---
title: "IL-8–positive CAFs drive chemotherapy‑induced tumor progression in breast cancer: multi‑omics inte"
id: "frontiers-in-immunology-1-integrating-multi-omics-data-reveals-il-8-positive-cancer-associated"
canonical_url: "https://medichelpline.com/clinical-feed/frontiers-in-immunology-1-integrating-multi-omics-data-reveals-il-8-positive-cancer-associated"
content_type: "clinical_feed_article"
specialty: "Oncology"
source_name: "Frontiers in Immunology"
source_url: "https://www.frontiersin.org/articles/10.3389/fimmu.2026.1878482"
published_at: "2026-07-20T00:00:00.000Z"
evidence_level: "Journal Feed"
license: "CC-BY-NC-4.0 / Informational Use"
---
# IL-8–positive CAFs drive chemotherapy‑induced tumor progression in breast cancer: multi‑omics inte
## Provenance & Clinical Metadata
- **Canonical URL:** https://medichelpline.com/clinical-feed/frontiers-in-immunology-1-integrating-multi-omics-data-reveals-il-8-positive-cancer-associated
- **Specialty:** [Oncology](https://medichelpline.com/clinical-feed/oncology.md)
- **Primary Source:** Frontiers in Immunology
- **Source URL:** [Original Journal Publication](https://www.frontiersin.org/articles/10.3389/fimmu.2026.1878482)
- **Published At:** 2026-07-20T00:00:00.000Z
- **Evidence Rating:** Journal Feed
## Executive GIST (TL;DR)
- Chemotherapy agents paclitaxel (PTX) and doxorubicin (DOX) can induce adaptive changes in the tumor microenvironment that promote tumor progression rather than only killing tumor cells. - Primary cancer‑associated fibroblasts (**CAFs**) exposed to PTX or DOX produced conditioned medium (CM) that increased proliferation, wound closure, invasion, and colony formation of MCF‑7 and MDA‑MB‑231 breast cancer cells compared with CM from untreated CAFs. - Bulk and single‑cell transcriptomic analyses (including GSE23399 and scRNA‑seq data GSE268662) and wet‑lab assays found that chemotherapy significantly **upregulated IL‑8** expression in CAFs in vitro and increased IL‑8 staining in tumor stroma after neoadjuvant chemotherapy (NAC) in patient samples; IL‑8 localized primarily to α‑SMA+ CAFs. - ELISA and qPCR confirmed dose‑ and time‑dependent increases of secreted IL‑8 from CAFs after PTX/DOX exposure; CAFs secreted substantially more IL‑8 than normal fibroblasts (NFs) or tumor cells. - Functional inhibition with reparixin, an IL‑8 receptor inhibitor, partly reversed the tumor‑promoting effects of chemotherapy‑exposed CAF CM, supporting a causal role for **CAF‑derived IL‑8** in promoting tumor cell migration, invasion, and chemoresistance. - Mendelian randomization (MR) using IL‑8 eQTL and circulating IL‑8 GWAS data suggested genetically predicted IL‑8 expression and plasma IL‑8 levels were significantly associated with breast cancer risk (methods: IVW, MR‑Egger, Cochran’s Q used for pleiotropy assessment). Exact MR effect estimates were reported in the source but are not restated here. - Deconvolution (BayesPrism) of TCGA‑BRCA and trait scoring (scPagwas) linked **IL‑8+ CAFs** to clinical prognosis and a potential role in breast cancer progression. - Mechanistically, CAF‑derived IL‑8 appears to activate the **NF‑κB** pathway in tumor cells, modulate apoptosis (increasing Bcl‑2/Bcl‑xL, decreasing Bax and TUNEL positivity), inhibit chemotherapy‑induced tumor cell apoptosis, and thereby promote chemoresistance. - Clinical correlations in patient samples showed high IL‑8 expression associated with chemoresistance and poorer survival (relapse‑free and distant metastasis‑free outcomes were analyzed using Kaplan‑Meier Plotter). - The study integrates in vitro experiments, patient tissues, single‑cell and bulk bioinformatics, MR, and mouse xenograft data to support targeting IL‑8 signaling in CAFs as a strategy to improve chemotherapy outcomes in breast cancer.
