Classification of rat mammary carcinoma with large scale in vivo microwave measurements.

Mammary carcinoma, breast cancer, is the most commonly diagnosed cancer type among women. Therefore, potential new technologies for the diagnosis and treatment of the disease are being investigated. One promising technique is microwave applications designed to exploit the inherent dielectric property discrepancy between the malignant and normal tissues. In theory, the anomalies can be characterized by simply measuring the dielectric properties. However, the current measurement technique is error-prone and a single measurement is not accurate enough to detect anomalies with high confidence. This work proposes to classify the rat mammary carcinoma, based on collected large-scale in vivo S[Formula: see text] measurements and corresponding tissue dielectric properties with a circular diffraction antenna. The tissues were classified with high accuracy in a reproducible way by leveraging a learning-based linear classifier. Moreover, the most discriminative S[Formula: see text] measurement was identified, and to our surprise, using the discriminative measurement along with a linear classifier an 86.92% accuracy was achieved. These findings suggest that a narrow band microwave circuitry can support the antenna enabling a low-cost automated microwave diagnostic system.

Differentiating breast cancer molecular subtypes using a DNA aptamer selected against MCF-7 cells.

Aptamers are single-stranded DNA or RNA oligonucleotides selected by systematic evolution of ligands by exponential enrichment (SELEX), which show great potential in the diagnosis and personalized therapy of cancers, due to their specific advantages over antibodies. In the past years, though great progress has been made in molecular subtyping of breast cancer, it remains a challenge in clinical medicine, which plays a crucial role in the treatment. In this study, a ssDNA aptamer MF3 against MCF-7 breast cancer cells was developed by Cell-SELEX for differentiating breast cancer molecular subtypes, which showed favorable specificity and binding affinity towards MCF-7 cells with a Kd value of 82.25 ± 25.14 nM. The aptamer could not only successfully distinguish MCF-7 breast cancer cells from MDA-MB-231 and SK-BR-3 breast cancer cells and MCF-10A human normal mammary epithelial cells, but also could differentiate MCF-7 cells from other cancer cells or normal cells. Moreover, both in vivo and in vitro fluorescence imaging studies demonstrated that aptamer MF3 was able to distinguish tumor-bearing mice and xenograft tissue sections of MCF-7 breast cancer cells from that of MDA-MB-231 and SK-BR-3 breast cancer cells. All these results suggested that aptamer MF3 is a potential tool for differentiating molecular subtypes and diagnosis of breast cancer.

Molecular imaging using PET and SPECT for identification of breast cancer subtypes.

Breast cancer is a major disease with high morbidity and mortality in women. As a highly heterogeneous tumor, it contains different molecular subtypes: luminal A, luminal B, human epidermal growth factor 2-positive, and triple-negative subtypes. As each subtype has unique features, it may not be universal to the optimal treatment and expected response for individual patients. Therefore, it is critical to identify different breast cancer subtypes. Targeting subcellular levels, molecular imaging, especially PET and single photon emission computed tomography, has become a promising means to identify breast cancer subtypes and monitor treatment. Different biological processes between various subtypes, including changes correlated with receptor expression, cell proliferation, or glucose metabolism, have the potential for imaging with PET and single photon emission computed tomography radiopharmaceuticals. Receptor imaging, with radiopharmaceuticals targeting estrogen receptor, progesterone receptor, or human epidermal growth factor 2, is available to distinguish receptor-positive tumors from receptor-negative ones. Cell proliferation imaging with fluorine-18 fluorothymidine PET aids identification of luminal A and B subtypes on the basis of the correlation with the immunohistochemical biomarker Ki-67. Glucose metabolism imaging with fluorine-18 fluorodeoxyglucose PET may have potential to discriminate triple-negative subtypes from others. With increasing numbers of novel radiopharmaceuticals, noninvasive molecular imaging will be applied widely for the identification of different subtypes and provide more in-vivo information on individualized management of breast cancer patients.

Common and distinct features of mammary tumors driven by Pten-deletion or activating Pik3ca mutation.

