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A Versatile Tumor Gene Deletion System Reveals a Crucial Role for FGFR1 in Breast Cancer Metastasis.

ABSTRACT: RCAS avian viruses have been used to deliver oncogene expression and induce tumors in transgenic mice expressing the virus receptor TVA. Here we report the generation and characterization of a novel RCAS-Cre-IRES-PyMT (RCI-PyMT) virus designed to specifically knockout genes of interest in tumors generated in appropriate mutant mouse hosts. FGF receptor 1 (FGFR1) is a gene that is amplified in human breast cancer, but there have been no definitive studies on its function in mammary tumorigenesis, progression, and metastasis in vivo in spontaneous tumors in mice. We used the retroviral tumor knockout, or TuKO, strategy to delete fgfr1 in PyMT-induced mammary tumors in K19-tva/fgfr1loxP/loxP mice. The similarly injected control K19-tva mice developed mammary tumors exhibiting high metastasis to lung, making this an ideal model for breast cancer metastasis. The fgfr1 TuKO tumors showed significantly decreased primary tumor growth and, most importantly, greatly reduced metastasis to lung. In contrast to previous reports, FGFR1 action in this spontaneous mammary tumor model does not significantly induce epithelial-to-mesenchymal transition. Loss of FGFR1 does generate a gene signature that is reverse correlated with FGFR1 gene amplification and/or upregulation in human breast cancer. Our results suggest that FGFR1 signaling is a key pathway driving breast cancer lung metastasis and that targeting FGFR1 in breast cancer is an exciting approach to inhibit metastasis.

SUBMITTER: Wang W

PROVIDER: S-EPMC5402631 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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A Versatile Tumor Gene Deletion System Reveals a Crucial Role for FGFR1 in Breast Cancer Metastasis.

Wang Wei W Meng Yanling Y Dong Bingning B Dong Jie J Ittmann Michael M MM Creighton Chad J CJ Lu Yang Y Zhang Hong H Shen Tao T Wang Jianghua J Rowley David R DR Li Yi Y Chen Fengju F Moore David D DD Yang Feng F

Neoplasia (New York, N.Y.) 20170420 5

RCAS avian viruses have been used to deliver oncogene expression and induce tumors in transgenic mice expressing the virus receptor TVA. Here we report the generation and characterization of a novel RCAS-Cre-IRES-PyMT (RCI-PyMT) virus designed to specifically knockout genes of interest in tumors generated in appropriate mutant mouse hosts. FGF receptor 1 (FGFR1) is a gene that is amplified in human breast cancer, but there have been no definitive studies on its function in mammary tumorigenesis, ...[more]

PMID: 28433771

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Project description:ObjectiveTriple-negative (PR-, ER-, HER-2-) breast cancer (TNBC) is regarded as more aggressive and more likely to recur after medical care. Emerging evidence has demonstrated that the circadian clock system regulates cell-signaling pathways critical to cancer cell proliferation, survival and metastasis, meaning that it could be a good candidate for TNBC treatment. As such, the aim of the current study was to examine the molecular mechanism by which the circadian clock system contributes to cancer progression in TNBC.MethodsCancer cells and primary breast cancer tissues were immunostained for the measurement of circadian clock proteins (CLOCK, BMAL1 and PER1) and acetylserotonin methyltransferase (ASMT). The association between ASMT and clock proteins was assessed using siRNA and Western blot. Transwell assays were used to detect cancer cell migration and invasion while MTT assays were utilized to evaluate cell proliferation.ResultsCircadian clock proteins (CLOCK, BMAL1, and PER1) and ASMT expression were higher in TNBC and triple positive breast cancer (TPBC) compared with para-carcinoma tissues (PCTs). Intriguingly, there was an obvious correlation between circadian clock proteins and ASMT expression in both TPBC and TNBC. Similarly, circadian clock proteins and ASMT were expressed to a greater extent in BT-474 (triple-positive) cells than in MDA-MB-231 (triple-negative) cells. The inhibition of ASMT reduced circadian clock protein levels in both breast cancer cell lines. Further analysis showed that the expression levels of ASMT and circadian clock proteins did not correlate with clinical parameters such as age, tumor size, histologic grade and CK5/6, but increased significantly with lymphatic invasion in TNBC. In agreement with this finding, knockdown of ASMT significantly leads to reductions in migration and invasion in MDA-MB-231 cells. However, over-expression of CLOCK reversed the decreases seen in ASMT inhibited cells.ConclusionOur study suggests that ASMT regulates the circadian clock system in breast cancer and inhibition of ASMT reduces the invasiveness of triple-negative breast cancer cells by downregulating clock protein in a certain extent, indicating the potential value of ASMT as a drug target for TNBC treatment.

Project description:Key pointsLAT4 is a broadly expressed uniporter selective for essential branched chain amino acids, methionine and phenylalanine, which are involved in epithelial transport. Its global deletion leads to an early malnutrition-like phenotype and death within 10 days after birth. Here, we tested the impact of deleting LAT4 selectively in the mouse intestine. This affected slightly the absorption of amino acids (AAs) and delayed gastrointestinal motility; however, it had no major phenotypic effect, even when combined with aromatic AA uniporter TAT1 knockout (KO). Conversely, kidney tubule-selective deletion of LAT4 led to a substantial aminoaciduria that strongly increased under a high protein diet. Combining a partial tubular LAT4 deletion with TAT1 KO implicated their synergistic action on AA reabsorption. These results show that LAT4 plays an important role for kidney AA reabsorption, but that its functional role in intestinal AA absorption is largely dispensable.AbstractAmino acid (AA) transporter LAT4 (Slc43a2) functions as facilitated diffusion uniporter for essential neutral AAs and is highly expressed at the basolateral membrane of small intestine (SI) and kidney tubule epithelia. Previously, we showed that LAT4 global knockout (KO) mice were born at the expected Mendelian ratio but died within 10 days. Their failure to gain weight and a severe malnutrition-like phenotype contrasted with apparently normal feeding, suggesting a severe intestinal AA absorption defect. In the present study, using conditional global and tissue-specific LAT4 KO mouse models, we nullified this hypothesis, demonstrating that the selective lack of intestinal LAT4 does not impair postnatal development, although it leads to an absorption defect accompanied by delayed gastrointestinal motility. Kidney tubule-specific LAT4 KO led to a substantial aminoaciduria as a result of a reabsorption defect of AAs transported by LAT4 and of other AAs that are substrates of the antiporter LAT2, demonstrating, in vivo, the functional co-operation of these two transporters. The major role played by basolateral uniporters in the kidney was further supported by the observation that, in mice lacking TAT1, another neutral AA uniporter, a partial LAT4 KO led to a synergistic increase of urinary AA loss. Surprisingly in the SI, the same combined KO induced no major effect, suggesting yet unknown compensatory mechanisms. Taken together, the lethal malnutrition-like phenotype observed previously in LAT4 global KO pups is suggested to be the consequence of a combinatorial effect of LAT4 deletion in the SI, kidney and presumably other tissues.

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A Versatile Tumor Gene Deletion System Reveals a Crucial Role for FGFR1 in Breast Cancer Metastasis.

Publications

A Versatile Tumor Gene Deletion System Reveals a Crucial Role for FGFR1 in Breast Cancer Metastasis.

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