Intratumour heterogeneity (ITH) is a hallmark of cancer that drives tumour evolution and disease progression. Technological and computational advances have enabled us to assess ITH at unprecedented depths, yet this accumulating knowledge has not had a substantial clinical impact. This is in part due to a limited understanding of the functional relevance of ITH and the inadequacy of preclinical experimental models to reproduce it. Here, we discuss progress made in these areas and illuminate future directions.
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Vendramin, R., Litchfield, K. & Swanton, C. Cancer evolution: Darwin and beyond. EMBO J. 40, e108389 (2021).
Marine, J. C., Dawson, S. J. & Dawson, M. A. Non-genetic mechanisms of therapeutic resistance in cancer. Nat. Rev. Cancer 20, 743–756 (2020).
Tomasetti, C. & Vogelstein, B. Cancer etiology. Variation in cancer risk among tissues can be explained by the number of stem cell divisions. Science 347, 78–81 (2015).
Wang, R. et al. Single-cell dissection of intratumoral heterogeneity and lineage diversity in metastatic gastric adenocarcinoma. Nat. Med. 27, 141–151 (2021).
Best, S. A. et al. Distinct initiating events underpin the immune and metabolic heterogeneity of KRAS-mutant lung adenocarcinoma. Nat. Commun. 10, 4190 (2019).
Baggiolini, A. et al. Developmental chromatin programs determine oncogenic competence in melanoma. Science 373, eabc1048 (2021).
Concepcion, C. P. et al. Smarca4 inactivation promotes lineage-specific transformation and early metastatic features in the lung. Cancer Discov. 12, 562–585 (2022).
Chetty, R. & Serra, S. SMARCA family of genes. J. Clin. Pathol. 73, 257–260 (2020).
Marjanovic, N. D. et al. Emergence of a high-plasticity cell state during lung cancer evolution. Cancer Cell 38, 229–246.e13 (2020).
Risom, T. et al. Differentiation-state plasticity is a targetable resistance mechanism in basal-like breast cancer. Nat. Commun. 9, 3815 (2018).
Negrini, S., Gorgoulis, V. G. & Halazonetis, T. D. Genomic instability—an evolving hallmark of cancer. Nat. Rev. Mol. Cell Biol. 11, 220–228 (2010).
Tubbs, A. & Nussenzweig, A. Endogenous DNA damage as a source of genomic instability in cancer. Cell 168, 644–656 (2017).
Ross, J. et al. Targeting MYC: from understanding its biology to drug discovery. Eur. J. Med. Chem. 213, 113137 (2021).
Roider, T. et al. Dissecting intratumour heterogeneity of nodal B-cell lymphomas at the transcriptional, genetic and drug-response levels. Nat. Cell Biol. 22, 896–906 (2020).
Maddipati, R. et al. MYC levels regulate metastatic heterogeneity in pancreatic adenocarcinoma. Cancer Discov. 12, 542–561 (2022).
Tirier, S. M. et al. Subclone-specific microenvironmental impact and drug response in refractory multiple myeloma revealed by single‐cell transcriptomics. Nat. Commun. 12, 6960 (2021).
Bakhoum, S. F. & Landau, D. A. Chromosomal instability as a driver of tumor heterogeneity and evolution. Cold Spring Harb. Perspect. Med. 7, a029611 (2017).
van Dijk, E. et al. Chromosomal copy number heterogeneity predicts survival rates across cancers. Nat. Commun. 12, 3188 (2021).
Hinohara, K. & Polyak, K. Intratumoral heterogeneity: more than just mutations. Trends Cell Biol. 29, 569–579 (2019).
Hinohara, K. et al. KDM5 histone demethylase activity links cellular transcriptomic heterogeneity to therapeutic resistance. Cancer Cell 34, 939–953.e9 (2018).
Sharma, S. V. et al. A chromatin-mediated reversible drug-tolerant state in cancer cell subpopulations. Cell 141, 69–80 (2010).
Hu, X., Biswas, A. & De, S. KMT2C deficiency promotes APOBEC mutagenesis and genomic instability in multiple cancers. Preprint at bioRxiv https://doi.org/10.1101/2022.02.04.478993 (2022).
Meir, Z., Mukamel, Z., Chomsky, E., Lifshitz, A. & Tanay, A. Single-cell analysis of clonal maintenance of transcriptional and epigenetic states in cancer cells. Nat. Genet. 52, 709–718 (2020).
Fennell, K. A. et al. Non-genetic determinants of malignant clonal fitness at single-cell resolution. Nature 601, 125–131 (2022).