PTEN loss and PIK3CA activation both promote the accumulation of phosphatidylinositol (3, 4, 5)-trisphosphate (PIP3). While these proteins also have distinct biochemical functions, beyond the regulation of PIP3, little is known about the consequences of these differences in vivo. Here, we directly compared cancer signalling in mammary tumors from MMTV-Cre:Ptenf/f and MMTV-Cre:Pik3ca(LSL-H1047R) mice. Using unsupervised hierarchical clustering we found that whereas MMTV-Cre:Pik3ca(LSL-H1047R)-derived tumors fall into two separate groups, designated squamous-likeEx and class14(Ex), MMTV-Cre:Ptenf/f tumors cluster as one group together with PIK3CA(H1047R) class14(Ex), exhibiting a 'luminal' expression profile. Gene Set Enrichment Analysis (GSEA) of Pten(Δ)ˆ† and PIK3CA(H1047R) class14(Ex) tumors revealed very similar profiles of signalling pathways as well as some interesting differences. Analysis of 18 signalling signatures revealed that PI3K signalling is significantly induced whereas EGFR signalling is significantly reduced in Pten(∆) versus PIK3CA(H1047R) tumors. Thus, Pten(∆) and PIK3CA(H1047R) tumors exhibit discernable differences that may impact tumorigenesis and response to therapy.

Genetic background may contribute to PAM50 gene expression breast cancer subtype assignments.

Recent advances in genome wide transcriptional analysis have provided greater insights into the etiology and heterogeneity of breast cancer. Molecular signatures have been developed that stratify the conventional estrogen receptor positive or negative categories into subtypes that are associated with differing clinical outcomes. It is thought that the expression patterns of the molecular subtypes primarily reflect cell-of-origin or tumor driver mutations. In this study however, using a genetically engineered mouse mammary tumor model we demonstrate that the PAM50 subtype signature of tumors driven by a common oncogenic event can be significantly influenced by the genetic background on which the tumor arises. These results have important implications for interpretation of "snapshot" expression profiles, as well as suggesting that incorporation of genetic background effects may allow investigation into phenotypes not initially anticipated in individual mouse models of cancer.

Transgenic IGF-IR overexpression induces mammary tumors with basal-like characteristics, whereas IGF-IR-independent mammary tumors express a claudin-low gene signature.

Molecular profiling has allowed a more precise classification of human cancers. With respect to breast cancer, this approach has been used to identify five subtypes; luminal A, luminal B, HER2-enriched, basal-like and claudin-low. In addition, this approach can be used to determine the type of tumor represented by particular cell lines or transgenic animal models. Therefore, this approach was utilized to classify the mammary tumors that develop in MTB-IGFIR transgenic mice. It was determined that the primary mammary tumors, which develop due to elevated expression of the type I insulin-like growth factor receptor (IGF-IR) in mammary epithelial cells, most closely resemble murine tumors with basal-like or mixed gene expression profiles and with human basal-like breast cancers. Downregulation of IGF-IR transgene in MTB-IGFIR tumor-bearing mice leads to the regression of most of the tumors, followed by tumor reappearance in some of the mice. These tumors that reappear following IGF-IR transgene downregulation do not express the IGF-IR transgene and cluster with murine mammary tumors that express a mesenchymal gene expression profile and with human claudin-low breast cancers. Therefore, IGF-IR overexpression in murine mammary epithelial cells induces mammary tumors with primarily basal-like characteristics, whereas tumors that develop following IGF-IR downregulation express a gene signature that most closely resembles human claudin-low breast tumors.

Gene expression profiling distinguishes between spontaneous and radiation-induced rat mammary carcinomas.

The ability to distinguish between spontaneous and radiation-induced cancers in humans is expected to improve the resolution of estimated risk from low dose radiation. Mammary carcinomas were obtained from Sprague-Dawley rats that were either untreated (n = 45) or acutely gamma-irradiated (1 Gy; n = 20) at seven weeks of age. Gene expression profiles of three spontaneous and four radiation-induced carcinomas, as well as those of normal mammary glands, were analyzed by microarrays. Differential expression of identified genes of interest was then verified by quantitative polymerase chain reaction (qPCR). Cluster analysis of global gene expression suggested that spontaneous carcinomas were distinguished from a heterogeneous population of radiation-induced carcinomas, though most gene expressions were common. We identified 50 genes that had different expression levels between spontaneous and radiogenic carcinomas. We then selected 18 genes for confirmation of the microarray data by qPCR analysis and obtained the following results: high expression of Plg, Pgr and Wnt4 was characteristic to all spontaneous carcinomas; Tnfsf11, Fgf10, Agtr1a, S100A9 and Pou3f3 showed high expression in a subset of radiation-induced carcinomas; and increased Gp2, Areg and Igf2 expression, as well as decreased expression of Ca3 and non-coding RNA Mg1, were common to all carcinomas. Thus, gene expression analysis distinguished between spontaneous and radiogenic carcinomas, suggesting possible differences in their carcinogenic mechanism.

Mixture model approach to tumor classification based on pharmacokinetic measures of tumor permeability.

