These findings define functionally distinct and pharmacologically targetable CAF programs in HNSCC and support stromal-directed interventions as a rational component of future combination treatment strategies.
Abstract
Head and neck squamous cell carcinoma (HNSCC) remains clinically challenging, with limited molecularly targeted options and a strong dependence on the tumor microenvironment. Cancer-associated fibroblasts (CAFs) are major stromal regulators that shape tumor progression, extracellular matrix remodeling, invasion, and therapeutic response. However, how CAF heterogeneity and plasticity translate into distinct tumor-promoting functions and targetable vulnerabilities remains insufficiently defined. Here, we integrated patient-matched primary CAFs and normal fibroblasts with 3D functional assays, tumor–stroma co-culture models, quantitative extracellular matrix analysis, whole-proteome profiling, and pharmacological perturbation. Primary fibroblast populations displayed marked interpatient heterogeneity and context-dependent plasticity in invasion, contractility, and responsiveness to tumor-derived signals, whereas enhanced fibronectin-rich matrix deposition and disorganization emerged as a conserved CAF-associated feature. Both normal fibroblasts and CAFs promoted HNSCC cell invasion in a population-dependent manner, whereas CAFs consistently induced less compact and more dispersed tumor nest architectures. Integrative functional analyses identified distinct CAF phenotypes characterized by either invasive and matrix-remodeling activity or high responsiveness to tumor-derived cues. Proteomic profiling revealed recurrent enrichment of adhesion, cytoskeletal, and extracellular matrix programs and guided the selection of pharmacological inhibitors aimed at modulating specific CAF-mediated pro-tumoral functions. Pharmacological targeting selectively altered these functions: CHI3L1 inhibition disrupted fibronectin matrix deposition, broad phosphodiesterase inhibition increased matrix alignment, and FZD7 inhibition consistently blocked tumor-induced CAF invasion across all tested populations. These findings define functionally distinct and pharmacologically targetable CAF programs in HNSCC and support stromal-directed interventions as a rational component of future combination treatment strategies.
Immune checkpoint inhibitors (ICIs) have achieved breakthroughs in various solid tumors; however, their efficacy remains limited in prostate cancer (PCa), which is closely associated with its highly immunosuppressive tumor microenvironment (TME). As the most abundant stromal component in the TME, cancer-associated fibroblasts (CAFs) exhibit remarkable heterogeneity and functional plasticity, playing a central role in orchestrating an immunologically "cold" tumor phenotype and driving resistance to immunotherapy. We have moved beyond simple marker-based classification of cancer-associated fibroblast (CAF) subtypes — a framework still widely used in routine pathological assessment — and now define these populations by their functional states and molecular profiles, enabled by advances in single-cell sequencing, spatial multi-omics, and computational biology. Such functional divergence across stromal subsets allows individual CAF populations to engage in complex, multilayered crosstalk with different immune cell types, and together they build an immunosuppressive network that drives tumor immune evasion. This review systematically catalogs the identification methods and signature gene panels for distinct CAF subtypes in prostate cancer, with particular focus on the multilayered regulatory circuits through which these stromal cells shape the immune microenvironment. We also highlight emerging CAF-targeted strategies designed to reverse immunosuppressive states. This integrated perspective frames CAF heterogeneity in prostate cancer immune evasion, and establishes theoretical underpinnings and translational pathways for next-generation precision combination immunotherapies.