Cejas, P. et al. Subtype heterogeneity and epigenetic convergence in neuroendocrine prostate cancer. Nat. Commun. 12, 5775 (2021).
Baron, M. et al. The stress-like cancer cell state is a consistent component of tumorigenesis. Cell Syst. 11, 536–546.e7 (2020).
Rutherford, S. L. & Lindquist, S. Hsp90 as a capacitor for morphological evolution. Nature 396, 336–342 (1998).
Gaglia, G. et al. HSF1 phase transition mediates stress adaptation and cell fate decisions. Nat. Cell Biol. 22, 151–158 (2020).
Campbell, N. R. et al. Cooperation between melanoma cell states promotes metastasis through heterotypic cluster formation. Dev. Cell 56, 2808–2825.e10 (2021).
Laconi, E., Marongiu, F. & DeGregori, J. Cancer as a disease of old age: changing mutational and microenvironmental landscapes. Br. J. Cancer 122, 943–952 (2020).
Li, C. M.-C. et al. Aging-associated alterations in mammary epithelia and stroma revealed by single-cell RNA sequencing. Cell Rep. 33, 108566 (2020).
Pradhan, R. N., Krishnamurty, A. T., Fletcher, A. L., Turley, S. J. & Muller, S. A bird’s eye view of fibroblast heterogeneity: a pan-disease, pan-cancer perspective. Immunol. Rev. 302, 299–320 (2021).
Costa, A. et al. Fibroblast heterogeneity and immunosuppressive environment in human breast cancer. Cancer Cell 33, 463–479.e10 (2018).
Pelon, F. et al. Cancer-associated fibroblast heterogeneity in axillary lymph nodes drives metastases in breast cancer through complementary mechanisms. Nat. Commun. 11, 404 (2020).
Tissot, T. et al. Do cell-autonomous and non-cell-autonomous effects drive the structure of tumor ecosystems? Biochim. Biophys. Acta 1865, 147–154 (2016).
Sun, Y.-F. et al. Dissecting spatial heterogeneity and the immune-evasion mechanism of CTCs by single-cell RNA-seq in hepatocellular carcinoma. Nat. Commun. 12, 4091 (2021).
Piersma, B., Hayward, M. K. & Weaver, V. M. Fibrosis and cancer: a strained relationship. Biochim. Biophys. Acta Rev. Cancer 1873, 188356 (2020).
Madan, E. et al. Cell competition boosts clonal evolution and hypoxic selection in cancer. Trends Cell Biol. 30, 967–978 (2020).
Zaidi, M., Fu, F., Cojocari, D., McKee, T. D. & Wouters, B. G. Quantitative visualization of hypoxia and proliferation gradients within histological tissue sections. Front. Bioeng. Biotechnol. 7, 397 (2019).
Nobre, A. R., Entenberg, D., Wang, Y., Condeelis, J. & Aguirre-Ghiso, J. A. The different routes to metastasis via hypoxia-regulated programs. Trends Cell Biol. 28, 941–956 (2018).
Dirkse, A. et al. Stem cell-associated heterogeneity in glioblastoma results from intrinsic tumor plasticity shaped by the microenvironment. Nat. Commun. 10, 1787 (2019).
Fluegen, G. et al. Phenotypic heterogeneity of disseminated tumour cells is preset by primary tumour hypoxic microenvironments. Nat. Cell Biol. 19, 120–132 (2017).
López-Carrasco, A. et al. Impact of extracellular matrix stiffness on genomic heterogeneity in MYCN-amplified neuroblastoma cell line. J. Exp. Clin. Cancer Res. 39, 226 (2020).
Ligorio, M. et al. Stromal microenvironment shapes the intratumoral architecture of pancreatic cancer. Cell 178, 160–175.e27 (2019).
Szczerba, B. M. et al. Neutrophils escort circulating tumour cells to enable cell cycle progression. Nature 566, 553–557 (2019).
Kok, S. Y. et al. Malignant subclone drives metastasis of genetically and phenotypically heterogenous cell clusters through fibrotic niche generation. Nat. Commun. 12, 863 (2021).
Li, B. et al. Therapy-induced mutations drive the genomic landscape of relapsed acute lymphoblastic leukemia. Blood 135, 41–55 (2020).
Tabassum, D. P. & Polyak, K. Tumorigenesis: it takes a village. Nat. Rev. Cancer 15, 473–483 (2015).
Cleary, A. S., Leonard, T. L., Gestl, S. A. & Gunther, E. J. Tumour cell heterogeneity maintained by cooperating subclones in Wnt-driven mammary cancers. Nature 508, 113–117 (2014).
Tammela, T. et al. A Wnt-producing niche drives proliferative potential and progression in lung adenocarcinoma. Nature 545, 355–359 (2017).
