Nonetheless, these models continue to serve a purpose in breast cancer research (Blaas et al., ;. The impact of transgenic mouse models on breast cancer research was the topic of recent conferences in Annapolis, Maryland (March , ) and Bar. Methods Mol Biol. ; doi: /_3. Mouse models of breast cancer. Sakamoto K(1), Schmidt JW, Wagner KU.
breast of cancer models Mouse
Here, a generalizable overview of the types of in vivo model systems, with an emphasis primarily on murine models that are widely deployed in preclinical breast cancer research, is provided; this overview encompasses the specific relationship of the models with the clinical disease and how imaging within the context of the models might be exploited to maximize translational gains to combat breast cancer.
A distinguishing feature of this article is that the key attributes of various preclinical breast cancer models and their utility are developed from the perspective of noninvasive molecular imaging. Despite major successes and lessons learned from the genomic landscape of cancer, it is now widely recognized that individual cancer genomes, like individual patients, are exquisitely heterogeneous; each contains a unique spectrum of drivers accompanied by passengers of less obvious significance.
Tools that illuminate the cellular and molecular underpinnings of tumors on a patient-by-patient basis, such as noninvasive molecular imaging, will be essential to bringing precision cancer therapy to fruition.
As such, preclinical imaging techniques relevant to mouse models of breast cancer, with an emphasis on molecular imaging, are also discussed in some detail. Regardless of the degree of sophistication, model systems are, by definition, not humans.
Rationales for late-phase clinical failures of new drugs are frequently based on a healthy skepticism of the translational value of certain preclinical models; much has been written about this issue already, and the value of model systems as a translational bridge to clinical applications is not debated in this article. However, in vivo modeling provides gains to the breast cancer field that complement what can be discovered at the laboratory bench. Indeed, the strongest experimental approaches will test hypotheses in multiple model systems.
Therefore, it is critical to understand both the strengths and the limitations of in vivo models of breast cancer to maximize what can be learned with this approach. The laboratory mouse Mus musculus represents a truly ideal model system for simulating the entire spectrum of events that lead to advanced breast cancer in humans. Mouse model systems enable elucidation of distinct facets of cancer biology that may not be frankly addressable in patients.
Some of the advantages of the mouse as a model system are as follows: Additionally, mice and other rodents share many physiologic similarities with humans 8 and therefore are commonly used in drug metabolism and pharmacokinetic and toxicity studies. Ironically, for imaging studies, the small size of the mouse can be a limitation, particularly when studies aim to image tumors whose diameters approximate or are smaller than the effective resolution of the imaging modality of choice.
Some notable differences between humans and mice include a higher metabolic rate in mice, an altered telomere length in inbred mouse strains, and an altered time frame for cancer onset 9. Several clinical and pathologic features of human breast cancer that allow stratification of patients on the basis of risk, prognosis, and likelihood of a response to certain types of therapy have been identified 10 ; in this light, for clinical breast cancer there are several impressive precision medicine—related success stories 11 and opportunities for future drug development Table 1.
Distinct molecular subtypes can be initially stratified on the basis of hormone receptor status; luminal breast cancers are typically hormone receptor—positive, whereas human epidermal growth factor receptor 2 HER2 and basallike breast cancers BLBCs are hormone receptor—negative. Other potential molecular subtypes, including luminal C and normallike tumors, have been reported; at present, however, little is known about these subtypes Luminal breast cancers are characterized by the expression of the estrogen receptor ER and the progesterone receptor PR , which are nuclear hormone receptors, and other associated genes Luminal A breast cancers tend to express greater quantities of hormone receptors, particularly the PR, than luminal B breast cancers.
In contrast, luminal B tumors tend to exhibit characteristics associated with higher-grade disease, are frequently more proliferative, are clinically more aggressive, and have a poorer prognosis than luminal A tumors.
