Herweck, F. | bioRxiv Science

bioRxiv2026

Macrophages target PVR/CD155 on colorectal cancer cells via REVERBalpha

Background&AimsPatients with microsatellite-instable (MSI) colorectal cancer (CRC) benefit from immune checkpoint therapy. For patients with microsatellite-stable (MSS) non-MSI tumors targeting alternative immune checkpoints, such as the Poliovirus receptor (PVR/CD155), may extend response to checkpoint inhibitors and thereby improve outcomes. The drugable transcription factor REVERB (NR1D1) is a master repressor of macrophage function. We hypothesized that regulation of PVR allows elimination of tumor cells by macrophage-directed precision therapy. MethodsHuman CRC cell lines (MSS+: HT29, SW480; MSI+: HCT116), patient-derived organoids (MSS+ PDOs) and tissues were assessed by PCR, Western blot, immunohistochemistry and flow cytometry. 3D co-cultures of CRC cells with macrophages derived from THP1 monocytic leukemia cells or peripheral blood of healthy donors were analysed by microscopy and viability assays. Functional perturbation of PVR and REVERB was achieved by CRISPR/Cas9-sgRNA gene modification, synthetic ligands or blocking antibodies (Abs). ResultsREVERB bound a cognate DNA-element in the -1 kb human PVR gene promoter, and its agonist (SR9009) super-repressed, whereas its antagonist (SR8278) de-repressed transcription of PVR mRNA in macrophages. Macrophages with CRISPR-modified REVERB were unresponsive to ligand, devoid of PVR protein, showed more phagocytosis, tumor cell efferocytosis and expression of genes related to host immunity (PDL1, TLR4 e.a.) than clones with the wild-type receptor. Macrophages lowered the viability of tumor cells, potentiated by PVR blocking Ab or PVR knock-out in tumor cells. Consistently, REVERB antagonist augmented tumor cell death in co-cultures with macrophages and PVR blocking Ab. ConclusionCo-addressing the checkpoint axis REVERB-PVR may represent a novel intervention strategy for patients with MSS+ CRC.

cancer biology↗

bioRxiv2019

Multiparametric phenotyping of compound effects on patient derived organoids

Patient derived organoids resemble the biology of tissues and tumors, enabling ex vivo modeling of human diseases from primary patient samples. Organoids can be used as models for drug discovery and are being explored to guide clinical decision making. Patient derived organoids can have heterogeneous morphologies with unclear biological causes and relationship to treatment response. Here, we used high-throughput, image-based profiling to quantify phenotypes of over 5 million individual colorectal cancer organoids after treatment with more than 500 small molecules. Integration of data using a joint multi-omics modelling framework identified organoid size and cystic vs. solid organoid architecture as axes of morphological variation across organoids. Mechanistically, we found that organoid size was linked to IGF1 receptor signaling, while a cystic organoid architecture was associated with an LGR5+ stemness program. Treatment-induced organoid morphology reflected organoid viability, drug mechanism of action, and was biologically interpretable using joint modelling. Inhibition of MEK led to cystic reorganization of organoids and increased expression of LGR5, while inhibition of mTOR induced IGF1 receptor signaling. In conclusion, we identified shared axes of variation for colorectal cancer organoid morphology, their underlying biological mechanisms, and pharmacological interventions with the ability to move organoids along them. Image-based profiling of patient derived organoids coupled with multi-omics integration facilitates drug discovery by linking drug responses with underlying biological mechanisms.

genomics↗