Kan-Kan He, Zhi-Te Zhao, Jian-Hui Bai et al.· International Journal on Bio...· 0 citations
Pancreatic ductal adenocarcinoma (PDAC) is characterized by a heavily fibrotic tumor microenvironment (TME), primarily driven by cancer-associated fibroblasts (CAFs). Due to the high heterogeneity and diverse biological behaviors of CAFs, the current strategies targeting CAFs have limited benefits in clinic. This study identifies a subset of CAFs marked by high expression of connective tissue growth factor (CTGF), which enhances tumor cell migration and correlates with advanced PDAC stages. By combining in silico screening and functional assays, we identified DP as a first-in-class small-molecule inhibitor targeting CTGF. Structural optimization of DP led to the synthesis of a series of derivatives, from which the optimal compound QX03-46 was identified. Targeting CTGF+ CAFs with QX03-46 significantly disrupted the CTGF/TGF-β1 signaling axis and suppressed CAF functions, resulting in decreased extracellular matrix deposition and oncogenic cytokine secretion. Furthermore, it enhanced the efficacy of anti-programmed cell death ligand 1 therapy, promoting a more favorable immune response. Our findings illuminate the critical role of CTGF+ CAFs in PDAC progression and position QX03-46 as a promising therapeutic candidate for targeting the fibrotic TME.
Yi-Wen Shen, Qing-Yan Sun, Yunzhe Wang et al.· Proceedings of the National...· 0 citations
An integrated analysis of EwS tumor samples reveals a dynamic tumor–CAF continuum that is uniquely preserved in primary tumors and establishes a framework for identifying clinically relevant tumor-associated surface TAs, providing a foundation for the rational development of next-generation immunotherapies and precision-targeted therapies for patients with EwS.
M. Sharif, Abdul S. Khan, R. Brentjens et al.· bioRxiv· 0 citations
Cancer-associated fibroblasts (CAFs) exhibit substantial heterogeneity in colorectal cancer (CRC), yet the upstream tumor-derived signals associated with their state remodeling during disease progression remain incompletely defined. Through integrative analysis of multi-cohort single-cell transcriptomics, proteomics, and bulk RNA sequencing data, we identified tumor-derived midkine (MDK) as a stage-associated candidate signal linked to CAF remodeling in CRC.
MDK cell-type specificity and stage-associated expression, together with CAF composition and stage-dependent remodeling, were evaluated in two large colorectal cancer single-cell RNA-sequencing cohorts, with the principal findings cross-validated between cohorts and further integrated with bulk transcriptomic, proteomic, and immunohistochemical datasets. Existing malignant-cell-to-CAF CellChat results were re-ranked using a joint breadth–strength framework to identify broadly activated and high-probability signaling pathways. CAF trajectories were reconstructed independently in both cohorts using Monocle 2 and rooted according to the same biological criterion, namely enrichment of the normal-like Fib-MFAP5 population. BayesPrism was used to deconvolve CAF subtype abundance from TCGA colorectal cancer bulk transcriptomes and to evaluate its association with overall survival.
MDK was consistently enriched in malignant epithelial cells across both single-cell cohorts. Within the same-platform bulk RNA cohort, median log2-normalized MDK expression increased from 7.93 in adenoma to 8.19 in carcinoma in situ and 9.02 in adenocarcinoma, supporting a significant ordered increase during colorectal cancer progression (Jonckheere–Terpstra
$$P=0.008$$
; 10,000-permutation
$$P=0.007$$
). Fib-WNT5A abundance likewise showed a stage-associated increase and was positioned significantly later than the normal-like Fib-MFAP5 population in patient-level pseudotime analyses of both the discovery and validation cohorts. Fib-WNT5A exhibited strong TGF-β and extracellular matrix programs, and integration of subtype-associated ECM signatures with bulk proteomics linked this population to collagen-enriched matrix remodeling. Joint breadth–strength analysis identified MK signaling as the top-ranked broadly activated malignant-cell-to-CAF pathway. CellChat further predicted COL1A1/COL1A2–CD44 interactions between Fib-WNT5A and multiple immune populations, supporting an association between Fib-WNT5A expansion and an ECM-rich, immune-restrictive microenvironment.
Patient-level pseudo-bulk analyses were used to evaluate BayesPrism recovery of major CAF states. In TCGA CRC, higher Fib-WNT5A abundance was associated with poorer overall survival in the prespecified median-based univariable analysis (HR = 1.64, 95% CI 1.09–2.49; log-rank
P
= 0.018), whereas higher Fib-MFAP5 abundance was associated with better survival (HR = 0.54, 95% CI 0.35–0.83; log-rank
P
= 0.0045).
