Project description:The gain-of-function mutation in the pleckstrin homology domain of AKT1 (AKT1E17K) occurs in lung and breast cancer. By use of human cellular models and of a AKT1E17K transgenic Cre-inducible murine strain (R26-AKT1E17K mice) we have demonstrated that AKT1E17K is a bona-fide oncogene for lung epithelial cells. However, the role of AKT1E17K in breast cancer remains to be determined. Here, we report the generation and the characterization of a MMTV-CRE;R26-AKT1E17K mouse strain that expresses the mutant AKT1E17K allele in the mammary epithelium. We observed that R26AKT1E17K;MMTV-Cre mice presented a variety of proliferative alterations of the mammary epithelium that were classified as adenosis with low-to-high grade dysplasia. In addition, AKT1E17K stimulates the development of mammary tumors with incidence of 43%. Tumors were morphologically classified as ductal adenocarcinoma of medium-high grade though subsequent immunoistochemical characterization suggested they were of basal-like origin, being PR-/HER2-/ERα+ and CK8-/CK10-/CK5+/CK14+. We also observed that tumors expressing mutant AKT1E17K presented activation of the downstream signaling axis GSK3/cyclin D1 in mammary epithelium, in parallel with increased proliferation rate as demonstrated by cell number count and measurement of Ki67. In conclusion, AKT1E17K is a bona fidae oncogene that is able to initiate tumors at high efficiency in murine mammary epithelium in vivo.
Project description:Canine mammary gland tumors can be used as predictive models for human breast cancer. There are several types of microRNAs common in human breast cancer and canine mammary gland tumors. The functions of microRNAs in canine mammary gland tumors are not well understood. In the present study, we compared the characterization of microRNA expression in two-dimensional and three-dimensional canine mammary gland tumor cell models. The expression of microRNA-210 in the three-dimensional-SNP cells was 10.19 times higher than that in the two-dimensional-SNP cells.
Project description:Emerging data indicate that breast epithelial stem cells and progenitors, particularly those in the luminal epithelial cell lineage, are the cells-of-origin of breast carcinomas, and factors that influence breast cancer risk may alter the number and/or properties of these cells. We hypothesize that a subset of p27+ cells represent hormone-responsive progenitors that are quiescent due to the high activity of TGFβ signaling in these cells. The Estrogen-induced mammary tumor model in ACI inbred rats is physiologically relevant rodent model of breast cancer. In the present study we successfully generated Cdkn1b knockout ACI rats and performed comprehensive phenotypic assessment and RNAseq profiling using FACS sorted basal (CD24+CD29high) and luminal (CD24+CD29low) cell populations to characterize Cdkn1b+/+ and Cdkn1b-/- females in prepubertal and adult cohorts. We found that p27KO rats exhibited mammary differentiation phenotype and reduced numbers of mammary epithelial progenitor pool, Interestingly, p27 ablation has the most pronounced effect on luminal progenitor cell gene expression, and milk protein genes and pStat5 were dramatically upregulated, while PR and FoxA1 were greatly downregulated in Cdkn1b-/- luminal cells. Further characterization of mammary glands of prepubertal Cdkn1b knockout rats by fat pad transplantation illustrated p27 deletion in the mammary cancer susceptible ACI rat strain induced mammary epithelial cell differentiation through cell non-autonomous mechanisms.
