{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,9]],"date-time":"2025-11-09T03:47:17Z","timestamp":1762660037001,"version":"3.37.3"},"reference-count":51,"publisher":"Oxford University Press (OUP)","issue":"14","license":[{"start":{"date-parts":[[2021,2,1]],"date-time":"2021-02-01T00:00:00Z","timestamp":1612137600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/journals\/pages\/open_access\/funder_policies\/chorus\/standard_publication_model"}],"funder":[{"name":"U.S. National Institutes of Health National Cancer Institute","award":["R01CA216101"],"award-info":[{"award-number":["R01CA216101"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2021,8,4]]},"abstract":"<jats:title>Abstract<\/jats:title>\n               <jats:sec>\n                  <jats:title>Motivation<\/jats:title>\n                  <jats:p>Cancer cell heterogeneity can manifest genetically and phenotypically. Bioinformatics methods have been used to analyze complex genomics and transcriptomics data, but have not been well-established for analyzing biophysical data of phenotypically heterogeneous tumor cells. Here, we take an informatics approach to analyze the biophysical data of MDA-MB-231 cells, a widely used breast cancer cell line, during their spontaneous migration through confined environments. Experimentally, we vary the constriction microchannel geometries (wide channel, short constriction, and long constriction) and apply drug treatments. We find that the cells in the short constriction are similar in morphology to the cells in the wide channel. However, their fluorescence profiles are comparable to those in the long constriction. We demonstrate that the cell migratory phenotype is correlated more to mitochondria in a non-confined environment and more to actin in a confined environment. We demonstrate that the cells\u2019 migratory phenotypes are altered by ciliobrevin D, a dynein inhibitor, in both confined and non-confined environments. Overall, our approach elucidates phenotypic heterogeneity in cancer cells under confined microenvironments at single-cell resolution.<\/jats:p>\n               <\/jats:sec>\n               <jats:sec>\n                  <jats:title>Results<\/jats:title>\n                  <jats:p>Here, we apply a bioinformatics approach to a single-cell invasion assay. We demonstrate that this method can determine distinctions in morphology, cytoskeletal activities, and mitochondrial activities under various geometric constraints and for cells of different speeds. Our approach can be readily adapted to various heterogeneity studies for different types of input biophysical data. In addition, this approach can be applied to studies related to biophysical changes due to differences in external stimuli, such as treatment effects on cellular and subcellular activities, at single-cell resolution. Finally, as similar bioinformatics methods have been widely applied in studies of genetic heterogeneity, biophysical information extracted using this approach can be analyzed together with the genetic data to relate genetic and phenotypic heterogeneity.<\/jats:p>\n               <\/jats:sec>\n               <jats:sec>\n                  <jats:title>Availability and implementation<\/jats:title>\n                  <jats:p>The data that support the findings of this study are available from the corresponding author upon reasonable request.<\/jats:p>\n               <\/jats:sec>\n               <jats:sec>\n                  <jats:title>Supplementary information<\/jats:title>\n                  <jats:p>Supplementary data are available at Bioinformatics online.