Biophysical and computational characterization identifies FSTL1 as a novel binding target of doxorubicin.
FSTL1 may be a relevant biomarker for doxorubicin response, but its clinical utility remains uncertain given the modest discriminative ability and reliance on preclinical models.
Where it sits
this study against the rest of the follistatin-344 corpusSummary and findings
This study investigated the binding landscape of doxorubicin (DOX) with a focus on follistatin-like 1 (FSTL1) in various cell lines. FSTL1 expression was suppressed in resistant cell lines and restored after DOX treatment in sensitive counterparts. The study also analyzed FSTL1 expression in tumor tissues and its association with overall survival.
Abstract
Mapping the protein-binding landscape of doxorubicin (DOX) is pivotal for optimizing therapeutic efficacy and overcoming resistance. In this study, we integrated data from prior protein microarray screenings that identified 363 DOX-binding proteins (with a signal-to-noise ratio (SNR) > 4) along with 224 Homo sapiens expression profiling series obtained from the Gene Expression Omnibus (GEO). Cross-analysis indicated that follistatin-like 1 (FSTL1) consistently demonstrated bidirectional expression across three GEO series (GSE202536, GSE13477, GSE155478), indicating sensitivity to DOX. Specifically, FSTL1 expression was suppressed in resistant cell lines (CAL-51/DOX, MCF-7/DOX) and restored after DOX treatment in their sensitive parental counterparts. Protein microarray analysis confirmed a strong interaction (SNR = 5.763), which was further corroborated by surface plasmon resonance (SPR) assays, indicating micromolar affinity. AlphaFold3 modeling combined with docking and 400 ns molecular dynamics simulations demonstrated that hydrophobic interactions and hydrogen bonding stabilize the FSTL1-DOX complex. TCGA-BRCA analysis revealed decreased FSTL1 expression in tumor tissues, modest diagnostic discrimination between tumor and normal samples (AUC = 0.76), and improved overall survival among patients with elevated FSTL1 expression. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses notably associated FSTL1 with extracellular matrix organization and the PI3K-Akt signaling pathway, supporting its potential relevance as a biomarker candidate and mechanistic hypothesis for anthracycline responsiveness.
Background
This paper addresses the binding interactions of doxorubicin (DOX) with proteins, specifically focusing on follistatin-like 1 (FSTL1). Prior studies have identified various DOX-binding proteins, but the role of FSTL1 in drug resistance and sensitivity remains less understood. Understanding these interactions may help optimize therapeutic efficacy and address resistance mechanisms in cancer treatment.
Methods
The study utilized prior protein microarray screenings that identified 363 DOX-binding proteins and analyzed 224 Homo sapiens expression profiling series from the Gene Expression Omnibus (GEO). Cross-analysis was performed to assess FSTL1 expression across different cell lines. Protein microarray analysis and surface plasmon resonance (SPR) assays were conducted to confirm binding interactions.
Results
FSTL1 expression was suppressed in resistant cell lines (CAL-51/DOX, MCF-7/DOX) and restored after DOX treatment in sensitive parental counterparts. The protein microarray analysis indicated a strong interaction with a signal-to-noise ratio (SNR) of 5.763. TCGA-BRCA analysis showed decreased FSTL1 expression in tumor tissues, with an area under the curve (AUC) of 0.76 for diagnostic discrimination between tumor and normal samples.
Interpretation
The findings suggest that FSTL1 may play a role in the response to DOX, aligning with previous literature that highlights the importance of protein interactions in drug efficacy. However, the clinical significance of the observed effect sizes, particularly the modest AUC, raises questions about the robustness of FSTL1 as a biomarker. Limitations include reliance on cell line models and the absence of direct clinical outcomes.
Key findings
- FSTL1 expression was suppressed in resistant cell lines (CAL-51/DOX, MCF-7/DOX) and restored after DOX treatment in sensitive parental counterparts.
- Protein microarray analysis confirmed a strong interaction (SNR = 5.763).
- TCGA-BRCA analysis revealed decreased FSTL1 expression in tumor tissues.
- Modest diagnostic discrimination between tumor and normal samples (AUC = 0.76).
- Improved overall survival among patients with elevated FSTL1 expression.
Limitations
- Not reported in abstract.
- Reliance on cell line models for resistance studies.
- Modest AUC of 0.76 for diagnostic discrimination.
- Lack of direct clinical outcome data.