PURPOSE: To categorize the disease severity of mammary tumors in an animal model through the application of a novel tumor permeability mixture model within a hierarchical modeling framework. MATERIALS AND METHODS: Thirty-six rats with mammary tumors of varying grade were imaged via dynamic contrast-enhanced (CE) MRI using albumin-(Gd-DTPA)30. Time-dependent contrast agent concentration curves for blood and tumor tissue were obtained and a mathematical model of microvascular blood-tissue exchange was developed under the hypothesis that endothelial integrity is disrupted in a manner proportional to the degree of malignancy, with benign tumors showing no disruption of the vasculature endothelium. This permeability model was incorporated into a statistical model for the benign and malignant tumor subgroups that enabled automatic subject classification. The structural and statistical models were implemented using the software Nonlinear Mixed Effects Modeling (NONMEM) to statistically separate subjects into the two subgroups. RESULTS: Individual tumor classifications (as benign or malignant) were evaluated against the Scarff-Bloom-Richardson microscopic scoring method as applied to the tumor histology of each subject. The model-based classification resulted in 90.9% sensitivity, 92.9% specificity, and 91.7% accuracy. CONCLUSION: Mixture model analysis provides a robust method for subject classification without user intervention and bias. Although the present results are promising, additional research is needed to further evaluate this technique for diagnostic purposes.

Characterization of medroxyprogesterone and DMBA-induced multilineage mammary tumors by gene expression profiling.

Mouse mammary tumors arising during medroxyprogesterone-DMBA-mediated mammary carcinogenesis comprised three distinct phenotypes: adenocarcinoma, squamous cell carcinoma, and myoepithelial carcinoma. The molecular signature for each of the three tumor subsets was characterized by gene microarray analysis, and three distinct sets of gene expression profiles were obtained that were corroborated in part by quantitative RT-PCR and immunohistochemistry. These results suggest that this carcinogenesis and gene expression model will be useful for rapidly assessing the histopathological differences arising in mammary carcinogenesis and the effects of tumor promoting or chemoprevention agents.

Proteotypic classification of spontaneous and transgenic mammary neoplasms.

INTRODUCTION: Mammary tumors in mice are categorized by using morphologic and architectural criteria. Immunolabeling for terminal differentiation markers was compared among a variety of mouse mammary neoplasms because expression of terminal differentiation markers, and especially of keratins, provides important information on the origin of neoplastic cells and their degree of differentiation. METHODS: Expression patterns for terminal differentiation markers were used to characterize tumor types and to study tumor progression in transgenic mouse models of mammary neoplasia (mice overexpressing Neu (Erbb2), Hras, Myc, Notch4, SV40-TAg, Tgfa, and Wnt1), in spontaneous mammary carcinomas, and in mammary neoplasms associated with infection by the mouse mammary tumor virus (MMTV). RESULTS: On the basis of the expression of terminal differentiation markers, three types of neoplasm were identified: first, simple carcinomas composed exclusively of cells with a luminal phenotype are characteristic of neoplasms arising in mice transgenic for Neu, Hras, Myc, Notch4, and SV40-TAg; second, 'complex carcinomas' displaying luminal and myoepithelial differentiation are characteristic of type P tumors arising in mice transgenic for Wnt1, neoplasms arising in mice infected by the MMTV, and spontaneous adenosquamous carcinomas; and third, 'carcinomas with epithelial to mesenchymal transition (EMT)' are a characteristic feature of tumor progression in Hras-, Myc-, and SV40-TAg-induced mammary neoplasms and PL/J and SJL/J mouse strains, and display de novo expression of myoepithelial and mesenchymal cell markers. In sharp contrast, EMT was not detected in papillary adenocarcinomas arising in BALB/cJ mice, spontaneous adenoacanthomas, neoplasms associated with MMTV-infection, or in neoplasms arising in mice transgenic for Neu and Wnt1. CONCLUSIONS: Immunohistochemical profiles of complex neoplasms are consistent with a stem cell origin, whereas simple carcinomas might originate from a cell committed to the luminal lineage. In addition, these results suggest that the initiating oncogenic events determine the morphologic features associated with cancer progression because EMT is observed only in certain types of neoplasm.

Differentiation and characterization of rat mammary fibroadenomas and 4T1 mouse carcinomas using quantitative ultrasound imaging.