Luminal A and luminal B tumors exhibit disparate responses to chemotherapy, with higher-grade luminal B tumors frequently responding more favorably to chemotherapy HER2-enriched tumors are frequently higher-grade tumors, with positive lymph node involvement. Precision medicine approaches to this cancer include the use of trastuzumab Tz Herceptin; Genentech , a monoclonal antibody that selectively targets the HER2 gene product, a receptor tyrosine kinase, as well as small-molecule kinase inhibitors lapatinib and everolimus 15 , HER2-enriched breast cancers with metastatic disease are additionally treated with anthracyclines doxorubicin and often display an initial response to treatment, although recurrence is seen in nearly all cases.
Other strategies targeting the HER2 receptor and its pathway include novel small-molecule inhibitors and HER2 antibodies, heat shock protein 90 inhibitors, agents targeting downstream components of the HER2 signaling pathway, and antibody—drug conjugates.
Certain molecular imaging strategies targeting HER2-enriched tumors have leveraged the selectivity of Tz labeled with a positron-emitting isotope 64 Cu or 89 Zr.
Promising clinical results in patients with metastatic breast cancer have been shown for these strategies 17 , BLBCs are especially common in African American women 10 and are generally associated with a poor prognosis. However, recent efforts to develop increasingly effective therapies against TNBC have led to the identification of several novel TNBC subtypes distinguishable by gene expression profiles and with potential vulnerabilities Rapidly increasing knowledge about breast cancer molecular subtypes may affect the genesis of, progression of, and therapeutic strategy for any given breast cancer and underscores the importance of mouse model selection in designing preclinical studies and coclinical trials.
Astounding growth in the reported number as well as the biologic elegance of mouse models for cancer research has been witnessed in the last decade. An extensive repertoire of mouse models with which to study breast cancer progression and treatment is now available. In genetically engineered mouse models GEMMs , the tumor develops through all stages of epithelial transformation with the native stroma, immune system, and microenvironment Another important contribution to the volume of mouse models recently described has come from the assembly of patient-derived xenograft PDX banks and, particularly for some cancer types, standardization of the infrastructure and protocols required to support these systems Here we describe 4 types of mouse model systems that can be used for breast cancer research, identifying both the strengths and the limitations of each Table 2.
Mouse models of breast cancer derived by transplanting immortalized human cancer cell lines into an immunocompromised murine host are among the simplest and most frequently deployed model systems in cancer research.
Most preclinical drug treatment studies performed in vivo have involved the use of immortalized human breast cancer cell lines growing within the subcutaneous dorsal flank of immunocompromised mice. Given the vast research history accumulated for many immortalized breast cancer cell lines and the numerous, diverse cell lines that represent all breast cancer molecular subtypes, xenografting breast cancer cell lines has become a staple in preclinical breast cancer research.
Although these models are technically simple to establish and are inexpensive to maintain over the short term, they have critical weaknesses that should be considered before larger programmatic efforts are based solely on them.
An insightful commentary suggested that cell line xenografts are useful as a bridge between in vitro and in vivo studies 3. Objectively, cell line xenograft models have clear strengths, especially for rapid hypothesis testing, including the following: These strengths are balanced by the following limitations of cell line xenografts: An often overlooked shortcoming of the cell line xenograft model is the fact that immortalized cell lines are developed through clonal attrition, resulting in cell populations that are propagated through multiple passages on a typically plastic surface.
Selective pressures and genetic drift give rise to genotypic and phenotypic changes that may irreversibly distinguish daughter clones from paternal tumors 26 ; this scenario may poorly recapitulate the original underlying cancer biology of the patient. Models developed from patient-derived tumors, otherwise known as PDX models—in which patient tumors are surgically implanted into recipient murine hosts without being cultured—overcome this limitation.
PDX models of various human tumors have been developed with great success, although breast cancer PDX models have historically been especially challenging The major strengths of the PDX approach include genetic diversity and heterogeneity that more accurately reflect human breast cancer; the ability to model various cancer subtypes; the incorporation of contextually correct human stroma within the tumor, including vascularity and inflammation; the documented ability to model metastasis; and easy interrogation of tumors, such as breast cancers, for correlative studies.
