Patient-derived organoids (PDOs) have demonstrated predictive value in prospective clinical trials supporting selection of personalized treatments. Because PDOs retain the organization and physiological functions of their source tissue, PDO biobanks could also be an ideal substrate to screen for novel therapeutic interventions. Here we describe a large-scale functional screen of dual targeting bispecific antibodies (bAbs) on a colorectal cancer (CRC) PDO biobank to target their dependency on cancer stem cells. A novel drug discovery pipeline was assembled where therapeutic bAb panels generated against WNT and receptor tyrosine kinases (RTK) targets were functionally evaluated by high content imaging to capture the complexity of PDO responses across a wide range of different CRCs and paired normal colonic mucosa samples. Our strategy resulted in the generation of MCLA-158, a bAb that specifically triggers EGFR degradation in LGR5+ cancer stem cells but shows minimal toxicity towards normal LGR5+ colon stem cells. MCLA-158 exhibits unique therapeutic properties such as potent growth inhibition of KRAS mutant CRCs, blockade of metastasis initiation and suppression of tumor outgrowth in preclinical models of different cancer types.
Niches in the bone marrow regulate hematopoietic stem and progenitor cell (HSPC) fate and behavior through cell–cell interactions and soluble factor secretion. The niche‐HSPC crosstalk is a very complex process not completely elucidated yet. To aid further investigation of this crosstalk, a functional in vitro 3D model that closely represents the main supportive compartments of the bone marrow is developed. Different combinations of human stromal cells and hydrogels are tested for their potential to maintain CD34+ HSPCs. Cell viability, clonogenic hematopoietic potential, and surface marker expression are assessed over time. Optimal HSPC support is obtained in presence of adipogenic and osteogenic cells, together with progenitor derived endothelial cells. When cultured in a bioactive hydrogel, the supportive cells self‐assemble into a hypoxic stromal network, stimulating CD34+CD38+ cell formation, while maintaining the pool of CD34+38− HSPCs. HSPC clusters colocalize with the stromal networks, in close proximity to sinusoidal clusters of CD31+ endothelial cells. Importantly, the primary in vitro niche model supports HSPCs with no cytokine addition. Overall, the engineered primary 3D bone marrow environment provides an easy and reliable model to further investigate interactions between HSPCs and their endosteal and perivascular niches, in the context of normal hematopoiesis or blood‐related diseases.
Introduction The pharmaceutical industry has mostly relied on 2D cancer cell lines and 3D spheroids for in vitro testing, but poor correlation of pre-clinical and clinical outcomes has driven the development of more predictive models. Patient-derived organoids (PDOs) have emerged as representative in vitro avatars of tumor biology, allowing the generation of biobanks covering a large variety in indications and genetic backgrounds. Here, we assess the robustness of our drug screening platform by testing the reproducibility of our organoid assays within and between organoid batches, read-outs, and different labs. We present an assay-ready organoid platform, allowing short timelines, repeated assays from a single batch of organoids, high throughput assays and large panel screens. Methods Colorectal, breast, lung, pancreatic, ovarian, cervix and melanoma PDO models were selected for subtype and driver mutation variety, banked in large batches, and preserved in assay-ready format. Outgrowth and drug responses of these batches were compared to those of organoids produced in the classic method. Experiments were executed using automated liquid handling equipment to standardize procedures and increase consistency. Performance of the drug screens were assessed by repeated drug sensitivity testing and by calculating intra- and interplate variability, control variability (CV), Z-factor, assay windows and IC50 values. Moreover, drug responses were tested in both celltiterglo-based and high content imaging-based assays. Results Assay performance of the organoid drug testing platform was high, with high Z-factors (>0.6), and low intra- and interplate variability (CV <15%) indicating reproducible assays. Both the classic and assay-ready organoid technologies resulted in highly reproducible IC50outputs, also when assays were executed months apart or in different labs. Timelines were shortened from months to weeks, and the reduced logistical burden allows screening of large organoid panels of >50 models. Drug sensitivity testing in our fast-paced panel screening platform distinguished sensitive from partially and insensitive models, illustrating how organoids allow patient stratification. Conclusion and Discussion The assay-ready organoid technology presented here has further improved our already highly predictive and reproducible organoid drug testing platform. The resulting short timelines and large panel screening capabilities will further unlock the great potential of PDO technology. Using large organoid panels in pre-clinical drug development and in combination with biomarker analysis will allow identification of responsive indications, subtypes, and genotypes, and for early patient stratification. Moreover, as most models presented in our panel screen platform are also available as PDX models, it allows for easy translation into in vivo follow-up studies. Citation Format: Liza Wijler, Jara Garcia Mateos, My Nguyen, Annelot Staes, Linda van Seters, Lama Alhaj Hasan, Victor Tiroille, Bram Herpers, Leo Price, Mariusz Madej, Michiel Fokkelman, Marrit Putker. Pan-cancer assay-ready organoid drug screening with robust, reproducible and clinically-relevant output [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 198.
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