1131. Generation of Whole Cell Vaccines for Acute Myeloid Leukaemia by Lentivirus Mediated IL-2/CD80 Transduction

Abstract

Despite recent treatment advances, the prognosis for the majority of AML patients remains bleak, particularly in the elderly, those with poor prognostic cytogenetics, and/or progression from myelodysplasia. We and others have previously shown that tumour cells engineered to express CD80 and IL-2 can stimulate T and NK cell responses against the unmodified tumour cells, and that vaccination with irradiated CD80/IL-2 expressing leukaemic cells can prolong survival in mouse models of leukaemia. In order to test the therapeutic potential of this approach, we have devised an efficient gene transfer protocol for primary human acute myeloid leukaemia blasts. Based on a previously established methodology for retroviral vectors, we show that disabled SIN-lentiviral vectors can be conjugated to streptavidin paramagnetic microparticles (PMP) after covalent linkage of biotin to 293T packaging cells. Biotin coated lentivirus is then captured and magnetically concentrated, resulting in up to 2600 fold titre enhancements after 100 fold volume reductions to achieve titres in excess of 109 cfu/ml for amphotropic or VSV-G pseudotypes. A self-inactivating lentiviral backbone, incorporating the HIV flap region and a myeloid efficient internal promoter, derived from spleen focus forming virus (SFFV), is used for the efficient co-expression of CD80 and IL-2 in primary AML blasts. These vectors express CD80 and IL-2 as a single fusion protein that incorporates the recognition sequence for a golgi/ER located ubiquitous endoprotease |[ndash]| furin between the CD80 and IL-2 moieties. This fusion protein is post-synthetically cleaved by furin to generate biologically active membrane anchored CD80 and secreted IL-2. Following a single round of infection of primary or established AML cells at an MOI of 10, between 40%-99% of AML blasts express CD80 and secrete 2-15 ng of IL-2/106/24hrs. The combined expression of IL-2 and CD80 enables the increased in vitro stimulation of allogeneic and autologous T cells obtained from AML patients, either before or following the induction of remission, with or without preceding bone marrow transplantation. The stimulated lymphocytes have a TH1 cytokine secretion profile and show evidence of specificity, as indicated by their increased proliferation in the presence of unmodified AML compared to remission bone marrow cells. The stimulated T cells also demonstrate cytotoxic activity against AML blasts. This data supports the notion that IL-2/CD80 whole cell vaccination could stimulate therapeutically useful immune responses in AML patients, particularly following chemotherapy induced remission and allogeneic bone marrow transplantation. We have recently been granted permission for the vaccination of poor prognosis AML patients with lentivirus mediated ex vivo transduced AML blasts expressing CD80 and IL-2.