Molecular Analysis of DCIS Progression in a Mouse Model

Institution: University of California, Davis
Investigator(s): Ruria Namba, Ph.D. -
Award Cycle: 2003 (Cycle IX) Grant #: 9FB-0212 Award: $80,000
Award Type: Postdoctoral Fellowship
Research Priorities
Biology of the Breast Cell>Pathogenesis: understanding the disease

Initial Award Abstract (2003)
Breast cancer development is a multi-step process, starting from a benign stage, through histologically and molecularly distinct intermediate proliferative stage of hyperplastic epithelium, and leading to invasive cancer and metastasis. The analysis of breast lesions indicates that early lesions are molecularly different from normal epithelium. DCIS (ductal carcinoma in situ) is a type of pre-malignant lesion in human breast tissue that has a high rate of transformation to invasive cancer. Our knowledge of this transformation process is still incomplete, especially at the molecular level, despite an increasing number of genes being associated with breast cancer. Understanding the molecular mechanism will lead to identification of biomarkers for early breast lesions and their prognosis, as well as molecular targets for preventive therapy.

The overall goal of this project is to study a well-characterized mouse model of breast cancer development by correlating the biology and molecular profiles of hyperplasia and tumor tissues. Our aim is to identify possible key regulators of this malignant transformation. The research will be based on a novel mouse model of DCIS. In this model, premalignant mouse mammary tissue has been maintained for many generations by serial transplantation into new host animals. These tissues, called hyperplastic outgrowths (HPO), were originally derived from hyperplastic epithelium of young, genetically engineered animals that expressed the polyomavirus middle T antigen in mammary glands (MMTV-PyVmT). They are histologically indistinguishable from human DCIS and will transform into malignant tumors, if allowed to grow. We will use normal, premalignant (HPO), and tumor tissues from the same mouse mammary fat pads (i.e., breasts) to study the gene expression differences. Since the mammary fat pads have been processed for histology after the sample extraction, the molecular differences can be correlated with the morphology and histology of each tissue. We also plan to develop cell lines from the tumor and HPO tissues. The purpose of these cell lines is to experimentally test the role of any suspected genes in breast cancer progression. For example, we will be able to employ the newly developed technique of “RNAi” inhibition to selectively block individual genes, and determine the effects on proliferation, migration, and invasion. Moreover, we will be able to treat these cells with chemical inhibitors for certain genes to block their function and assess the effects on tumor development before testing them on the animal model.

Initial studies have shown that HPO tissues in our model are similar to human DCIS in their histology, biology, and molecular alterations. Development and thorough characterization of this model and cell culture lines will be beneficial because they can be used as tools to design treatment and preventive therapy for breast cancer.

Final Report (2005)
DCIS (ductal carcinoma in situ) is a type of pre-malignant lesion in human breast tissue that has a high rate of transformation to invasive cancer. Our knowledge of breast cancer progression and the biology of pre-malignant breast lesions is still incomplete, partly due to the lack of experimentally amenable in vivo DCIS models. We have developed a pre-malignant mouse mammary model that progresses to invasive carcinoma. Individual premalignant lesions, also called, Mammary Intraepithelial Neoplasia (MIN), from a Tg: MMTV-PyVmT mammary fat pad were isolated and transplanted into a wild type host mammary fat pad to establish a distinct outgrowth line (MIN-Outgrowth, or MIN-Os). These lines were verified to be pre-malignant and transplanted repeatedly to establish stable and predictable biological properties (e.g., morphology, growth rate and tumor latency). Although the lesions originated from the mammary fat pad of the same genotype (MMTV-PyVmT), each MIN-O line seems to have distinct biological characteristics. This also resembles the heterogeneity associated with human DCIS lesions. The overall goal of this project was to study the changes associated with tumor progression in this MIN-O model to further understand breast cancer progression in humans and eventually to be able to apply this MIN-O model for preclinical studies.

From gene expression analysis of the MIN-O lines, we found that MIN-Os and the corresponding tumors that arised from the MIN-Os have highly similar the global gene expression profiles despite their different malignant potential. The expression profiles of the MIN-Os are very similar within each line, reflecting the stable, but distint, biological characteristics. Moreover, the major gene expression changes are already in place at the MIN-O stage. These gene expression results are in agreement with the findings from human studies of DCIS and further supports the notion that the MIN-O model effectively mimics the biology of early stages of human breast cancer. In particular, in the pre-malignant MIN-O stage, we found that Ccnd1, Cbfb, Serpine2, and Pdk3 are upregulated. In contrast, regulators of transcription (Atbf1, Nfe213, Hoxb4, Ifrd1) and fatty acid synthesis and transport related genes (Pdk3, Pdhb, Dlat, CD36, Fabp3) are down-regulated, compared to the normal proliferating mammary glands. The transition from the MIN-Os to invasive tumors, on the other hand, has approximately 10-fold fewer genes differentially expressed between the transition from normal developing mammary gland to the MIN-Os.

Since the estrogen receptor (ER) status is a very important prognostic indicator in human breast cancer, we have also characterized the estrogen dependency and ER sensitivity of the MIN-O lines. The estrogen dependence was heterogeneous, since the effect of ovariectomy on the MIN-O growth rate and tumor latency varied among the lines. We were also able to show growth inhibition of one of the MIN-O lines by Selective Estrogen Receptor Modulators (SERMs), tamoxifen and ospemifene.

These results demonstrated that this transplantable pre-malignant mouse mammary lesion model recapitulates the biology of human DCIS. The heterogeneity of the MIN-O lines provides an excellent platform to investigate various characteristics relevant for breast cancer, such as estrogen status and metastasis. Since the MIN-O lines have stable biological characteristics and their molecular profiles have been documented, this model provides an experimental platform that can be used to assess functional genomics, molecular pharmacology, and to test promising chemoprevention strategies.

Syngeneic mouse mammary carcinoma cell lines: Two closely related cell lines with divergent metastatic behavior.
Periodical:Clinical & Experimental Metastasis
Index Medicus: Clin. Exp. Metastasis
Authors: Borowsky AD, Namba R, et al., and Gregg JP
Yr: 2005 Vol: 22 Nbr: Abs: Pg:47-58

Molecular characterization of the transition to malignancy in a genetically engineered mouse-based model of ductal carcinoma in situ.
Periodical:Molecular Cancer Research
Index Medicus: Mol Cancer Res
Authors: Namba R, Maglione JE, Young LJ, Borowsky AD, Cardiff RD, MacLeod CL, Gregg JP.
Yr: 2004 Vol: 2 Nbr: 8 Abs: Pg:453-63

The molecular biology of mammary intraepithelial neoplasia outgrowths
Periodical:Breast Cancer Research
Index Medicus: Breast Cancer Res
Authors: Namba R et al.
Yr: 2003 Vol: 5 Nbr: Suppl. 1 Abs: Pg:S7

Rapamycin inhibits growth of premalignant and malignant mammary lesions in a mouse model of ductal carcinoma in situ.
Periodical:Clinical Cancer Research
Index Medicus: Clin Cancer Res
Authors: Namba R, Young LJ, et al. and , MacLeod CL, and Gregg JP
Yr: 2006 Vol: 12 Nbr: 8 Abs: Pg:2613-21

Selective estrogen receptor modulators inhibit growth and progression of premalignant lesions in a mouse model of ductal carcinoma in situ.
Periodical:Breast Cancer Research
Index Medicus: Breast Cancer Res
Authors: Namba R, Young LJ, et al, and Gregg JP
Yr: 2005 Vol: 7 Nbr: Abs: Pg:881-889