establishment-histological-analysis-esophageal-organoids-modeling

Establishment and Histological Analysis of Esophageal Organoids Modeling the Progression from Normal to Cancerous Tissues

Organoids have emerged as a pivotal tool for advancing the understanding of tumorigenesis and cancer therapy. By generating human organoid models ...

Chinese Academy of Medical Sciences (CAMS) and Peking Union Medical College (PUMC)

Esophageal squamous-cell carcinoma (ESCC) is commonly treated with radiotherapy; however, radioresistance hinders its clinical effectiveness, and the underlying mechanism remains elusive. Here, we develop patient-derived xenografts (PDXs) from 19 patients with ESCC to investigate the mechanisms driving radioresistance. Using RNA sequencing, cytokine arrays, and single-cell RNA sequencing, we reveal an enrichment of cancer-associated fibroblast (CAF)-derived collagen type 1 (Col1) and tumor-cell-derived CXCL1 in non-responsive PDXs. Col1 not only promotes radioresistance by augmenting DNA repair capacity but also induces CXCL1 secretion in tumor cells. Additionally, CXCL1 further activates CAFs via the CXCR2-STAT3 pathway, establishing a positive feedback loop. Directly interfering with tumor-cell-derived CXCL1 or inhibiting the CXCL1-CXCR2 pathway effectively restores the radiosensitivity of radioresistant xenografts in vivo. Collectively, our study provides a comprehensive understanding of the molecular mechanisms underlying radioresistance and identifies potential targets to improve the efficacy of radiotherapy for ESCC.

Collagen 1-mediated CXCL1 secretion in tumor cells activates fibroblasts to promote radioresistance of esophageal cancer.

Chemotherapy remains the first-line treatment for advanced esophageal cancer. However, durable benefits are achieved by only a limited subset of individuals due to the elusive chemoresistance. Here, we utilize patient-derived xenografts (PDXs) from esophageal squamous-cell carcinoma to investigate chemoresistance mechanisms in preclinical settings. We observe that activated cancer-associated fibroblasts (CAFs) are enriched in the tumor microenvironment of PDXs resistant to chemotherapy. Mechanistically, we reveal that cancer-cell-derived S100A8 triggers the intracellular RhoA-ROCK-MLC2-MRTF-A pathway by binding to the CD147 receptor of CAFs, inducing CAF polarization and leading to chemoresistance. Therapeutically, we demonstrate that blocking the S100A8-CD147 pathway can improve chemotherapy efficiency. Prognostically, we found the S100A8 levels in peripheral blood can serve as an indicator of chemotherapy responsiveness. Collectively, our study offers a comprehensive understanding of the molecular mechanisms underlying chemoresistance in esophageal cancer and highlights the potential value of S100A8 in the clinical management of esophageal cancer.

Alarmin S100A8 imparts chemoresistance of esophageal cancer by reprogramming cancer-associated fibroblasts.

Multiple sampling strategies to cover different or dynamic stages of malignant continuum with organoid cultures provide a valuable platform for epithelium homeostasis, transformation, and cancer progression. Here, we present a protocol to initiate, culture, passage, and characterize organoids from normal and cancer-prone human esophageal tissues. We describe steps for multiple sampling of malignant continuum and the initiation and maintenance of multi-stage organoids. We then detail procedures for the histological characterization of organoids and co-culture systems based on organoids and stromal cells. For complete details on the use and execution of this protocol, please refer to Chen et al..

Protocol for the generation, characterization, and functional assays of organoid cultures from normal and cancer-prone human esophageal tissues.

Cancer-associated fibroblasts (CAF) contribute to cancer initiation and progression and play a pivotal role in therapeutic response and patient prognosis. CAFs exhibit functional and phenotypic heterogeneity, highlighting the need to clarify the specific subtypes of CAFs to facilitate the development of targeted therapies against protumorigenic CAFs. In this study, using single-cell RNA sequencing on patient samples of esophageal squamous cell carcinoma (ESCC), we identified a CAF subcluster associated with tumor stemness that was enriched in genes associated with hypoxia and senescence. The CAF subpopulation, termed as hypoxia-induced senescent fibroblasts (hsCAF), displayed high secretion of insulin-like growth factor 1 (IGF1). The hsCAFs inhibited AMP-activated protein kinase (AMPK) activity in cancer cells via IGF1 to promote tumor stemness. The formation of hsCAFs was induced by the synergetic effect of hypoxia and cancer cells. Activation of nuclear factor erythroid 2-related factor 2 (NRF2) in cancer cells under hypoxia drove IL1α production to trigger CAF senescence and IGF1 secretion via nuclear factor I A. Knockout of IGF1 in CAFs or nuclear factor erythroid 2-related factor 2 in ESCC cells suppressed the tumor growth and chemotherapy resistance induced by CAFs in vivo. Importantly, patients with high proportions of hsCAFs showed poor survival and a worse response to chemotherapy. In summary, these findings identify a hsCAF subpopulation generated by interplay between cancer cells and CAFs under hypoxic conditions that promotes ESCC stemness and reveal targeting hsCAFs as an effective therapeutic strategy against chemotherapy-resistant ESCC. Significance: A hypoxic microenvironment and cancer cells cooperate to induce a senescent fibroblast subset that supports tumor stemness, suggesting that targeting this cancer-associated fibroblast subpopulation is a potential therapeutic strategy to overcome chemoresistance.

Hypoxia-Induced Senescent Fibroblasts Secrete IGF1 to Promote Cancer Stemness in Esophageal Squamous Cell Carcinoma.

Cancer development involves the co-evolution of cancer cells and their surrounding microenvironment, yet the dynamics of this interaction within the physical architecture remains poorly understood. Here, we present a spatial transcriptomic map at single-cell resolution, encompassing 127 multi-stage fields of view from 43 patients, to chart the evolutionary trajectories of human esophageal squamous cell carcinoma (ESCC). By analyzing 6.4 million cells, we reveal that ESCC progression is driven by a proliferative epithelial cell subpopulation that acquires dedifferentiated and invasive characteristics. At the late precancerous stage, these cells disrupt the epithelial-stromal interface and recruit normal fibroblasts via JAG1-NOTCH1 signaling, transforming them into cancer-associated fibroblasts (CAFs). This interaction leads to the formation of a "CAF-Epi" (CAF and epithelial cell) niche at the tumor edge that shields the tumor from immune surveillance. The CAF-Epi niche formation is a key indicator of progression in ESCC and other squamous cell carcinomas and patient outcomes.

Article	Year
Collagen 1-mediated CXCL1 secretion in tumor cells activates fibroblasts to promote radioresistance of esophageal cancer. Cell reports\| PubMed ID: 37851572	2023
Alarmin S100A8 imparts chemoresistance of esophageal cancer by reprogramming cancer-associated fibroblasts. Cell reports. Medicine\| PubMed ID: 38776909	2024
Protocol for the generation, characterization, and functional assays of organoid cultures from normal and cancer-prone human esophageal tissues. STAR protocols\| PubMed ID: 39277865	2024
Hypoxia-Induced Senescent Fibroblasts Secrete IGF1 to Promote Cancer Stemness in Esophageal Squamous Cell Carcinoma. Cancer research\| PubMed ID: 39661488	2025
Single-cell multi-stage spatial evolutional map of esophageal carcinogenesis. Cancer cell\| PubMed ID: 40068596	2025

Guoyu Cheng

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