## Clinical Analysis & Structured Key Points
About us All journals All articles Submit your research Search Login Frontiers in Immunology Sections Articles Research Topics Editorial board About journal Published in Frontiers in Immunology Inflammation 7 impact factor 11.3 citescore Editor & Reviewers Edited by Zhi Chen Reviewed by Uzma Saqib Farnaz Sedghy Outline Abstract 1 Introduction 2 Materials and methods 3 Results 4 Discussion 5 Conclusion Data availability statement Ethics statement Author contributions Funding Acknowledgments Conflict of interest Generative AI statement Publisher’s note Supplementary material Abbreviations References Figures and Tables Figure 1 View in article Figure 2 View in article Figure 3 View in article Figure 4 View in article Figure 5 View in article Figure 6 View in article Figure 7 View in article Figure 8 View in article Figure 9 View in article Figure 10 View in article Figure 11 View in article Table 1 Heterogeneity and pleiotropy of individual SNPs in IL-8 eQTL. View in article Table 2 Heterogeneity and pleiotropy of individual SNPs in the inflammatory factor IL-8. View in article Table 3 IL-8 expression levels correlate with the clinicopathological characteristics of BRCA NAC patients. View in article ORIGINAL RESEARCH article Front. Immunol., 20 July 2026 Sec. Inflammation Volume 17 - 2026 | https://doi.org/10.3389/fimmu.2026.1878482 Integrating multi-omics data reveals IL-8 positive cancer-associated fibroblasts as mediators of chemotherapy-induced tumor progression in breast cancer H L Huifeng Liao 1,2† H L Huayan Li 1† J S Jin Song 3† J D Junhua Dong 2 Xue Bai 1,4* 1. Department of General Surgery, The First Medical Center of Chinese People’s Liberation Army General Hospital, Beijing, China 2. The Second School of Clinical Medicine, Southern Medical University, Guangzhou, China See more Article metrics View details 75 Views Abstract Background: Recent studies have shown that while chemotherapy kills tumor cells, it may also induce adaptive changes in cells within the tumor microenvironment, particularly cancer-associated fibroblasts (CAFs), which could paradoxically promote tumor progression. This study aimed to investigate the role of CAFs exposed to paclitaxel (PTX) or doxorubicin (DOX) in tumor progression and explore the underlying mechanisms. Methods: We examined the effects of PTX- or DOX-exposed CAFs on tumor migration and invasion through in vitro experiments. Bioinformatics analyses including Mendelian randomization, BayesPrism, and scPagwas were performed to elucidate the clinical relevance of IL-8 + CAFs in breast cancer. Additionally, in vitro experiments were conducted to confirm the tumor-promoting effects of CAFs-derived IL-8 and explore the underlying molecular mechanisms. Results: Our findings showed that CAFs exposed to PTX or DOX further promoted tumor migration and invasion compared to untreated CAFs. Notably, PTX and DOX significantly increased IL-8 expression levels in CAFs, suggesting that IL-8 may be a key cytokine mediating the tumor-promoting effect of CAFs exposed to chemotherapeutic agents. Interestingly, this tumor-promoting effect could be partly reversed by reparixin, an IL-8 receptor inhibitor. Mendelian randomization analysis confirmed that both IL-8 eQTL and plasma IL-8 levels were significantly associated with breast cancer risk. Further insights from BayesPrism and scPagwas indicated that IL-8 + CAFs were associated with clinical prognosis and may play a critical role in breast cancer progression. Furthermore, we confirmed the tumor-promoting effect of CAFs-derived IL-8. Mechanistically, IL-8 may modulate apoptosis by activating the NF-κB pathway, thereby inhibiting tumor cell apoptosis and leading to chemoresistance. Clinically, high IL-8 expression correlated with chemoresistance and poor survival in breast cancer patients. Conclusion: Our study suggests that the upregulation of IL-8 in CAFs following PTX or DOX treatment may contribute to chemotherapy-mediated tumor progression. These findings provide potential avenues for improving chemotherapy outcomes. 1 Introduction Chemotherapy plays an instrumental role in the pre-operative, post-operative, and palliative treatment of breast cancer (BRCA), but the recurrence and metastasis caused by chemoresistance remain major obstacles preventing patients from achieving a durable remission and eventual cure. Most of the previous studies on chemoresistance and metastasis have focused on the genetic alterations that occur within tumor cells, but in recent years, chemotherapy-induced chemoresistance and metastasis have gradually emerged as a new research hotspot (1–4). Despite the effectiveness of chemotherapy as a first-line cancer treatment, accumulating evidence from animal models indicates that while chemotherapy exerts antitumor effects, it also promotes several off-target