Purpose Daratumumab treatment results in a marked reduction of CD38 expression on multiple myeloma (MM) cells. The aim of this study was to investigate the clinical implications and the underlying mechanisms of daratumumab-mediated CD38 reduction. Experimental design We evaluated the effect of daratumumab alone or in combination with lenalidomide-dexamethasone, on CD38 levels of MM cells and non-tumor immune cells in the GEN501 study (daratumumab monotherapy) and the GEN503 study (daratumumab combined with lenalidomide-dexamethasone). In vitro assays were also performed. Results In both trials daratumumab reduced CD38 expression on MM cells within hours after starting the first infusion, regardless of depth and duration of the response. In addition, CD38 expression on non-tumor immune cells, including NK-, T- B-cells and monocytes, was also reduced irrespective of alterations in their absolute numbers during therapy. In-depth analyses revealed that CD38 levels of MM cells were only reduced in the presence of complement or effector cells, suggesting that the rapid elimination of CD38high MM cells can contribute to CD38 reduction. In addition, we discovered that daratumumab-CD38 complexes and accompanying cell membrane were actively transferred from MM cells to monocytes and granulocytes. This process of trogocytosis was also associated with reduced surface levels of some other membrane proteins including CD49d, CD56, and CD138. Conclusion Daratumumab rapidly reduced CD38 expression levels, at least in part, through trogocytosis. Importantly, all these effects also occurred in patients with deep and durable responses, thus excluding CD38 reduction alone as a mechanism of daratumumab resistance.

In healthy humans, lymphocyte populations are maintained at a relatively constant size throughout life, reflecting a balance between lymphocyte production and loss. Given the profound immunological changes that occur during healthy aging, including a significant decline in T-cell production by the thymus, lymphocyte maintenance in the elderly is generally thought to require homeostatic alterations in lymphocyte dynamics. Surprisingly, using in vivo 2H2O labeling, we find similar dynamics of most lymphocyte subsets between young adult and elderly healthy individuals. As the contribution of thymic output to T-cell production is only minor from young adulthood onward, compensatory increases in peripheral T-cell division rates are not required to maintain the T-cell pool, despite a tenfold decline in thymic output. These fundamental insights will aid the interpretation of further research into aging and clinical conditions related to disturbed lymphocyte dynamics.

Interactions within the hematopoietic niche in the BM microenvironment are essential for maintenance of the stem cell pool. In addition, this niche is thought to serve as a sanctuary site for malignant progenitors during chemotherapy. Therapy resistance induced by interactions with the BM microenvironment is a major drawback in the treatment of hematologic malignancies and bone-metastasizing solid tumors. To date, studying these interactions was hampered by the lack of adequate in vivo models that simulate the human situation. In the present study, we describe a unique human-mouse hybrid model that allows engraftment and outgrowth of normal and malignant hematopoietic progenitors by implementing a technology for generating a human bone environment. Using luciferase gene marking of patient-derived multiple myeloma cells and bioluminescent imaging, we were able to follow pMM cells outgrowth and to visualize the effect of treatment. Therapeutic interventions in this model IntroductionIn the BM, specialized microenvironments such as hematopoietic niches regulate hematopoiesis. Within these niches, hematopoietic stem cells (HSCs) are present in a complex network consisting of mesenchymal stromal cells (MSCs), osteoblasts, osteoclasts, endothelial cells, and adipocytes embedded in an extracellular matrix. The bidirectional interactions with the hematopoietic niche are essential for HSC maintenance and function. [1][2][3][4] The BM niche is also thought to serve as a sanctuary site for leukemic stem cells (LSCs), which, in addition to their immortalizing genetic events, highly depend on interaction with the microenvironment to survive and proliferate. 5,6 Although the majority of leukemias initially respond to therapeutic intervention, relapse rates are high. [7][8][9] There is increasing evidence that the tumor niche plays a crucial role in the survival and drug resistance of LSCs. Interactions with the niche provide signals protecting the LSCs from apoptosis and eventually leading to the selection and outgrowth of a resistant cell. 10-13 Therefore, it is apparent that the hematopoietic niche plays an important role in hematopoietic development and in chemotherapy resistance of BM-localized leukemic and solid tumors.Although our understanding of how the BM niche regulates HSC self-renewal and confers therapy resistance has advanced greatly over the past years, most of this knowledge is based on genetic loss-of-function or gain-of-function murine models. 1,2,10,11,14 However, these murine models do not simulate human physiology and much of the constituents of the human hematopoietic niche remain largely unclear. [14][15][16] This emphasizes the need for more suitable models that recapitulate the human BM microenvironment and, very importantly, facilitate the engraftment and outgrowth of normal HSCs and patient-derived tumor cells within these protected sites.In the present study, we describe a unique humanized model that implements a novel scaffold-based technology for generating a human bone environment in RAG 2 Ϫ/Ϫ ...

The immune competent abdominal adipose tissue, either stored viscerally [visceral adipose tissue (VAT)] or sc [sc adipose tissue (SAT)], has been identified as a source of IL-1β and IL-18. To become active, the proforms of these cytokines require processing by caspase-1, which itself is mediated by the inflammasome. In this descriptive study, we investigate the expression of inflammasome components and caspase-1 in human fat and determine whether caspase-1 activity contributes to the enhanced inflammatory status of VAT. Paired SAT and VAT biopsies from 10 overweight subjects (body mass index, 25-28 kg/m(2)) were used to study the cellular composition and the intrinsic inflammatory capacity of both adipose tissue depots. The percentage of CD8(+) T cells within the lymphocyte fraction was significantly higher in VAT compared with SAT (41.6 vs. 30.4%; P < 0.05). Adipose tissue cultures showed a higher release of IL-1β (10-fold; P < 0.05), IL-18 (3-fold; P < 0.05), and IL-6 and IL-8 (3-fold, P < 0.05; and 4-fold, P < 0.05, respectively) from VAT compared with SAT that was significantly reduced by inhibiting caspase-1 activity. In addition, caspase-1 activity was 3-fold (P < 0.05) higher in VAT compared with SAT, together with an increase in the protein levels of the inflammasome members apoptosis-associated speck-like protein containing a C-terminal caspase-recruitment domain (2-fold; P < 0.05) and nucleotide-binding oligomerization domain-like receptor pyrin domain containing 3 (2-fold; nonsignificant). Finally, caspase-1 activity levels were positively correlated with the percentage of CD8(+) T cells present in adipose tissue. Our results show that caspase-1 and nucleotide-binding oligomerization domain-like receptor pyrin domain containing 3 inflammasome members are abundantly present in human VAT. The increased intrinsic caspase-1 activity in VAT represents a novel and specific inflammatory pathway that may determine the proinflammatory character of this specific depot.

To begin to understand the mechanisms that regulate self-renewal, differentiation, and transformation of human hematopoietic stem cells or to evaluate the efficacy of novel treatment modalities, stem cells need to be studied in their own species-specific microenvironment. By implanting ceramic scaffolds coated with human mesenchymal stromal cells into immune-deficient mice, we were able to mimic the human bone marrow niche. Thus, we have established a human leukemia xenograft mouse model in which a large cohort of patient samples successfully engrafted, which covered all of the important genetic and risk subgroups. We found that by providing a humanized environment, stem cell self-renewal properties were better maintained as determined by serial transplantation assays and genome-wide transcriptome studies, and less clonal drift was observed as determined by exome sequencing. The human leukemia xenograft mouse models that we have established here will serve as an excellent resource for future studies aimed at exploring novel therapeutic approaches.