After COVID-19, some patients develop long-term symptoms. Whether such symptoms correlate with immune responses, and how long immunity persists, is not yet clear. This study focused on mild COVID-19 and investigated correlations of immunity with persistent symptoms and immune longevity. Persistent complications, including headache, concentration difficulties and loss of smell/taste, were reported by 51 of 83 (61%) participants and decreased over time to 28% one year after COVID-19. Specific IgA and IgG antibodies were detectable in 78% and 66% of participants, respectively, at a 12-month follow-up. Median antibody levels decreased by approximately 50% within the first 6 months but remained stable up to 12 months. Neutralizing antibodies could be found in 50% of participants; specific INFgamma-producing T-cells were present in two thirds one year after COVID-19. Activation-induced marker assays identified specific T-helper cells and central memory T-cells in 80% of participants at a 12-month follow-up. In correlative analyses, older age and a longer duration of the acute phase of COVID-19 were associated with higher humoral and T-cell responses. A weak correlation between long-term loss of taste/smell and low IgA levels was found at early time points. These data indicate a long-lasting immunological memory against SARS-CoV-2 after mild COVID-19.

Background: Recently, our group introduced Stroma AReactive Invasion Front Areas (SARIFA) as an independent prognostic predictor for a poorer outcome in colon cancer patients, which is probably based on immunologic alterations combined with a direct tumor-adipocyte interaction: the two together reflecting a distinct tumor biology. Considering it is already known that peripheral immune cells are altered in colorectal cancer (CRC) patients, this study aims to investigate the changes in lymphocyte subsets in SARIFA-positive cases and correlate these changes with the local immune response. Methods: Flow cytometry was performed to analyze B, T, and natural killer (NK) cells in the peripheral blood (PB) of 45 CRC patients. Consecutively, lymphocytes in PB, tumor-infiltrating lymphocytes (TILs), and CD56+ and CD57+ lymphocytes at the invasion front and the tumor center were compared between patients with SARIFA-positive and SARIFA-negative CRCs. Results: Whereas no differences could be observed regarding most PB lymphocyte populations as well as TILs, NK cells were dramatically reduced in the PB of SARIFA-positive cases. Moreover, CD56 and CD57 immunohistochemistry suggested SARIFA-status-dependent changes regarding NK cells and NK-like lymphocytes in the tumor microenvironment. Conclusion: This study proves that our newly introduced biomarker, SARIFA, comes along with distinct immunologic alterations, especially regarding NK cells.

Dendritic cells (DCs) and leukemia-derived DC (DC_leu) are potent stimulators of various immunoreactive cells and they play a pivotal role in the (re-) activation of the immune system. As a potential treatment tool for patients with acute myeloid leukemia, we developed and analyzed two new PGE₁-containing protocols (Pici-_PGE1, Kit M) to generate DC/DC_leu ex vivo from leukemic peripheral blood mononuclear cells (PBMCs) or directly from leukemic whole blood (WB) to simulate physiological conditions. Pici-_PGE1 generated significantly higher amounts of DCs from leukemic and healthy PBMCs when compared to control and comparable amounts as the already established protocol Pici-_PGE2. The proportions of sufficient DC-generation were even higher after DC/DC_leu-generation with Pici-_PGE1. With Kits, it was possible to generate DCs and DC_leu directly from leukemic and healthy WB without induction of blast proliferation. The average amounts of generated DCs and DC_leu-subgroups were comparable with all Kits. The PGE₁ containing Kit M generated significantly higher amounts of mature DCs when compared to the PGE₂-containing Kit K and increased the anti-leukemic-activity. In summary PGE₁-containing protocols were suitable for generating DC/DC_leu from PBMCs as well as from WB, which reliably (re-) activated immunoreactive cells, improved the overall ex vivo anti-leukemic activity, and influenced cytokine-release-profiles.

Background: Extracellular vesicles (EV) are cell-derived vesicles released by all cells in health and disease. Accordingly, EVs are also released by cells in acute myeloid leukemia (AML), a hematologic malignancy characterized by uncontrolled growth of immature myeloid cells, and these EVs likely carry markers and molecular cargo reflecting the malignant transformation occurring in diseased cells. Monitoring antileukemic or proleukemic processes during disease development and treatment is essential. Therefore, EVs and EV-derived microRNA (miRNA) from AML samples were explored as biomarkers to distinguish disease-related patterns ex vivo or in vivo. Methodology: EVs were purified from serum of healthy (H) volunteers and AML patients by immunoaffinity. EV surface protein profiles were analyzed by multiplex bead-based flow cytometry (MBFCM) and total RNA was isolated from EVs prior to miRNA profiling via small RNA sequencing. Results: MBFCM revealed different surface protein patterns in H versus AML EVs. miRNA analysis showed individual as well as highly dysregulated patterns in H and AML samples. Conclusions: In this study, we provide a proof-of-concept for the discriminative potential of EV derived miRNA profiles as biomarkers in H versus AML samples.

Dendritic cells (DC) and leukaemia derived DC (DC_leu) are potent stimulators of anti-leukaemic activity in acute myeloid leukaemia (AML) and can be generated from mononuclear cells in vitro following standard DC/DC_leu-generating protocols. With respect to future clinical applications though, DC/DC_leu-generating protocols specifically designed for application in a whole-blood-(WB)-environment must be established. Therefore, we developed ten new DC/DC_leu-generating protocols (kits; Kit-A/-C/-D/-E/-F/-G/-H/-I/-K/-M) for the generation of DC/DC_leu from leukaemic WB, containing calcium-ionophore, granulocyte-macrophage-colony-stimulating-factor (GM-CSF), tumour-necrosis-factor-alpha, prostaglandin-E₁ (PGE₁), prostaglandin-E₂ (PGE₂) and/or picibanil (OK-432). All protocols were evaluated regarding their performance in generating DC/DC_leu using refined classification and/or ranking systems; DC/DC_leu were evaluated regarding their performance in stimulating anti-leukaemic activity using a cytotoxicity fluorolysis assay. Overall, we found the new kits capable to generate (mature) DC/DC_leu from leukaemic WB. Through refined classification and ranking systems, we were able to select Kit-I (GM-CSF + OK-432), -K (GM-CSF + PGE₂) and -M (GM-CSF + PGE₁) as the most efficient kits in generating (mature) DC/DC_leu, which are further competent to stimulate immunoreactive cells to show an improved anti-leukaemic cytotoxicity as well. This great performance of Kit-I, -K and -M in mediating DC/DC_leu-based anti-leukaemic immunity in a WB-environment in vitro constitutes an important and directive step for translating DC/DC_leu-based immunotherapy of AML into clinical application.