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Single Cell Proteomics Analysis of Drug Response Shows its Potential as Drug Discovery Platform

ABSTRACT: Recently, single cell proteomics techniques that allow quantification of thousands of proteins from single mammalian cells, have been introduced. Here we present an improved single cell proteomics platform named SCREEN (Single Cell pRotEomE aNalysis) mass spectrometry platform for deeper high throughput single cell proteome coverage with higher efficiency, less time and with an improved ability to see protein quantitation across more cells. We applied this new platform to analyze single cell proteomic responses upon different drug treatment times to uncover heterogeneity in cancer cell responses that is done for the first time based on mass spectrometry analysis. We discuss the challenges in single cell proteomics and future improvements in technique and general trends with the view to encourage forthcoming technical developments.

INSTRUMENT(S): Orbitrap HFX

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Bogdan Budnik

PROVIDER: MSV000088243 | MassIVE | Sun Oct 17 16:39:00 BST 2021

REPOSITORIES: MassIVE

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Project description:The traditional method for studying cancer in vitro is to grow immortalized cancer cells in two-dimensional (2D) monolayers on plastic. However, many cellular features are impaired in these unnatural conditions and big alterations in gene expression in comparison to tumors have been reported. Three-dimensional (3D) cell culture models have become increasingly popular and are suggested to be better models than 2D monolayers due to improved cell-to-cell contacts and structures that resemble in vivo architecture. The aim of this study was to develop a simple high-throughput 3D drug screening method and to compare drug responses in JIMT1 breast cancer cells when grown in 2D, in polyHEMA coated anchorage independent 3D models and in Matrigel on-top 3D cell culture models. We screened 102 compounds with multiple concentrations and biological replicates for their effects on cell proliferation. The cells were either treated immediately upon plating or they were allowed to grow in 3D for four days prior to the drug treatment. Big variations in drug responses were observed between the models indicating that comparisons of culture model influenced drug sensitivities cannot be made based on effects of a single drug. However, we show with the 63 most prominent drugs that, in general, JIMT1 cells grown on Matrigel were significantly more sensitive to drugs than cells grown in 2D cultures, while responses of cells grown in polyHEMA resembled those of 2D. Furthermore, comparison of gene expression profiles of the cell culture models to xenograft tumors indicated that cells cultured in Matrigel and as xenografts most closely resembled each other. In this study we also suggest that 3D cultures can provide a platform for systematic experimentation of larger compound collections in a high-throughput mode and be used as alternatives for traditional 2D screens towards better comparability to in vivo state. Gene expression analysis of JIMT1 breast cancer cells cultured as xenografts for 43 days, in two dimensional cultures for seven days (2D7d), in polyHEMA three dimensional cell culture models for four and seven days (PH7d and PH7d), and in Matrigel three dimensional cultures for four and seven days (MG4d and MG7d). Two biological replicates was included for each sample.

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Single Cell Proteomics Analysis of Drug Response Shows its Potential as Drug Discovery Platform

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