Significant advancements in the field of preclinical in vitro blood-brain barrier (BBB) models have been achieved in recent years, by developing monolayer-based culture systems towards complex multi-cellular assays. The coupling of those models with other relevant organoid systems to integrate the investigation of blood-brain barrier permeation in the larger picture of drug distribution and metabolization is still missing. Here, we report for the first time the combination of a human induced pluripotent stem cell (hiPSC)-derived blood-brain barrier model with a cortical brain and a liver spheroid model from the same donor in a closed microfluidic system (MPS). The two model compounds atenolol and propranolol were used to measure permeation at the blood–brain barrier and to assess metabolization. Both substances showed an in vivo-like permeation behavior and were metabolized in vitro. Therefore, the novel multi-organ system enabled not only the measurement of parent compound concentrations but also of metabolite distribution at the blood-brain barrier.

Hematologic adverse events are common dose-limiting toxicities in drug development. Classical animal models for preclinical safety assessment of immunotherapies are often limited due to insufficient cross-reactivity with non-human homologous proteins, immune system differences, and ethical considerations. Therefore, we evaluate a human bone marrow (BM) microphysiological system (MPS) for its ability to predict expected hematopoietic liabilities of immunotherapeutics. The BM-MPS consists of a closed microfluidic circuit containing a ceramic scaffold covered with human mesenchymal stromal cells and populated with human BM-derived CD34+ cells in chemically defined growth factor-enriched media. The model supports on-chip differentiation of erythroid, myeloid and NK cells from CD34+ cells over 31 days. The hematopoietic lineage balance and output is responsive to pro-inflammatory factors and cytokines. Treatment with a transferrin receptor-targeting IgG1 antibody results in inhibition of on-chip erythropoiesis. The immunocompetence of the chip is established by the addition of peripheral blood T cells in a fully autologous setup. Treatment with T cell bispecific antibodies induces T cell activation and target cell killing consistent with expected on-target off-tumor toxicities. In conclusion, this study provides a proof-of-concept that this BM-MPS is applicable for in vitro hematopoietic safety profiling of immunotherapeutics.

Microphysiological systems are fundamental for progressing towards a global paradigm shift in drug development through the generation of patient-on-a-chip models. An increasing number of single- and multi-organ systems have been adopted by the pharmaceutical and cosmetic industries for predictive substance testing. These models run on heterogeneous tissues and cell types from different donors. However, a patient is an individual. Therefore, patient-on-a-chip systems need to be built from tissues from one autologous source. Individual on-chip organ differentiation from a single induced pluripotent stem cell source could provide a solution to this challenge. We designed a four-organ chip based on human physiology. It enables the interconnection of miniaturized human intestine, liver, brain and kidney equivalents. All four organ models were predifferentiated from induced pluripotent stem cells from the same healthy donor and integrated into the microphysiological system. The cross talk led to further differentiation over a 14-day cultivation period under pulsatile blood flow conditions in one common medium deprived of growth factors. This model platform will pave the way for disease induction and subsequent drug testing.

Introduction: The human bone marrow is a complex organ that is critical for self-renewal and differentiation of hematopoietic progenitor cells into various lineages of blood cells. Perturbations of the hematopoietic system have been reported to cause numerous diseases. Yet, understanding the fundamental biology of the human bone marrow in health and disease and during the preclinical stages of drug development is challenging due to the complexity of studying or manipulating the human bone marrow. Human cell-based microfluidic bone marrow models are promising research tools to explore multi-lineage differentiation of human stem and progenitor cells over long periods of time.Methods: Human hematopoietic stem and progenitor cells were cultured with mesenchymal stromal cells on a zirconium oxide ceramic scaffold in a microfluidic device recapitulating the human bone marrow. NK cell differentiation was induced by the application of a lymphoid cultivation medium containing IL-15. The kinetics of differentiation into mature NK cells was traced by flow cytometry over a period of up to seven weeks, and functionality was measured by stimulation with phorbol myristate acetate (PMA) and ionomycin. The effect of an anti-IL-15 monoclonal antibody (TEV-53408) on different NK cell subtypes was tested at different time points.Results: Our data shows that within 28 days of culture, differentiation into all developmental stages of NK cells was accomplished in this system. Alongside with the NK cells, myeloid cells developed in the system including granulocytes, monocytes and dendritic cells. The differentiated NK cells could be activated after stimulation with PMA and ionomycin indicating the functionality of the cells. Treatment with an anti-IL-15 antibody induced a reduction in proliferation of late-stage NK cells as shown by EdU staining. This led to significantly dose dependent reduction in the number of circulating stage 4 - 6 NK cells in the system after one week of treatment. This effect was partially reversible after a two-week treatment-free period.Discussion: In summary, the presented model enables investigation of human NK cell development in the bone marrow and provides a basis to study related diseases and drug response effects in a microenvironment that is designed mimic human physiology.