Generation of Leukaemia-Derived Dendritic Cells (DCleu) to Improve Anti-Leukaemic Activity in AML: Selection of the Most Efficient Response Modifier Combinations

Abstract

Dendritic cells (DC) and leukaemia derived DC (DCleu) are potent stimulators of anti-leukaemic activity in acute myeloid leukaemia (AML) and can be generated from mononuclear cells in vitro following standard DC/DCleu-generating protocols. With respect to future clinical applications though, DC/DCleu-generating protocols specifically designed for application in a whole-blood-(WB)-environment must be established. Therefore, we developed ten new DC/DCleu-generating protocols (kits; Kit-A/-C/-D/-E/-F/-G/-H/-I/-K/-M) for the generation of DC/DCleu from leukaemic WB, containing calcium-ionophore, granulocyte-macrophage-colony-stimulating-factor (GM-CSF), tumour-necrosis-factor-alpha, prostaglandin-E1 (PGE1), prostaglandin-E2 (PGE2) and/or picibanil (OK-432). All protocols were evaluated regarding their performance in generating DC/DCleu using refined classification and/or ranking systems; DC/DCleu were evaluated regarding their performance in stimulating anti-leukaemic activity using a cytotoxicity fluorolysis assay. Overall, we found the new kits capable to generate (mature) DC/DCleu from leukaemic WB. Through refined classification and ranking systems, we were able to select Kit-I (GM-CSF + OK-432), -K (GM-CSF + PGE2) and -M (GM-CSF + PGE1) as the most efficient kits in generating (mature) DC/DCleu, which are further competent to stimulate immunoreactive cells to show an improved anti-leukaemic cytotoxicity as well. This great performance of Kit-I, -K and -M in mediating DC/DCleu-based anti-leukaemic immunity in a WB-environment in vitro constitutes an important and directive step for translating DC/DCleu-based immunotherapy of AML into clinical application.

Dendritic Cells as a Therapeutic Strategy in Acute Myeloid Leukemia: Vaccines

“…The manufacture of DC vaccines mainly begins with the isolation of monocytes, CD34+ hematopoietic stem cells (HSC), and myeloid leukemic blasts to obtain imDCs or leukemia-derived DCs (DCleu) by adding granulocyte colony-stimulating factor- macrophages (GM-CSF) plus IL-4 or Prostaglandin E1 (PGE1) and Fms-related receptor tyrosine kinase 3 ligand (Flt3L), respectively. Then, once imDCs are obtained, they are ex vivo matured by the addition of a cocktail of several cytokines, such as TNF-α, prostaglandin-E1 (PGE1), prostaglandin-E2 (PGE2) and/or picibanil (OK-432) or CD40 ligand (CD40L) [ 54 , 55 ], and loaded with antigens through several pathways: whole cell antigens or tumor lysates, synthetic antigenic peptides, tumor DNA or mRNA, exosomes derived from cancer cells [ 54 , 56 ], Figure 2 .…”

Volatile Phases Derived from Serum, DC, or MLC Culture Supernatants to Deduce a VOC-Based Diagnostic Profiling Strategy for Leukemic Diseases

“…They are activated and maturate after danger-signaling adhesions (e.g., nucleic acids, infectious particles), resulting in upregulated chemokine receptors (e.g., CCR7), MHC-antigens and other costimulatory factors [ 8 , 9 , 10 , 11 ]. Ex vivo DCs can be generated from CD14+ monocytes (and loaded with tumor antigens) or from myeloid blasts (DC leu , leukemia-derived DC; without induction of blast proliferation) from AML patients’ WB in the presence of different combinations of response modifiers (Kits) [ 9 , 12 , 13 ]; the resulting DCs express costimulatory molecules together with individual patients’ leukemic antigens, and gain the capacity to activate the cells of the immune system against blasts. Ex vivo-generated (and manipulated) DCs or DC leu could be used for an adoptive transfer; applying Kits directly to AML patients could induce DC leu from blasts, leading to an antileukemic immunoreaction in vivo [ 9 ].…”

“…Ex vivo-generated (and manipulated) DCs or DC leu could be used for an adoptive transfer; applying Kits directly to AML patients could induce DC leu from blasts, leading to an antileukemic immunoreaction in vivo [ 9 ]. Immunomodulatory Kit M, composed of granulocyte macrophage colony-stimulating factor (GM-CSF) and prostaglandin E1 (PGE 1 ), has proved to be one of the best combinations of immune response modifiers to generate DC/DC leu from leukemic WB [ 9 , 13 ]; therefore, it was used in this ex vivo study.…”

“…The key players of the innate immune system are APCs (e.g., monocytes, macrophages, DC), cytokine-induced killer (CIK), invariant natural killer T cells (iNKT cells) and natural killer (NK) cells; they mediate the earliest interactions with pathogens/tumors [ 12 , 13 , 14 ]. Antigen-specific response and generation of immunologic memory are the central tasks of the adaptive immune system [ 12 , 13 ].…”

“…The key players of the innate immune system are APCs (e.g., monocytes, macrophages, DC), cytokine-induced killer (CIK), invariant natural killer T cells (iNKT cells) and natural killer (NK) cells; they mediate the earliest interactions with pathogens/tumors [ 12 , 13 , 14 ]. Antigen-specific response and generation of immunologic memory are the central tasks of the adaptive immune system [ 12 , 13 ]. After activation, naive T cells (T naive , CD3+CD45RO−) are probably converted to non-naive T cells (T non-naive , CD3+CD45RO+), which mediate various immune responses or develop into long-living central memory cells (T cm , CD3+CD45RO+CD197+) or effector memory cells (T em/eff , CD3+CD45RO+CD197−) to facilitate a faster reactivation of the immune system against recurring antigens [ 8 , 15 ].…”