Joyce A Schroeder

Interests

Research

My laboratory investigates the normal biology of the Epidermal Growth Factor Receptor (EGFR, and its family members, HER2 and ErbB3), as well as their role in transformation and metastasis. We focus on the role of LLGL1, MUC1 and CD44 to modulate EGFR function through alterations in receptor trafficking, signal transduction and non-canonical activities such as nuclear translocation and gene regulation. We have identified the loss of apicobasal polarity as a driver in EGFR-driven breast cancer, through its mislocalization and interaction with the apical protein MUC1. Interaction with MUC1 drives EGFR-dependent breast cancer by altering EGFR trafficking and driving EGFR nuclear translocation and induction of the Met Receptor to drive metastasis. Additionally, we have identified LLGL1 as a polarity protein that suppresses EGFR-dependent migration and survival. We also investigate CD44-dependent migration and metastasis, and the role of Hyaluronic Acid in extracellular matrix-driven activation of EGFR. Based on these studies, we have developed peptide-based therapeutics for cancer that block protein-protein interactions between EGFR and MUC1, as well as peptides that promote the multimerization of inactive ErbB receptors. We are now focused on the development of these peptide-based therapeutics for clinical applications, as well as investigating the mechanisms of polarity as a suppressor of transformation and metastasis.

Teaching

In my laboratory, I have trained over 50 undergraduate students and 13 graduate students.  I am also actively engaged in classroom teaching at the graduate and undergraduate level, including cell biology and cancer biology (for which I have written a text book).

Courses

Scholarly Contributions

Journals/Publications

• Atwell, B., Chen, C., Christofferson, M., Montfort, W. R., & Schroeder, J. A. (2022).

  Sorting nexin-dependent therapeutic targeting of oncogenic epidermal growth factor receptor

  . Cancer Gene Therapy. doi:10.1038/s41417-022-00541-7
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  Overexpression and/or overactivation of the Epidermal Growth Factor Receptor (EGFR) is oncogenic in several tumor types yet targeting the kinase domain of wildtype EGFR has had limited success. EGFR has numerous kinase-independent roles, one of which is accomplished through the Sorting Nexin-dependent retrotranslocation of EGFR to the nucleus, which is observed in some metastatic cancers and therapeutically resistant disease. Here, we have utilized the BAR domain of Sorting Nexin 1 to create a peptide-based therapeutic (cSNX1.3) that promotes cell death in EGFR-expressing cancer. We evaluated the efficacy of cSNX1.3 in tumor-bearing WAP-TGFα transgenic mice (an EGFR-dependent model of breast cancer), where cSNX1.3 treatment resulted in significant tumor regression without observable toxicity. Evaluation of remaining tumor tissues found evidence of increased PARP cleavage, suggesting apoptotic tumor cell death. To evaluate the mechanism of action for cSNX1.3, we found that cSNX1.3 binds the C-terminus of the EGFR kinase domain at an interface site opposite the ATP binding domain with a Kd of ~4.0 µM. In vitro analysis found that cSNX1.3 inhibits the nuclear localization of EGFR. To determine specificity, we evaluated cancer cell lines expressing wildtype EGFR (MDA-MB-468, BT20 and A549), mutant EGFR (H1975) and non-transformed lines (CHO and MCF10A). Only transformed lines expressing wildtype EGFR responded to cSNX1.3, while mutant EGFR and normal cells responded better to an EGFR kinase inhibitor. Phenotypically, cSNX1.3 inhibits EGF-, NRG-, and HGF-dependent migration, but not HA-dependent migration. Together, these data indicate that targeting retrotranslocation of EGFR may be a potent therapeutic for RTK-active cancer.
• Schroeder, J. A., Stricklin, J., Tsutsumi, E., Peterson, E. A., & Kim, S. (2022). Cxcl10 Chemokine Induces Migration of ING4-Deficient Breast Cancer Cells via a Novel Cross Talk Mechanism between the Cxcr3 and Egfr Receptors. Molecular and Cellular Biology, 42(2). doi:10.1128/mcb.00382-21
• Tsutsumi, E., Stricklin, J., Peterson, E. A., Schroeder, J. A., & Kim, S. (2022). Cxcl10 Chemokine Induces Migration of ING4-Deficient Breast Cancer Cells via a Novel Cross Talk Mechanism between the Cxcr3 and Egfr Receptors. Molecular and cellular biology, 42(2), e0038221.
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  The chemokine Cxcl10 has been associated with poor prognosis in breast cancer, but the mechanism is not well understood. Our previous study has shown that was repressed by the ING4 tumor suppressor, suggesting a potential inverse functional relationship. We thus investigated a role for Cxcl10 in the context of ING4 deficiencies in breast cancer. We first analyzed public gene expression data sets and found that patients with -high/-low expressing tumors had significantly reduced disease-free survival in breast cancer. , Cxcl10 induced migration of -deleted breast cancer cells but not of -intact cells. Using inhibitors, we found that Cxcl10-induced migration of -deleted cells required Cxcr3, Egfr, and the Gβγ subunits downstream of Cxcr3 but not Gαi. Immunofluorescent imaging showed that Cxcl10 induced early transient colocalization between Cxcr3 and Egfr in both -intact and -deleted cells, which recurred only in -deleted cells. A peptide agent that binds to the internal juxtamembrane domain of Egfr inhibited Cxcr3/Egfr colocalization and cell migration. Taken together, these results presented a novel mechanism of Cxcl10 that elicits migration of -deleted cells, in part by inducing a physical or proximal association between Cxcr3 and Egfr and signaling downstream via Gβγ. These results further indicated that ING4 plays a critical role in the regulation of Cxcl10 signaling that enables breast cancer progression.
