in large-scale Phase 3 trials. These immunogenicity endpoints also can be harnessed in immunobridging studies that can expand the indication of an already approved vaccine in populations lacking trial participants, such as pediatrics, a practice that gained traction during the Covid pandemic. 

Research to characterize the unique cell subtypes in various cancers has proven fruitful in the development of many targeted cancer immunotherapies that function to block immune checkpoints, boost T cell function or deliver monoclonal antibodies to target specific antigens on cancer cells.(5) On the other hand, in autoimmune diseases, where certain cells of the immune system attack ‘self’-identifying cells, immunotherapies exist to repress or block this activity to control the disease.(5) Even with these successes, many cancers and patients with autoimmune diseases remain resistant to immunotherapy or ultimately develop resistance following treatment. Further research into understanding the full repertoire of immune cell subsets activated during cancer development will help bridge the existing gap in potential therapeutic avenues.    

Overall, understanding the role of these various cell types in infection responses, cancer development and autoimmunity can yield valuable insights into how we can modulate and target the immune system to more effectively clear infections, eliminate or control tumor and cancer progression, and protect the body from being attacked by its own immune cells.

References

  1. Flower DR. The Immune System as Drug Target. Immunology and Immunogenetics Insights. (2013). https://doi:10.4137/III.S12145
  2. Cameron MJ, Kelvin DJ. Cytokines, Chemokines and Their Receptors. In: Madame Curie Bioscience Database [Internet]. Austin (TX): Landes Bioscience; 2000-2013. Available from: https://www.ncbi.nlm.nih.gov/books/NBK6294/
  3. Vivier E, Tomasello E, Baratin M, Walzer T, Ugolini S. Functions of natural killer cells. Nat Immunol 9, 503–510 (2008). https://doi.org/10.1038/ni1582
  4. Mousset CM, Hobo W, Woestenenk R, Preijers F, Dolstra H, van der Waart AB. Comprehensive Phenotyping of T Cells Using Flow Cytometry. Cytometry. 2019 Jun;95(6):647–54. 
  5. Melief CJM. Special Review: The future of Immunotherapy. Immunotherapy Advances. 2021 Jan 1;1(1):ltaa005.