<\/jats:p>\n               <\/jats:sec>","DOI":"10.1093\/bioinformatics\/btab053","type":"journal-article","created":{"date-parts":[[2021,1,27]],"date-time":"2021-01-27T04:13:32Z","timestamp":1611720812000},"page":"2042-2052","source":"Crossref","is-referenced-by-count":14,"title":["Biophysical informatics approach for quantifying phenotypic heterogeneity in cancer cell migration in confined microenvironments"],"prefix":"10.1093","volume":"37","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1369-6556","authenticated-orcid":false,"given":"Xingjian","family":"Zhang","sequence":"first","affiliation":[{"name":"Department of Biomedical Engineering, Yale University , New Haven, CT 06510, USA"}]},{"given":"Michael","family":"Mak","sequence":"additional","affiliation":[{"name":"Department of Biomedical Engineering, Yale University , New Haven, CT 06510, USA"}]}],"member":"286","published-online":{"date-parts":[[2021,2,1]]},"reference":[{"key":"2023061310304658300_btab053-B1","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1002\/cm.21150","article-title":"Microtubules and actin crosstalk in cell migration and division","volume":"71","author":"Akhshi","year":"2014","journal-title":"Cytoskeleton"},{"key":"2023061310304658300_btab053-B2","doi-asserted-by":"crossref","first-page":"846","DOI":"10.1038\/nm.3915","article-title":"Toward understanding and exploiting tumor heterogeneity","volume":"21","author":"Alizadeh","year":"2015","journal-title":"Nat. Med"},{"key":"2023061310304658300_btab053-B3","doi-asserted-by":"crossref","first-page":"5615","DOI":"10.1158\/1078-0432.CCR-06-0169","article-title":"Most early disseminated cancer cells detected in bone marrow of breast cancer patients have a putative breast cancer stem cell phenotype","volume":"12","author":"Balic","year":"2006","journal-title":"Clin. Cancer Res"},{"key":"2023061310304658300_btab053-B4","doi-asserted-by":"crossref","first-page":"617","DOI":"10.1038\/s41567-019-0493-9","article-title":"Publisher Correction: stochastic nonlinear dynamics of confined cell migration in two-state systems","volume":"15","author":"Br\u00fcckner","year":"2019","journal-title":"Nat. Phys"},{"key":"2023061310304658300_btab053-B5","doi-asserted-by":"crossref","first-page":"155","DOI":"10.1038\/nbt.3102","article-title":"Computational analysis of cell-to-cell heterogeneity in single-cell RNA-sequencing data reveals hidden subpopulations of cells","volume":"33","author":"Buettner","year":"2015","journal-title":"Nat. Biotechnol"},{"key":"2023061310304658300_btab053-B6","doi-asserted-by":"crossref","first-page":"2879","DOI":"10.1084\/jem.20061877","article-title":"Orchestration of lymphocyte chemotaxis by mitochondrial dynamics","volume":"203","author":"Campello","year":"2006","journal-title":"J. Exp. Med"},{"key":"2023061310304658300_btab053-B7","doi-asserted-by":"crossref","first-page":"7296","DOI":"10.1039\/C9AN01358J","article-title":"Single-cell RNA-sequencing of migratory breast cancer cells: discovering genes associated with cancer metastasis","volume":"144","author":"Chen","year":"2019","journal-title":"Analyst"},{"key":"2023061310304658300_btab053-B8","doi-asserted-by":"crossref","first-page":"15081","DOI":"10.1038\/ncomms15081","article-title":"Single-cell RNA-seq enables comprehensive tumour and immune cell profiling in primary breast cancer","volume":"8","author":"Chung","year":"2017","journal-title":"Nat. Commun"},{"key":"2023061310304658300_btab053-B9","doi-asserted-by":"crossref","first-page":"313","DOI":"10.1038\/nm.2304","article-title":"The cancer stem cell: premises, promises and challenges","volume":"17","author":"Clevers","year":"2011","journal-title":"Nat. Med"},{"key":"2023061310304658300_btab053-B10","doi-asserted-by":"crossref","first-page":"316","DOI":"10.1016\/0014-5793(87)81513-2","article-title":"Inhibition of actin polymerization by latrunculin A","volume":"213","author":"Cou\u00e9","year":"1987","journal-title":"FEBS Lett"},{"key":"2023061310304658300_btab053-B11","doi-asserted-by":"crossref","first-page":"353","DOI":"10.1126\/science.aad7297","article-title":"Nuclear envelope rupture and repair during cancer cell migration","volume":"352","author":"Denais","year":"2016","journal-title":"Science"},{"key":"2023061310304658300_btab053-B12","doi-asserted-by":"crossref","first-page":"2077","DOI":"10.1016\/j.bpj.2013.03.025","article-title":"Mitochondrial localization and the persistent migration of epithelial cancer cells","volume":"104","author":"Desai","year":"2013","journal-title":"Biophys. J"},{"key":"2023061310304658300_btab053-B13","doi-asserted-by":"crossref","first-page":"4793","DOI":"10.1182\/blood-2008-08-077941","article-title":"Stem cell concepts renew cancer research","volume":"112","author":"Dick","year":"2008","journal-title":"Blood"},{"key":"2023061310304658300_btab053-B14","doi-asserted-by":"crossref","first-page":"780","DOI":"10.1038\/nature07733","article-title":"Association