Immunological substance testing on human lymphatic micro-organoids in vitro

Abstract

Pharmaceutical drugs and compounds used for consumer products may bear the risk of unexpected immuno-toxicological side effects, such as sensitization, allergy, anaphylaxis or immunogenicity. Modern biopharmaceuticals with high potency and target specificity, like antibodies and cytokines need to be tested for their therapeutical doses, their exposition regimens and their immune functionality prior to first-in-man applications. For the latter, existing in vitro tests and animal models do not sufficiently reflect the complexity and specificity of the human immune system. Even novel humanised animal models have limitations in their systemic reactions. Monolayer or suspended cell culture possesses neither tissue functionality nor organ physiology, and also cannot be used for long term culture and experiments. In contrast, solid tissue biopsies, e.g. tonsil preparations of tonsillitis patients typically show inflammatory artefacts and degrade in long term culture due to preparation-induced damage. The construction of tissue-like structures in vitro, so-called "micro-organoids", can overcome these limitations. Key structures of secondary lymphatic organs, e.g. lymph nodes or the spleen are the primary lymphatic follicles and germinal centres, in particular during the "activated state" of an inflammation or infection. To remodel lymphatic follicles, functional and structural cells, e.g. lymphoid cells derived from peripheral blood mononuclear cells (PBMCs) and stromal cells need to be combined with biogenic or artificial matrices and scaffolds to produce a suitable 3D tissue-mimicking environment. Therefore, a unique human lymph node model (HuALN) was designed to operate over several weeks, and allow long term and repeated drug exposure to induce and monitor both cellular and humoral immune responses. Cellular immunity is monitored, for example, by cytokine release patterns; humoral immunity is analysed, for example, by B cell activation, plasma cell formation and antibody secretion profiles (IgM and IgG). Moreover, cellular composition and micro-organoid formation are analysed by flow cytometry, histology and in situ imaging.