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Updated: May 29, 2026
Three-Dimensional Bone Extracellular Matrix Model for Osteosarcoma
Published on: April 12, 2019
Integrative single-cell and bulk analyses reveal ATF4-associated immune suppression and WNT/β-catenin signaling in
Xiaomin Ding1, Feihu Chen2, Jin Zhou1
1Orthopaedics Department, Nantong First People's Hospital, Nantong, Jiangsu, China.
Background:
Osteosarcoma (OS) is highly heterogeneous and often exhibits an immunosuppressive tumor microenvironment, contributing to poor outcomes. The cell-type-resolved role of the ATF/CREB stress-response network, particularly ATF4, remains unclear in OS.
Methods:
We integrated public OS single-cell RNA-seq and bulk transcriptome cohorts. Single-cell analyses included malignant program discovery (cNMF), regulon inference (SCENIC), and cell-cell communication (CellChat). Bulk cohorts (TARGET and GEO) were used to build and validate a prognostic model. ATF4-associated immune features were assessed using ESTIMATE, immune deconvolution, and TIDE-related metrics. WNT/β-catenin activity was evaluated by gene-set scoring. ATF4-related drugs were screened using DSigDB/Enrichr and assessed by docking. Sorafenib was selected for molecular dynamics simulation. ATF4 function was validated by siRNA knockdown (MG63, U2OS) and overexpression (HOS).
Results:
A stress-associated malignant program showed enriched ATF/CREB activity and prominent signaling interactions. Intersecting its markers with ATF/CREB genes identified ATF3/ATF4/CREB5, which were used to construct an ATF/CREB-associated risk score (ACS) that stratified survival in TARGET and external cohorts. ATF4 showed the most consistent adverse prognostic association and remained an independent factor in multivariate analyses. ATF4-high tumors were associated with a more immunosuppressive profile and stronger WNT/β-catenin signaling, and these features tracked with the ACS. Among the compounds tested in silico, sorafenib ranked highest in docking, and subsequent molecular dynamics simulations suggested that the ATF4-sorafenib interaction can remain stable. In cell-based assays, silencing ATF4 curtailed proliferation, colony formation, and migration, accompanied by reduced β-catenin, CyclinD1, and c-MYC. By contrast, ATF4 overexpression led to the opposite pattern.
Conclusion:
Our results place ATF4 at the intersection of stress-related tumor programs, immune suppression, and WNT/β-catenin activation in OS, and this pattern is associated with poorer clinical outcomes. The ACS offers a practical way to stratify risk, and sorafenib warrants further evaluation as a potential ATF4-oriented therapeutic lead.
Insights
The study reveals that ATF4 is a key factor in osteosarcoma (OS) progression, linking stress responses to immune suppression and WNT/β-catenin signaling. An ATF/CREB-associated risk score (ACS) can predict patient survival, and sorafenib shows potential as an ATF4-targeted therapy.
Area of Science:
- Oncology
- Molecular Biology
- Immunology
Background:
- Osteosarcoma (OS) is characterized by heterogeneity and an immunosuppressive tumor microenvironment, leading to poor patient outcomes.
- The specific role of the ATF/CREB stress-response network, particularly ATF4, in OS remains incompletely understood.
Purpose of the Study:
- To elucidate the role of the ATF/CREB network in osteosarcoma.
- To develop a prognostic model for OS based on ATF/CREB activity.
- To identify potential therapeutic targets and strategies for OS.
Main Methods:
- Integrated analysis of public OS single-cell RNA-seq and bulk transcriptome data.
- Malignant program discovery, regulon inference, and cell-cell communication analysis.
- Development and validation of an ATF/CREB-associated risk score (ACS) using bulk cohorts.
- In silico drug screening and molecular dynamics simulation for ATF4-targeting agents.
- Experimental validation of ATF4 function via siRNA knockdown and overexpression.
Main Results:
- A stress-associated malignant program with enriched ATF/CREB activity was identified.
- The developed ACS effectively stratified patient survival in independent cohorts.
- ATF4 expression was consistently associated with adverse prognosis, immunosuppression, and WNT/β-catenin signaling.
- Sorafenib demonstrated potential as an ATF4 inhibitor through in silico and molecular dynamics simulations.
- Experimental validation confirmed ATF4's role in promoting OS cell proliferation, colony formation, and migration.
Conclusions:
- ATF4 integrates stress-related programs, immune suppression, and WNT/β-catenin activation in osteosarcoma, correlating with poor clinical outcomes.
- The ACS provides a practical tool for risk stratification in OS patients.
- Sorafenib is a promising therapeutic lead for ATF4-targeted treatment strategies in OS.
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