This approach maintains the genetic and phenotypic integrity of the tumor cells, without the clonal selection or inadvertent genetic drift seen in immortalized breast cancer cell lines. Nevertheless, there are several potential drawbacks of PDX models, including the requirement to use a severely immunodeficient murine host; the fact that the surgical procedure for implanting tumors into mice is invasive and requires skill 29 ; a species disconnect between the implanted tumor cells and stroma human and subsequently infiltrating stroma mouse ; and the time required to generate the models, which can require several months simply for the establishment of engraftment Technical issues aside, the fact that establishing and maintaining PDX model systems require major capital investments in supporting infrastructure and personnel must not be overlooked.
The requirement for the use of immunocompromised mice in xenograft models fails to incorporate the impact of the immune system on the tumor response. This area of cancer research is in its early stages, with rapid progress and vast promise that underscores the need for immunocompetent models of breast cancer for more rigorous analyses. Adequately modeling cancer immunology requires a propagating tumor within an immunocompetent host.
One approach is to use mouse mammary tumors or mouse mammary tumor cell lines implanted into syngeneic immunocompetent murine hosts. Devoid of the species constraints inherent in xenografts and xenotransplants, allografted mouse tumors are not typically rejected by the murine host, given the similar genetic backgrounds. Syngeneic model systems offer the distinct advantage of studying cancer biology within the context of an intact immune system and species-specific tumor microenvironment.
However, mouse tumor cell lines are limited and annotated to various degrees, and although small-molecule therapies may be adequately evaluated within these models, the species specificity of antibody imaging agents and therapies generally precludes their evaluation in syngeneic model systems.
GEMMs are the most sophisticated in vivo platforms used to simulate human cancer. These models are capable of not only accurately mimicking many relevant pathophysiologic features of human cancer but also recapitulating the sequence of molecular events that give rise to cancer.
The transgenic expression of an oncogene specifically within the mouse mammary epithelium under the control of a strong mammary epithelial promoter is frequently used to induce mammary tumor formation. This is a clinically relevant model of tumor initiation and progression, enforcing the stepwise procession of cells from hyperplasia to ductal carcinoma in situ and then to invasive ductal carcinoma.
Importantly, this process occurs within the context of the native stromal matrix requiring stromal remodeling and angiogenesis and the native immune system requiring immune system evasion. The genetic manipulations can drive oncogene expression in a reversible or irreversible manner, in a tissue-specific manner 3 or, more broadly, throughout an entire organism. Frequently, GEMMs that harbor oncogenic driver genes e.
The diverse array of oncogenes used to generate transgenic models of breast cancer has resulted in a multitude of models that mimic many of the specific molecular subtypes seen in clinical breast cancers, as confirmed by comparative expression analyses of mouse and human breast tumor samples The advantages of GEMMs include tumor formation in the contextually appropriate tissue and potentially cell of origin through the use of tissue-specific or cell-specific promoters; an intact immune system; and a native tumor microenvironment that more accurately reflects human disease, including stromal components, vascularity, and inflammation.
However, GEMMs are limited by the time, expense, and resources required to derive, establish, and maintain them; these demands can be overly burdensome given the potentially low experimental throughput of GEMMs. Few GEMMs of breast cancer truly harbor ER expression, despite commonalities in expression profiles between mouse and human luminal breast cancers.
Although metastases in mouse breast cancer models are hematogenous and almost exclusively pulmonary, human breast cancer metastases occur though lymphatic spread that often precedes hematogenous metastasis to the lungs, liver, bone, brain, and elsewhere. Molecular imaging is an indispensable tool uniquely poised to address major challenges obstructing the delivery of personalized cancer therapy.
Capable of noninvasively quantifying the cellular and molecular underpinnings of tumors on a patient-by-patient basis, molecular imaging enables the detection of tumors at early, potentially curable stages and provides a means to accurately predict the response of a tumor to therapy well before conventional means of assessment.
