单细胞测序在口腔鳞状细胞癌研究中的价值

doi:10.19401/j.cnki.1007-3639.2024.05.007

摘要/Abstract

摘要：

单细胞测序（single-cell sequencing，SCS）在口腔鳞状细胞癌（oral squamous cell carcinoma，OSCC）研究中具有巨大潜力。随着SCS技术的发展，其灵敏度和准确度不断提高且成本逐渐降低，SCS将成为肿瘤研究的重要技术工具。SCS技术通过在单个细胞分辨率下识别基因组突变所导致的差异基因表达和表观遗传学信息改变，为发现新的细胞特异性标志物和细胞类型提供巨大帮助。在OSCC研究中，SCS不仅有助于揭示癌细胞的异质性以及更准确地理解肿瘤微环境，也有助于深入探讨恶性细胞、免疫细胞及基质细胞三者间的相互作用，揭示它们在肿瘤发生、发展中的相互影响及作用。利用SCS对肿瘤中的免疫细胞进行分类、理解免疫逃逸机制，将为免疫治疗的有效开展提供关键支持。本综述介绍SCS技术的发展现状，并回顾和讨论该技术在OSCC领域中的最新研究进展和应用前景。

关键词: 口腔鳞状细胞癌, 单细胞测序, 肿瘤异质性, 肿瘤免疫反应, 肿瘤微环境

Abstract:

Single-cell sequencing (SCS) has great potential in oral squamous cell carcinoma (OSCC) research. With the development of SCS technology, its sensitivity and accuracy are gradually increasing, while its cost is gradually decreasing. SCS is poised to become a crucial technological tool in cancer research. SCS technology provides significant assistance in the discovery of new cell-specific markers and cell types by identifying differential gene expression and epigenetic information alterations caused by genomic mutations at the resolution of a single cell. In OSCC studies, SCS not only helps unveil the heterogeneity of cancer cells and provides more accurate understanding of the tumor microenvironment, but also facilitates a deeper exploration of the interactions between OSCC cells, immune cells and stromal cells. This sheds light on their mutual influences and roles in tumor initiation and progression. Utilizing SCS to classify immune cells in tumors and comprehend immune escape mechanisms is pivotal for the effective development of immunotherapy. This comprehensive review outlined the current status of SCS technology development and discussed its latest research advancements and prospective applications in the field of OSCC.

Key words: Oral squamous cell carcinoma, Single-cell sequencing, Tumor heterogeneity, Tumor immune response, Tumor microenvironment

中图分类号:

R739.85

王小聪, 李明. 单细胞测序在口腔鳞状细胞癌研究中的价值[J]. 中国癌症杂志, 2024, 34(5): 501-508.

WANG Xiaocong, LI Ming. The value of single-cell sequencing in oral squamous cell carcinoma research[J]. China Oncology, 2024, 34(5): 501-508.

