Rapid normal fibroblast‐tumor crosstalk promotes cancer‐associated fibroblast‐like activation and attenuates mitomycin C cytotoxicity in bladder cancer
Aug 2026· Journal of cell communication and signaling· Vol 20· 0 citations· 20 references
Medicine
TL;DR
Results show that NF‐tumor crosstalk is rapid, bidirectional and functionally relevant to MMC response in vitro, and NFs can rapidly acquire CAF‐like activation features following tumor‐derived stimulation and contribute to a stromal context associated with EMT‐like plasticity and reduced MMC‐induced cytotoxicity in vitro.
Abstract
Abstract Normal fibroblasts (NFs) are among the earliest stromal cells encountered by bladder cancer cells during invasion into the lamina propria, yet their contribution to tumor progression and intravesical chemotherapy response remains poorly defined. Here, we investigated whether NFs are passive stromal bystanders or active participants in early bladder cancer microenvironmental remodeling. Analysis of an early‐stage non‐muscle‐invasive bladder cancer transcriptomic cohort revealed that fibroblast‐associated scores were higher in T1 than Ta tumors, enriched in European Organisation for Research and Treatment of Cancer high‐risk disease, positively correlated with epithelial‐mesenchymal transition (EMT) scores, and associated with increased expression of resistance‐linked genes. Experimentally, fibroblast‐conditioned medium reduced bladder cancer cell proliferation while accelerating wound closure, indicating a less proliferative but more motile phenotype. This was accompanied by EMT‐like cadherin remodeling, including reduced E‐cadherin and increased N‐cadherin staining. Reciprocally, tumor‐derived signals rapidly induced cancer‐associated fibroblast (CAF)‐like activation features in NFs within 48 h, including increased α‐smooth muscle actin, fibroblast activation protein, and PDGFRβ expression. Functionally, increasing fibroblast‐to‐tumor cell ratios progressively attenuated mitomycin C (MMC)‐induced cytotoxicity, demonstrating that stromal context can modify chemotherapy response in vitro. Together, these findings show that NF‐tumor crosstalk is rapid, bidirectional and functionally relevant to MMC response in vitro. Rather than acting only after stable CAF formation, NFs can rapidly acquire CAF‐like activation features following tumor‐derived stimulation and contribute to a stromal context associated with EMT‐like plasticity and reduced MMC‐induced cytotoxicity in vitro.
ABSTRACT Therapy resistance in prostate cancer arises from coordinated remodeling of malignant and stromal compartments, yet the mechanisms orchestrating this ecosystem adaptation remain elusive. Here, single‐cell RNA sequencing of longitudinal biopsies obtained before and after androgen‐deprivation therapy (ADT) delineated a therapy‐induced stromal lineage bifurcation toward APOD+ and DPT+ fibroblast states. DPT+ fibroblasts activated a C3‐ITGAX/ITGB2 complement signaling axis targeting macrophages, coinciding with suppression of M1 inflammatory programs, amplification of immune‐checkpoint signaling, and a shift of CD8+ T cells from cytotoxic to exhausted phenotypes. Concomitantly, we identified pre‐existing malignant epithelial subpopulations characterized by reduced AR/KLK3 activity and heightened chromosomal instability that preferentially persisted following therapy. Integrative multi‐omic analyses nominated TSPAN1 as a functional effector of castrate resistant prostate cancer (CRPC) and NRXN1 as a regulator of neuroendocrine plasticity through calcium‐dependent signaling programs. Genetic silencing of either gene suppressed proliferation, clonogenicity, migration, and tumor growth, while attenuating neuroendocrine features in vitro and in vivo. Spatial mapping, functional perturbation, and stromal‐epithelial co‐culture experiments mechanistically established a therapy‐induced DPT+ fibroblast‐complement circuit that enforced immune evasion and channels epithelial trajectories toward CRPC or neuroendocrine prostate cancer. Collectively, these findings defined the DPT+‐complement‐macrophage axis as an actionable vulnerability and position TSPAN1 and NRXN1 as therapeutic entry points to disrupt ADT‐driven tumor ecosystem remodeling in prostate cancer.