Numerous excellent review articles that thoroughly discuss the attributes of various molecular imaging modalities in both patients and preclinical animal models have been disseminated. Rather than recapitulate a description of specific imaging systems and methods, we simply suggest that interested readers consult specific articles that already relate directly to this topic 32 — However, as an introduction to preclinical molecular imaging in breast cancer models, it is worth noting that a range of imaging modalities can be entirely suitable for this purpose; such modalities include optical techniques bioluminescence and fluorescence , ultrasound, MRI, MR spectroscopy, and nuclear imaging techniques that use ionizing radiation, namely, PET and SPECT Table 3.
The modalities can be generally parsed into 2 major categories: The choice of imaging modality for addressing in vivo hypotheses depends largely on the biologic question of interest and is often guided by the strengths and limitations inherent in the modality. Although all have been used in preclinical studies, only a select few are considered eminently translational. Once the modality and the model have been selected, numerous clinically unmet needs can potentially be addressed in the laboratory through the marriage of noninvasive molecular imaging and preclinical mouse models of breast cancer.
For example, the development of inhibitors targeting various portions of the ErbB signaling axis is an active and clinically important area of breast cancer research. Tz is a Food and Drug Administration—approved, recombinant, humanized monoclonal antibody that selectively binds to the extracellular domain of HER2, yet objective means to assess the treatment response to Tz therapy remain undeveloped. To this end, Whisenant et al. The development of noninvasive imaging methods that could identify nonresponders earlier during therapeutic intervention is of great clinical interest because of the desire to spare patients any delay in the initiation of effective combination therapies.
For example, Kramer-Marek et al. Moreover, molecular imaging with PET could serve as a valuable strategy for predicting the tumor response to Tz. Our laboratory has also evaluated a suite of translational, noninvasive molecular imaging metrics in an attempt to predict the response to Tz in preclinical mouse models of HER2-overexpressing breast cancer The results of that study suggested that molecular imaging of apoptosis accurately predicted Tz-induced regression of HER2-positive tumors and warranted clinical exploration as a means to predict an early response to neoadjuvant Tz Fig.
In an analogous study seeking the elucidation of mechanisms that affect Tz efficacy, Miller et al. B NIRannexin V did not accumulate in nonresponding tumors, and overall uptake of imaging probe decreased as tumors progressed. Nonlinear inverse correlation was observed between change in NIRannexin V uptake from baseline and change in tumor volume from baseline when all imaging time points and all mice were considered.
Reprinted with permission of Because of de novo and acquired resistance of luminal breast cancers to endocrine therapy, there remains a need to identify which ER- or PR-positive tumors are most likely to respond to therapy fulvestrant. Noninvasive imaging of baseline tumoral 18 F-FES uptake and initial changes in 18 F-fluorofuranylnorprogesterone uptake could be used as a prognostic strategy to identify responders and nonresponders to endocrine therapy at an early stage of disease. Another research area in which noninvasive molecular imaging and preclinical mouse models have natural synergy is the coclinical trial concept.
Mice can be a good representation of diseases in humans because:. Mice may not be an ideal model for breast cancer. This is mainly due to the lack of precision in many of the models.
When looking at metastasis, it is difficult to determine the precise location as well as its' frequency. Another issue revolves around the epithelial sub types and the inability to specifically target them when targeting a mutation. In a standard case, the excision of BRCA2 resulted in no tumorgenesis, but if p53 was mutated and inactivated, tumorgenesis would occur. Therefore, there is not a definitive answer in terms of the origin of the tumor, due to the extra mutation in p Transplantation of tumor cells into immunodeficient mice is a tool to study breast cancer and its metastatic effects.
The transplantation occurs as either allotransplants or xenographic transplants. Inoculating cells through intra ductal transplantations,  by cleared mammary fat pad injections   or by transplantations into the tail vein.
These mutations allow for the integration of new xenograft tissue. Without this injection, the human mammary epithelial cells en-grafted onto the pad are unable to colonize and grow. After 4 weeks of development, the newly en-grafted human mammary epithelial cells expanded within the fat pad.