• Zohar, Y., Schroeder, J. A., Jiang, L., & Frankman, Z. D. (2022). Application of Microfluidic Systems for Breast Cancer Research.. Micromachines, 13(2), 152. doi:10.3390/mi13020152
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  Cancer is a disease in which cells in the body grow out of control; breast cancer is the most common cancer in women in the United States. Due to early screening and advancements in therapeutic interventions, deaths from breast cancer have declined over time, although breast cancer remains the second leading cause of cancer death among women. Most deaths are due to metastasis, as cancer cells from the primary tumor in the breast form secondary tumors in remote sites in distant organs. Over many years, the basic biological mechanisms of breast cancer initiation and progression, as well as the subsequent metastatic cascade, have been studied using cell cultures and animal models. These models, although extremely useful for delineating cellular mechanisms, are poor predictors of physiological responses, primarily due to lack of proper microenvironments. In the last decade, microfluidics has emerged as a technology that could lead to a paradigm shift in breast cancer research. With the introduction of the organ-on-a-chip concept, microfluidic-based systems have been developed to reconstitute the dominant functions of several organs. These systems enable the construction of 3D cellular co-cultures mimicking in vivo tissue-level microenvironments, including that of breast cancer. Several reviews have been presented focusing on breast cancer formation, growth and metastasis, including invasion, intravasation, and extravasation. In this review, realizing that breast cancer can recur decades following post-treatment disease-free survival, we expand the discussion to account for microfluidic applications in the important areas of breast cancer detection, dormancy, and therapeutic development. It appears that, in the future, the role of microfluidics will only increase in the effort to eradicate breast cancer.
• Maisel, S. A., Broka, D., Atwell, B., Bunch, T., Kupp, R., Singh, S. K., Mehta, S., & Schroeder, J. (2019). Stapled EGFR peptide reduces inflammatory breast cancer and inhibits additional HER-driven models of cancer. Journal of translational medicine, 17(1), 201.
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  The human epidermal growth factor receptor (HER) family of transmembrane tyrosine kinases is overexpressed and correlates with poor prognosis and decreased survival in many cancers. The receptor family has been therapeutically targeted, yet tyrosine kinase inhibitors (TKIs) do not inhibit kinase-independent functions and antibody-based targeting does not affect internalized receptors. We have previously demonstrated that a peptide mimicking the internal juxtamembrane domain of HER1 (EGFR; EJ1) promotes the formation of non-functional HER dimers that inhibit kinase-dependent and kinase-independent functions of HER1 (ERBB1/EGFR), HER2 (ERBB2) and HER3 (ERBB3). Despite inducing rapid HER-dependent cell death in vitro, EJ1 peptides are rapidly cleared in vivo, limiting their efficacy.
• Schroeder, J. A., & Maisel, S. (2018). Wrong Place at the Wrong Time: How Retrograde Trafficking of Receptor Tyrosine Kinases Drives Cancer Metastasis. Journal of Cancer Metastasis and Treatment.
• Schroeder, J. A., & Maisel, S. A. (2019). Wrong place at the wrong time: how retrograde trafficking drives cancer metastasis through receptor mislocalization. Journal of Cancer Metastasis and Treatment, 2019. doi:10.20517/2394-4722.2018.82
• Daniel, S. G., Russ, A. D., Guthridge, K. M., Raina, A. I., Estes, P. S., Parsons, L. M., Richardson, H. E., Schroeder, J. A., & Zarnescu, D. C. (2018). mediates the role of Lethal giant larvae as an epithelial growth inhibitor in. Biology open, 7(1).
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  () encodes a conserved tumor suppressor with established roles in cell polarity, asymmetric division, and proliferation control. Lgl's human orthologs, HUGL1 and HUGL2, are altered in human cancers, however, its mechanistic role as a tumor suppressor remains poorly understood. Based on a previously established connection between Lgl and Fragile X protein (FMRP), a miRNA-associated translational regulator, we hypothesized that Lgl may exert its role as a tumor suppressor by interacting with the miRNA pathway. Consistent with this model, we found thatis a dominant modifier of Argonaute1 overexpression in the eye neuroepithelium. Using microarray profiling we identified a core set of ten miRNAs that are altered throughout tumorigenesis inmutants. Among these are several miRNAs previously linked to human cancers including, which we found to be downregulated inneuroepithelial tissues. To determine whethercan act as an effector of Lgl, we overexpressed it in the context ofknock-down by RNAi and found it able to reduce the overgrowth phenotype caused by Lgl loss in epithelia. Furthermore, cross-comparisons between miRNA and mRNA profiling inmutant tissues and human breast cancer cells identified() as a common factor altered in both fly and human breast cancer tumorigenesis models. Our work provides the first evidence of a functional connection between Lgl and the miRNA pathway, demonstrates thatmediates Lgl's role in restricting epithelial proliferation, and provides novel insights into pathways controlled by Lgl during tumor progression.
• Maisel, S., Broka, D., & Schroeder, J. (2018). Intravesicular epidermal growth factor receptor subject to retrograde trafficking drives epidermal growth factor-dependent migration. Oncotarget, 9(5), 6463-6477.
