Simple cell geometry metrics reveal cancer cell diversity and plasticity and guide combination therapy

Simple cell geometry metrics reveal cancer cell diversity and plasticity and guide combination therapy | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Biological Sciences - Article Simple cell geometry metrics reveal cancer cell diversity and plasticity and guide combination therapy Hadrien De Blander, Christos Gkemisis, Niccolo Roda, Carlos Ariel Pulido-Vincuna, and 17 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8278743/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted You are reading this latest preprint version Abstract Intra-tumour heterogeneity (ITH) and cellular plasticity contribute to therapy resistance. Single-cell transcriptomics is the gold standard for assessing ITH, but this approach remains costly and slow. Here, we show that distinct melanoma transcriptomic states exhibit unique cellular morphologies, and introduce ORIGAMI, a scalable flow cytometry assay leveraging cell surface area (S) and volume (V) measurements to resolve melanoma cell state diversity and to track therapy-induced transitions. MAPK- and CDK4/6-targeted therapies induce predictable shifts in cell geometry: drug-tolerant persister cancer cells increase cell surface area along a de-differentiation trajectory and/or increase volume while acquiring senescent-like features. Using ORIGAMI-guided screening, we identified brefeldin A (BFA) and other ER-Golgi modulators as agents that reduce surface area, promote differentiation, and sensitize melanoma cells to targeted therapies. Therapy-induced high-volume cells exhibit lysosomal scaling and increased sensitivity to nigericin, a natural polyether related to the lysosomal iron sequestering compounds and ferroptosis inducers salinomycin and narasin. Nigericin and BFA combined with MEK- and CDK4/6-inhibitors are highly potent in vitro across cell states and genotypes, and significantly extend survival in NRAS-mutant melanoma patient-derived xenografts without overt toxicity. Moreover, this combination induces hallmarks of immunogenic cell death and enhances anti–PD-1 efficacy in immunotherapy-refractory preclinical models. Beyond melanoma, the combination shows broad anti-tumour activity across a large panel of cancer lines derived from varied tissues. These findings establish a generalizable, geometry-based framework to monitor ITH, anticipate plasticity, and inform genotype-agnostic combination strategies that negate non-genetic adaptive resistance while potentiating immunotherapy. Biological sciences/Cancer/Tumour heterogeneity Biological sciences/Cancer/Cancer therapy/Cancer therapeutic resistance cancer cell plasticity cell geometry therapy resistance melanoma Full Text Additional Declarations There is NO Competing Interest. Supplementary Files SupplementaryTables.pdf Supplementary Table 1-9 ExtendedFigures.pdf Extended Data Figure 1-11 Cite Share Download PDF Status: Under Review Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Single-cell transcriptomics is the gold standard for assessing ITH, but this approach remains costly and slow. Here, we show that distinct melanoma transcriptomic states exhibit unique cellular morphologies, and introduce ORIGAMI, a scalable flow cytometry assay leveraging cell surface area (S) and volume (V) measurements to resolve melanoma cell state diversity and to track therapy-induced transitions. MAPK- and CDK4/6-targeted therapies induce predictable shifts in cell geometry: drug-tolerant persister cancer cells increase cell surface area along a de-differentiation trajectory and/or increase volume while acquiring senescent-like features. Using ORIGAMI-guided screening, we identified brefeldin A (BFA) and other ER-Golgi modulators as agents that reduce surface area, promote differentiation, and sensitize melanoma cells to targeted therapies. 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