of reactive oxygen species levels and radioresistance in cancer stem cells","volume":"458","author":"Diehn","year":"2009","journal-title":"Nature"},{"key":"2023061310304658300_btab053-B15","doi-asserted-by":"crossref","first-page":"716","DOI":"10.1016\/j.molcel.2014.05.015","article-title":"Cancer epigenetics: tumor heterogeneity, plasticity of stem-like states, and drug resistance","volume":"54","author":"Easwaran","year":"2014","journal-title":"Mol. Cell"},{"key":"2023061310304658300_btab053-B16","doi-asserted-by":"crossref","first-page":"883","DOI":"10.1056\/NEJMoa1113205","article-title":"Intratumor heterogeneity and branched evolution revealed by multiregion sequencing","volume":"366","author":"Gerlinger","year":"2012","journal-title":"N. Engl. J. Med"},{"key":"2023061310304658300_btab053-B17","doi-asserted-by":"crossref","first-page":"646","DOI":"10.1016\/j.cell.2011.02.013","article-title":"Hallmarks of cancer: the next generation","volume":"144","author":"Hanahan","year":"2011","journal-title":"Cell"},{"key":"2023061310304658300_btab053-B18","doi-asserted-by":"crossref","first-page":"714","DOI":"10.1038\/nrc3599","article-title":"Cancer drug resistance: an evolving paradigm","volume":"13","author":"Holohan","year":"2013","journal-title":"Nat. Rev. Cancer"},{"key":"2023061310304658300_btab053-B19","doi-asserted-by":"crossref","first-page":"e56910","DOI":"10.1371\/journal.pone.0056910","article-title":"In vitro model of tumor cell extravasation","volume":"8","author":"Jeon","year":"2013","journal-title":"PLoS ONE"},{"key":"2023061310304658300_btab053-B20","doi-asserted-by":"crossref","first-page":"500","DOI":"10.3389\/fgene.2019.00500","article-title":"Identification of breast cancer stem cell related genes using functional cellular assays combined with single-cell RNA sequencing in MDA-MB-231 cells","volume":"10","author":"Jonasson","year":"2019","journal-title":"Front. Genet"},{"key":"2023061310304658300_btab053-B21","doi-asserted-by":"crossref","first-page":"2343","DOI":"10.21873\/anticanres.11572","article-title":"Heterogeneous cell types in single-cell-derived clones of MCF7 and MDA-MB-231 cells","volume":"37","author":"Khan","year":"2017","journal-title":"Anticancer Res"},{"key":"2023061310304658300_btab053-B22","doi-asserted-by":"crossref","first-page":"127","DOI":"10.1186\/s13059-015-0692-3","article-title":"Single-cell mRNA sequencing identifies subclonal heterogeneity in anti-cancer drug responses of lung adenocarcinoma cells","volume":"16","author":"Kim","year":"2015","journal-title":"Genome Biol"},{"key":"2023061310304658300_btab053-B23","doi-asserted-by":"crossref","first-page":"48","DOI":"10.1016\/j.ab.2017.10.008","article-title":"Microvalve controlled multi-functional microfluidic chip for divisional cell co-culture","volume":"539","author":"Li","year":"2017","journal-title":"Anal. Biochem"},{"key":"2023061310304658300_btab053-B24","doi-asserted-by":"crossref","first-page":"e20825","DOI":"10.1371\/journal.pone.0020825","article-title":"Microfabricated physical spatial gradients for investigating cell migration and invasion dynamics","volume":"6","author":"Mak","year":"2011","journal-title":"PLoS One"},{"key":"2023061310304658300_btab053-B25","doi-asserted-by":"crossref","first-page":"340","DOI":"10.1039\/C2LC41117B","article-title":"Elucidating mechanical transition effects of invading cancer cells with a subnucleus-scaled microfluidic serial dimensional modulation device","volume":"13","author":"Mak","year":"2013","journal-title":"Lab. Chip"},{"key":"2023061310304658300_btab053-B26","doi-asserted-by":"crossref","first-page":"1374","DOI":"10.1039\/c3ib40128f","article-title":"A serial micropipette microfluidic device with applications to cancer cell repeated deformation studies","volume":"5","author":"Mak","year":"2013","journal-title":"Integr. Biol"},{"key":"2023061310304658300_btab053-B27","doi-asserted-by":"crossref","first-page":"704","DOI":"10.1016\/j.cell.2008.03.027","article-title":"The epithelial-mesenchymal transition generates cells with properties of stem