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  The Epidermal Growth Factor Receptor (EGFR) is frequently mutated and overexpressed in metastatic cancer. Although EGFR is a transmembrane tyrosine kinase localized to the basolateral membrane in normal epithelium, it is frequently found intracellularly localized in transformed cells. We have previously demonstrated the epithelial adaptor protein mucin 1 (MUC1) alters trafficking of EGFR, inhibiting its degradation and promoting its translocation to the nucleus, where it can directly modulate gene transcription. Here, we demonstrate that MUC1 promotes the retention of EGF-bound EGFR in Early Endosome Antigen1 (EEA1)-positive vesicles while preventing its trafficking to the lysosome. These events result in the accumulation of endosomal vesicles harboring active receptor throughout the cell and a reorganization of the actin cytoskeleton. EGF-dependent cell migration and filopodia formation is reliant upon this altered trafficking, and can be prevented by blocking retrograde trafficking. Together, these results indicate that intracellular EGFR may play an essential role in cancer metastasis and a potential mechanism for the failure of therapeutic antibodies in EGFR-driven metastatic breast cancer.
• Chalasani, P., Segar, J., Pandey, R., Nagle, R. B., Lebeau, L. G., MacKerricher, W., Viscusi, R. K., Gonzalez, V., Schroeder, J. A., Livingston, R. B., & Baker, A. F. (2017). Clinicopathological and Molecular Characteristics of Pleomorphic Invasive Lobular Carcinoma. BMC Disease Markers.
• Chalasani, P., Segar, J., Pandey, R., Nagle, R. B., Lebeau, L. G., MacKerricher, W., Viscusi, R. K., Gonzalez, V., Schroeder, J. A., Livingston, R. B., & Baker, A. F. (2017). Clinicopathological and Molecular Characteristics of Pleomorphic Invasive Lobular Carcinoma. Breast Cancer Research.
• Schroeder, J., Maisel, S., & Broka, D. (2017). Intravesicular epidermal growth factor receptor subject to retrograde trafficking drives epidermal growth factor-dependent migration. Oncotarget, 9(5), 6463-6477. doi:10.18632/oncotarget.23766
• Zarnescu, D. C., Schroeder, J. A., Daniel, S. G., Russ, A. D., Guthridge, K. M., Raina, A. I., Estes, P. S., Parsons, L. M., & Richardson, H. E. (2017). miR-9a mediates the role of Lethal giant larvae as an epithelial growth inhibitor in Drosophila. Biology Open. doi:10.1242/bio.027391
• Buehler, J., Zeltzer, S., Reitsma, J., Petrucelli, A., Umashankar, M., Rak, M., Zagallo, P., Schroeder, J., Terhune, S., & Goodrum, F. (2016). Opposing Regulation of the EGF Receptor: A Molecular Switch Controlling Cytomegalovirus Latency and Replication. PLoS pathogens, 12(5), e1005655.
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  Herpesviruses persist indefinitely in their host through complex and poorly defined interactions that mediate latent, chronic or productive states of infection. Human cytomegalovirus (CMV or HCMV), a ubiquitous β-herpesvirus, coordinates the expression of two viral genes, UL135 and UL138, which have opposing roles in regulating viral replication. UL135 promotes reactivation from latency and virus replication, in part, by overcoming replication-suppressive effects of UL138. The mechanism by which UL135 and UL138 oppose one another is not known. We identified viral and host proteins interacting with UL138 protein (pUL138) to begin to define the mechanisms by which pUL135 and pUL138 function. We show that pUL135 and pUL138 regulate the viral cycle by targeting that same receptor tyrosine kinase (RTK) epidermal growth factor receptor (EGFR). EGFR is a major homeostatic regulator involved in cellular proliferation, differentiation, and survival, making it an ideal target for viral manipulation during infection. pUL135 promotes internalization and turnover of EGFR from the cell surface, whereas pUL138 preserves surface expression and activation of EGFR. We show that activated EGFR is sequestered within the infection-induced, juxtanuclear viral assembly compartment and is unresponsive to stress. Intriguingly, these findings suggest that CMV insulates active EGFR in the cell and that pUL135 and pUL138 function to fine-tune EGFR levels at the cell surface to allow the infected cell to respond to extracellular cues. Consistent with the role of pUL135 in promoting replication, inhibition of EGFR or the downstream phosphoinositide 3-kinase (PI3K) favors reactivation from latency and replication. We propose a model whereby pUL135 and pUL138 together with EGFR comprise a molecular switch that regulates states of latency and replication in HCMV infection by regulating EGFR trafficking to fine tune EGFR signaling.
• Greenwood, E., Maisel, S., Ebertz, D., Russ, A., Pandey, R., & Schroeder, J. (2016). Llgl1 prevents metaplastic survival driven by epidermal growth factor dependent migration. Oncotarget, 7(38), 60776-60792.
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  We have previously demonstrated that Llgl1 loss results in a gain of mesenchymal phenotypes and a loss of apicobasal and planar polarity. We now demonstrate that these changes represent a fundamental shift in cellular phenotype. Llgl1 regulates the expression of multiple cell identity markers, including CD44, CD49f, and CD24, and the nuclear translocation of TAZ and Slug. Cells lacking Llgl1 form mammospheres, where survival and transplantability is dependent upon the Epidermal Growth Factor Receptor (EGFR). Additionally, Llgl1 loss allows cells to grow in soft-agar and maintain prolonged survival as orthotopic transplants in NOD-SCIDmice. Lineage tracing and wound healing experiments demonstrate that mammosphere survival is due to enhanced EGF-dependent migration. The loss of Llgl1 drives EGFR mislocalization and an EGFR mislocalization point mutation (P667A) drives these same phenotypes, including activation of AKT and TAZ nuclear translocation. Together, these data indicate that the loss of Llgl1 results in EGFR mislocalization, promoting pre-neoplastic changes.
