Figure 6. RASAL2 expression is lost/low in primary human breast cancers and low levels are associated with metastasis and recurrence.

(A) RASAL2 dot blot of whole cell RIPA extracts from human breast cancer cell lines with high or low RASAL2 expression (MDA-MB-231 “231” and T47D, respectively) (top) or MCF10ADCIS cells infected with control or RASAL2-targeting shRNAs (bottom).

(B) Dot blot images from human breast tumor lysate array. Six sets of RASAL2 and total protein stains are shown. Each set contains triplicate spots of tumor lysate (right) and triplicate spots of paired normal tissue lysate (left).

(C) Quantification of RASAL2 expression in tumor lysate arrays. Each bar depicts the change in RASAL2 expression in one sample as compared to its matched normal control as described in Experimental Procedures. Shaded bars indicate metastatic samples.

(D) RASAL2 protein expression in tumor versus normal in non-metastatic (Stages I, II, III) versus metastatic (Stage IV) tumors. Graph shows the Log2 fold change in RASAL2 protein expression in tumor versus normal. Data are reported as average ± 95% CI. p=0.006.

(E) Heatmap of RASAL2 gene expression as a function of robust molecular subtype predictor classification, which is based on patients classified in the same tumor subtype. Percentages of tumors with high, intermediate, and low RASAL2 expression per molecular subtype are given in the gene expression table.

(F) RASAL2 expression table. For each breast cancer subtype, the number of samples and percentage of samples with low, intermediate, or high RASAL2 mRNA expression are indicated.

(G) Kaplan-Meier curve showing recurrence-free survival of luminal B tumors with high or low RASAL2 expression (logrank p = 0.013).

(H) Kaplan-Meier curve showing overall survival of luminal B tumors with high or low RASAL2 expression (logrank p = 0.013).