Acute myeloid leukemia (AML) is characterized by the proliferation of clonal neoplastic myeloid hematopoietic precursor cells and impaired normal hematopoiesis. Although 70–80% of patients (<60 years) achieve complete remission after intensive chemotherapy, AML frequently relapses due to the persistence of minimal residual disease (MRD) [
1]. Escape of leukemic cells from immune surveillance has been associated with the poor clinical outcome. For instance, a high occupancy of HLA-DR molecules with the class II-associated invariant chain peptide (CLIP) instead of an antigenic peptide is correlated with a shortened disease-free and overall survival [
2,
3]. In contrast, immune control of leukemia, as shown for instance by the graft-versus-leukemia effect induced by allogeneic stem cell transplantation (SCT) or the reinduction of complete remission after donor lymphocyte infusion following allogeneic SCT, demonstrates the potential of exploiting the immune system in aid of anti-AML therapy [
1].
The induction, regulation, and maintenance of primary immune responses, including specific anti-tumor T cell responses are coordinated by dendritic cells (DC). Vaccination with DC has been recognized as a promising investigational therapy due to the uniquely powerful antigen (Ag)-presenting capacity of DC and its potential to circumvent immunosuppressive features of leukemia [
4]. The first steps down the road to DC vaccination in AML have been taken and results from small clinical trials have been reported [
5]. The general lack of clinical responses evoked important questions concerning the optimal methodologies for DC vaccine preparation as well as the design of clinical vaccination protocols. Many strategies are explored in the preparation of DC vaccines ex vivo; among these, autologous monocyte-derived DC (MoDC) loaded with leukemia-associated Ag (LAA) are promising [
6]. Various sources of LAA and different methods of loading LAA onto DC have been explored in an attempt to optimize anti-tumor responses [
7]. For AML, several relevant LAA have been identified including PRAME, RHAMM, WT1, and PR1. Unfortunately, overexpression of these LAA is common, but not uniform in leukemia [
8]. Moreover, HLA restriction of LAA-derived peptides limits application of such vaccines to patients with certain HLA profiles. These restrictions inherent to the use of defined LAA or LAA-derived peptides may be overcome using the whole AML cells as a source of LAA, for instance by generation of AML lysates or apoptotic leukemic cells. Among other whole AML cell-derived antigen loading strategies that have been explored is electroporation of DC with AML-derived RNA [
9]. Also vaccination with modified AML cells, such as AML-derived DC or fusions between AML cells and DC, has been investigated; further modification of DC with 4-1BB-L or CD40 might enhance the efficacy of such vaccines [
10‐
13]. It has long been assumed that the apoptotic cell death is poorly immunogenic or even tolerogenic, whereas necrotic cell death is considered to be immunogenic. However, stress-induced heat shock protein (HSP)-peptide complexes (commonly induced during apoptosis) are more efficiently taken up via scavenger receptors and Toll-like receptors (TLR) on the DC surface and induce efficient cross-priming and skewing of the immune response towards a Th1-type profile [
14,
15], whereas necrosis has been associated with the local immune suppression in solid tumors [
16]. Furthermore, apoptosis induction after irradiation with UV light or by treatment with chemotherapeutic drugs results in upregulation of calreticulin, a scavenger receptor class-A ligand associated with immunogenic apoptosis as demonstrated for colon carcinoma cells [
17,
18]. For AML, it is not yet clear whether cell lysates or apoptotic cells are preferable for Ag loading onto MoDC [
9,
19‐
25]. Next to enhancing immunogenicity of tumor Ag sources by, for instance, heat shock, addition of DC-maturing stimuli, such as Toll-like receptor ligands (TLR-L), is explored; e.g., electroporation of TLR3-L Poly(I:C) into AML cells results in enhanced uptake of leukemic cells by DC and improves their subsequent maturation and cytokine production [
26]. Furthermore, intracellular binding of TLR8 by its ligand R848 has been reported to result in enhanced cross-priming of exogenous Ag by MoDC [
27]. Various whole AML cell preparations loaded onto MoDC in combination with DC maturation-inducing cocktails have been explored [
9,
21]; however, the quantitative effects of TLR-L and cytokines on uptake of leukemic cells are still unclear.
In this study, we compared the uptake of allogeneic apoptotic leukemic cells with lysates derived from leukemic cells. We investigated the uptake of heat shock-induced apoptotic leukemic cells by MoDC and DC maturation by combining a standard cytokine cocktail (CC) with the clinically applicable TLR7/8-L R848.