2017 KhalafWSSPhD.pdf (10.73 MB)
Download file

In vitro generation of cytotoxic T cells with potential for adoptive tumour immunotherapy

Download (10.73 MB)
posted on 25.09.2017, 13:24 by Wafaa Seifalnaser Sedik Khalaf
Multiple myeloma (MM) is a life-threatening haematological malignancy, which is rarely curable by conventional therapies. Immunotherapy, using autologous antigen specific cytotoxic T-lymphocytes (ASCTL), may represent a useful adjunct therapy for MM. In this study, I assessed the ability of previously described hybrid cell lines, generated by chemical fusion of myeloma tumour cells and the EBV B-lymphoblastoid cell line (EBV B-LCL) HMy2, to induce ASCTL in vitro from peripheral blood lymphocytes from patients with MM (and from healthy individuals). The tumour associated antigens (TAAs) hTERT, MUC1, MAGE-C1 and CS1 were selected as potential inducers of ASCTL, based on their prevalence of expression in MM patients. Expression of these TAAs was assessed in four B-LCL/myeloma hybrid cell lines, using real time PCR and flow cytometry, and two of the hybrid cell lines were selected as in vitro stimulator cell lines in long-term activated T-cell cultures, using PBMCs from HLA-A2+ healthy donors and multiple myeloma patients. Induction of ASCTLs was assessed by HLA-A2-peptide pentamer staining and flow cytometry, Europium release cytotoxicity assays, and interferon-gamma and perforin ELIspot assays, using known HLA-A2 restricted peptide epitopes of the TAAs. The hybrid cell lines induced ASCTLs to the 4 selected TAAs, after 4 rounds of in vitro stimulation with the hybrid cell lines, in PBMCs from both (HLA-A2+) healthy donors and MM patients. In contrast, the (HLA-A2+) myeloma cell line, U266, failed to activate ASCTL in vitro. Hybrid cell lines, generated by fusion of EBV B-LCL and myeloma tumour cells, can induce ASCTL in PBMCs from healthy individuals and multiple myeloma patients in vitro, and may represent a novel strategy for use in immunotherapy of MM.



Browning, Michael; Stover, Cordula

Date of award


Author affiliation

Department of Infection, Immunity and Inflammation

Awarding institution

University of Leicester

Qualification level


Qualification name




Usage metrics