Ex vivo cultures and drug testing of primary acute myeloid leukemia samples: Current techniques and implications for experimental design and outcome.

New treatment options of acute myeloid leukemia (AML) are rapidly emerging. Pre-clinical models such as ex vivo cultures are extensively used towards the development of novel drugs and to study synergistic drug combinations, as well as to discover biomarkers for both drug response and anti-cancer drug resistance. Although these approaches empower efficient investigation of multiple drugs in a multitude of primary AML samples, their translational value and reproducibility are hampered by the lack of standardized methodologies and by culture system-specific behavior of AML cells and chemotherapeutic drugs. Moreover, distinct research questions require specific methods which rely on specific technical knowledge and skills. To address these aspects, we herein review commonly used culture techniques in light of diverse research questions. In addition, culture-dependent effects on drug resistance towards commonly used drugs in the treatment of AML are summarized including several pitfalls that may arise because of culture technique artifacts. The primary aim of the current review is to provide practical guidelines for ex vivo primary AML culture experimental design.

Modeling BCR-ABL and MLL-AF9 leukemia in a human bone marrow-like scaffold-based xenograft model.

Although NOD-SCID IL2Rγ-/- (NSG) xenograft mice are currently the most frequently used model to study human leukemia in vivo, the absence of a human niche severely hampers faithful recapitulation of the disease. We used NSG mice in which ceramic scaffolds seeded with human mesenchymal stromal cells were implanted to generate a human bone marrow (huBM-sc)-like niche. We observed that, in contrast to the murine bone marrow (mBM) niche, the expression of BCR-ABL or MLL-AF9 was sufficient to induce both primary acute myeloid leukemia (AML) and acute lymphocytic leukemia (ALL). Stemness was preserved within the human niches as demonstrated by serial transplantation assays. Efficient engraftment of AML MLL-AF9 and blast-crisis chronic myeloid leukemia patient cells was also observed, whereby the immature blast-like phenotype was maintained in the huBM-sc niche but to a much lesser extent in mBM niches. We compared transcriptomes of leukemias derived from mBM niches versus leukemias from huBM-like scaffold-based niches, which revealed striking differences in the expression of genes associated with hypoxia, mitochondria and metabolism. Finally, we utilized the huBM-sc MLL-AF9 B-ALL model to evaluate the efficacy of the I-BET151 inhibitor in vivo. In conclusion, we have established human niche models in which the myeloid and lymphoid features of BCR-ABL+ and MLL-AF9+ leukemias can be studied in detail.

Upregulation of CD38 expression on multiple myeloma cells by all-trans retinoic acid improves the efficacy of daratumumab.

Daratumumab is an anti-CD38 monoclonal antibody with lytic activity against multiple myeloma (MM) cells, including ADCC (antibody-dependent cellular cytotoxicity) and CDC (complement-dependent cytotoxicity). Owing to a marked heterogeneity of response to daratumumab therapy in MM, we investigated determinants of the sensitivity of MM cells toward daratumumab-mediated ADCC and CDC. In bone marrow samples from 144 MM patients, we observed no difference in daratumumab-mediated lysis between newly diagnosed or relapsed/refractory patients. However, we discovered, next to an expected effect of effector (natural killer cells/monocytes) to target (MM cells) ratio on ADCC, a significant association between CD38 expression and daratumumab-mediated ADCC (127 patients), as well as CDC (56 patients). Similarly, experiments with isogenic MM cell lines expressing different levels of CD38 revealed that the level of CD38 expression is an important determinant of daratumumab-mediated ADCC and CDC. Importantly, all-trans retinoic acid (ATRA) increased CD38 expression levels but also reduced expression of the complement-inhibitory proteins CD55 and CD59 in both cell lines and primary MM samples. This resulted in a significant enhancement of the activity of daratumumab in vitro and in a humanized MM mouse model as well. Our results provide the preclinical rationale for further evaluation of daratumumab combined with ATRA in MM patients.