Yang Chen, Dandan Dong, Jinling Liao et al.· Advancement of science· 0 citations
Tumor progression and metastasis in bladder cancer are driven by epithelial cell heterogeneity and dynamic interactions with the tumor microenvironment. To elucidate epithelial subpopulations associated with cancer progression, we performed single-cell transcriptomic profiling of bladder cancer tissues and identified a distinct epithelial subset, termed Meta-program 6 (MP6), that was significantly enriched in samples exhibiting lymphovascular invasion or lymph node metastasis. The MP6 gene signature correlated strongly with advanced tumor stage, lymph node involvement, and lymphovascular infiltration. CNV analysis revealed extensive chromosomal alterations, particularly chromosome 19 deletion, indicating genomic instability. Cell–cell communication analysis demonstrated active crosstalk between MP6 tumor cells and cancer-associated fibroblasts, including WNT and BMP signaling pathways, indicating that stromal crosstalk may contribute to the establishment of a pro-tumorigenic microenvironment. Transcription factor network inference further identified elevated activity of regulators such as TFCP2 and ELF1, implicating them in the acquisition of aggressive phenotypes. Immunohistochemical validation supported the clinical relevance of selected MP6-associated targets. Collectively, these findings define a clonally evolved epithelial subtype associated with lymphatic invasion and provide mechanistic insights into tumor cell plasticity and stromal–epithelial interactions underlying bladder cancer progression.
Yongxiang Luo, Xiaoping Liu, Sihua Zhu et al.· Frontiers in Cell and Develo...· 0 citations
Abstract Brain metastases (BrM) are a frequent and life‐threatening complication of solid tumours, with lung cancer representing their most common source. While cancer‐associated fibroblasts (CAFs) are well‐established contributors to tumour progression in many extracranial malignancies, their presence and function within the brain tumour microenvironment, where fibroblasts have long been considered scarce or absent, remain poorly understood. Here, we isolated and characterised fibroblast‐like cells from 13 human BrMs of diverse origins. These BrM‐associated CAFs (BrM‐CAFs) expressed canonical CAF markers and showed reduced proliferation and increased senescence compared to normal fibroblasts. Their transcriptome was enriched for extracellular matrix (ECM)‐related genes, including multiple collagens, fibronectin, and matrix‐remodelling enzymes. In vitro, BrM‐CAFs produced a fibrillar ECM, and in BrM tissues, their abundance was associated with collagen I and fibronectin deposition. Transcriptomic, proteomic, and secretome analyses further revealed that BrM‐CAFs produce multiple cytokines, chemokines, and growth factors that promote cell motility. BrM‐CAF conditioned medium promoted both monocyte migration and the migration of cancer cells, including established cell lines and patient‐derived lung cancer BrM cells; monocyte migration was partially reduced by inhibition of CCL2/CCR2 signalling, whereas blocking CXCL12, CXCL16, or CX3CL1 attenuated BrM‐CAF‐induced cancer cell migration. Beyond these effects on migration, exposure to BrM‐CAFs increased cancer cell invasion in transwell and heterotypic 3D spheroid assays. In contrast, their effects on cancer cell proliferation were limited and did not indicate a growth‐promoting role. Exposure to BrM‐CAFs was also associated with increased expression of interferon‐stimulated genes in cancer cells. Together, our findings support a role for BrM‐CAFs in shaping the brain metastatic microenvironment through ECM remodelling and the secretion of pro‐migratory mediators, promoting monocyte and cancer cell migration and enhancing cancer cell invasion. These data identify BrM‐CAFs as active stromal participants in BrM biology and support further investigation of their biological, diagnostic, and therapeutic relevance.
B. Vymolova, L. Pfeiferová, T. Smetana et al.· Brain Pathology· 0 citations
Bone metastases (BoMs) are a major clinical challenge across cancer types, yet mechanistic insights remain limited by small cohorts and insufficient profiling depth. Here, we present a single‐cell RNA sequencing atlas of 95 BoMs, 22 healthy bone marrows (hBMs) and 129 primary tumors (PTs) spanning 10 cancer types, comprising 895,475 high‐quality transcriptomes, to map cellular remodeling during bone metastatic colonization. Malignant cells in BoMs converged on chromosomal instability—high proliferative states with enhanced angiogenic programs and suppressed immune‐inflammatory and metabolic pathways. The BoM immune landscape featured reduced cytotoxic lymphoid populations and expanded exhausted T‐cell states, with pronounced cancer type‐specific heterogeneity. BoM‐resident myeloid cells showed marked suppression of antigen presentation and phagocytosis, indicating immunosuppressive reprogramming. Stromal remodeling was characterized by enrichment of immunosuppressive CAFs, depletion of antigen‐presenting fibroblasts, reduced mesenchymal MHC expression, and expanded angiogenic endothelial programs. Cell–cell communication analyses predicted strengthened CXCL12–CXCR4 stromal–immune signaling. In vitro, the CXCL12–CXCR4 axis recruited CD8
+
T cells and enhanced their adhesion, spatially sequestering them from the tumor parenchyma, while CXCR4 blockade or CXCL12 knockdown significantly reduced tumor‐cell migration and invasion in vitro. This atlas defines convergent hallmarks of bone metastasis and highlights shared stromal–immune dependencies as therapeutic vulnerabilities.