• Schroeder, J., Pandey, R., Greenwood, E., Maisel, S., Ebertz, D., & Russ, A. (2016). Llgl1 prevents metaplastic survival driven by epidermal growth factor dependent migration. Oncotarget, 7(38), 60776-60792. doi:10.18632/oncotarget.11320
• Zheng, X., Jiang, L., Schroeder, J., Stopeck, A., & Zohar, Y. (2014). Isolation of viable cancer cells in antibody-functionalized microfluidic devices. Biomicrofluidics, 8(2), 024119.
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  Microfluidic devices functionalized with EpCAM antibodies were utilized for the capture of target cancer cells representing circulating tumor cells (CTCs). The fraction of cancer cells captured from homogeneous suspensions is mainly a function of flow shear rate, and can be described by an exponential function. A characteristic shear rate emerges as the most dominant parameter affecting the cell attachment ratio. Utilizing this characteristic shear rate as a scaling factor, all attachment ratio results for various combinations of receptor and ligand densities collapsed onto a single curve described by the empirical formula. The characteristic shear rate increases with both cell-receptor and surface-ligand densities, and empirical formulae featuring a product of two independent cumulative distributions described well these relationships. The minimum detection limit in isolation of target cancer cells from binary mixtures was experimentally explored utilizing microchannel arrays that allow high-throughput processing of suspensions about 0.5 ml in volume, which are clinically relevant, within a short time. Under a two-step attachment/detachment flow rate, both high sensitivity (almost 1.0) and high specificity (about 0.985) can be achieved in isolating target cancer cells from binary mixtures even for the lowest target/non-target cell concentration ratio of 1:100 000; this is a realistic ratio between CTCs and white blood cells in blood of cancer patients. Detection of CTCs from blood samples was also demonstrated using whole blood from healthy donors spiked with cancer cells. Finally, the viability of target cancer cells released after capture was confirmed by observing continuous cell growth in culture.
• Hart, M. R., Su, H., Broka, D., Goverdhan, A., & Schroeder, J. A. (2013). Inactive ERBB receptors cooperate with reactive oxygen species to suppress cancer progression. Molecular Therapy, 21(11), 1996-2007.
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  PMID: 24081029;PMCID: PMC3831044;Abstract: The ERBB receptors are a family of heterodimerization partners capable of driving transformation and metastasis. While the therapeutic targeting of single receptors has proven efficacious, optimal targeting of this receptor family should target all oncogenic members simultaneously. The juxtamembrane domains of ERBB1, ERBB2, and ERBB3 are highly conserved and control various aspects of ERBB-dependent biology. In an effort to block those functions, we have targeted this domain with decoy peptides synthesized in tandem with a cell-penetrating peptide, termed EJ1. Treatment with EJ1 induces cell death, promotes the formation of inactive ERBB multimers, and results in simultaneous reduction of ERBB1, ERBB2, and ERBB3 activation. Treatment also results in the activation of myosin light chain-dependent cell blebbing while inactivating CaMKII signaling, coincident with the induction of cell death. EJ1 also directly translocates to mitochondria, correlating with a loss of mitochondrial membrane potential and production of reactive oxygen species. Finally, treatment of a mouse model of breast cancer with EJ1 results in the inhibition of tumor growth and metastasis without associated toxicities in normal cells. Overall, these data demonstrate that a portion of the ERBB jxm domain, when used as an intracellular decoy, can inhibit tumor growth and metastasis, representing a novel anticancer therapeutic. © The American Society of Gene and Cell Therapy.
• Horm, T. M., & Schroeder, J. A. (2013). MUC1 and metastatic cancer: Expression, function and therapeutic targeting. Cell Adhesion and Migration, 7(2), 187-198.
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  PMID: 23303343;PMCID: PMC3954031;Abstract: MUC1 is a transmembrane mucin that is often overexpressed in metastatic cancers and often used as a diagnostic marker for metastatic progression. The extracellular domain of MUC1 can serve as a ligand for stromal and endothelial cell adhesion receptors, and the cytoplasmic domain engages in several interactions that can result in increased migration and invasion, as well as survival. In this review, we address the role of MUC1 in metastatic progression by assessing clinical studies reporting MUC1 levels at various disease stages, reviewing mouse models utilized to study the role of MUC1 in metastatic progression, discuss mechanisms of MUC1 upregulation, and detail MUC1 protein interactions and signaling events. We review interactions between MUC1 and the extracellular environment, with proteins colocalized in the plasma membrane and/or cytoplasmic proteins, and summarize the role of MUC1 in the nucleus as a transcriptional cofactor. Finally, we review recent publications describing current therapies targeting MUC1 in patients with advanced disease and the stage of these therapies in preclinical development or clinical trials. © 2013 Landes Bioscience.
• Schroeder, J., Hart, M. R., Su, H., Broka, D., Goverdhan, A., & Schroeder, J. A. (2013). Inactive ERBB Receptors Cooperate With Reactive Oxygen Species To Suppress Cancer Progression. Molecular therapy : the journal of the American Society of Gene Therapy.