Shimokawa, M. et al. Visualization and targeting of LGR5+ human colon cancer stem cells. Nature 545, 187–192 (2017).
Lim, J. S. et al. Intratumoural heterogeneity generated by Notch signalling promotes small-cell lung cancer. Nature 545, 360–364 (2017).
Lim, B., Lin, Y. & Navin, N. Advancing cancer research and medicine with single-cell genomics. Cancer cell 37, 456–470 (2020).
Nam, A. S., Chaligne, R. & Landau, D. A. Integrating genetic and non-genetic determinants of cancer evolution by single-cell multi-omics. Nat. Rev. Genet. 22, 3–18 (2021).
Eirew, P. et al. Dynamics of genomic clones in breast cancer patient xenografts at single-cell resolution. Nature 518, 422–426 (2015).
Bhang, H. E. et al. Studying clonal dynamics in response to cancer therapy using high-complexity barcoding. Nat. Med. 21, 440–448 (2015).
Snippert, H. J. et al. Lgr6 marks stem cells in the hair follicle that generate all cell lineages of the skin. Science 327, 1385–1389 (2010).
Gutierrez, C. et al. Multifunctional barcoding with ClonMapper enables high-resolution study of clonal dynamics during tumor evolution and treatment. Nat. Cancer 2, 758–772 (2021).
Quinn, J. J. et al. Single-cell lineages reveal the rates, routes, and drivers of metastasis in cancer xenografts. Science 371, eabc1944 (2021).
Simeonov, K. P. et al. Single-cell lineage tracing of metastatic cancer reveals selection of hybrid EMT states. Cancer Cell 39, 1150–1162.e9 (2021).
Garcia-Marques, J. et al. A programmable sequence of reporters for lineage analysis. Nat. Neurosci. 23, 1618–1628 (2020).
Oren, Y. et al. Cycling cancer persister cells arise from lineages with distinct programs. Nature 596, 576–582 (2021).
Zdraljevic, S. & Andersen, E. C. Natural diversity facilitates the discovery of conserved chemotherapeutic response mechanisms. Curr. Opin. Genet. Dev. 47, 41–47 (2017).
Godet, I. et al. Fate-mapping post-hypoxic tumor cells reveals a ROS-resistant phenotype that promotes metastasis. Nat. Commun. 10, 4862 (2019).
Ong, T. T. et al. pHLuc, a ratiometric luminescent reporter for in vivo monitoring of tumor acidosis. Front. Bioeng. Biotechnol. 8, 412 (2020).
Ombrato, L. et al. Metastatic-niche labelling reveals parenchymal cells with stem features. Nature 572, 603–608 (2019).
Tang, R. et al. A versatile system to record cell–cell interactions. eLife 9, e61080 (2020).
Morris, L. G. et al. Pan-cancer analysis of intratumor heterogeneity as a prognostic determinant of survival. Oncotarget 7, 10051–10063 (2016).
McGranahan, N. & Swanton, C. Clonal heterogeneity and tumor evolution: past, present, and the future. Cell 168, 613–628 (2017).
Turajlic, S. et al. Deterministic evolutionary trajectories influence primary tumor growth: TRACERx renal. Cell 173, 595–610.e11 (2018).
Marusyk, A. & Polyak, K. Tumor heterogeneity: causes and consequences. Biochim. Biophys. Acta 1805, 105–117 (2010).
Martinez, P. et al. Parallel evolution of tumour subclones mimics diversity between tumours. J. Pathol. 230, 356–364 (2013).
McMurray, H. R. et al. Synergistic response to oncogenic mutations defines gene class critical to cancer phenotype. Nature 453, 1112–1116 (2008).
Caswell, D. R. & Swanton, C. The role of tumour heterogeneity and clonal cooperativity in metastasis, immune evasion and clinical outcome. BMC Med. 15, 133 (2017).
Marusyk, A. et al. Non-cell-autonomous driving of tumour growth supports sub-clonal heterogeneity. Nature 514, 54–58 (2014).
Dey, P. et al. Oncogenic KRAS-driven metabolic reprogramming in pancreatic cancer cells utilizes cytokines from the tumor microenvironment. Cancer Discov. 10, 608–625 (2020).
Seehawer, M. et al. Necroptosis microenvironment directs lineage commitment in liver cancer. Nature 562, 69–75 (2018).
Peinado, H. et al. Pre-metastatic niches: organ-specific homes for metastases. Nat. Rev. Cancer 17, 302–317 (2017).
Kang, Y. et al. A multigenic program mediating breast cancer metastasis to bone. Cancer Cell 3, 537–549 (2003).