effects that induce tumor metastasis (5–7). Usually, chemotherapy aims to destroy tumor cells alone, but the systemic administration of cytotoxic drugs inevitably affects non-tumor cells, thus promoting the host response (8). The host response to chemotherapy is a result of the systemic release of multiple cytokines and the mobilization of various host cells, including macrophages, endothelial cells, and fibroblasts, which influence the response to therapeutics and the overall disease outcome to some extent (9–13). As cancer research continues to advance, more and more studies have reported that chemotherapy promotes cellular and molecular responses in host cells which promote chemoresistance and tumor progression. Generally, the undesired side effects of chemotherapy on the tumor microenvironment (TME) are the major factors that induce metastasis (14, 15). Indeed, there is a dynamic interaction between tumor cells and non-tumor cells in the TME during chemotherapy, which allows tumor cells to escape from the primary tumor site to the peripheral blood, thereby promoting chemoresistance and metastasis (16–18). For example, DNA damage caused by DOX induces thymic endothelial cells to release IL-6 and tissue inhibitor of metallopeptidase 1, which protects tumor cells from the genotoxic effects of chemotherapy (19). PTX induces the infiltration of tumor-associated macrophages into the tumor, which then release cathepsin proteases, thereby preventing tumor cells from chemotherapy-induced cell death (20). Similarly, platinum drugs induce the activation of bone marrow mesenchymal stem cells, causing them to secrete unique fatty acids that lead to the development of chemoresistance (21). In conclusion, chemotherapy induces various cells in the TME to secrete cytokines through different pathways, which protects the tumor cells from the cytotoxic effects of chemotherapeutic agents and then promotes tumor progression. CAFs are one of the most important cell subpopulations in the TME, however, only a few studies have focused on the effects of CAFs exposed to chemotherapeutic agents on tumor cells. Therefore, it is clinically relevant to reveal the potential mechanism of CAFs in chemotherapy-mediated tumor progression. Neoadjuvant chemotherapy (NAC) provides long-term benefits for the clinical treatment of BRCA patients. However, in addition to direct effects on tumor cells, the administration of cytotoxic drugs alters the phenotype and characteristics of non-tumor cells, especially CAFs, thus promoting therapy resistance and metastasis. Our study highlights the important function of CAFs in chemotherapy-induced progression and proposes that chemotherapy combined with targeted IL-8 may be an effective treatment strategy for inhibiting tumor progression. 2 Materials and methods 2.1 Human serum and tissue samples We collected serum samples from 50 patients during NAC treatment, and tumor tissue from 100 patients pre- and post-NAC. All patients were diagnosed with primary invasive ductal carcinoma of the breast by core needle biopsy. Inclusion criteria were: (1) histologically confirmed primary BRCA; (2) received PTX− or DOX−based NAC; (3) available pre− and post−treatment tissue samples. Exclusion criteria were: (1) prior history of other malignancies; (2) receipt of radiotherapy or targeted therapy before NAC; (3) distant metastasis at initial diagnosis. Hormone receptor (ER/PR) and HER-2 status were determined by IHC. Molecular subtypes were classified according to the St. Gallen criteria. The sensitivity of patients to chemotherapy was determined according to the Response Evaluation Criteria in Solid Tumors (RECIST) guidelines. 2.2 Primary fibroblast isolation and culture Primary CAFs were isolated from the tumor tissue, while normal fibroblasts (NFs) were isolated from normal breast tissue. The fresh tissue samples were minced into pieces, centrifuged to remove the supernatant, and were added to the digestion solution (0.01 mg/ml collagenase II + 0.01 mg/ml hyaluronidase) for 15 min. The homogenate was collected and passed through a 100 μm sieve and centrifuged (1500 rpm, 5 min) to remove the supernatant. The cells were then resuspended in DMEM/F12 media and placed in an incubator (37 °C, 5% CO2). We seeded 1×106 of CAFs in a 6-well plate and allowed them to grow up to 80%. The conditioned medium (CM) was collected from serum-free media after culturing for 24 or 48 h and was stored at -80 °C for further use after centrifugation. 