Figure 4: TNBC metastasis shows increased IRAK1 expression/activity and gain of IRAK1 dependency.
From: IRAK1 is a therapeutic target that drives breast cancer metastasis and resistance to paclitaxel
(a) Scatter plots showing the IRAK1 mRNA levels in Schmidt Breast Cohort (GSE11121) composed of 200 breast cancer tumours of different clinical grades (Grade 1, n=29; Grade 2, n=136; Grade 3, n=35) (left), and IRAK1 IHC staining results in TMA-BR1505 with 150 breast tumour cores of different clinical grades (Grade 1, n=14; Grade 2, n=104; Grade 3, n=28) (right). (b) Representative IHC images of IRAK1 protein expression in matched normal, primary and metastatic tumours from two patients (left). Scale bars, 100 μm. Scatter plot showing IRAK1 protein expression in TMA-IMH364, including nine paired primary and matched metastatic tumours, and indicated by the black lines (right). (c) Phase-contrast microscopic images of MDA231 parental and MDA231-LM2 cells cultured in monolayer, mammospheres and 3D Matrigel conditions. Scale bars, 100 μm. (d) Western blotting showing the expression of indicated proteins. (e) Western blotting showing the expression of indicated proteins in the absence or presence of IL-1β (10 ng ml−1) for 24 h. (f) Quantitative PCR analysis showing the relative expression of indicated cytokines in cells treated in e. (g) Mammosphere formation assay of MDA231 and MDA231-LM2 cells expressing vector, shIRAK1, or together with ectopic IRAK1 for rescue. (h) Representative phase-contrast images in mammosphere and 3D Matrigel gel, treated with or without IRAK-inh (5μM) for 7 days. Scale bars, 100 μm. (i) Mammosphere growth of MDA231-LM2 cells treated with 5 μM IRAK-inh, with and without adding conditioned medium (CM) from mock-treated MDA231-LM2 cells. (j) ELISA quantification of indicated cytokine levels in i. (k) 3D Matrigel growth of MCF10A and HMLE cells expressing vector control or ecotopic IRAK1. Scale bars, 100 μm. (l) Mammosphere assay of cells in k. (m) Invasion assay of cells in k treated with recombinant IL-1b (10 ng ml−1) for 3 days. Error bars represent s.e.m. n=3. *P<0.05, **P<0.01. P-values were calculated with two-tailed t-test.
