Figure 4

Decreased SNAI1, SEMA4A, VEGFR3 and PLXNA1 PBMC gene expression in patients with large tumor size and advanced disease stage. (a) The mean relative Log₁₀ SNAI1 expression ( ± SEM), as quantified by real time PCR, is significantly downregulated in PBMCs from breast cancer cases with tumor size ≥ 2 cm (T2, T3 and T4) compared to <2 cm (T1) (median value is represented by black line inside boxplot). Similarly, (b) Log₁₀ SEMA4A, (c) Log₁₀ VEGFR3 and (d) Log₁₀ PLXNA1 mean relative gene expression decreases as the tumor size increases. (e) Mean relative Log₁₀ SNAI1 expression and (f) Log₁₀ SEMA4A (±SEM) is significantly downregulated in advanced disease stages 2, 3 and 4 compared to Stage 1. (g) ROC analysis confirms that the expression of Log₁₀ SNAI1 in PBMCs differentiates between Stage 1 cases and all other advanced breast cancer stages (Stage 2–4) (AUC = 0.870 ± 0.056, p = 0.003, 95% CI: 0.760–0.979). Similarly, mean relative gene expression of (h) Log₁₀ PLXNA1 and (i) Log₁₀ VEGFR3 decreases in advancing disease with lowest expression in Stage 4 compared to the other stages. Target gene expression levels normalized against GUSB gene expression and scaled to expression levels in healthy controls. p < 0.05 considered to indicate statistical significance. *p < 0.05, **p < 0.01, ***p < 0.001, Multivariate ANOVA followed by Fisher’s LSD post hoc test.