Ligand dependent and independent roles of interleukin-13 receptor alpha 2 in breast cancer brain metastasis
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2020-02
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Cancer Research, 1538-7445, Vol 80, Sup. 4, 2020
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American Association for Cancer Research
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Abstract
Introduction: Interleukin-13 receptor alpha 2 (IL13Rα2) is a high affinity receptor for IL-13, best known as a decoy receptor that lacks intracellular kinase activity. Yet, IL13Rα2 has been shown to play an important role in the pathobiology of glioblastomas, colon, ovarian and pancreatic cancer, through the recruitment and activation of FAK, SRC and other intracellular signaling pathways. In breast cancer, the function of IL13Rα2 remains controversial. Increased IL13Rα2 expression is associated with poor prognosis for high grade tumors but not low-grade tumors, while was also reported to be associated with poor prognosis in luminal subtypes only. Increased levels of IL13Rα2 were reported in breast cancer cells with brain metastatic tropism, yet the function of IL13Rα2 during brain metastatic colonization remains unexplored. Since IL-13 is expressed by various cells within the brain niche, this study tests the hypothesis that ligand dependent and independent mechanisms modulate IL13Rα2 function during brain metastatic progression.
Methods and results: qPCR, Western blot and IHC analysis demonstrated that IL13Rα2 was expressed at various levels in HER2+ and TN brain-trophic breast cancer cell lines (231BR, JmT1BR3, 4T1BR5, F2-7), brain-metastasis patient-derived xenografts (BM-PDXs) and a cohort of 26 clinical breast cancer brain metastasis (BCBM) samples. To assess how IL13Rα2 influences pro-metastatic abilities independent of ligand, inducible-shRNAs and CRISPR/cas9 were used to downregulate IL13Rα2 expression in 231BR cells. Knockdown of IL13Rα2 reduced the ability of 231BR cells to migrate (41.7% ± 4.5 vs 71.5% ± 9.7; p=0.001) and invade (49.5% ± 6.5 vs 73.1% ± 2.9; p=0.001) compared to the empty vector (EV) control cells. Furthermore, inducible-IL13Rα2 downregulation decreased significantly the proliferation (-22.1% at 96 hours, p<0.0001) and migration (-10.6% at 24 hours, p<0.0001) as compared to EV controls. Consistently, inducible overexpression of IL13Rα2 in HER2+ BT474 cells (which lack endogenous IL13Rα2) increased proliferation by 22.8% (p<0.0005, at 5 days) compared to parental cells. Knockdown of IL13Rα2 in 231BR cells impaired their ability to colonize the brain in mice following intracardiac injection as compared to EV 231Br cells (median number of metastatic clusters: 11.8 ± 5.5 vs 30.5 ± 13.9, respectively, p=0.2671), suggesting that upregulation of IL13Rα2 facilitates early brain metastatic colonization. To assess whether levels of IL13Rα2 remain high at late stages of brain metastasis, a publicly available transcriptome profile of 21 matched primary tumor and brain metastases was analyzed. Surprisingly, high IL13RA2 mRNA expression in brain metastasis (but not in primary tumors) predicted better brain-metastasis free survival (n=21, p=0.0026). Since the brain has several sources of IL-13, we assessed whether ligand-dependent mechanisms alter IL13Rα2 function, leading to a loss of expression in more aggressive brain metastasis. Western blot and IF analyses showed IL-13 expressed in reactive astrocytes. IL-13 treatment of serum-deprived IL13Rα2+ 231BR and JmT1Br cells resulted in decreased proliferation, a moderate increase in invasion, and phosphorylation of FAK and SRC. Knockdown of IL13Rα2 blocked the ability of IL-13 to phosphoactive FAK and SRC, and prevented the antiproliferative effect of IL-13 on 231BR cells.
Conclusions: These results suggest that ligand-dependent and independent mechanisms modulate IL13Rα2 through brain metastatic colonization. Ligand-independent IL13Rα2 promotes proliferation and invasion of cancer cells, favoring early dissemination and brain colonization. Loss of IL13Rα2 at later stages of BM, may prevent IL-13-induced blockage of proliferation and promote aggressiveness of established BM
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Keywords
Temáticas
Neoplasias de la mama
Adrenérgicos
Metástasis de la neoplasia
Adrenérgicos
Metástasis de la neoplasia