cells","volume":"133","author":"Mani","year":"2008","journal-title":"Cell"},{"key":"2023061310304658300_btab053-B28","doi-asserted-by":"crossref","first-page":"328","DOI":"10.1038\/nature12624","article-title":"Tumour heterogeneity and cancer cell plasticity","volume":"501","author":"Meacham","year":"2013","journal-title":"Nature"},{"key":"2023061310304658300_btab053-B29","doi-asserted-by":"crossref","first-page":"35","DOI":"10.1016\/j.coisb.2019.09.002","article-title":"Systems biology approaches to measure and model phenotypic heterogeneity in cancer","volume":"17","author":"Meyer","year":"2019","journal-title":"Curr. Opin. Syst. Biol"},{"key":"2023061310304658300_btab053-B30","doi-asserted-by":"crossref","first-page":"11246","DOI":"10.1038\/ncomms11246","article-title":"Highly variable cancer subpopulations that exhibit enhanced transcriptome variability and metastatic fitness","volume":"7","author":"Nguyen","year":"2016","journal-title":"Nat. Commun"},{"key":"2023061310304658300_btab053-B31","doi-asserted-by":"crossref","first-page":"35","DOI":"10.1038\/nri.2017.76","article-title":"Single-cell RNA sequencing to explore immune cell heterogeneity","volume":"18","author":"Papalexi","year":"2018","journal-title":"Nat. Rev. Immunol"},{"key":"2023061310304658300_btab053-B32","doi-asserted-by":"crossref","first-page":"1396","DOI":"10.1126\/science.1254257","article-title":"Single-cell RNA-seq highlights intratumoral heterogeneity in primary glioblastoma","volume":"344","author":"Patel","year":"2014","journal-title":"Science"},{"key":"2023061310304658300_btab053-B33","doi-asserted-by":"crossref","first-page":"176","DOI":"10.1016\/j.biomaterials.2017.04.017","article-title":"Advanced biomaterials and microengineering technologies to recapitulate the stepwise process of cancer metastasis","volume":"133","author":"Peela","year":"2017","journal-title":"Biomaterials"},{"key":"2023061310304658300_btab053-B34","doi-asserted-by":"crossref","first-page":"a022038","DOI":"10.1101\/cshperspect.a022038","article-title":"Mechanical properties of the cytoskeleton and cells","volume":"9","author":"Pegoraro","year":"2017","journal-title":"Cold Spring Harbor Perspectives in Biology"},{"key":"2023061310304658300_btab053-B35","doi-asserted-by":"crossref","first-page":"453","DOI":"10.1016\/S0092-8674(03)00120-X","article-title":"Cellular motility driven by assembly and disassembly of actin filaments","volume":"112","author":"Pollard","year":"2003","journal-title":"Cell"},{"key":"2023061310304658300_btab053-B36","doi-asserted-by":"crossref","first-page":"777","DOI":"10.1038\/nbt.2282","article-title":"Full-length mRNA-Seq from single-cell levels of RNA and individual circulating tumor cells","volume":"30","author":"Ramsk\u00f6ld","year":"2012","journal-title":"Nat. Biotechnol"},{"key":"2023061310304658300_btab053-B37","doi-asserted-by":"crossref","first-page":"105","DOI":"10.1038\/35102167","article-title":"Stem cells, cancer, and cancer stem cells","volume":"414","author":"Reya","year":"2001","journal-title":"Nature"},{"key":"2023061310304658300_btab053-B38","doi-asserted-by":"crossref","first-page":"757","DOI":"10.1002\/dneu.22246","article-title":"The dynein inhibitor ciliobrevin D inhibits the bidirectional transport of organelles along sensory axons and impairs NGF-mediated regulation of growth cones and axon branches","volume":"75","author":"Sainath","year":"2015","journal-title":"Dev. Neurobiol"},{"key":"2023061310304658300_btab053-B39","doi-asserted-by":"crossref","first-page":"1009","DOI":"10.1083\/jcb.147.5.1009","article-title":"Rac downregulates Rho activity: reciprocal balance between both GTPases determines cellular morphology and migratory behavior","volume":"147","author":"Sander","year":"1999","journal-title":"J. Cell Biol"},{"key":"2023061310304658300_btab053-B40","doi-asserted-by":"crossref","first-page":"912","DOI":"10.1039\/c7ib00128b","article-title":"A biomaterial screening approach reveals microenvironmental mechanisms of drug resistance","volume":"9","author":"Schwartz","year":"2017","journal-title":"Integr. Biol"},{"key":"2023061310304658300_btab053-B41","doi-asserted-by":"crossref","first-page":"822","DOI":"10.1016\/j.cell.2009.08.017","article-title":"Heterogeneity in cancer: cancer stem cells versus clonal