参考文献 60

[1]	ONG Y L R, TIVEY D, HUANG L, et al. Factors affecting surgical mortality of oral squamous cell carcinoma resection[J]. Int J Oral Maxillofac Surg, 2021, 50(1): 1-6.
[2]	HOWARD A, AGRAWAL N, GOOI Z. Lip and oral cavity squamous cell carcinoma[J]. Hematol Oncol Clin North Am, 2021, 35(5): 895-911.
[3]	MROZ E A, TWARD A D, PICKERING C R, et al. High intratumor genetic heterogeneity is related to worse outcome in patients with head and neck squamous cell carcinoma[J]. Cancer, 2013, 119(16): 3034-3042. doi: 10.1002/cncr.28150 pmid: 23696076
[4]	BOXBERG M, LEISING L, STEIGER K, et al. Composition and clinical impact of the immunologic tumor microenvironment in oral squamous cell carcinoma[J]. J Immunol, 2019, 202(1): 278-291. doi: 10.4049/jimmunol.1800242 pmid: 30530592
[5]	CANCER GENOME ATLAS NETWORK. Comprehensive genomic characterization of head and neck squamous cell carcinomas[J]. Nature, 2015, 517(7536): 576-582.
[6]	STRANSKY N, EGLOFF A M, TWARD A D, et al. The mutational landscape of head and neck squamous cell carcinoma[J]. Science, 2011, 333(6046): 1157-1160. doi: 10.1126/science.1208130 pmid: 21798893
[7]	RIVERA C. Essentials of oral cancer[J]. Int J Clin Exp Pathol, 2015, 8(9): 11884-11894.
[8]	REN X W, KANG B X, ZHANG Z M. Understanding tumor ecosystems by single-cell sequencing: promises and limitations[J]. Genome Biol, 2018, 19(1): 211.
[9]	HAQUE A, ENGEL J, TEICHMANN S A, et al. A practical guide to single-cell RNA-sequencing for biomedical research and clinical applications[J]. Genome Med, 2017, 9(1): 75.
[10]	LEE M N, HWANG H S, OH S H, et al. Elevated extracellular calcium ions promote proliferation and migration of mesenchymal stem cells via increasing osteopontin expression[J]. Exp Mol Med, 2018, 50(11): 1-16. doi: 10.1038/s12276-018-0170-6 pmid: 30393382
[11]	YAMADA S, NOMURA S. Review of single-cell RNA sequencing in the heart[J]. Int J Mol Sci, 2020, 21(21): 8345.
[12]	GAO C X, ZHANG M N, CHEN L. The comparison of two single-cell sequencing platforms: BD rhapsody and 10x genomics chromium[J]. Curr Genomics, 2020, 21(8): 602-609. doi: 10.2174/1389202921999200625220812 pmid: 33414681
[13]	TANG F C, BARBACIORU C, WANG Y Z, et al. mRNA-Seq whole-transcriptome analysis of a single cell[J]. Nat Methods, 2009, 6(5): 377-382. doi: 10.1038/nmeth.1315 pmid: 19349980
[14]	MACOSKO E Z, BASU A, SATIJA R, et al. Highly parallel genome-wide expression profiling of individual cells using nanoliter droplets[J]. Cell, 2015, 161(5): 1202-1214. doi: S0092-8674(15)00549-8 pmid: 26000488
[15]	JAITIN D A, KENIGSBERG E, KEREN-SHAUL H, et al. Massively parallel single-cell RNA-seq for marker-free decomposition of tissues into cell types[J]. Science, 2014, 343(6172): 776-779. doi: 10.1126/science.1247651 pmid: 24531970
[16]	BRANTON D, DEAMER D W, MARZIALI A, et al. The potential and challenges of nanopore sequencing[J]. Nat Biotechnol, 2008, 26(10): 1146-1153. doi: 10.1038/nbt.1495 pmid: 18846088
[17]	RHOADS A, AU K F. PacBio sequencing and its applications[J]. Genomics Proteomics Bioinformatics, 2015, 13(5): 278-289.
[18]	GUPTA I, COLLIER P G, HAASE B, et al. Single-cell isoform RNA sequencing characterizes isoforms in thousands of cerebellar cells[J]. Nat Biotechnol, 2018.[Online ahead of print]
[19]	LEBRIGAND K, MAGNONE V, BARBRY P, et al. High throughput error corrected Nanopore single cell transcriptome sequencing[J]. Nat Commun, 2020, 11(1): 4025.
[20]	BYRNE A, BEAUDIN A E, OLSEN H E, et al. Nanopore long-read RNAseq reveals widespread transcriptional variation among the surface receptors of individual B cells[J]. Nat Commun, 2017, 8: 16027. doi: 10.1038/ncomms16027 pmid: 28722025
[21]	PICELLI S, BJÖRKLUND Å K, FARIDANI O R, et al. Smart-seq2 for sensitive full-length transcriptome profiling in single cells[J]. Nat Methods, 2013, 10(11): 1096-1098. doi: 10.1038/nmeth.2639 pmid: 24056875
