Development of a Unique Anti-AML Immune Therapy Consisting of Cord Blood HSCT and Cord Blood Stem Cell-Derived Dendritic Cell (CB-DC) Vaccination

Background: Development of novel (immune) therapies is of utmost importance to improve survival in relapsed pediatric-AML (acute myeloid leukemia). We aim to develop a powerful and safe therapy consisting of two potentially synergistic components: Cord Blood (CB) HSCT and vaccination with CB-derived Wilms Tumor-1 (WT1) mRNA-electroporated dendritic cells (DCs). Materials & Methods: After isolation, the CD34+ CB stem cells were cultured using a two-step protocol. First, they were expanded using a combination of (growth) factors (Flt3L, SCF, IL-3 and IL-6). Next, the cells were differentiated towards DCs for one week using medium containing Flt3L, SCF, GM-CSF, IL-4 and human serum followed by a CYTOMIX (IL-1beta, IL-6, TNF-alpha and PGE2)-induced maturation for the last 24 hours. Finally, the CB-DC culture was electroporated with WT1-mRNA and their phenotype (cell surface markers) and function (migration and antigen presentation) were assessed. Results: Using the two-step protocol a total cell expansion of 300-500 fold was achieved. Based on surface marker expression, different DC subsets could be distinguished in our CB-DC cultures. Since no differences in antigen presentation capacity between the DC subsets were detected, the whole CB-DC culture was used in all phenotypic and functional assays. The maturation using CYTOMIX induced upregulation of costimulatory molecules (CD80, CD83 and CD86) and the chemokine receptor CCR7 on the CB-DCs. These matured CB-DCs showed enhanced CCR7-dependent migration towards CCL21 in a trans-well migration assay. The presentation of WT1 peptides by the CB-DC culture, matured using CYTOMIX and electroporated with WT1 mRNA, was confirmed by the stimulation of T cells expressing a recombinant WT-1 peptide specific TCR. Furthermore, this same CB-DC culture was able to induce expansion of the WT-1 specific T cells from their autologous pool of na•ve T cells in cord blood. Conclusion: We have developed and tested an in vitro system for culturing large amounts of DCs from the CD34+ CB stem cells. Both the phenotypic and functional data support the use of the whole CB-DC culture as vaccine. We are currently translating our preclinical protocol to GMP production of a clinical grade vaccine.