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  The ERBB receptors are a family of heterodimerization partners capable of driving transformation and metastasis. While the therapeutic targeting of single receptors has proven efficacious, optimal targeting of this receptor family should target all oncogenic members simultaneously. The juxtamembrane domains of ERBB1, ERBB2, and ERBB3 are highly conserved and control various aspects of ERBB-dependent biology. In an effort to block those functions, we have targeted this domain with decoy peptides synthesized in tandem with a cell-penetrating peptide, termed EJ1. Treatment with EJ1 induces cell death, promotes the formation of inactive ERBB multimers, and results in simultaneous reduction of ERBB1, ERBB2, and ERBB3 activation. Treatment also results in the activation of myosin light chain-dependent cell blebbing while inactivating CaMKII signaling, coincident with the induction of cell death. EJ1 also directly translocates to mitochondria, correlating with a loss of mitochondrial membrane potential and production of reactive oxygen species. Finally, treatment of a mouse model of breast cancer with EJ1 results in the inhibition of tumor growth and metastasis without associated toxicities in normal cells. Overall, these data demonstrate that a portion of the ERBB jxm domain, when used as an intracellular decoy, can inhibit tumor growth and metastasis, representing a novel anticancer therapeutic.Molecular Therapy (2013); doi:10.1038/mt.2013.196.
• Schroeder, J. A., Horm, T. M., Bitler, B. G., Broka, D. M., & Louderbough, J. M. (2012). MUC1 Drives c-Met–Dependent Migration and Scattering. Molecular Cancer Research, 10(12), 1544-1554. doi:10.1158/1541-7786.mcr-12-0296
• Schroeder, J., Horm, T. M., Bitler, B. G., Broka, D. M., Louderbough, J. M., & Schroeder, J. A. (2012). MUC1 drives c-Met-dependent migration and scattering. Molecular cancer research : MCR, 10(12).
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  The transmembrane mucin MUC1 is overexpressed in most ductal carcinomas, and its overexpression is frequently associated with metastatic progression. MUC1 can drive tumor initiation and progression via interactions with many oncogenic partners, including β-catenin, the EGF receptor (EGFR) and Src. The decoy peptide protein transduction domain MUC1 inhibitory peptide (PMIP) has been shown to inhibit the tumor promoting activities of MUC1 in breast and lung cancer, including cell growth and invasion, and its usage suppresses metastatic progression in mouse models of breast cancer. To further characterize the reduced metastasis observed upon PMIP treatment, we conducted motility assays and observed that PMIP inhibits cell motility of breast cancer cells. To determine the mechanism by which PMIP inhibits motility, we evaluated changes in global gene transcription upon PMIP treatment, and identified a number of genes with altered expression in response to PMIP. Among these genes is the metastatic mediator, c-Met, a transmembrane tyrosine kinase that can promote cell scattering, migration, and invasion. To further investigate the role of c-Met in MUC1-dependent metastatic events, we evaluated the effects of MUC1 expression and EGFR activation on breast cancer cell scattering, branching, and migration. We found that MUC1 strongly promoted all of these events and this effect was further amplified by EGF treatment. Importantly, the effect of MUC1 and EGF on these phenotypes was dependent upon c-Met activity. Overall, these results indicate that PMIP can block the expression of a key metastatic mediator, further advancing its potential use as a clinical therapeutic.
• Schroeder, J., Russ, A., Louderbough, J. M., Zarnescu, D., & Schroeder, J. A. (2012). Hugl1 and Hugl2 in mammary epithelial cells: polarity, proliferation, and differentiation. PloS one, 7(10).
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  Loss of epithelial polarity is described as a hallmark of epithelial cancer. To determine the role of Hugl1 and Hugl2 expression in the breast, we investigated their localization in human mammary duct tissue and the effects of expression modulation in normal and cancer cell lines on polarity, proliferation and differentiation. Expression of Hugl1 and Hugl2 was silenced in both MCF10A cells and Human Mammary Epithelial Cells and cell lines were grown in 2-D on plastic and in 3-D in Matrigel to form acini. Cells in monolayer were compared for proliferative and phenotypic changes while acini were examined for differences in size, ability to form a hollow lumen, nuclear size and shape, and localization of key domain-specific proteins as a measure of polarity. We detected overlapping but distinct localization of Hugl1 and Hugl2 in the human mammary gland, with Hugl1 expressed in both luminal and myoepithelium and Hugl2 largely restricted to myoepithelium. On a plastic surface, loss of Hugl1 or Hugl2 in normal epithelium induced a mesenchymal phenotype, and these cells formed large cellular masses when grown in Matrigel. In addition, loss of Hugl1 or Hugl2 expression in MCF10A cells resulted in increased proliferation on Matrigel, while gain of Hugl1 expression in tumor cells suppressed proliferation. Loss of polarity was also observed with knockdown of either Hugl1 or Hugl2, with cells growing in Matrigel appearing as a multilayered epithelium, with randomly oriented Golgi and multiple enlarged nuclei. Furthermore, Hugl1 knock down resulted in a loss of membrane identity and the development of cellular asymmetries in Human Mammary Epithelial Cells. Overall, these data demonstrate an essential role for both Hugl1 and Hugl2 in the maintenance of breast epithelial polarity and differentiated cell morphology, as well as growth control.
• Louderbough, J. M., & Schroeder, J. A. (2011). Understanding the dual nature of CD44 in breast cancer progression. Molecular cancer research : MCR, 9(12), 1573-86.
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  CD44 has been the subject of extensive research for more than 3 decades because of its role in breast cancer, in addition to many physiological processes, but interestingly, conflicting data implicate CD44 in both tumor suppression and tumor promotion. CD44 has been shown to promote protumorigenic signaling and advance the metastatic cascade. On the other hand, CD44 has been shown to suppress growth and metastasis. Histopathological studies of human breast cancer have correlated CD44 expression with both favorable and unfavorable clinical outcomes. In recent years, CD44 has garnered significant attention because of its utility as a stem cell marker and has surfaced as a potential therapeutic target, necessitating a greater understanding of CD44 in breast cancer. In this review, we attempt to unify the literature implicating CD44 in both tumor promotion and suppression, and explain its dualistic nature.
