Mutated Ras-Transfected, EBV-Transformed Lymphoblastoid Cell Lines as a Model Tumor Vaccine for Boosting T-Cell Responses Against Pancreatic Cancer: A Pilot Trial

Abstract

Genetically modified lymphoblastoid cell lines (LCL) have been shown to be an attractive alternative source of antigen-presenting cells for cancer vaccination in vitro. We tested their application in patients with pancreatic cancer in a phase I clinical trial. As a model tumor antigen, we selected the point-mutated (codon 12) Ki-Ras p21 oncogene (muRas) frequently (∼85%) present in pancreatic adenocarcinoma. Autologous LCLs were established in vitro by spontaneous outgrowth from peripheral blood lymphocytes of seven pancreatic carcinoma patients and were genetically modified with an episomal Epstein-Barr virus (EBV)-based expression vector to express muRas (muRas-LCL). Weekly vaccinations with subcutaneous injection of 5×10(6) muRas-LCL were done. In six of seven patients, therapeutic vaccination elicited a T-cell response with an increase in the frequency of muRas-specific precursor cytotoxic T lymphocytes in the peripheral blood and positive delayed-type hypersensitivity reactions at the injection site. Besides local reactions and flu-like symptoms, there were no signs of toxicity and no acute EBV infection, onset of EBV-associated lymphoma, or other severe complications. A clinical response (stable disease) was observed for a short time period (2-4 months) in four of seven patients (57%), mostly in earlier tumor stages. Our results indicate that LCL presenting genetically modified antigen represent a valuable and easily available tool for in vivo autologous tumor vaccination. LCL can be transfected with any known tumor antigen and therefore should be further clinically investigated.

Mutation analysis of the EBV‐lymphoblastoid cell line cautions their use as antigen‐presenting cells

“…LCLs have a high immunostimulatory capacity and can be easily and largely obtained by rapid and long‐lasting growth in vitro , and thus they are potentially unlimited sources of potent APCs to prime either CD4 + or CD8 + for immunotherapy. Indeed, several studies reported that LCLs pulsed ex vivo with neoantigens or other tumor‐associated antigens could induce specific anti‐tumor immunity . However, in addition to EBV‐associated antigens, nonsynonymous mutations arising in high‐passage LCLs could also induce activation and proliferation of nonspecific T cells, which theoretically result in obscuring the detection of tumor‐specific T‐cell response.…”

“…However, the expense and complexity of obtaining sufficient dendritic cells due to low frequency in peripheral blood and in vitro proliferation capacity of dendritic cells has limited its wide application, particularly in critically ill cancer patients . Alternatively, Epstein–Barr virus (EBV)‐transformed B‐lymphoblastoid cell lines (LCLs) have been demonstrated to be excellent professional APCs and most importantly, LCLs could be easily obtained by culturing peripheral blood lymphocytes from EBV‐positive humans in vitro or by incubating with B95‐8 cell supernatant . However, neoantigen‐loaded LCLs could stimulate not only neoantigen‐specific T cells but also nonspecific T cells, which could be due to expression of both EBV antigens and endogenous nonsynonymous mutations arising in LCLs .…”

Gene Therapy for Pancreatic Cancer: Specificity, Issues and Hopes

“…Main indication is PDAC at an advanced stage (locally advanced and/or metastatic) with phases 1 and 2 trials and one phase 3 [101,120,121,122,123,124,125,126,127,128,129,130,131,132,133,134,135,136,137,138,139,140,141,142,143,144,145,146]. Three types of strategies have been conducted: in situ gene therapy product injection (by means of EUS), vaccination (intradermal or subcutaneous route) and oncolytic virus administration.…”

Functional analysis and molecular characterization of spontaneously outgrown human lymphoblastoid cell lines

“…LCL have been used as APC for the presentation of tumor antigens in vaccination studies [11,17]. In these studies, spontaneously outgrown LCL have been used.…”