The molecular classification of cancer‐associated fibroblasts on a pan‐cancer single‐cell transcriptional atlas

Abstract Background Cancer‐associated fibroblasts (CAFs), integral to the tumour microenvironment, are pivotal in cancer progression, exhibiting either pro‐tumourigenic or anti‐tumourigenic functions. Their inherent phenotypic and functional diversity allows for the subdivision of CAFs into various subpopulations. While several classification systems have been suggested for different cancer types, a unified molecular classification of CAFs on a single‐cell pan‐cancer scale has yet to be established. Methods We employed a comprehensive single‐cell transcriptomic atlas encompassing 12 solid tumour types. Our objective was to establish a novel molecular classification and to elucidate the evolutionary trajectories of CAFs. We investigated the functional profiles of each CAF subtype using Single‐Cell Regulatory Network Inference and Clustering and single‐cell gene set enrichment analysis. The clinical relevance of these subtypes was assessed through survival curve analysis. Concurrently, we employed multiplex immunofluorescence staining on tumour tissues to determine the dynamic changes of CAF subtypes across different tumour stages. Additionally, we identified the small molecule procyanidin C1 (PCC1) as a target for matrix‐producing CAF (matCAF) using molecular docking techniques and further validated these findings through in vitro and in vivo experiments. Results In our investigation of solid tumours, we identified four molecular clusters of CAFs: progenitor CAF (proCAF), inflammatory CAF (iCAF), myofibroblastic CAF (myCAF) and matCAF, each characterised by distinct molecular traits. This classification was consistently applicable across all nine studied solid tumour types. These CAF subtypes displayed unique evolutionary pathways, functional roles and clinical relevance in various solid tumours. Notably, the matCAF subtype was associated with poorer prognoses in several cancer types. The targeting of matCAF using the identified small molecule, PCC1, demonstrated promising antitumour activity. Conclusions Collectively, the various subtypes of CAFs, particularly matCAF, are crucial in the initiation and progression of cancer. Focusing therapeutic strategies on targeting matCAF in solid tumours holds significant potential for cancer treatment.

[1]  Jun Tian,et al.  Procyanidin C1 inhibits tumor growth and metastasis in colon cancer via modulating miR-501-3p/HIGD1A axis , 2023, Journal of advanced research.

[2]  K. To,et al.  The E2F1–HOXB9/PBX2–CDK6 axis drives gastric tumorigenesis and serves as a therapeutic target in gastric cancer , 2023, The Journal of pathology.

[3]  Yao Fu,et al.  NNMT enriches for AQP5+ cancer stem cells to drive malignant progression in early gastric cardia adenocarcinoma , 2023, Gut.

[4]  C. Li,et al.  Pan-cancer single-cell analysis reveals the heterogeneity and plasticity of cancer-associated fibroblasts in the tumor microenvironment , 2022, Nature Communications.

[5]  B. Parvin,et al.  CD36+ Fibroblasts Secrete Protein Ligands That Growth-Suppress Triple-Negative Breast Cancer Cells While Elevating Adipogenic Markers for a Model of Cancer-Associated Fibroblast , 2022, International journal of molecular sciences.

[6]  S. Herrell,et al.  Single cell analysis of cribriform prostate cancer reveals cell intrinsic and tumor microenvironmental pathways of aggressive disease , 2022, Nature Communications.

[7]  Zongfu Pan,et al.  CREB3L1 promotes tumor growth and metastasis of anaplastic thyroid carcinoma by remodeling the tumor microenvironment , 2022, Molecular Cancer.

[8]  Jordan F Hastings,et al.  Temporal profiling of the breast tumour microenvironment reveals collagen XII as a driver of metastasis , 2022, Nature Communications.

[9]  R. Kalluri,et al.  Oncogenic collagen I homotrimers from cancer cells bind to α3β1 integrin and impact tumor microbiome and immunity to promote pancreatic cancer. , 2022, Cancer cell.

[10]  H. Crawford,et al.  Hypoxia promotes an inflammatory phenotype of fibroblasts in pancreatic cancer , 2022, bioRxiv.

[11]  Di Huang,et al.  Turning cold tumors hot: from molecular mechanisms to clinical applications. , 2022, Trends in immunology.

[12]  R. Sullivan,et al.  STAG2 regulates interferon signaling in melanoma via enhancer loop reprogramming , 2022, Nature Communications.

[13]  E. Cukierman The Few yet Fabp4ulous Pancreatic Stellate Cells Give Rise to Protumoral CAFs. , 2022, Cancer discovery.

[14]  J. Campisi,et al.  The flavonoid procyanidin C1 has senotherapeutic activity and increases lifespan in mice , 2021, Nature Metabolism.

[15]  Hong-Jian Zhu,et al.  Cancer associated-fibroblast-derived exosomes in cancer progression , 2021, Molecular Cancer.

