Multiplexed single-cell analysis of organoid signaling networks

Abstract

This multiplexed mass cytometry protocol uses Thiol-reactive Organoid Barcoding in situ (TOBis) and a CyTOF siGNalling AnaLysis pipeline (CyGNAL) to enable 126-plex single-cell analysis of cell-type, cell-state, and posttranslational modification signalling network in organoids.Tweet A multiplexed mass cytometry protocol using Thiol-reactive Organoid Barcoding in situ (TOBis) and a CyTOF siGNalling AnaLysis pipeline (CyGNAL) for 126-plex single-cell analysis of cell-type-specific PTM signalling in organoids (from @QinXiao1990 @FerranC96 and @christophertape).

Oncogenic signalling is coupled to colorectal cancer cell differentiation state

“…Several studies analysed single-cell transcriptomes of CRC organoids and tumours, showing that cell-state heterogeneity exists in the transcriptome of tumours with implications for therapy. 14,40,41 However, the study of underlying signalling networks has only recently gained attention 9,42 as MC was previously mainly limited to the study of extracellular markers of immune cells. Application of MC on primary cancer tissue holds the promise of dissecting signalling heterogeneity in cell-state gradients of tumour tissue.…”

Everything You Always Wanted to Know About Organoid-Based Models (and Never Dared to Ask)

“… 45 , 188 , 189 , 190 Recently, a multiplex single-cell analysis pipeline was developed on organoids co-cultured with fibroblast and leukocytes to establish post-translational modification signaling networks that can be altered in diseases. 191 For example, growing organoids from patient-derived stem cell aggregates in preformed U-shaped microcavities imprinted in the hydrogel achieves highly homogenous cultures, both in size and maturation level. In addition, in this high-throughput single organoid model, cells will be positioned on the same Z plane, thus facilitating the automated live bioimaging screening of various drugs for the development of personalized medicine.…”

Preclinical platforms to study therapeutic efficacy of human γδT‐cells for oncology indications

“…In the design of ex‐vivo models, we next consider the model matrix. While often centring on generalizable, all‐purpose materials such as Matrigel, 4 substantial recent advances have been made in studying the significance of the physical parameters of 3D cancer models, including stiffness and compartmentalisation 5 . Work from Bakkalci et al., 6 for example, describes a model of bone cancer, ameloblastoma, which consisted of engineering active bone‐forming stroma to probe the interaction of ameloblastoma with its native bony microenvironment.…”

“…While ideally autologous to the tumour, bystander cells allogeneic to the tumour can be used for modelling non‐alloreactive therapeutics like γδT‐cells. Depending on availability and tolerance to various culture conditions, a mixture of primary and immortalized bystanders may be used, such as in colorectal carcinoma organoid modelling by Qin and colleagues 4 . When investigating the homing potential of cell‐based therapies to a tumour and its microenvironment following intravenous infusion, ‘organ‐on‐a‐chip’ microfluidic systems offer suitable advanced modelling strategies.…”