2.3 Cell culture and transfection MCF-7 cells were cultured in DMEM medium with 10% fetal bovine serum (FBS), and MDA-MB-231 cells (Boster, Wuhan, China) were cultured in L-15 media with 10% FBS. The cell lines were authenticated through STR analyses and mycoplasma tests were performed each month through qPCR. Patient-derived tumor-like cell clusters (PTCs) were isolated from BRCA tissues and cultured in a low-attachment-surface petri-dish with advanced DMEM (22). According to the manufacturer’s protocol, CAFs were transfected with the IL-8-pcDNA3.1 (Sangon, Shanghai, China) using Lipofectamine 3000 (Invitrogen, USA). Tumor cells were transfected with Bcl-2 or Bcl-xL siRNA (Genepharma, China) using Lipofectamine 3000 (Invitrogen, USA). The transfection efficiency was shown in Supplementary Figures 2A, B. 2.4 Data acquisition and processing To investigate the causal association of IL-8 with BRCA risk, we performed Mendelian randomization (MR) analysis. The exposure data for genetically predicted IL-8 expression levels (eQTL) were obtained from the GWAS database (ukb-a-213), and the circulating IL-8 protein levels were sourced from the GWAS Catalog (GCST90274817). The survival information for BRCA was obtained from the TCGA database. The GSE23399 and GSE268662 datasets were downloaded from the GEO database. GSE23399 was used to study the expression of IL-8 in CAFs before and after chemotherapy. Single-cell RNA-sequencing data from 5 BRCA patients (GSE268662) were analyzed using the Seurat package (v 4.3) to comprehensively characterize the CAFs signature. Quality control was performed to filter out low-quality cells. Specifically, cells expressing fewer than 200 genes or more than 10,000 genes were excluded. Additionally, cells with a mitochondrial gene percentage greater than 20% were removed to eliminate dying cells. After filtering, a total of 14,825 cells and 25,603 genes were retained for subsequent normalization and dimensionality reduction. CAFs were annotated with 4 marker genes, including Vimentin, FAP, PDGFRα, and PDGFRβ. 2.5 BayesPrism deconvolution analysis and scPagwas analysis BayesPrism (23) was employed to compute the score for each cell subpopulation by adding the deconvolution scores integrals of each cell state for each subpopulation. Utilizing BayesPrism, we performed deconvolution on the bulk RNA-seq data from TCGA-BRCA to investigate the correlation between IL-8 + CAFs and clinical outcomes. Based on the annotations of different cell types from scRNA-seq, scPagwas (24) was used to calculate the Trait Related Score (TRS) for each cell type to evaluate their potential role in BRCA. 2.6 Cell viability assay The Cell Counting Kit-8 (CCK-8) assay was used to measure the viability of tumor cells. The tumor cells were seeded in a 96-well culture plate and co-cultured with CAFs CM for 48 hours. For the drug sensitivity assay, the tumor cells were treated with PTX or DOX for 48 hours after co-culture with CAFs CM for 48 hours. 10 μL of CCK-8 solution (Boster, Wuhan, China) was added to each well for 1 hour and then the absorbance was measured at 450 nm with a microplate reader. The area of ​​all PTCs clusters in the wells was measured to assess the drug effect. The clusters were photographed on days 0 and 7, and only those with diameters greater than 40 μm at both time points were selected for area calculation. The formula for calculating PTCs cell viability after chemotherapy was performed as previously described (22). 2.7 Wound healing, invasion, and colony formation assays Wound healing, invasion, and colony formation assays were performed as previously described (25). 2.8 Enzyme-linked immunosorbent assay and quantitative real-time PCR The IL-8 concentration in human serum samples and CAFs CM was quantified by the ELISA kit (Boster, Wuhan, China). The mRNA levels of IL-8 were quantified by qRT-PCR. 2.9 TUNEL formation assay The TUNEL Apoptosis Assay Kit (Beyotime, Shanghai, China) was used to detect apoptosis in the tumor cells and xenograft tissues. Five areas were randomly selected under the fluorescence microscope and the percentage of apoptotic cells was calculated. 2.10 Western blotting Cells were lysed with RIPA buffer. Following SDS-PAGE, transfer, and blocking, the polyvinylidene difluoride membranes were incubated with the respective primary antibodies overnight at 4 °C. The membranes were washed and incubated with the secondary antibodies. Enhanced chemiluminescence reagent was used to detect the protein bands. Protein levels were normalized to GAPDH levels. 2.11 Immunohistochemistry Immunohistochemistry (IHC) staining was performed to detect the expression of IL-8, α-smooth muscle actin (α-SMA), Bcl-2, Bcl-xL, and Bax in human and mouse tumor tissues, according to standard protocols. The IL-8, α-SMA, Bcl-2, and Bcl-xL were purchased from CST, USA. The Bax and p-p65 were purchased from Beyotime, Shanghai, China. For IL-8 staining evaluation, cells with more than 5% positivity were considered positive, while those with less than 5% were considered negative. For evaluation of α-SMA, Bcl-2, Bcl-xL, and Bax staining, the percentage of positive cells was calculated as follows: 75% = 4. There were four levels of staining intensity: 0, negative; 1, weak; 2, medium; and 3, strong. Multiplying the two scores resulted in the following combined score: 0, negative; 1-4, weak expression; 5-8, moderate expression; and 9-12, strong expression. 