[16]  M. Samuel,et al.  Differentiation of the tumor microenvironment: are CAFs the Organizer? , 2021, Trends in cell biology.

[17]  Jun Yu,et al.  STK3 promotes gastric carcinogenesis by activating Ras-MAPK mediated cell cycle progression and serves as an independent prognostic biomarker , 2021, Molecular cancer.

[18]  C. Perou,et al.  A multi-omic single-cell landscape of human gynecologic malignancies. , 2021, Molecular cell.

[19]  M. Teh,et al.  Single-Cell Atlas of Lineage States, Tumor Microenvironment, and Subtype-Specific Expression Programs in Gastric Cancer , 2021, Cancer discovery.

[20]  M. Mino‐Kenudson,et al.  Three subtypes of lung cancer fibroblasts define distinct therapeutic paradigms. , 2021, Cancer cell.

[21]  R. Kalluri,et al.  Clinical and therapeutic relevance of cancer-associated fibroblasts , 2021, Nature Reviews Clinical Oncology.

[22]  C. Jørgensen,et al.  Single-cell analysis defines a pancreatic fibroblast lineage that supports anti-tumor immunity , 2021, Cancer cell.

[23]  J. Visvader,et al.  A single‐cell RNA expression atlas of normal, preneoplastic and tumorigenic states in the human breast , 2021, The EMBO journal.

[24]  S. Hingorani,et al.  Mesenchymal Lineage Heterogeneity Underlies Nonredundant Functions of Pancreatic Cancer–Associated Fibroblasts , 2021, bioRxiv.

[25]  R. Schwabe,et al.  Promotion of cholangiocarcinoma growth by diverse cancer-associated fibroblast subpopulations. , 2021, Cancer cell.

[26]  S. Mandrup,et al.  Transcriptional networks controlling stromal cell differentiation , 2021, Nature Reviews Molecular Cell Biology.

[27]  Ruhong Li,et al.  Cancer-associated fibroblasts: overview, progress, challenges, and directions , 2021, Cancer Gene Therapy.

[28]  R. Kalluri,et al.  Type I collagen deletion in αSMA+ myofibroblasts augments immune suppression and accelerates progression of pancreatic cancer. , 2021, Cancer cell.

[29]  D. Zheng,et al.  Molecular Features of Cancer-associated Fibroblast Subtypes and their Implication on Cancer Pathogenesis, Prognosis, and Immunotherapy Resistance , 2021, Clinical Cancer Research.

[30]  Wei Xue,et al.  Single-cell analysis supports a luminal-neuroendocrine transdifferentiation in human prostate cancer , 2020, Communications biology.

[31]  R. Yin,et al.  Biomarkers for cancer-associated fibroblasts , 2020, Biomarker research.

[32]  Lihua Zhang,et al.  Inference and analysis of cell-cell communication using CellChat , 2020, Nature Communications.

[33]  Yumin Li,et al.  The advances in immunotherapy for hepatocellular carcinoma , 2020 .

[34]  Uri Alon,et al.  Cancer-associated fibroblast compositions change with breast cancer progression linking the ratio of S100A4+ and PDPN+ CAFs to clinical outcome , 2020, Nature Cancer.

[35]  Yuan Zhang,et al.  Single cell transcriptomic architecture and intercellular crosstalk of human intrahepatic cholangiocarcinoma. , 2020, Journal of hepatology.

[36]  D. Tuveson,et al.  DIVERSITY AND BIOLOGY OF CANCER-ASSOCIATED FIBROBLASTS. , 2020, Physiological reviews.

[37]  Y. Cho,et al.  Lineage-dependent gene expression programs influence the immune landscape of colorectal cancer , 2020, Nature Genetics.

[38]  K. Tarte,et al.  Single-cell analysis reveals fibroblast clusters linked to immunotherapy resistance in cancer. , 2020, Cancer discovery.

[39]  Nayoung K. D. Kim,et al.  Single-cell RNA sequencing demonstrates the molecular and cellular reprogramming of metastatic lung adenocarcinoma , 2020, Nature Communications.

[40]  Michael A. Durante,et al.  Single-cell analysis reveals new evolutionary complexity in uveal melanoma , 2020, Nature Communications.

[41]  A. Vincent-Salomon,et al.  Cancer-associated fibroblast heterogeneity in axillary lymph nodes drives metastases in breast cancer through complementary mechanisms , 2020, Nature Communications.

[42]  Gustavo Stolovitzky,et al.  Intratumoral heterogeneity and clonal evolution in liver cancer , 2020, Nature Communications.

[43]  Ashley M. Laughney,et al.  Regenerative lineages and immune-mediated pruning in lung cancer metastasis , 2019, Nature Medicine.

[44]  R. Bourgon,et al.  Single-cell RNA sequencing reveals stromal evolution into LRRC15+ myofibroblasts as a determinant of patient response to cancer immunotherapy. , 2019, Cancer discovery.