• Schroeder, J. A., & Louderbough, J. M. (2011). Understanding the Dual Nature of CD44 in Breast Cancer Progression. Molecular Cancer Research, 9(12), 1573-1586. doi:10.1158/1541-7786.mcr-11-0156
• Schroeder, J., Louderbough, J. M., Brown, J. A., Nagle, R. B., & Schroeder, J. A. (2011). CD44 Promotes Epithelial Mammary Gland Development and Exhibits Altered Localization during Cancer Progression. Genes & cancer, 2(8).
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  The basal cell layer has emerged as a critical player in cancer progression, and understanding the molecular contribution of specific cell types is important in treatment and prevention. The adhesion receptor CD44, which mediates epithelial-stromal and cell-cell interactions, has been shown to both promote and suppress tumor progression. To better understand the normal function of CD44, we have investigated its role in mouse mammary gland development and its expression in human breast and prostate cancer. We have found that CD44 is expressed in the myoepithelium of the developing mammary gland and modulates ductal development of FVB/N mice. The loss of CD44 results in defective luminal-myoepithelial cell-cell adhesion and promotes the mixing of luminal and myoepithelial layers, disrupting epithelial bilayer organization, and CD44-null mice experience delayed ductal outgrowth and impaired terminal end bud formation. The myoepithelial expression of CD44 is also relevant to its expression in cancer, as CD44 is expressed in the basal cells of early-stage breast and prostate cancer but exhibits altered localization with increasing tumorigenicity and is strongly expressed by tumor epithelium.
• Zheng, X., Cheung, L. S., Schroeder, J. A., Jiang, L., & Zohar, Y. (2011). A high-performance microsystem for isolating circulating tumor cells. Lab on a chip, 11(19), 3269-76.
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  A unique flow field pattern in a bio-functional microchannel is utilized to significantly enhance the performance of a microsystem developed for selectively isolating circulating tumor cells from cell suspensions. For high performance of such systems, disposal of maximum non-target species is just as important as retention of maximum target species; unfortunately, most studies ignore or fail to report this aspect. Therefore, sensitivity and specificity are introduced as quantitative criteria to evaluate the system performance enabling a direct comparison among systems employing different techniques. The newly proposed fluidic scheme combines a slow flow field, for maximum target-cell attachment, followed by a faster flow field, for maximum detachment of non-target cells. Suspensions of homogeneous or binary mixtures of circulating breast tumor cells, with varying relative concentrations, were driven through antibody-functionalized microchannels. Either EpCAM or cadherin-11 transmembrane receptors were targeted to selectively capture target cells from the suspensions. Cadherin-11-expressing MDA-MB-231 cancer cells were used as target cells, while BT-20 cells were used as non-target cells as they do not express cadherin-11. The attachment and detachment of these two cell lines are characterized, and a two-step attachment/detachment flow field pattern is implemented to enhance the system performance in capturing target cells from binary mixtures. While the system sensitivity remains high, above 0.95, the specificity increases from about 0.85 to 0.95 solely due to the second detachment step even for a 1 : 1000 relative concentration of the target cells.
• Zheng, X., Cheung, L. S., Schroeder, J. A., Jiang, L., & Zohar, Y. (2011). Cell receptor and surface ligand density effects on dynamic states of adhering circulating tumor cells. Lab on a chip, 11(20), 3431-9.
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  Dynamic states of cancer cells moving under shear flow in an antibody-functionalized microchannel are investigated experimentally and theoretically. The cell motion is analyzed with the aid of a simplified physical model featuring a receptor-coated rigid sphere moving above a solid surface with immobilized ligands. The motion of the sphere is described by the Langevin equation accounting for the hydrodynamic loadings, gravitational force, receptor-ligand bindings, and thermal fluctuations; the receptor-ligand bonds are modeled as linear springs. Depending on the applied shear flow rate, three dynamic states of cell motion have been identified: (i) free motion, (ii) rolling adhesion, and (iii) firm adhesion. Of particular interest is the fraction of captured circulating tumor cells, defined as the capture ratio, via specific receptor-ligand bonds. The cell capture ratio decreases with increasing shear flow rate with a characteristic rate. Based on both experimental and theoretical results, the characteristic flow rate increases monotonically with increasing either cell-receptor or surface-ligand density within certain ranges. Utilizing it as a scaling parameter, flow-rate dependent capture ratios for various cell-surface combinations collapse onto a single curve described by an exponential formula.
• Flowers, M., Schroeder, J. A., Borowsky, A. D., Besselsen, D. G., Thomson, C. A., Pandey, R., & Thompson, P. A. (2010). Pilot study on the effects of dietary conjugated linoleic acid on tumorigenesis and gene expression in PyMT transgenic mice. Carcinogenesis, 31(9), 1642-1649.
