Addressing Cellular Heterogeneity in Cancer through Precision Proteomics

Abstract

Cells exhibit a broad spectrum of functions driven by differences in molecular phenotype. Understanding the heterogeneity between and within cell types has led to advances in our ability to diagnose and manipulate biological systems. Heterogeneity within and between tumors still poses a challenge to the development and efficacy of therapeutics. In this Perspective we review the toolkit of protein-level experimental approaches for investigating cellular heterogeneity. We describe how innovative approaches and technical developments have supported the advent of bottom-up singlecell proteomic analysis and present opportunities and challenges within cancer research. Finally, we introduce the concept of "precision proteomics" and discuss how the advantages and limitations of various experimental approaches render them suitable for different biological systems and questions.

Droplet based low input proteomic platform for rare cell populations

“…These results show that most of the observed protein variance is due to biological differences between cell lines rather than experimental variance. Importantly, proteomic profile differences acquired by DROPPS were highly similar to proteomes acquired in two independent bulk proteomic datatsets 13,14 (Spearman’s ρ of 0.62 and 0.64, both P < 2.2 x 10 -16 ) (Extended Data Figure 2d-e). Furthermore, our data also successfully clustered breast cancer cell lines into molecular subtypes of TNBC 21 indicating that the profiles generated by DROPPS accurately reflect previously described phenotypic differences (Figure 2f, Extended Data Figure 2f, Supporting Information Table 1).…”

“…[13][14][15] Due to the extensive sample processing required prior to data acquisition, such proteomic methods often require 10,000s -1,000,000s of cells (roughly, 10 -100's µg) as starting material, which can preclude the analysis of rare cell populations or other sample-limited systems. 16 Recent single-cell or low-input proteomic methods typically leverage low-volumes and minimal manual manipulation to reduce losses during sample preparation. [17][18][19] Despite these technical advances, many low-input methods only support specific study design (e.g., isobaric labeling with booster channel) or require specialized equipment (e.g., microfabrication capabilities, microfluidic cell isolation systems).…”

Translational Animal Models Provide Insight Into Mesenchymal Stromal Cell (MSC) Secretome Therapy

“…Our group used single-cell transcriptomics to analyze donor-matched equine MSCs isolated from three different tissue sources and we found inter-and intra-source genetic heterogeneity that resulted in functional heterogeneity in immune function and cell motility (Harman et al, 2020). The emerging technology of high-resolution precision proteomics is currently only being used to evaluate cancer cellular heterogeneity (Waas and Kislinger, 2020), but will certainly be transferrable to MSCs, where this technique can provide additional insights into the heterogeneity of MSC populations to allow for the purification of MSC subpopulations with high secretory potential.…”

Biological mass spectrometry enables spatiotemporal ‘omics: From tissues to cells to organelles

“…Thousands of different proteins can be identified using this approach, even from limited samples. Bottom‐up proteomics was applied to study complex spatiotemporal processes at the organismal to subcellular levels in development (Baxi et al, 2021; L. W. Yang et al, 2020), neuroscience (Hobson et al, 2022; Velasquez et al, 2019), and various disease states (Vo & Palsson, 2007; Waas & Kislinger, 2020). The information from these measurements provides the foundation for hypothesis‐driven and functional studies.…”