Minn, A. J. et al. Genes that mediate breast cancer metastasis to lung. Nature 436, 518–524 (2005).
Janiszewska, M. et al. Subclonal cooperation drives metastasis by modulating local and systemic immune microenvironments. Nat. Cell Biol. 21, 879–888 (2019).
Wu, S. Z. et al. A single-cell and spatially resolved atlas of human breast cancers. Nat. Genet. 53, 1334–1347 (2021).
Yarchoan, M., Johnson, B. A. 3rd, Lutz, E. R., Laheru, D. A. & Jaffee, E. M. Targeting neoantigens to augment antitumour immunity. Nat. Rev. Cancer 17, 209–222 (2017).
O’Donnell, J. S., Teng, M. W. L. & Smyth, M. J. Cancer immunoediting and resistance to T cell-based immunotherapy. Nat. Rev. Clin. Oncol. 16, 151–167 (2019).
Matsushita, H. et al. Cancer exome analysis reveals a T-cell-dependent mechanism of cancer immunoediting. Nature 482, 400–404 (2012).
Waldman, A. D., Fritz, J. M. & Lenardo, M. J. A guide to cancer immunotherapy: from T cell basic science to clinical practice. Nat. Rev. Immunol. 20, 651–668 (2020).
Le, D. T. et al. PD-1 blockade in tumors with mismatch-repair deficiency. N. Engl. J. Med. 372, 2509–2520 (2015).
Wolf, Y. et al. UVB-induced tumor heterogeneity diminishes immune response in melanoma. Cell 179, 219–235.e21 (2019).
Peng, W. et al. Loss of PTEN promotes resistance to T cell-mediated immunotherapy. Cancer Discov. 6, 202–216 (2016).
Miranda, A. et al. Cancer stemness, intratumoral heterogeneity, and immune response across cancers. Proc. Natl Acad. Sci. USA 116, 9020–9029 (2019).
Ho, D. W. et al. Single-cell RNA sequencing shows the immunosuppressive landscape and tumor heterogeneity of HBV-associated hepatocellular carcinoma. Nat. Commun. 12, 3684 (2021).
Rosenthal, R. et al. Neoantigen-directed immune escape in lung cancer evolution. Nature 567, 479–485 (2019).
Filho, O. M. et al. Impact of HER2 heterogeneity on treatment response of early-stage HER2-positive breast Cancer: Phase II neoadjuvant clinical trial of T-DM1 combined with pertuzumab. Cancer Discov. 11, 2474–2487 (2021).
Jaber, M. I. et al. A deep learning image-based intrinsic molecular subtype classifier of breast tumors reveals tumor heterogeneity that may affect survival. Breast Cancer Res. 22, 12 (2020).
Xing, Q. R. et al. Parallel bimodal single-cell sequencing of transcriptome and chromatin accessibility. Genome Res. 30, 1027–1039 (2020).
Liu, L. et al. Deconvolution of single-cell multi-omics layers reveals regulatory heterogeneity. Nat. Commun. 10, 470 (2019).
Chen, S., Lake, B. B. & Zhang, K. High-throughput sequencing of the transcriptome and chromatin accessibility in the same cell. Nat. Biotechnol. 37, 1452–1457 (2019).
Macaulay, I. C. et al. G&T-seq: parallel sequencing of single-cell genomes and transcriptomes. Nat. Methods 12, 519–522 (2015).
Stoeckius, M. et al. Simultaneous epitope and transcriptome measurement in single cells. Nat. Methods 14, 865–868 (2017).
Zhu, C. et al. Joint profiling of histone modifications and transcriptome in single cells from mouse brain. Nat. Methods 18, 283–292 (2021).
Mimitou, E. P. et al. Scalable, multimodal profiling of chromatin accessibility, gene expression and protein levels in single cells. Nat. Biotechnol. 39, 1246–1258 (2021).
Niemöller, C. et al. Bisulfite-free epigenomics and genomics of single cells through methylation-sensitive restriction. Commun. Biol. 4, 153 (2021).
Pott, S. Simultaneous measurement of chromatin accessibility, DNA methylation, and nucleosome phasing in single cells. eLife 6, e23203 (2017).
Hu, Y. et al. Simultaneous profiling of transcriptome and DNA methylome from a single cell. Genome Biol. 17, 88 (2016).
Chung, H. et al. Joint single-cell measurements of nuclear proteins and RNA in vivo. Nat. Methods 18, 1204–1212 (2021).
Moncada, R. et al. Integrating microarray-based spatial transcriptomics and single-cell RNA-seq reveals tissue architecture in pancreatic ductal adenocarcinomas. Nat. Biotechnol. 38, 333–342 (2020).