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  PMID: 20624750;PMCID: PMC2930807;Abstract: Conjugated linoleic acid (CLA) is a class of commercially available fatty acids that have been associated with anticancer properties in rodent models of chemical carcinogenesis. We conducted a pilot study to examine the antitumor effect of dietary CLA in a polyoma virus-middle T antigen (PyMT) mouse model of invasive breast cancer. Virgin 4-week-old PyMT mice were administered a mixed-isomer CLA diet (1% wt/wt) or control AIN-93G diet for 4 weeks (N=6 and 5, respectively) and tumor burden was assessed at 8 weeks of age. Thoracic mammary glands were prepared as whole mounts with other glands being formalin fixed and paraffin embedded for histology and immunohistochemistry (IHC). Total RNA was prepared for microarray and real-time reverse transcription-polymerase chain reaction analysis. Western blots were performed for protein expression analysis. Tumor incidence was significantly increased in CLA-treated animals compared with controls (P=0.009) and occurred with extensive lobular-alveolar expansion and loss of mammary adipose tissue. More than 100 genes were downregulated ≥-2-fold in the CLA-treated group compared with controls, including adipose-specific markers, as wells as cytoskeletal and adhesion-related genes. This was supported by dramatic decreases in the epithelial adherens E-cadherin and β-catenin as demonstrated by IHC. Taken together, these results suggest that dietary CLA affects the mammary stromal environment, leading to tumor progression and cellular expansion in the PyMT mouse model. Further studies of the potential for cancer promotion are needed, especially because mixed-isomer CLA formulations are sold commercially as a nutritional supplement. © The Author 2010. Published by Oxford University Press. All rights reserved.
• M., J., Lopez, J. I., & Schroeder, J. A. (2010). Matrix hyaluronan alters epidermal growth factor receptor-dependent cell morphology. Cell Adhesion and Migration, 4(1), 26-31.
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  PMID: 20009574;PMCID: PMC2852554;Abstract: EGFR, a critical regulator of oncogenic signaling during cancer progression, is capable of integrating multireceptor signaling pathways that promote metastasis. EGFRr is subject to regulatory cues from the extracellular matrix (ECM), of which hyaluronan (HA) is a major component. in mammary tumors, HA is deposited in the ECM where it functions in biomechanical support and modulates intracellular signaling. We utilized a 3D collagen system in which is either polymerized in collagen matrix or provided soluble in the media (sHA). Here we report that collagen-embedded (eHA) inhibits EGFR activation, filopodia formation, and cell spreading on a collagen matrix. These findings demonstrate a novel role for eHA as a protective molecule when encountered in the collagen matrix during cancer progression. © 2010 Landes Bioscience.
• Schroeder, J. A., Bitler, B. G., & Goverdhan, A. (2010). MUC1 regulates nuclear localization and function of the epidermal growth factor receptor. Journal of Cell Science, 123(10), 1716-1723. doi:10.1242/jcs.062661
• Schroeder, J., Bitler, B. G., & Schroeder, J. A. (2010). Anti-cancer therapies that utilize cell penetrating peptides. Recent patents on anti-cancer drug discovery, 5(2).
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  Cell penetrating peptides (CPPs) are 9-35mer cationic and/or amphipathic peptides that are rapidly internalized across cell membranes. Importantly, they can be linked to a variety of cargo, including anti-cancer therapeutics, making CPPs an efficient, effective and non-toxic mechanism for drug delivery. In this review, we discuss a number of CPP conjugated therapies (CTTs) that are either patented are in the progress of patenting, and show strong promise for clinical efficacy. The CTTs discussed here target a number of different processes specific to cancer progression, including proliferation, survival and migration. In addition, many of these CTTs also increase sensitivity to current anti-cancer therapy modalities, including radiation and other DNA damaging chemotherapies, thereby decreasing the toxic dosage required for effective treatment. Mechanistically, these CTTs function in a dominant-negative manner by blocking tumor-specific protein-protein interactions with the CPP-conjugated peptide or protein. The treatment of both cell lines and mouse models demonstrates that this method of molecular targeting results in equal if not greater efficacy than current standards of care, including DNA damaging agents and topoisomerase inhibitors. For the treatment of invasive carcinoma, these CTTs have significant clinical potential to deliver highly targeted therapies without sacrificing the patient's quality of life.
• Schroeder, J., Bitler, B. G., Goverdhan, A., & Schroeder, J. A. (2010). MUC1 regulates nuclear localization and function of the epidermal growth factor receptor. Journal of cell science, 123(Pt 10).
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  Alteration of protein trafficking and localization is associated with several diseases, including cystic fibrosis, breast cancer, colorectal cancer, leukemia and diabetes. Specifically, aberrant nuclear localization of the epidermal growth factor receptor (EGFR), a receptor tyrosine kinase, is a poor prognostic indicator in several epithelial carcinomas. It is now appreciated that in addition to signaling from the plasma membrane, EGFR also trafficks to the nucleus, and can directly bind the promoter regions of genes encoding cyclin D1 (CCND1) and B-Myb (MYBL2). We have previously established that loss of MUC1 in an EGFR-dependent transgenic mouse model of breast cancer correlates with the loss of cyclin D1 expression. Here, we provide evidence for a novel regulatory function of MUC1 in the trafficking and nuclear activity of EGFR. We found that MUC1 and EGFR interact in the nucleus of breast cancer cells, which promotes the accumulation of chromatin-bound EGFR. Additionally, the presence of MUC1 results in significant colocalization of EGFR and phosphorylated RNA polymerase II, indicating that MUC1 influences the association of EGFR with transcriptionally active promoter regions. Importantly, we found that the loss of MUC1 expression resulted in a decrease in the interaction between EGFR and the CCND1 promoter, which translated to a significant decrease in cyclin D1 protein expression. This data offers insights into a novel regulatory mechanism of EGFR nuclear function and could have important implications for evaluating nuclear localization in cancer.