Genetically engineered mice are constructed to model human phenotypes and pathologies. Mutant mice may include transgenes using different delivery methods:. The mice undergoing the process of transgenesis are known as transgenic mice. A basic transgene has a promoter region, Protein coding sequence, Intron and a stop codon. Mouse mammary tumor virus MMTV , is a retro virus that has been a known promoter to cause breast tumors once activated. It harbors a regulatory DNA sequence called the long terminal repeat LTR , which promotes steroid-hormone-inducible transcription.
The sites of integration have been known to be critical genes for cellular regulation. For a list of other mammary gland specific promoters and mouse models see.
MMTV-PyMT mice are then crossed bred with other genetically modified mice to generate various types of breast cancer models, including:. The mice harbouring this oncogene develop multifocal adenocarcinomas with lung metastases at about 15 weeks after pregnancy. This addresses the issue in terms of modeling the amplification of HER2 in mice development. In the non-fused mouse, the mammary gland would revert to a near virgin, but with this addition the mammary gland maintained the developed function.
Mouse models having two transgenes are called bi transgenic. Tri-transgenic mouse models constitute of more than two genes.
Multiple combinations and genetic modifications are made in such a way that either one or all the genes are put into a continuously expressed status, or in a controlled fashion to activate them at different time points.
They can also both be activated or deactivated by adding doxycycline. Metastatic cascade can be studied by keeping the gene activation under control or by adding a reporter gene e. The quantitative lineage-tracing strategies have proven to be successful in resolving cell fates in normal epithelial tissues either using tissue —specific or stem-cell -specific transgenes.
To conduct an inducible lineage-tracing experiment two components must be engineered into the mouse genome: The switch is commonly a drug-regulated form of the bacterial enzyme Cre-recombinase. This enzyme recognizes specific sequences, called LoxP sites.
After harvesting all the ten mouse mammary glands from the transgenic mice, single cell suspension is usually made and transplanted either in tail vein of non transgenic recipient mice  or in cleared fat pad of non-transgenic mice repopulating the mammary fat pad. Another tool to study breast cancer metastasis is to look for circulating tumor cells in transgenic mice e.
In the absence of specific markers for mammary cells, models with genetic marking of tumor cells gives the best experimental advantage, however the low volume of peripheral blood that can be obtained from live animals limits the application of this technique. Bioluminescence imaging relies on the detection of light produced by the enzymatic oxidation of an exogenous substrate.
The substrate luciferin, is oxidized to oxyluciferin in the presence of luciferase and emits light, which can be detected using an IVIS system such as a Xenogen machine. Luc; MTB Internal ribosome entry site: Luciferin animals which were not exposed to doxycycline can be injected into the lateral tail veins of immunodeficient mice on a doxycycline-free diet.
No bioluminescence signal will be observed in the lungs of recipient mice until they are given doxycycline food.
Bioluminescence can then be detected in the chest within 2 weeks of the start of doxycycline exposure. Intravital microscopy with multi photon excitation is a technique to visualize genetically engineered cells directly in vivo. Multi step metastatic cascades can be visualized by labelling with unique fluorescent colour under fluorescence microscope. Positron emission tomography PET , single photon emission computed tomography SPECT and computed tomography CT have been used to compare the efficiency of these in vivo imaging for detecting lesions at an early stage and to evaluate the response to chemotherapy.
Magnetic resonance imaging requires the use of nano-particles liposomes and an MRI contrast agent called gadolinium. The particles were then placed in vesicles via a polycarbonate membrane filter. The nano-particles are injected into the metastases evolved mice, and left there for twenty-four hours.
These mice are then scanned, and in the imaging software there are accumulations of these particles in certain areas where cells have metastasized. From Wikipedia, the free encyclopedia.
Mouse models of breast cancer metastasis
The impact of transgenic mouse models on breast cancer research was the topic of a recent conference in Annapolis. This conference (March. As such, preclinical imaging techniques relevant to mouse models of breast cancer, with an emphasis on molecular imaging, are also. Breast cancer metastatic mouse models are experimental approaches in which mice are genetically manipulated to develop a mammary tumor leading to distant .