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In acute lymphoblastic leukemia (ALL), conventional cell lines do not recapitulate the clonal diversity and microenvironment. Orthotopic patient-derived xenograft models (PDX) overcome these limitations and mimic the clinical situation, but molecular stability and engraftment patterns have not yet been thoroughly assessed. We herein describe and characterize the PDX generation in NSG mice. In vivo tumor cell proliferation, engraftment and location were monitored by flow cytometry and bioluminescence imaging. Leukemic cells were retransplanted for up to four passages, and comparative analyses of engraftment pattern, cellular morphology and genomic hotspot mutations were conducted. Ninety-four percent of all samples were successfully engrafted, and the xenograft velocity was dependent on the molecular subtype, outcome of the patient and transplantation passage. While BCR::ABL1 blasts were located in the spleen, KMT2A-positive cases had higher frequencies in the bone marrow. Molecular changes appeared in most model systems, with low allele frequency variants lost during primary engraftment. After the initial xenografting, however, the PDX models demonstrated high molecular stability. This protocol for reliable ALL engraftment demonstrates variability in the location and molecular signatures during serial transplantation. Thorough characterization of experimentally used PDX systems is indispensable for the correct analysis and valid data interpretation of preclinical PDX studies.

The requirements for bone substitute materials are multifaceted. Beside biomechanical stability, these materials should provide osteoconductive and osteoinductive properties to promote integration into the host tissue. So far, autologous bone is the only material, which combines all properties, but is naturally limited. Allogenic bone grafts have to be decellularized prior to implantation. This causes the reduction of biomechanical properties and the loss of osteoinductive qualities. High hydrostatic pressure (HHP) offers a gentle alternative for processing and supply of allogenic bone substitute materials while preserving biomechanical integrity. To determine whether osteogenic properties are retained by HHP treatment, mesenchymal stem cells (MSCs) were cultured with HHP‐treated and untreated allogenic trabecular bone blocks up to 28 days. Both, gene expression and protein analysis showed that HHP‐treated bone positively influenced differentiation of MSCs into osteoblasts and mineralization of bone matrix. This effect was greater in samples cultivated with HHP‐treated bone blocks. The present study shows that HHP treatment does not result in the reduction of osteoinductivity, thus serving as an alternative approach for processing allogeneic bone substitute materials.