2.12 H&E staining The paraffin sections were baked at 60 °C until the wax melted, and washed with xylene, ethanol, and running water for dewaxing. Hematoxylin staining was performed for 10 min, followed by differentiation for 20 s, blueing for 5 min, and eosin staining for 3 min, with running water washes between steps. Finally, the sections were dehydrated and sealed with neutral gum seal. 2.13 Animal experiments 16 four-week-old female NSG nude mice were purchased from the Medical Discovery Leader (MDL, Beijing, China). They were randomly divided into four groups and injected with tumor cells (1 × 107) mixed with Matrigel subcutaneously into the left armpit. Compared with the control group, the experimental group was injected with IL-8 (100 ng/ml) continuously for 5 days, beginning on the 12th day. Tumor diameter was measured every 3 days and tumor volume (mm3) was estimated by volume = (shortest diameter) (mm)2 × (longest diameter) (mm) × 0.5. 2.14 Kaplan-Meier plotter database Kaplan-Meier Plotter was used for survival analysis including relapse-free survival (RFS) and distant metastasis-free survival (DMFS) in BRCA patients (26). “Auto select best cutoff” and “only JetSet best probe set” were chosen in the analysis. 2.15 Statistical analysis All continuous variables were subjected to normality testing using the Shapiro-Wilk (n 0.05, LSD post-hoc test was used for pairwise comparisons, and when P ≤ 0.05, Dunnett’s T3 post-hoc test was applied. Statistical analyses were performed using SPSS 24.0 software. For single-cell differential expression analysis, P values were adjusted using the Benjamini-Hochberg method to control the False Discovery Rate (FDR). A threshold of FDR < 0.05 was considered statistically significant. In MR analysis, the IVW method was used for causal estimation, supported by MR-Egger and Cochran’s Q tests to assess pleiotropy. 3 Results 3.1 Chemotherapy-pretreated CAFs exhibit superior pro-tumorigenic capabilities compared to CAFs CAFs in the TME may undergo adaptive responses during chemotherapy, which subsequently lead to tumor progression. To investigate the effect of chemotherapy-pretreated CAFs on tumor cells, we exposed CAFs to PTX (0.01 μM) or DOX (0.01 μM) for 24 hours, then replaced the medium with serum-free culture medium for 24 hours before collecting the CM. As expected, CM from CAFs promoted the proliferation, wound healing, and invasion of MCF-7 and MDA-MB-231 cells. It was worth noting that CAFs treated with PTX or DOX further promoted the proliferation, wound healing, and invasion of tumor cells compared to CAFs (Figures 1A–D). Figure 1 Effects of CAFs exposed to chemotherapeutic agents on tumor cells. (A-D) By using CCK-8 assay (A, B), wound healing assay (C), and invasion assay (D), we detected the proliferation, wound healing, and invasion abilities of MCF-7 and MDA-MB-231 cells that co-culture with CM from CAFs or CAFs treated with chemotherapeutic agents. The data were expressed as mean ± SD for at least triplicate experiments. *P < 0.05, **P < 0.01, ***P < 0.001. 3.2 Chemotherapy promoted IL-8 expression in vitro and in vivo To analyze the underlying mechanisms of this phenomenon, we downloaded the GSE23399 dataset from the GEO database. GSE23399 was used to analyze the gene expression changes of CAFs following PTX or DOX treatment. Interestingly, both PTX and DOX treatment upregulated IL-8 expression in CAFs (Figures 2A, B). We then used qPCR and ELISA assay to examine the expression of IL-8 in CAFs before and after chemotherapy. We found that PTX and DOX significantly promoted IL-8 expression in CAFs, and IL-8 levels in the CM rose in a dose- and time-dependent manner (Figures 2C, D). To further verify whether chemotherapy promoted IL-8 expression in vivo, we performed IHC on samples from BRCA patients who received PTX- or DOX-based NAC. IHC staining showed that IL-8 expression level was significantly upregulated in breast tumor stroma post-NAC in comparison to the pre-NAC samples. Moreover, IL-8 was mostly located in the CAFs expressing α-SMA (Figures 2E, F). The above data indicated that chemotherapy promoted IL-8 expression from the CAFs both in vitro and in vivo. Additionally, IL-8 levels in CAFs were significantly higher than in tumor cells and NFs, and after co-culturing with tumor cells, the expression of IL-8 in CAFs CM was significantly increased (Figure 2G). Interestingly, chemotherapy also promoted IL-8 expression in NFs, but the IL-8 concentration in supernatant from NFs was considerably lower than that from CAFs (Figure 2H). Figure 2 PTX and DOX promoted IL-8 expression in CAFs in vitro and in vivo. (A, B) The IL-8
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