Background: Chronic lymphocytic leukaemia (CLL) is an incurable but highly treatable malignancy of mature monoclonal B cells. Patients with CLL suffer from immune dysfunction and experience frequent opportunistic infections. Natural killer (NK) cells are critical for effective responses to bacterial and viral infections. The pro- and anti-tumour properties of NK cells have been studied in patients with CLL, where these cells demonstrate reduced cytotoxicity and increased expression of inhibitory receptors. However, the capacity of NK cells to respond to bacterial and viral stimulation has not been studied in this patient cohort.
Aims/objectives: To optimise flow cytometry panels for the measurement of NK cell activation in patients with CLL. To measure NK cell responses to heat-killed bacteria and viral mimetics in this cohort.
Methods: Antibody titrations will be performed using frozen peripheral blood mononuclear cell (PBMC) samples from 10 healthy donors and treatment-naïve 10 patients with CLL at the existing biobank at University Hospital Limerick. The optimal antibody staining index will be determined for NK cell surface markers (CD56, CD16, TRAIL, NKp44, NKp30, NKG2D, CD161, CD69) and intracellular cytokines (IFN-g, TNF-a, IL-22, granzyme B). NK cells will be stimulated for 24 hours using 10 million heat-killed Staphylococcus aureus or Escherichia coli cells per million NK cells (MOI 10:1) or 500 ng/ml CL097 viral mimetic. Cytokine production will be validated orthogonally by enzyme-linked immunosorbent assay (ELISA).
Outputs/Impact: This project will reveal the capacity of NK cells to respond to infectious agents in patients with CLL. The resulting flow cytometry data will reveal the composition and range of NK cell activation states across healthy donors and patients with CLL. This may be useful to identify patients at heightened risk of infection, allowing for early therapeutic intervention such as antibiotic administration to reduce hospitalisation and improve patients’ quality of life.