[45]  Chi-kong Li,et al.  CD9 blockade suppresses disease progression of high-risk pediatric B-cell precursor acute lymphoblastic leukemia and enhances chemosensitivity , 2019, Leukemia.

[46]  Hanlee P. Ji,et al.  Single-Cell Genomic Characterization Reveals the Cellular Reprogramming of the Gastric Tumor Microenvironment , 2019, Clinical Cancer Research.

[47]  Siwei Wang,et al.  Cancer-associated fibroblasts: an emerging target of anti-cancer immunotherapy , 2019, Journal of Hematology & Oncology.

[48]  Yun-Gui Yang,et al.  Single-cell RNA-seq highlights intra-tumoral heterogeneity and malignant progression in pancreatic ductal adenocarcinoma , 2019, Cell Research.

[49]  Qihao Ren,et al.  Unmasking senescence: context-dependent effects of SASP in cancer , 2019, Nature Reviews Cancer.

[50]  M. Rojas,et al.  Single-cell analysis reveals fibroblast heterogeneity and myofibroblasts in systemic sclerosis-associated interstitial lung disease , 2019, Annals of the Rheumatic Diseases.

[51]  P. Chiao,et al.  Two Birds with One Stone: Therapeutic Targeting of IL1α Signaling Pathways in Pancreatic Ductal Adenocarcinoma and the Cancer-Associated Fibroblasts. , 2019, Cancer discovery.

[52]  Åsa K. Björklund,et al.  Spatially and functionally distinct subclasses of breast cancer-associated fibroblasts revealed by single cell RNA sequencing , 2018, Nature Communications.

[53]  Wenbin Lin,et al.  Nanoparticle-Mediated Immunogenic Cell Death Enables and Potentiates Cancer Immunotherapy. , 2018, Angewandte Chemie.

[54]  Paul J. Hoffman,et al.  Comprehensive Integration of Single-Cell Data , 2018, Cell.

[55]  D. Tuveson,et al.  IL1-Induced JAK/STAT Signaling Is Antagonized by TGFβ to Shape CAF Heterogeneity in Pancreatic Ductal Adenocarcinoma. , 2018, Cancer discovery.

[56]  E. Song,et al.  Turning foes to friends: targeting cancer-associated fibroblasts , 2018, Nature Reviews Drug Discovery.

[57]  E. Puré,et al.  FAP Delineates Heterogeneous and Functionally Divergent Stromal Cells in Immune-Excluded Breast Tumors , 2018, Cancer Immunology Research.

[58]  Dongyuan Lü,et al.  Effects of mechanical stretching on the morphology of extracellular polymers and the mRNA expression of collagens and small leucine-rich repeat proteoglycans in vaginal fibroblasts from women with pelvic organ prolapse , 2018, PloS one.

[59]  Paul Hoffman,et al.  Integrating single-cell transcriptomic data across different conditions, technologies, and species , 2018, Nature Biotechnology.

[60]  J. Aerts,et al.  SCENIC: Single-cell regulatory network inference and clustering , 2017, Nature Methods.

[61]  D. Tuveson,et al.  Br Ief Definitive Repor T , 2022 .

[62]  R. Schwabe,et al.  The Role of Cancer-Associated Fibroblasts and Fibrosis in Liver Cancer. , 2017, Annual review of pathology.

[63]  R. Kalluri The biology and function of fibroblasts in cancer , 2016, Nature Reviews Cancer.

[64]  T. Welling,et al.  GM-CSF Mediates Mesenchymal-Epithelial Cross-talk in Pancreatic Cancer. , 2016, Cancer discovery.

[65]  Hans Clevers,et al.  Long-Term Culture of Genome-Stable Bipotent Stem Cells from Adult Human Liver , 2015, Cell.

[66]  Qi Zhang,et al.  Myocyte enhancer factor 2C regulation of hepatocellular carcinoma via vascular endothelial growth factor and Wnt/β-catenin signaling , 2014, Oncogene.

[67]  Cole Trapnell,et al.  Pseudo-temporal ordering of individual cells reveals dynamics and regulators of cell fate decisions , 2014, Nature Biotechnology.

[68]  H. Sakurai,et al.  Procyanidin C1 from Cinnamomi Cortex inhibits TGF-β-induced epithelial-to-mesenchymal transition in the A549 lung cancer cell line. , 2013, International journal of oncology.

[69]  B. Alman,et al.  Identification of IGFBP-6 as a significantly downregulated gene by β-catenin in desmoid tumors , 2004, Oncogene.

[70]  M. Rifkin,et al.  Echogenicity of prostate cancer correlated with histologic grade and stromal fibrosis: endorectal US studies. , 1989, Radiology.

[71]  R. Gronostajski,et al.  Nuclear Factor One X in Development and Disease. , 2019, Trends in cell biology.