evolution","volume":"138","author":"Shackleton","year":"2009","journal-title":"Cell"},{"key":"2023061310304658300_btab053-B42","doi-asserted-by":"crossref","first-page":"431","DOI":"10.1038\/nature22794","article-title":"Rare cell variability and drug-induced reprogramming as a mode of cancer drug resistance","volume":"546","author":"Shaffer","year":"2017","journal-title":"Nature"},{"key":"2023061310304658300_btab053-B43","first-page":"2313","article-title":"Mitochondria dynamism: of shape, transport and cell migration","volume":"71","author":"da Silva","year":"2014","journal-title":"Cell. Mol. Life Sci"},{"key":"2023061310304658300_btab053-B44","doi-asserted-by":"crossref","first-page":"611","DOI":"10.1016\/j.cell.2014.02.052","article-title":"Water permeation drives tumor cell migration in confined microenvironments","volume":"157","author":"Stroka","year":"2014","journal-title":"Cell"},{"key":"2023061310304658300_btab053-B45","doi-asserted-by":"crossref","first-page":"131","DOI":"10.1038\/nrm1835","article-title":"Complex networks orchestrate epithelial\u2013mesenchymal transitions","volume":"7","author":"Thiery","year":"2006","journal-title":"Nat. Rev. Mol. Cell Biol"},{"key":"2023061310304658300_btab053-B46","doi-asserted-by":"crossref","first-page":"911","DOI":"10.1073\/pnas.1118910109","article-title":"Mechanical compression drives cancer cells toward invasive phenotype","volume":"109","author":"Tse","year":"2012","journal-title":"Proc. Natl. Acad. Sci. U.S.A"},{"key":"2023061310304658300_btab053-B47","doi-asserted-by":"crossref","first-page":"1069","DOI":"10.1083\/jcb.201210152","article-title":"Physical limits of cell migration: control by ECM space and nuclear deformation and tuning by proteolysis and traction force","volume":"201","author":"Wolf","year":"2013","journal-title":"J. Cell Biol"},{"key":"2023061310304658300_btab053-B48","doi-asserted-by":"crossref","first-page":"eaaw6938","DOI":"10.1126\/sciadv.aaw6938","article-title":"Single-cell morphology encodes metastatic potential","volume":"6","author":"Wu","year":"2020","journal-title":"Sci. Adv"},{"key":"2023061310304658300_btab053-B49","doi-asserted-by":"crossref","first-page":"642","DOI":"10.1016\/j.bbamcr.2006.07.001","article-title":"Regulation of the actin cytoskeleton in cancer cell migration and invasion","volume":"1773","author":"Yamaguchi","year":"2007","journal-title":"Biochim. Biophys. Acta (BBA) \u2013 Mol. Cell Res"},{"key":"2023061310304658300_btab053-B50","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1007\/s10555-008-9169-0","article-title":"EMT, the cytoskeleton, and cancer cell invasion","volume":"28","author":"Yilmaz","year":"2009","journal-title":"Cancer Metastasis Rev"},{"key":"2023061310304658300_btab053-B51","doi-asserted-by":"crossref","first-page":"497","DOI":"10.1093\/carcin\/21.3.497","article-title":"Tumor progression and metastasis","volume":"21","author":"Yokota","year":"2000","journal-title":"Carcinogenesis"}],"container-title":["Bioinformatics"],"original-title":[],"language":"en","link":[{"URL":"http:\/\/academic.oup.com\/bioinformatics\/advance-article-pdf\/doi\/10.1093\/bioinformatics\/btab053\/37910573\/btab053.pdf","content-type":"application\/pdf","content-version":"am","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/bioinformatics\/article-pdf\/37\/14\/2042\/50579033\/btab053.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/bioinformatics\/article-pdf\/37\/14\/2042\/50579033\/btab053.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,6,13]],"date-time":"2023-06-13T10:34:07Z","timestamp":1686652447000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/bioinformatics\/article\/37\/14\/2042\/6125378"}},"subtitle":[],"editor":[{"given":"Jinbo","family":"Xu","sequence":"additional","affiliation":[]}],"short-title":[],"issued":{"date-parts":[[2021,2,1]]},"references-count":51,"journal-issue":{"issue":"14","published-print":{"date-parts":[[2021,8,4]]}},"URL":"https:\/\/doi.org\/10.1093\/bioinformatics\/btab053","relation":{},"ISSN":["1367-4803","1367-4811"],"issn-type":[{"type":"print","value":"1367-4803"},{"type":"electronic","value":"1367-4811"}],"subject":[],"published-other":{"date-parts":[[2021,7,15]]},"published":{"date-parts":[[2021,2,1]]}}}