Macrophages and dendritic cells are heterogenous and highly plastic bone marrow-derived cells that play major roles in renal diseases. We characterized these cells using immunohistochemistry in 55 renal biopsies from control patients or patients with glomerulonephritis as an initial step towards postulating specific roles for these cells in kidney disease. In proliferative glomerulonephritis numerous CD68 positive (pan monocyte, macrophage and dendritic marker) cells were found in both glomeruli and the tubulointerstitial space, however, a myeloid dendritic cell marker (DC-SIGN) was identified only in the tubulointerstitium. A significant number of plasmacytoid dendritic cells (identified as BDCA-2 positive cells) were seen at sites of interstitial inflammation, including follicular aggregates of inflammatory cells. Langerin positive cells (a marker of Langerhans' cells) were detectable but rare. The area of either CD68 or DC-SIGN positive interstitial cells correlated with serum creatinine. Low levels of DC-SIGN, DC-LAMP and MHC class II mRNA were present in the tubulointerstitial space in controls and increased only in that region in proliferative glomerulonephritis. We demonstrate that the CD68 positive cells infiltrating the glomerulus lack dendritic cell markers (reflecting macrophages), whereas in the tubulointerstitial space the majority of CD68 positive cells are also DC-SIGN positive (reflecting myeloid dendritic cells). Their number correlated with serum creatinine, which further emphasizes the significance of interstitial DCs in progressive glomerular diseases.

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SummaryPrevious studies have shown that immature mouse class I molecules transiently associate with a resident endoplasmic reticulum protein of 88 kD that has been proposed to act as a chaperone for class I assembly. Subsequently, this protein was demonstrated to be identical to calnexin and to associate with immature forms of the T cell receptor complex, immunoglobulin, and human class I HLA heavy chains. In this paper we define further the interaction of human dass I HLA heavy chains with chaperone proteins and find key differences with the complexes observed in the mouse system. First, calnexin and immunoglobulin binding protein (BiP) both associate with immature HLA class I heavy chains. The two chaperones are not found within the same molecular complex, suggesting that calnexin and BiP do not interact simultaneously with the same HLA class I heavy chain. Second, only free HLA class I heavy chains, and not 32-microglobulin (Bzm)-associated heavy chains are found associated with the chaperones. Indeed, addition of free 32m in vitro induces dissociation of chaperone-class I HLA heavy chain complexes. The kinetics for dissociation of the class I HLA heavy chain-chaperone complexes and for formation of the class I HLA heavy chain-32m complex display a reciprocity that suggests the interactions with chaperone and 32m are mutually exclusive. Mouse class I heavy chains expressed in human cells exhibit the mouse pattern of interaction with human chaperones and human 32m and not the human pattern, showing the difference in behavior is purely a function of the class I heavy chain sequence.

The fms-related tyrosine kinase 3 (FLT3) receptor has been extensively studied over the past two decades with regard to oncogenic alterations that do not only serve as prognostic markers but also as therapeutic targets in acute myeloid leukemia (AML). Internal tandem duplications (ITDs) became of special interest in this setting as they are associated with unfavorable prognosis. Because of sequence-dependent protein conformational changes FLT3-ITD tends to autophosphorylate and displays a constitutive intracellular localization. Here, we analyzed the effect of tyrosine kinase inhibitors (TKIs) on the localization of the FLT3 receptor and its mutants. TKI treatment increased the surface expression through upregulation of FLT3 and glycosylation of FLT3-ITD and FLT3-D835Y mutants. In T cell-mediated cytotoxicity (TCMC) assays, using a bispecific FLT3 × CD3 antibody construct, the combination with TKI treatment increased TCMC in the FLT3-ITD-positive AML cell lines MOLM-13 and MV4-11, patient-derived xenograft cells and primary patient samples. Our findings provide the basis for rational combination of TKI and FLT3-directed immunotherapy with potential benefit for FLT3-ITD-positive AML patients.

Cell-based immunotherapy in settings of allogeneic stem cell transplantation or donor leukocyte infusion has curative potential, especially in hematologic malignancies. However, this approach is severely restricted due to graft-versus-host disease (GvHD). This limitation may be overcome if target antigens are molecularly defined and effector cells are specifically selected. We chose formin-related protein in leukocytes 1 (FMNL1) as a target antigen after intensive investigation of its expression profile at the mRNA and protein levels. Here, we confirm re- IntroductionAlthough it is controversially discussed whether adaptive immune responses in tumor-bearing hosts play a role in controlling growth and recurrence of human tumors, 1,2 T cells can be converted to highly efficient killers of tumors. Donor leukocyte infusions (DLIs) are responsible for a graft-versus-leukemia effect (GvL) after allogeneic stem-cell transplantation (SCT), representing a therapeutic option with curative potential in different diseases at advanced stages. 3 Although GvL responses have also been shown for low-grade lymphomas, improvement of long-term survival after allogeneic transplantation has not been demonstrated in these patients. [4][5][6][7][8] This can be attributed primarily to the high treatmentassociated mortality due to transfer of T cells recognizing allogeneic minor histocompatibility (MHC) antigens, thereby causing potentially life-threatening graft-versus-host disease (GvHD). For broader applicability of this therapeutic option, it is therefore essential to reduce the risk of detrimental GvHD.One approach to gain a tumor-specific effect while significantly reducing the risk of GvHD is to generate allorestricted T cells with specificity for epitopes derived from tumor-associated antigens (TAAs). 9 Such allorestricted peptide-specific T cells may display high avidity toward MHC-presented TAAs, because they have not been negatively selected in the thymus. Moreover, they can be isolated by tetramers and cloned by limiting dilution to identify the specific TCR responsible for tumor-selective killing. 10 The isolation of a TCR with defined specificity for TAAs facilitates genetic TCR transfer into T-cell lines, 11-14 allowing major expansion of tumor-specific T cells.The choice of an appropriate target antigen is of fundamental importance for the success of an antitumor immunotherapeutic approach. So far, there is no universal antigen that serves as a target antigen in a broad range of malignancies. Many tumor-specific antigens are derived from aberrant expression of cell type-specific proteins and are expressed only in a restricted number of tumor types, limiting the tumors that can be treated. However, even the presence of a suitable antigen does not guarantee effective antigen recognition, because tumor cells evade immune recognition by down-regulation of the specific target or by inhibition of MHC class I-restricted antigen presentation. Thus, the level of epitope presentation in the context of classical or nonclassical MHC...