Significance of frequencies, compositions and/or antileukemic activity of (DC-stimulated) invariant NKT, NK and CIK cells on the outcome of patients with AML, ALL and CLL

Böck, Corinna Lesley (2018): Significance of frequencies, compositions and/or antileukemic activity of (DC-stimulated) invariant NKT, NK and CIK cells on the outcome of patients with AML, ALL and CLL. Dissertation, LMU München: Medizinische Fakultät

Abstract

Invariant natural killer cells (iNKT cells), natural killer cells (NKT cells) and cytokine induced killer cells (CIK cells) and their subsets are important for immune-surveillance. Recently, a new monoclonal antibody ‘6B11’ was developed for iNKT cell detection, which targets the Vα24Jα18 invariant T cell receptor (TCR) in the CDR3-region of iNKT cells. In the first part of my work iNKT, NK and CIK cells and their subsets were analysed in PB-samples from healthy donors (n=9), patients with acute myeloid leukemia (AML; n=23), patients with acute lymphoid leukemia (ALL; n=20) and patients with chronic lymphoid leukemia (CLL; n=21) in acute disease-stages and findings were correlated with prognosis. The main results were: I.1) Compared to healthy mononuclear cells (MNC) (significantly) lower proportions of iNKT cells (6B11+/ 6B11+CD3+/ 6B11+CD161+), NK cells (CD3-CD56+/ CD3-CD161+) and CIK cells (CD3+CD56+/ CD3+CD161+) were found in MNC from AML, ALL and CLL patients. I.2) Compared to healthy MNC (significantly) higher proportions of 6B11+iNKT cells were found in the CD3+ T cell- (‘T cell-like’ iNKT cells) and in the CD161+NK cell fraction (‘NK cell-like’ iNKT cells) in AML, ALL and CLL patients. I.3) Prognostically favorable AML subgroups (patients younger than 18 years, primary disease-status, no extramedullary disease, achievement and maintenance of complete remission (CR) after induction-chemotherapy) showed higher proportions of iNKT, NK and CIK cells. Comparable correlations were seen in (adult) ALL and CLL patients. In the second part of my work heparinized whole blood-(WB) and MNC-samples from healthy donors (n=10), AML patients (n=5) and one patient with myelodysplastic syndrome (MDS; n=1) were analysed. Samples were pre-treated with different ‘cocktails’ in the cell culture, which induce the conversion of leukemic cells to dendritic cells of leukemic origin (DCleu, leukemia-antigen-presenting cells). Not pre-treated, cultured WB- or MNC-samples served as control. iNKT, NK and CIK cells and their subsets were quantitatively and qualitatively analysed before and after mixed-lymphocyte-cultures (MLC) of T cell enriched WB- or MNC-samples under normoxia (21% O2) as well as under physiological hypoxia (approx. 10% O2). ‘Cocktail’ pre-treated or not pre-treated WB- or MNC-samples served as a ‘stimulator-suspension’, T cell enriched immune-reactive WB- or MNC-samples served as effector cells. Blast-lytic activity of iNKT, NK, CIK and T cells after MLC was examined using a cytotoxicity (fluorolysis) assay. The main results were: II.1) iNKT, NK and CIK cells increased after MLC independent of the applied ‘cocktail’ / ‘stimulator cell suspension’ (under the influence of IL-2). II.2) Pretreatment of MNC- or WB-samples with ‘cocktails’ in the cell culture increased frequencies of iNKT cells and led to a shift in the composition of iNKT, NK and CIK cell subsets after MLC, what might correlate with an improved antileukemic potential. II. 3) Individual samples showed varying, however higher iNKT and CIK cell frequencies after pretreatment with different (especially prostaglandin-containing) ‘cocktails’. II.4) DC, iNKT, NK and CIK cell values after MLC were comparable in physiological hypoxia vs normoxia. II.5) Frequencies of T, iNKT, NK and CIK cells after MLC correlated directly with antileukemic, blast-lytic activity – pointing to an involvement of these cells in antileukemic reactions. In summary, healthy MNC presented with significantly higher iNKT, NK and CIK cells compared to leukemic AML, ALL and CLL MNC. Moreover, subtypes of iNKT cells differ in healthy vs leukemic samples. Furthermore, frequencies of iNKT, NK and CIK cells correlated with prognosis of patients with AML, ALL and CLL. In addition, ‘cocktail’ pre-treated AML-blasts (resulting in DCleu) after MLC with effector cells (T cell enriched WB / MNC) led to a shift in T, iNKT, NK and CIK cell counts and compositions. These findings correlated with improved antileukemic activity against AML-blasts - pointing to a cross-talk of these cells. Proportions of iNKT, NK and CIK-cells (based on detection with 6B11/ CD161/ CD56/ CD3 antibodies) should regularly be evaluated in AML, ALL and CLL diagnosis-panels for quantitative, qualitative and prognostically relevant estimation of individual pts’ antileukemic potential in detail and to learn about their role in DC/DCleu triggered immune-surveillance.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: iNKT, NK, CIK, DCleu, AML, ALL, CLL
• Themengebiete: 600 Technik, Medizin, angewandte Wissenschaften ; 600 Technik, Medizin, angewandte Wissenschaften > 610 Medizin und Gesundheit
• Fakultäten: Medizinische Fakultät
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 14. Juni 2018
• 1. Bericht­erstatter:in: Schmetzer, Helga