[22]	HAGEMANN-JENSEN M, ZIEGENHAIN C, CHEN P, et al. Single-cell RNA counting at allele and isoform resolution using Smart-seq3[J]. Nat Biotechnol, 2020, 38(6): 708-714.
[23]	FAN X Y, YANG C, LI W, et al. SMOOTH-seq: single-cell genome sequencing of human cells on a third-generation sequencing platform[J]. Genome Biol, 2021, 22(1): 195.
[24]	PURAM S V, TIROSH I, PARIKH A S, et al. Single-cell transcriptomic analysis of primary and metastatic tumor ecosystems in head and neck cancer[J]. Cell, 2017, 171(7): 1611-1624.e24. doi: S0092-8674(17)31270-9 pmid: 29198524
[25]	TIROSH I, IZAR B, PRAKADAN S M, et al. Dissecting the multicellular ecosystem of metastatic melanoma by single-cell RNA-seq[J]. Science, 2016, 352(6282): 189-196. doi: 10.1126/science.aad0501 pmid: 27124452
[26]	DARMANIS S, SLOAN S A, CROOTE D, et al. Single-cell RNA-seq analysis of infiltrating neoplastic cells at the migrating front of human glioblastoma[J]. Cell Rep, 2017, 21(5): 1399-1410. doi: S2211-1247(17)31462-6 pmid: 29091775
[27]	AZIZI E, CARR A J, PLITAS G, et al. Single-cell map of diverse immune phenotypes in the breast tumor microenvironment[J]. Cell, 2018, 174(5): 1293-1308.e36. doi: S0092-8674(18)30723-2 pmid: 29961579
[28]	GUO X Y, ZHANG Y Y, ZHENG L T, et al. Global characterization of T cells in non-small-cell lung cancer by single-cell sequencing[J]. Nat Med, 2018, 24(7): 978-985. doi: 10.1038/s41591-018-0045-3 pmid: 29942094
[29]	BELLI C, TRAPANI D, VIALE G, et al. Targeting the microenvironment in solid tumors[J]. Cancer Treat Rev, 2018, 65: 22-32. doi: S0305-7372(18)30014-8 pmid: 29502037
[30]	WEVER O D, DEMETTER P, MAREEL M, et al. Stromal myofibroblasts are drivers of invasive cancer growth[J]. Int J Cancer, 2008, 123(10): 2229-2238. doi: 10.1002/ijc.23925 pmid: 18777559
[31]	MARSH D, SUCHAK K, MOUTASIM K A, et al. Stromal features are predictive of disease mortality in oral cancer patients[J]. J Pathol, 2011, 223(4): 470-481.
[32]	GASCARD P, TLSTY T D. Carcinoma-associated fibroblasts: orchestrating the composition of malignancy[J]. Genes Dev, 2016, 30(9): 1002-1019.
[33]	KUZET S E, GAGGIOLI C. Fibroblast activation in cancer: when seed fertilizes soil[J]. Cell Tissue Res, 2016, 365(3): 607-619.
[34]	LI H P, COURTOIS E T, SENGUPTA D, et al. Reference component analysis of single-cell transcriptomes elucidates cellular heterogeneity in human colorectal tumors[J]. Nat Genet, 2017, 49(5): 708-718. doi: 10.1038/ng.3818 pmid: 28319088
[35]	QI Z T, BARRETT T, PARIKH A S, et al. Single-cell sequencing and its applications in head and neck cancer[J]. Oral Oncol, 2019, 99: 104441.
[36]	RAMAZZOTTI D, ANGARONI F, MASPERO D, et al. Variant calling from scRNA-seq data allows the assessment of cellular identity in patient-derived cell lines[J]. Nat Commun, 2022, 13(1): 2718.
[37]	HARA T, TANEGASHIMA K. Pleiotropic functions of the CXC-type chemokine CXCL14 in mammals[J]. J Biochem, 2012, 151(5): 469-476. doi: 10.1093/jb/mvs030 pmid: 22437940
[38]	SJÖBERG E, AUGSTEN M, BERGH J, et al. Expression of the chemokine CXCL14 in the tumour stroma is an independent marker of survival in breast cancer[J]. Br J Cancer, 2016, 114(10): 1117-1124.
[39]	LIN K Z, ZOU R M, LIN F, et al. Expression and effect of CXCL14 in colorectal carcinoma[J]. Mol Med Rep, 2014, 10(3): 1561-1568. doi: 10.3892/mmr.2014.2343 pmid: 24938992
[40]	NAKAYAMA R, ARIKAWA K, BHAWAL U K. The epigenetic regulation of CXCL14 plays a role in the pathobiology of oral cancers[J]. J Cancer, 2017, 8(15): 3014-3027. doi: 10.7150/jca.21169 pmid: 28928893
[41]	OZAWA S, KATO Y, KOMORI R, et al. BRAK/CXCL14 expression suppresses tumor growth in vivo in human oral carcinoma cells[J]. Biochem Biophys Res Commun, 2006, 348(2): 406-412.
[42]	WESTRICH J A, VERMEER D W, SILVA A, et al. CXCL14 suppresses human papillomavirus-associated head and neck cancer through antigen-specific CD8⁺ T-cell responses by upregulating MHC-I expression[J]. Oncogene, 2019, 38(46): 7166-7180.
