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T cell bispecific antibodies (TCBs) are the focus of intense development for cancer immunotherapy. Recently, peptide-MHC (major histocompatibility complex)-targeted TCBs have emerged as a new class of biotherapeutics with improved specificity. These TCBs simultaneously bind to target peptides presented by the polymorphic, species-specific MHC encoded by the human leukocyte antigen (HLA) allele present on target cells and to the CD3 coreceptor expressed by human T lymphocytes. Unfortunately, traditional models for assessing their effects on human tissues often lack predictive capability, particularly for "on-target, off-tumor" interactions. Here, we report an immune-infiltrated, kidney organoid-on-chip model in which peripheral blood mononuclear cells (PBMCs) along with nontargeting (control) or targeting TCB-based tool compounds are circulated under flow. The target consists of the RMF peptide derived from the intracellular tumor antigen Wilms' tumor 1 (WT1) presented on HLA-A2 via a bivalent T cell receptor-like binding domain. Using our model, we measured TCB-mediated CD8 + T cell activation and killing of RMF-HLA-A2-presenting cells in the presence of PBMCs and multiple tool compounds. DP47, a non-pMHC-targeting TCB that only binds to CD3 (negative control), does not promote T cell activation and killing. Conversely, the nonspecific ESK1-like TCB (positive control) promotes CD8 + T cell expansion accompanied by dose-dependent T cell-mediated killing of multiple cell types, while WT1-TCB* recognizing the RMF-HLA-A2 complex with high specificity, leads solely to selective killing of WT1-expressing cells within kidney organoids under flow. Our 3D kidney organoid model offers a platform for preclinical testing of cancer immunotherapies and investigating tissue-immune system interactions.

Traditional drug safety assessment often fails to predict complications in humans, especially when the drug targets the immune system. Here, we show the unprecedented capability of two human Organs-on-Chips to evaluate the safety profile of T-cell bispecific antibodies (TCBs) targeting tumor antigens. Although promising for cancer immunotherapy, TCBs are associated with an on-target, off-tumor risk due to low levels of expression of tumor antigens in healthy tissues. We leveraged in vivo target expression and toxicity data of TCBs targeting folate receptor 1 (FOLR1) or carcinoembryonic antigen (CEA) to design and validate human immunocompetent Organs-on-Chips safety platforms. We discovered that the Lung-Chip and Intestine-Chip could reproduce and predict target-dependent TCB safety liabilities, based on sensitivity to key determinants thereof, such as target expression and antibody affinity. These novel tools broaden the research options available for mechanistic understandings of engineered therapeutic antibodies and assessing safety in tissues susceptible to adverse events., Competing Interests: SK Is a current employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). Is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers 'Method for Assessing a Compound Interacting with a Target on Epithelial Cells', CB, HG, KK Is a former employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). Is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers 'Method for Assessing a Compound Interacting with a Target on Epithelial Cells', DP Is a former employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). Is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers Method for Assessing a Compound Interacting with a Target on Epithelial Cells,, MK Is a current or former employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). RB Is a former employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). Is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers, Method for Assessing a Compound Interacting with a Target on Epithelial Cells', JS, TW, TF, AF, JE, MG, AS, AG, TS, CK, MB Employment, patents and ownership of stock with Roche. CH, VM, RG, MB, EB Employment and ownership of stock with Roche. PN is a former employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). WT is a current employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). DM Is a current employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). CB, AR Is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers 'Method for Assessing a Compound Interacting with a Target on Epithelial Cells'. Employment, patents and ownership of stock with Roche. GH Is a current or former employee of and hold equity interests or options to obtain equity interests in (Emulate Inc). Is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers 'Method for Assessing a Compound Interacting with a Target on Epithelial Cells', AM is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers Method for Assessing a Compound Interacting with a Target on Epithelial Cells'. PB No competing interests declared, NG, LC is an inventor on a patent application (P16451EP00/16/912,391) submitted by Hoffmann-LaRoche and Emulate that covers Method for Assessing a Compound Interacting with a Target on Epithelial Cells'. Employment, patents and ownership of stock with Roche., (© 2021, Kerns et al.)

For low molecular weight drugs, lipid bilayer permeation is considered the major route for in vivo cell barrier passage. We recently introduced a fluorescence assay with liposomes to determine permeation kinetics of ionisable compounds across the lipid bilayer by monitoring drug-induced pH changes inside the liposomes. Here, we determined the permeability coefficients (P FLipP , FLipP for "Fluorescence Liposomal Permeability") across 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) bilayers of 35 ionisable drugs at pH6.0 and compared them to available in vivo human jejunal permeability (P eff ) data. P FLipP values were furthermore compared with published Caco-2 cell permeability coefficients (P Caco-2 ), permeability coefficients determined with the parallel artificial membrane permeability assay (PAMPA) and with log D (pH6.0). The log P FLipP , corrected for predicted para-cellular diffusion, and log P Caco-2 correlated best with log P eff , with similar adjusted R 2 (0.75 and 0.74, n=12). Our results suggest that transporter-independent intestinal drug absorption is predictable from liposomal permeability., (Copyright © 2017 Elsevier B.V. All rights reserved.)