Yitong Pan, Zhou Yang, Junyuan Deng et al.· iMetaMed· 0 citations
Non‐small cell lung cancer (NSCLC) progression is strongly influenced by tumor‐associated macrophages (TAMs), which frequently acquire an M2‐like phenotype that promotes proliferation, epithelial‐mesenchymal transition (EMT), angiogenesis, stemness, and therapy resistance. Previous studies from our group established the direct cytostatic and macrophage‐polarizing effects of the synthetic coumarin derivative 4‐fluorophenylacetamide‐acetyl coumarin (4‐FPAC), including ROS‐mediated apoptosis, G0/G1 cell‐cycle arrest, suppression of EMT‐associated signaling in A549 cells, and promotion of M1‐like macrophage polarization. In the present study, we investigated whether 4‐FPAC‐modulated TAM‐derived conditioned media could suppress tumor‐promoting behavior in A549 NSCLC cells. THP‐1 monocytes were differentiated into macrophages and polarized using A549‐conditioned medium to generate TAM‐like macrophages. These TAMs were subsequently treated with 4‐FPAC, and conditioned media from control or treated TAMs were applied to A549 cells to evaluate macrophage‐associated anti‐tumor effects. MTT analysis further showed that direct exposure of A549 cells to 4‐FPAC under TAM‐CM‐supported conditions reduced cell viability in a dose‐dependent manner. Functional assays demonstrated that conditioned media derived from 4‐FPAC‐treated TAMs significantly reduced A549 proliferation, inhibited migration and invasion, and impaired angiogenic responses in a chick chorioallantoic membrane model. Flow cytometry revealed G0/G1 cell‐cycle arrest and reduced NANOG‐positive stem‐like cells, while DNA fragmentation and TUNEL assays confirmed enhanced apoptosis. Mechanistically, qRT‐PCR and immunoblotting demonstrated EMT reversal, suppression of AKT signaling, VEGFα, IL8, OCT3/4, and NANOG, and upregulation of p21, p53, and caspase‐3/cleaved caspase‐3. Collectively, these findings demonstrate that 4‐FPAC directly suppresses A549 viability under TAM‐CM‐supported conditions and modulates TAM‐derived conditioned‐media activity, highlighting its potential to target both tumor cells and tumor‐macrophage interactions within the NSCLC microenvironment.
Anjali Singh, Dhanush Danes, Suresh Balakrishnan· Journal of biochemical and m...· 0 citations
Introduction Cervical cancer is a common gynecological malignancy, having high incidence and mortality rates, especially in developing regions. The tumor microenvironment (TME) plays a crucial role in disease progression, and cancer‐associated fibroblasts (CAFs) are key components of the TME. However, the roles of CAFs in tumor‐adjacent normal tissue remain unclear. Methods Herein, we used single‐cell RNA sequencing to generate high‐resolution cellular atlases of cervical cancer and tumor‐adjacent normal tissue, systematically profiling fibroblast–cancer cell interactions and comparing fibroblasts from distinct origins. Pseudotime analysis and CytoTRACE scoring were employed to investigate the developmental trajectory of fibroblasts. Immunohistochemistry validated the inflammatory CAF (iCAF) subpopulation of C7 fibroblasts and assessed its correlations with clinicopathological features. Results We found that C7 fibroblasts secrete CXCL12 to activate the ACKR3 (CXCR7) receptor in cancer cells, thereby promoting tumor cell proliferation, invasion, and metastasis. C7 fibroblasts also highly expressed multiple oncogenic genes, including IL6, CXCL3, CXCL2, CCL2, GPC3, PLAU, TNFAIP6, PTX3, and EIF4A3, and genes associated with poor prognosis, including CXCL3, TNFAIP6, PTX3, IGSF10, and LDLR. High C7 fibroblast abundance was associated with advanced International Federation of Gynecology and Obstetrics stage, lymph node metastasis, and postmenopausal status. Discussion Our findings suggest that C7 fibroblasts being predominantly distributed in tumor‐adjacent normal tissues is significantly associated with cervical cancer progression, and that C7 fibroblasts in these tissues may serve as potential biomarkers and therapeutic targets for tumor microenvironment modulation.
Yi Liu, D. Yin, Lu-guang Wu et al.· Frontiers in Cell and Develo...· 0 citations