[43]	PARIKH A, SHIN J, FAQUIN W, et al. Malignant cell-specific CXCL14 promotes tumor lymphocyte infiltration in oral cavity squamous cell carcinoma[J]. J Immunother Cancer, 2020, 8(2): e001048.
[44]	KONDO T, OZAWA S, IKOMA T, et al. Expression of the chemokine CXCL14 and cetuximab-dependent tumour suppression in head and neck squamous cell carcinoma[J]. Oncogenesis, 2016, 5(7): e240.
[45]	EYQUEM J, MANSILLA-SOTO J, GIAVRIDIS T, et al. Targeting a CAR to the TRAC locus with CRISPR/Cas9 enhances tumour rejection[J]. Nature, 2017, 543(7643): 113-117.
[46]	WALDMAN A D, FRITZ J M, LENARDO M J. A guide to cancer immunotherapy: from T cell basic science to clinical practice[J]. Nat Rev Immunol, 2020, 20(11): 651-668.
[47]	BURTNESS B, HARRINGTON K J, GREIL R, et al. Pembrolizumab alone or with chemotherapy versus cetuximab with chemotherapy for recurrent or metastatic squamous cell carcinoma of the head and neck (KEYNOTE-048): a randomised, open-label, phase 3 study[J]. Lancet, 2019, 394(10212): 1915-1928. doi: S0140-6736(19)32591-7 pmid: 31679945
[48]	HARRINGTON K J, BURTNESS B, GREIL R, et al. Pembrolizumab with or without chemotherapy in recurrent or metastatic head and neck squamous cell carcinoma: updated results of the phase III KEYNOTE-048 study[J]. J Clin Oncol, 2023, 41(4): 790-802.
[49]	CHEN J T, YANG J F, LI H, et al. Single-cell transcriptomics reveal the intratumoral landscape of infiltrated T-cell subpopulations in oral squamous cell carcinoma[J]. Mol Oncol, 2021, 15(4): 866-886. doi: 10.1002/1878-0261.12910 pmid: 33513276
[50]	TOGASHI Y, SHITARA K, NISHIKAWA H. Regulatory T cells in cancer immunosuppression-implications for anticancer therapy[J]. Nat Rev Clin Oncol, 2019, 16(6): 356-371.
[51]	BELTRA J C, MANNE S, ABDEL-HAKEEM M S, et al. Developmental relationships of four exhausted CD8⁺ T cell subsets reveals underlying transcriptional and epigenetic landscape control mechanisms[J]. Immunity, 2020, 52(5): 825-841.e8.
[52]	IM S J, HASHIMOTO M, GERNER M Y, et al. Defining CD8⁺ T cells that provide the proliferative burst after PD-1 therapy[J]. Nature, 2016, 537(7620): 417-421.
[53]	HUDSON W H, GENSHEIMER J, HASHIMOTO M, et al. Proliferating transitory T cells with an effector-like transcriptional signature emerge from PD-1⁺ stem-like CD8⁺ T cells during chronic infection[J]. Immunity, 2019, 51(6): 1043-1058.e4.
[54]	ZEHN D, THIMME R, LUGLI E, et al. ‘Stem-like’ precursors are the fount to sustain persistent CD8⁺ T cell responses[J]. Nat Immunol, 2022, 23(6): 836-847.
[55]	RAHIM M K, OKHOLM T L H, JONES K B, et al. Dynamic CD8⁺ T cell responses to cancer immunotherapy in human regional lymph nodes are disrupted in metastatic lymph nodes[J]. Cell, 2023, 186(6): 1127-1143.e18.
[56]	RAGHU D, XUE H H, MIELKE L A. Control of lymphocyte fate, infection, and tumor immunity by TCF-1[J]. Trends Immunol, 2019, 40(12): 1149-1162. doi: S1471-4906(19)30215-7 pmid: 31734149
[57]	PENG Y, XIAO L P, RONG H X, et al. Single-cell profiling of tumor-infiltrating TCF1/TCF7⁺ T cells reveals a T lymphocyte subset associated with tertiary lymphoid structures/organs and a superior prognosis in oral cancer[J]. Oral Oncol, 2021, 119: 105348.
[58]	HUYNH N C, HUANG T T, NGUYEN C T, et al. Comprehensive integrated single-cell whole transcriptome analysis revealed the p-EMT tumor cells-CAFs communication in oral squamous cell carcinoma[J]. Int J Mol Sci, 2022, 23(12): 6470.
[59]	YOSHITOMI H, UENO H. Shared and distinct roles of T peripheral helper and T follicular helper cells in human diseases[J]. Cell Mol Immunol, 2021, 18(3): 523-527.
[60]	GU-TRANTIEN C, MIGLIORI E, BUISSERET L, et al. CXCL13-producing TFH cells link immune suppression and adaptive memory in human breast cancer[J]. JCI Insight, 2017, 2(11): e91487.