Project description:Breast cancer is the most frequent cancer among women causing the greatest number of cancer-related deaths. Cancer heterogeneity is a main obstacle to therapies. Around 96% of the drugs fail from discovery to the clinical trial phase probably because of the current unreliable preclinical models. New models emerge such as companion dogs who develop spontaneous mammary tumors resembling human breast cancer in many clinical and molecular aspects. The present work aimed at developing a robust canine mammary tumor model in the form of tumoroids which recapitulate the tumor diversity and heterogeneity. We conducted a complete characterization of these canine mammary tumoroids through histologic, molecular and proteomic analysis, demonstrating their strong similarity to the primary tumor. We demonstrated that these tumoroids can be used as a drug screening model. Due to easy tissue availability, tumoroids can be produced at larger scale and cryopreserved to constitute a biobank. We have demonstrated that cryopreserved tumoroids keep the same histologic and molecular features (ER, PR and HER2 expression) as fresh tumoroids. Two techniques of cryopreservation were compared demonstrating that tumoroids made from frozen tumor material allowed to maintain a higher molecular diversity. These findings revealed that canine mammary tumoroids can be easily generated at large scale and can represent a more reliable preclinical model to investigate tumorigenesis mechanisms and develop new treatments for both veterinary and human medicine.
Project description:We profiled androgen receptor (AR) genomic targets using high-throughput sequencing of chromatin-immunoprecipitated (ChIP) DNA from TMPRSS2-ERG fusion gene positive DUCaP prostate cancer cells. ChIp-seq and microarray gene expression profiling datasets were integrated with the NHGRI GWAS PCa risk SNPs catalog to identify disease susceptibility SNPs localized within functional androgen receptor binding sites (ARBSs). Eighty GWAS index or linked SNPs were found to be localized in ARBSs. Among these rs11891426:T>G in the 7th intron of the melanophilin gene was found located within a novel putative auxiliary AR binding motif, which we found enriched in the neighborhood of canonical androgen responsive elements. T→G exchange attenuated the transcriptional activity of the ARBS in an AR reporter gene assay of prostate cancer cell models. It went also in line with decreased melanophilin protein level in primary prostate tumors with G allele.These results unravel a hidden link between androgen receptor and a functional PCa risk SNP, whose allele alteration affects androgen regulation of its host gene melanophilin . Genomic profile of androgen receptor binding sites of androgen or vehicle treated DUCaP cells using ChIP-seq. IgG precipiated DNAs from both treatments served as controls.
Project description:Porcine mammary fatty tissues represent an abundant source of biomaterial for generation of breast-specific extracellular matrix (ECM). Here we report the extraction of total ECM proteins from pig breast fatty tissues, the fabrication of hydrogel scaffolds from the extracted ECM proteins, the structural properties of the scaffolds, and the applications of the hydrogel in human mammary epithelial cell spatial cultures for cell surface receptor expression, metabolomics characterization, acini formation, proliferation, migration between different scaffolding compartments, and in vivo tumor formation. This model system provides an additional option for studying human breast diseases such as breast cancer.
Project description:MicroRNAs (miRNAs) are small noncoding RNAs that typically inhibit the translation and stability of messenger RNAs (mRNAs), controlling genes involved in a variety of cellular processes. miRNA dysregulation is recognized to play an essential role in the development and progression of cancer. MMTV-PyMT mice (Jax Strain: FVB/N-Tg(MMTV-PyVT)634Mul/J) are a well-characterized transgenic mouse model of breast cancer. Upon activation of the MMTV-PyVT transgene (mouse mammary tumor virus (MMTV) long terminal repeat upstream of a cDNA sequence encoding the Polyoma Virus middle T antigen (PyVT)) female carriers develop palpable mammary tumors as early as 5 weeks of age. We performed miRNA microarrays on samples from the MMTV-PyMT transgenic mouse model to investigate the differential expression of miRNAs during development of malignant disease in this model.
Project description:Breast cancer brain metastasis remains largely incurable. While several mouse models have been developed to investigate the genes and mechanisms regulating breast cancer brain metastasis, these models often lack clinical relevance since they require the use of immune-compromised mice and/or are poorly metastatic to brain from the mammary gland. We describe the development and characterization of an aggressive brain metastatic variant of the 4T1 syngeneic model (4T1Br4) that spontaneously metastasises to lung, bone and brain but is selectively more metastatic to the brain from the mammary gland than parental 4T1 tumors. The 4T1Br4 model will provide a clinically relevant tool to evaluate novel therapies against brain metastasis.