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. 2020 Jul 16;12(7):1918. doi: 10.3390/cancers12071918

Figure 2.

Figure 2

Knockdown of MALAT1 increases HER2-cells sensitivity of trastuzumab. (A). SKBR3/100-8 and BT474/100-2 cells were treated with siRNA MALAT1 or negative sequences (mock) for 72 h as described in Methods, and RNA extracted. The bar graphs indicate the relative levels of MALAT1 (mean ± SEM) as determined by RT-qPCR from four repeated experiments and show MALAT1 knockdown cells have downregulated MALAT1, * p < 0.05, and ** p < 0.01 compared to mock cell lines. The ANOVA test determined the significance; (B). SKBR3/100-8 and BT474/100-2 treated with siRNA MALAT1 or negative sequences (mock) for 24 h, were co-treated with trastuzumab at the indicated doses either with siRNA MALAT1 or negative sequences for an additional 48 h. MTT assay determined cell viability. Each data point was from six measurements, and the experiments were independently performed four times. Red color indicates SKBR3/100-8 and blue color indicates BT474. The graph shows the mean ± SEM from four repeated tests. A statistically significant change in cell viability was observed between the siRNA-treated SKBR3/100-8 (red dotted line) and BT474/100-2 (blue dotted line). siRNA treatment decreased cell viability in the trastuzumab-resistant HER2+, * p < 0.05, and ** p < 0.01 compared to their untreated cells, respectively. The ANOVA test determined the significance; (C,D). SKBR3/100-8 and BT474/1002 cells treated with siRNA MALAT1 or negative sequence (mock) for 24 h, were co-treated with or without trastuzumab at 20µg/mL for an additional 48 h. The invaded cells were measured by the Boyden Chamber Invasion assay, as described in the Methods section. The mean invaded cells were counted from five different areas first for each time of the experiment. The bar in the figure showed the mean ± SEM from four independent experiments. The number of invading cells was reduced significantly upon combination treatment of siRNA and trastuzumab in the resistant cells, * p < 0.05, and ** p < 0.01 compared to their respective untreated cells. The ANOVA test determined the significance.