Metastatic dissemination is the principal cause of mortality in cervical cancer (CC). The matricellular protein connective tissue growth factor (CTGF/CCN2) modulates cell–extracellular matrix interactions, but its specific role in CC invasion and metastasis had not been clearly defined. This study aimed to investigate whether CTGF contributes to an invasive phenotype in CC using 3D spheroid models and to determine CTGF prevalence in clinical cervical lesion specimens, including metastatic tissue.
The investigators used two cervical cancer cell lines, C33A and HT3, grown as three-dimensional spheroids to better approximate tumor cell–matrix interactions than two-dimensional culture. Spheroids were treated either with recombinant human CTGF (rhCTGF) to augment CTGF signaling or with a function-blocking anti-CTGF antibody (IgG CTGF) to inhibit CTGF activity. These interventions were designed to evaluate whether exogenous CTGF or CTGF blockade modifies invasive behavior in a 3D context.
Invasive growth from spheroids was quantified using a 3D spheroid invasion assay combined with imaging on a Celigo imaging cytometer. The assay measured progressive invasion into the surrounding matrix following treatment conditions. The abstract reports a statistically significant reduction in spheroid invasive growth after CTGF blockade in both cell lines (p < 0.0001), indicating a measurable functional effect on invasive behavior when CTGF is inhibited.
To explore cellular correlates of the invasive phenotype, immunofluorescence analysis was performed for cancer stem cell markers (CD133, CD44) and epithelial–mesenchymal transition (EMT) markers (E-cadherin, N-cadherin). Modulation of CTGF altered the spatial distribution of these proteins within spheroids: treatment with rhCTGF induced surface clustering of CD133 and peripheral enrichment of N-cadherin, features consistent with enhanced invasive potential. In contrast, CTGF blockade was associated with apparent nuclear or perinuclear enrichment of CD133 and E-cadherin, accompanied by a reduced N-cadherin signal.
The clinical component used a tissue microarray containing 69 cervical lesion cases represented in triplicate. Cores included pre-invasive lesions, invasive lesions staged FIGO I through III, and metastatic cervical lesions. CTGF expression in the tissue cores was quantified by immunofluorescence scoring, producing an immunoreactivity score (IRS). The abstract reports categorical results for CTGF intensity and the frequency of high expression (IRS ≥ 6) across FIGO stages and metastatic samples.
Functionally, blocking CTGF produced a significant decrease in 3D spheroid invasive growth for both C33A and HT3 models (p < 0.0001). Spatial reorganization of stemness and EMT markers accompanied CTGF modulation: rhCTGF promoted CD133 surface clustering and peripheral N-cadherin, while CTGF blockade shifted CD133 and E-cadherin to nuclear/perinuclear regions and diminished N-cadherin signal.
In the tissue microarray, metastatic samples exhibited the highest CTGF fluorescence intensity. When stratified by FIGO stage, a greater proportion of FIGO I cases demonstrated high CTGF expression (IRS ≥ 6; 35.5%) than FIGO III cases (10.0%). These tissue data suggest a complex relationship between CTGF expression, tumor stage, and metastatic status.
The authors report that higher CTGF mRNA expression was associated with significantly reduced recurrence-free survival, as shown by Kaplan–Meier analysis (log-rank p = 0.0032). This finding supports a potential prognostic role for CTGF in cervical cancer outcomes, specifically recurrence risk.
Based on the 3D in vitro data and tissue analyses, the authors conclude that CTGF appears to regulate an invasive phenotype in cervical cancer spheroids, likely by influencing aberrant subcellular localization of stemness and EMT markers. The apparently elevated CTGF expression in some early-stage carcinomas (FIGO I) and in metastatic lesions, together with its association with poorer recurrence-free survival, led the authors to propose CTGF as a candidate early prognostic biomarker that may be involved in initiating metastatic potential.
The abstract provides core findings but omits several details that would inform interpretation and reproducibility. Specifics not reported in the abstract include full quantitative tissue scoring distributions beyond the percentages given for high IRS, breakdown of case numbers by lesion category, detailed imaging quantification methods and thresholds, exact concentrations and exposure times for rhCTGF or IgG CTGF, and mechanistic signaling pathways linking CTGF to marker relocalization. These details would be required from the full text to assess experimental rigor and to plan follow-up validation studies.
Overall, the study supplies evidence from complementary 3D models and a clinical tissue series that CTGF/CCN2 influences invasive behavior in cervical cancer and associates with metastatic tissue and worse recurrence-free survival, supporting further investigation of CTGF as a prognostic biomarker and potential therapeutic target.