Scatterer properties like the average effective scatterer diameter and acoustic concentration were determined in vivo using a quantitative ultrasound (QUS) technique from two tumor phenotypes grown in animal models. These tumor models included spontaneously occurring mammary fibroadenomas in rats and transplanted 4T1 mammary carcinomas in mice. The scatterer properties of average scatterer diameter and acoustic concentration were estimated using a Gaussian form factor from the backscattered ultrasound measured from both types of tumors. QUS images of the tumors were constructed utilizing estimated scatterer properties from regions in the tumors. The QUS images showed a clear distinction between the two types of tumors and a statistically significant difference existed between their estimated scatterer properties. The average scatterer diameter and acoustic concentration for the mammary fibroadenomas were estimated to be 105 +/- 25 microm and -15.6 +/- 5 dB(mm(-3)), respectively. The average scatterer diameter and acoustic concentration for the carcinomas was estimated to be 28 +/- 4.6 microm and 10.6 +/- 6.9 dB(mm(-3)), respectively. The distinctions in the scattering properties are clearly seen in the QUS images of the tumors and indicate that QUS imaging can be useful in differentiating between different types of mammary tumors.

The C-neu mammary carcinoma in Oncomice; characterization and monitoring response to treatment with herceptin by magnetic resonance methods.

To characterize spontaneously occurring c-neu/HER2 overexpressing tumours in oncomice and their response to herceptin by non-invasive magnetic resonance spectroscopy (MRS) and magnetic resonance imaging (MRI). Oncomice were monitored by localized 31P MRS during unperturbed growth and before and after treatment with 10 mg/kg herceptin (Hoffman La Roche) intraperitoneally for up to 21 days post-treatment. Vascular morphology and function was assessed by quantitation of tumour magnetic resonance (MR) relaxation rates R2* and R2 prior to and either during carbogen (95% O2/5% CO2) breathing or following administration of the blood-pool contrast agent NC100150 (Clariscan, Amersham Health). Immunohistochemistry showed strong membrane staining for HER2 protein overexpression. The 31P MRS showed only a significant (p<0.01) increase of phosphomonoester / total phosphate ratio over 21 days of growth. Herceptin increased the tumour volume doubling time compared to untreated tumours and significantly increased the phosphomonoester / beta-nucleoside triphosphate ratio 2 days after treatment (p=0.01). Tumours showed a highly heterogeneous yet significant (p<0.01) decrease or increase in R2* in response to carbogen or NC100150 respectively. The absence of a decline in tumour bioenergetics with growth, commonly seen in 31P MRS studies of transplanted rodent tumour models, coupled with the heterogeneous blood volume revealed by 1H MRI, suggest a metabolic and vascular phenotype similar to that found in human tumours.

cDNA microarray profiling of rat mammary gland carcinomas induced by 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and 7,12-dimethylbenz[a]anthracene.

cDNA microarray analysis was used to examine gene expression profiles in normal female Sprague-Dawley rat mammary gland and in carcinomas induced by the cooked meat-derived carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and the potent experimental carcinogen 7,12-dimethylbenz[a]anthracene (DMBA). Nine tubulopapillary carcinomas (five from PhIP-treated rats and four from DMBA-treated rats) and normal mammary gland from virgin, pregnant and lactating rats were examined on a rat 6.9k cDNA microarray. Although histologically identical, PhIP- and DMBA-induced carcinomas could be distinguished by hierarchical clustering and multi-dimensional scaling analyses of cDNA expression. In addition, the expression of 21 clones was statistically different between PhIP- and DMBA-induced carcinomas (F-test, P < 0.05). The data indicate that distinct chemical carcinogens induce unique gene expression patterns in mammary gland carcinomas. The specific chemical carcinogen-associated cDNA array profiles found in carcinomas may ultimately be applicable to better understanding cancer etiology. PhIP- and DMBA-induced carcinomas also shared similarities in cDNA expression profiles. By comparing the expression in carcinomas (PhIP plus DMBA induced) with normal rat mammary gland (at any stage of differentiation), 172 clones were found to be differentially expressed. Genes showing increased expression in carcinomas by cDNA microarray analysis (and further validated by immunohistochemistry and western blot analysis) include cyclin D1, PDGF-A chain, retinol binding protein 1, prohibitin and the transcription factor STAT5A. The similarities in gene expression between PhIP- and DMBA-induced carcinomas raise the possibility that several molecular pathways for rat mammary gland transformation are maintained irrespective of the carcinogenic initiating agent.

The mammary pathology of genetically engineered mice: the consensus report and recommendations from the Annapolis meeting.

NIH sponsored a meeting of medical and veterinary pathologists with mammary gland expertise in Annapolis in March 1999. Rapid development of mouse mammary models has accentuated the need for definitions of the mammary lesions in genetically engineered mice (GEM) and to assess their usefulness as models of human breast disease. The panel of nine pathologists independently reviewed material representing over 90% of the published systems. The GEM tumors were found to have: (1) phenotypes similar to those of non-GEM; (2) signature phenotypes specific to the transgene; and (3) some morphological similarities to the human disease. The current mouse mammary and human breast tumor classifications describe the majority of GEM lesions but unique morphologic lesions are found in many GEM. Since little information is available on the natural history of GEM lesions, a simple morphologic nomenclature is proposed that allows direct comparisons between models. Future progress requires rigorous application of guidelines covering pathologic examination of the mammary gland and the whole animal. Since the phenotype of the lesions is an essential component of their molecular pathology, funding agencies should adopt policies ensuring careful morphological evaluation of any funded research involving animal models. A pathologist should be part of each research team.