Lee, Y. et al. XYZeq: spatially resolved single-cell RNA sequencing reveals expression heterogeneity in the tumor microenvironment. Sci. Adv. 7, eabg4755 (2021).
Clark, S. J. et al. scNMT-seq enables joint profiling of chromatin accessibility DNA methylation and transcription in single cells. Nat. Commun. 9, 781 (2018).
Swanson, E. et al. Simultaneous trimodal single-cell measurement of transcripts, epitopes, and chromatin accessibility using TEA-seq. eLife 10, e63632 (2021).
Berthelet, J. et al. The site of breast cancer metastases dictates their clonal composition and reversible transcriptomic profile. Sci. Adv. 7, eabf4408 (2021).
Pei, W. et al. Polylox barcoding reveals haematopoietic stem cell fates realized in vivo. Nature 548, 456–460 (2017).
Biase, F. H. et al. Rainbow-Seq: combining cell lineage tracing with single-cell RNA sequencing in preimplantation embryos. iScience 7, 16–29 (2018).
Jacob, F. et al. A patient-derived glioblastoma organoid model and biobank recapitulates inter-and intra-tumoral heterogeneity. Cell 180, 188–204.e22 (2020).
Kopper, O. et al. An organoid platform for ovarian cancer captures intra- and interpatient heterogeneity. Nat. Med. 25, 838–849 (2019).
Sachs, N. et al. A living biobank of breast cancer organoids captures disease heterogeneity. Cell 172, 373–386.e10 (2018).
Kim, M. et al. Patient-derived lung cancer organoids as in vitro cancer models for therapeutic screening. Nat. Commun. 10, 3991 (2019).
Broutier, L. et al. Human primary liver cancer–derived organoid cultures for disease modeling and drug screening. Nat. Med. 23, 1424–1435 (2017).
Calandrini, C. et al. An organoid biobank for childhood kidney cancers that captures disease and tissue heterogeneity. Nat. Commun. 11, 1310 (2020).
Tuveson, D. & Clevers, H. Cancer modeling meets human organoid technology. Science 364, 952–955 (2019).
Lo, Y.-H., Karlsson, K. & Kuo, C. J. Applications of organoids for cancer biology and precision medicine. Nat. Cancer 1, 761–773 (2020).
Neal, J. T. et al. Organoid modeling of the tumor immune microenvironment. Cell 175, 1972–1988.e16 (2018).
Simpson, K. L. et al. A biobank of small cell lung cancer CDX models elucidates inter- and intratumoral phenotypic heterogeneity. Nat. Cancer 1, 437–451 (2020).
Guillen, K. P. et al. A human breast cancer-derived xenograft and organoid platform for drug discovery and precision oncology. Nat. Cancer 3, 232–250 (2022).
Sun, H. et al. Comprehensive characterization of 536 patient-derived xenograft models prioritizes candidates for targeted treatment. Nat. Commun. 12, 5086 (2021).
Reiter, J. G. et al. An analysis of genetic heterogeneity in untreated cancers. Nat. Rev. Cancer 19, 639–650 (2019).
Fearon, E. R. & Vogelstein, B. A genetic model for colorectal tumorigenesis. Cell 61, 759–767 (1990).
Gerlinger, M. et al. Intratumor heterogeneity and branched evolution revealed by multiregion sequencing. N. Engl. J. Med. 366, 883–892 (2012).
Gao, R. et al. Punctuated copy number evolution and clonal stasis in triple-negative breast cancer. Nat. Genet. 48, 1119–1130 (2016).
Sottoriva, A. et al. A Big Bang model of human colorectal tumor growth. Nat. Genet. 47, 209–216 (2015).
We thank A. Marusyk (Moffit Cancer Center) and members of our laboratory for critical reading of the manuscript and helpful suggestions. The authors are funded by the National Cancer Institute R35 CA197623 (K.P.) and EMBO (M.S.).
K.P. serves on the Scientific Advisory Board of Acrivon Therapeutics, Vividion Therapeutics, Scorpion Therapeutics and the Novartis Institute for BioMedical Research, holds equity options in Scorpion Therapeutics, is a consultant to Aria Pharmaceuticals, received honorarium from AstraZeneca and New Equilibrium Biosciences, and has an institutional research agreement with Novartis. The remaining authors declare no competing interests.
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Li, Z., Seehawer, M. & Polyak, K. Untangling the web of intratumour heterogeneity. Nat Cell Biol 24, 1192–1201 (2022). https://doi.org/10.1038/s41556-022-00969-x