Collectively, the data support a progression-linked association among tumor-cell-enriched MDK, CAF state remodeling, an ECM-high Fib-WNT5A program, and predicted collagen-mediated immune interactions in CRC. Functional perturbation studies are required to establish the direction and causality of the proposed MDK-CAF axis.
Laryngeal squamous cell carcinoma (LSCC) remains a clinically challenging malignancy of the head and neck. Because organ preservation is a major therapeutic objective in LSCC, stromal biology is of particular clinical relevance. Stromal factors influence local disease control, functional outcomes, and treatment resistance. Cancer-associated fibroblasts (CAFs) are dynamic regulators of extracellular matrix remodeling, immune exclusion, angiogenesis, and therapeutic resistance. However, LSCC-specific evidence remains fragmented, and many mechanistic concepts are still extrapolated from head and neck squamous cell carcinoma (HNSCC) and hypopharyngeal squamous cell carcinoma (HPSCC). This review adopts an evidence-stratified framework that distinguishes direct LSCC evidence from translatable cross-site evidence and hypothesis-generating inferences. We propose a provisional five-state model for LSCC that includes myofibroblastic, inflammatory, antigen-presenting, extracellular matrix-remodeling, and putative immune-trapping CAF programs. Direct LSCC studies support the presence of early stromal activation along the leukoplakia-to-carcinoma continuum and the enrichment of FAP-positive and POSTN-positive fibroblastic programs in metastatic disease. Associations involving stromal PD-L1 expression and altered CAF-derived exosomal microRNA cargo have also been reported. Translatable evidence from HNSCC implicates TGF-β- and CXCL12-dependent T-cell exclusion together with Gal9-mediated T-cell dysfunction. Additional mechanisms include IL-6/JAK/STAT3-driven myeloid skewing and NOX4-dependent stabilization of the myofibroblastic phenotype. We integrate these findings into a spatially informed conceptual framework encompassing the invasive front, the perivascular compartment, the cartilage interface, and the lymphovascular niche. We also discuss biomarker-guided therapeutic opportunities, including CAF normalization, FAP-directed approaches, stromal signaling blockade, extracellular matrix-targeted strategies, and interference with extracellular vesicle-mediated signaling. This review thereby establishes an evidence-stratified framework for CAF heterogeneity in LSCC and clarifies the translational value and current limitations of stroma-targeted strategies. It further provides a structured foundation for hypothesis-driven clinical and experimental investigation in laryngeal cancer.
Wei-Zhe Hong, Guan-Jiang Huang, Qing Luo et al.· Frontiers in Immunology· 0 citations
Gastric cancer (GC) remains a major global health challenge characterized by poor prognosis and high mortality rates, despite advances in anticancer therapies. Increasing evidence indicates that GC is a highly stromal-rich tumor, with cancer-associated fibroblasts (CAFs) serving as key stromal components of the tumor microenvironment (TME). CAFs exhibit substantial metabolic heterogeneity, such that distinct CAF subtypes demonstrate intrinsic differences and dynamic remodeling in glucose, lipid and amino acid metabolism. This metabolic reprogramming represents not only functional diversity but also an adaptive response to microenvironmental stressors, including tumor-derived signals, hypoxia, nutrient fluctuations and oxidative stress. Through these adaptations, CAFs engage in extensive crosstalk with tumor and immune cells, thereby contributing to microenvironmental remodeling. Due to their critical role and abundance in GC, CAFs have emerged as promising therapeutic targets. The present review summarizes the origins, characteristics and heterogeneity of CAFs in GC and discusses their interactions with the TME. In addition, it summarizes the metabolic heterogeneity of CAFs, explores potential metabolism-targeted therapeutic strategies and outlines the broader impact of CAFs on the TME.
Qi-Lu Fang, Neng Lv, Shuwei Yu et al.· Oncology Letters· 1 citation
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