Atlas and histologic classification of tumors of the rat mammary gland.

Studies performed in experimental animal models have demonstrated that mammary cancer is a complex multistep process that can be induced either by chemicals, radiation, viruses, or genetic factors. Rodent models have been useful for dissecting the initiation, promotion, and progression steps of mammary carcinogenesis. Chemically induced mammary tumors, such as those induced by 7,12-dimethylbenz(a)anthracene and N-methyl-N-nitrosourea, are, in general, hormone-dependent adenocarcinomas whose incidence, number of tumors per animal, tumor latency, and tumor type are influenced by the age, reproductive history, and endocrinologic milieu of the host at the time of carcinogen exposure as well as by diet and the dose of carcinogen administered. There is a need to classify tumors according to their histopathological type because those characteristics have implications in the interpretation of experimental data. In the classification presented here we attempt to provide a working framework for diagnosis of the type of lesions found in the mammary glands of rats treated with chemical carcinogens or radiation and to clarify criteria for establishing the basic biological characteristics of tumors.

Classification of premalignant and malignant lesions developing in the rat mammary gland after injection of sexually immature rats with 1-methyl-1-nitrosourea.

Premalignant and malignant stages of mammary carcinogenesis can be rapidly induced by injecting female rats i.p. with 1-methyl-1-nitrosourea (MNU)3 at 21 days of age. In this paper, the characteristics of this model are briefly reviewed and the histology of the lesions induced is presented and compared to those that occur in humans. Malignant mammary lesions induced in rats injected with MNU at 21 days of age are compared with the lesions that develop when MNU is administered to 50-day-old female rats.

Biopsia mamaria por aspiración con aguja fina / Biopsy of the breast with fine needle by aspiration

Se evaluaron 43 casos de pacientes que asistieron a la Unidad de Patología Mamaria de la Maternidad Concepción Palacios, a las cuales se les detectó una masa palpable y se les realizó biopsia por aspiración con aguja fina. La sensibilidad del método fue de 83,33 por ciento, obteniendo una especificidad de 78,4 por ciento valor predictivó positivo y negativo del 100 por ciento. Se calcularon límites de confianza del 95 por ciento de verosimilitud

Insertional mutagenesis identifies a member of the Wnt gene family as a candidate oncogene in the mammary epithelium of int-2/Fgf-3 transgenic mice.

Transgenic mice harboring the int-2/Fgf-3 protooncogene under transcriptional control of the mouse mammary tumor virus (MMTV) promoter/enhancer exhibit a dramatic, benign hyperplasia of the mammary gland. In one int-2 transgenic line (TG.NX), this growth disturbance is evoked by pregnancy and regresses after parturition. Regression of hyperplastic mammary epithelium is less complete after successive pregnancies, and, within 10 months, most TG.NX mice stochastically develop mammary carcinomas that are transplantable in virgin, syngeneic mice. To identify genes that cooperate with int-2 in cell transformation, we infected TG.NX transgenic mice with MMTV. In a cohort of 14 animals, most mammary tumors represented clonal or oligoclonal outgrowths harboring one to five proviral MMTV integrants. Eight of 35 (23%) MMTV+ tumors exhibited proviral insertion at the Wnt-1 locus. No provirus was detected at the int-2, int-3, or Wnt-3 loci. By Southern analysis, two tumors had proviral insertions at the same genomic location, which was mapped to chromosome 15. Cloning of this int locus identified an additional member of the Wnt gene family. The predicted 389-amino acid protein is most closely related to zebrafish Wnt-10a (58% amino acid identity over 362 residues) and, based on homology analysis, was designated Wnt-10b. This newly discovered Wnt family member was expressed in the embryo and mammary gland of virgin but not pregnant mice and represents a candidate collaborating oncogene of int-2/Fgf-3 in the mammary epithelium.

Comparative pathology of mammary tumorigenesis in transgenic mice.

Mammary tumors arise in transgenic mice bearing growth factors, proto-oncogenes, oncogenes and tumor suppressor genes. The tumors arise from hyperplasias. The tumor natural history and histogenesis are oncogene specific. Interactions between oncogenes may impede or accelerate tumorigenesis.