The relationship between the tumor population and the host organism can be seen as a competition of two genetically distinct entities that compete for resources and space for growth in accordance with Richard Dawkins' theory of the selfish gene. The host organism tries to recognize and eliminate malignant cells, or, at a later stage, at least to regulate their number. The tumor population, on the other hand, seeks to increase in size, gain access to resources, and expand to other appropriate sites in the host organism. To escape the host's immune response, tumor cells use a variety of passive and active measures that increase the likelihood of their survival. One of the clinically significant interactions between tumor and immunity is the functional impairment of T-lymphocytes, resulting in a failure of anti-tumor immunity as well as reduced resistance to bacterial and viral pathogens. This involvement is caused by a complex of events, mainly by the interaction of the PD-1 receptor (CD279), expressed mainly on B- and T-lymphocytes with its ligands PD-L1 (CD274, B7-H1) and PD-L2 (CD273, B7-DC), the so-called programmed cell death pathway.
PD-1 and its ligands are, due to their location on the cell surface, an ideal target for analysis by multiparametric flow cytometry, while suitable antibodies are already available in the form of conjugates for multicolor assays.
Within the programmed cell death pathway, the interaction between PD-1 and PD-L1 and L2 particularly affects T-lymphocytes, in which they cause a phenomenon called functional exhausting, by attenuating TCR-mediated T-cell proliferation by negative costimulation control of activation of T-lymphocytes during the presentation of antigens by dendritic cells. In this way, they effectively suppress the host's immunity and thus allow tumor cells to escape cytotoxic response. The effect of PD-1 on NK cells, i.e. NK exhaustion, has also been described in haematological malignancies, which may affect the use of therapeutic monoclonal antibodies acting through NK cell-mediated cytotoxicity (ADCC).
This efect does not only affects direct interactions between malignant cells and immunocompetent effector cells, but expression on non-malignant tumor cells significantly affects the microenvironment of tumor and creates a suitable niche for the survival and growth of the malignant population. PD-1 positive tumor cells are also able to induce T-regulatory lymphocytes and control their expansion and function, with PD-1 positive tumor infiltrating dendritic cells showing the same effect. Last but not least, tumor cells with higher PD-1 expression show resistance to apoptoptic stimuli induced by both the Fas pathway and the effect of chemotherapy.
Exhausting of effector T-cells induced by chronic activation of the programmed cell death signaling pathway leads not only to induction of tumor cell immunotolerance, but to alteration of T-cell immunity as a whole, which is ultimately seen in higher incidence and more severe bacterial and viral infections. Functional tests in patients with CLL revealed that markers of T-cell dysfunction (PD-1, CD244 and CD160) are most strongly expressed on TEM cells (T effector memory, CCR7- / CD45RA-) and TEMRA (CD45RA +), while on naive ( CCR7 + CD45 + RA +) and T-central memory cells (CCR7 + / CD45RA-) expression is significantly lower. These results indicate a shift in the circulating T-cell compartment toward differentiated forms after contact with an antigen bearing markers of exhaustion. Furthermore, a reduction in the number of CD8 + T-cells capable of proliferating after polyclonal activation and cell cycle prolongation was found in the mitotic forms. Finally, a reduced ability of the cytotoxic effect on pathological cells has been demonstrated experimentally. In contrast to T-cell exhaustion in chronic viral infections, no loss of effector cytokines was observed in patients with CLL, intracellular interferon-γ (IFNγ) and tumor necrosis factor-inducing factor (TNFα) levels were even increased compared to healthy donors, corresponding to higher proportion of TEM and TEMRA cells, producing higher amounts of IFNγ and TNFα than naive T-lymphocytes and TCM.
In light of current knowledge, PD-1 appears to be a tumor-inducible negative costimulatory receptor whose interaction with PD-L1 and PD-L2 ligands affects T-cell subpopulations, inhibits their proliferation and cytokine production, and reduces cytotoxic activity, particularly by inducing functional inactivity of cytotoxic CD8 + T-cells. The tumor clone is able to influence the state of the anti-tumor immune response via the PD-1 / PD-L pathway, and the PD-1 marker can therefore be considered a sign of systemic immune dysregulation in CLL in relation to healthy individuals.
Increased PD-1 and PD-L1, L2 expression has been demonstrated not only in hematological malignancies but also in many solid tumors, where it also inhibits cytokine production and the activity of tumor infiltrating CD4 + and CD8 + T cells.
The PD-L1 / PD-1 pathway is a valuable target for biological therapy. At the end of 2014, the first two PD-1 / PD-L1 interaction inhibitors, pembrolizumab and nivolumab, were approved by U.S. Pat. Food and Drug Administration (FDA). Today, FDA and EMEA have approved 7 anti–PD-1/PD-L1 immune checkpoint inhibitors (ICIs) covering more than 15 cancer indications as monotherapy or in combination with other drugs. Therapeutic blockade of the interaction of the PD-1 receptor with its ligands increases the level of the immune response and is directly involved in restoring the effectiveness of the anti-tumor activity of the immune system.
FIG. 1: In our laboratory practice, we use PD-1 expression assays to assess the state of alteration of the T-cell immune response in peripheral blood in patients with CLL. Using the anti-CD45 Pacific Orange antibody, we define the leukocyte population, from which immunocompetent subpopulations of interest (anti-CD3 APC, anti CD4 Pacific Blue, anti CD8 PerCP) are excluded. We then measure the percentage of PD-1 positive events (CD4 + / PD-1 + and CD8 + / PD-1 +), which reflect the degree of influence on the patient's immune response by the programmed cell death pathway.
Martin Novak, Ph.D.
Laboratory of Flow Cytometry / Department of Hematooncology
Olomouc University Hospital
EXBIO PRODUCTS RELATED TO THIS BLOG:
EXBIO GATING ANTIBODIES:
CD45 Pacific Orange™
CD4 Pacific Blue™
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