• Cheung, L. S., Zheng, X., Stopa, A., Baygents, J. C., Guzman, R., Schroeder, J. A., Heimark, R. L., & Zohar, Y. (2009). Detachment of captured cancer cells under flow acceleration in a bio-functionalized microchannel. Lab on a chip, 9(12), 1721-31.
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  Attachment, deformation and detachment of N-cadherin expressing prostate and breast cancer cell lines in a functionalized microchannel under hydrodynamic loading have been studied. N-cadherin antibodies are immobilized on the microchannel surface to capture the target cancer cells, PC3N and MDA-MB-231-N, from a homogeneous cell suspension. Although difficult, a significant fraction of moving cells can be captured under a low flow rate. More than 90% of the target cells are captured after a certain incubation time under no flow condition. The mechanical response of a captured cancer cell to hydrodynamic flow field is investigated and, in particular, the effect of flow acceleration is examined. The observed cell deformation is dramatic under low acceleration, but is negligible under high acceleration. Consequently, the detachment of captured cells depends on both flow rate and flow acceleration. The flow rate required for cell detachment is a random variable that can be described by a log-normal distribution. Two flow acceleration limits have been identified for proper scaling of the flow rate required to detach captured cells. A time constant for the mechanical response of a captured cell, on the order of 1 min, has been identified for scaling the flow acceleration. Based on these acceleration limits and time constant, an exponential-like empirical model is proposed to predict the flow rate required for cell detachment as a function of flow acceleration.
• Schroeder, J., Bitler, B. G., Menzl, I., Huerta, C. L., Sands, B., Knowlton, W., Chang, A., & Schroeder, J. A. (2009). Intracellular MUC1 peptides inhibit cancer progression. Clinical cancer research : an official journal of the American Association for Cancer Research, 15(1).
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  During cancer progression, the oncoprotein MUC1 binds beta-catenin while simultaneously inhibiting the degradation of the epidermal growth factor receptor (EGFR), resulting in enhanced transformation and metastasis. The purpose of this study was to design a peptide-based therapy that would block these intracellular protein-protein interactions as a treatment for metastatic breast cancer.
• Schroeder, J. A., Bitler, B. G., Menzl, I., Huerta, C. L., Sands, B., Knowlton, W., & Chang, A. (2008). Intracellular MUC1 Peptides Inhibit Cancer Progression. Clinical Cancer Research, 15(1), 100-109. doi:10.1158/1078-0432.ccr-08-1745
• Schroeder, J. A., Louderbough, J. V., & Lopez, J. I. (2008). Go or Grow: How HA Controls the EGF Receptor. The FASEB Journal, 22(S1). doi:10.1096/fasebj.22.1_supplement.387.1
• Schroeder, J. A., Pochampalli, M. R., & Bitler, B. G. (2007). Transforming Growth Factor α–Dependent Cancer Progression Is Modulated by Muc1. Cancer Research, 67(14), 6591-6598. doi:10.1158/0008-5472.can-06-4518
• Schroeder, J., Pochampalli, M. R., Bitler, B. G., & Schroeder, J. A. (2007). Transforming growth factor alpha dependent cancer progression is modulated by Muc1. Cancer research, 67(14).
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  Transforming growth factor alpha (TGFalpha) is a potent inducer of cellular transformation, through its binding and activation of the epidermal growth factor receptor (EGFR). Previous studies in our laboratory showed that EGFR could also be affected by the glycoprotein MUC1, which inhibits ligand-stimulated degradation of EGFR in breast epithelial cell lines. To determine the effect of Muc1 expression on TGFalpha/EGFR-dependent breast transformation, we crossed the WAP-TGFalpha transgenic mouse model of breast cancer onto a Muc1-null background. We found that the loss of Muc1 expression dramatically affects mammary gland transformation and progression. Although 100% of WAP-TGFalpha/Muc1(+/+) mice form mammary gland tumors by 1 year, only 37% of WAP-TGFalpha/Muc1(-/-) form tumors by this time. This difference is also associated with a delay in onset, with a doubling of onset time observed in the WAP-TGFalpha/Muc1(-/-) compared with the WAP-TGFalpha/Muc1(+/+) mice. Analysis of signal transduction pathways revealed that activation of cyclin D1 expression is significantly suppressed in tumors derived from WAP-TGFalpha/Muc1(-/-) animals compared with those expressing Muc1. The loss of Muc1 expression also results in a significant inhibition in the formation of hyperplastic lesions during tumor progression. On the C57Bl/6 inbred background, pulmonary lesions were observed in 28 of 29 WAP-TGFalpha/Muc1(+/+) animals (including one metastatic pulmonary adenocarcinoma and multiple perivascular lymphomas), although none were detected in the WAP-TGFalpha/Muc1(-/-) animals. Together, these data indicate that Muc1 is an important modulator of TGFalpha-dependent tumor progression.
• Schroeder, J., Pochampalli, M. R., el Bejjani, R. M., & Schroeder, J. A. (2007). MUC1 is a novel regulator of ErbB1 receptor trafficking. Oncogene, 26(12).
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  ErbB receptors are key regulators of cell survival and growth in normal and transformed tissues. The oncogenic glycoprotein MUC1 is a binding partner and substrate for erbB1 and MUC1 expression can potentiate erbB-dependent signal transduction. After receptor activation, erbB1 is typically downregulated via an endocytic pathway that results in receptor degradation or recycling. We report here that MUC1 expression inhibits the degradation of ligand-activated erbB1. Through the use of both RNAi-mediated knock down and overexpression constructs of MUC1, we show that MUC1 expression inhibits erbB1 degradation after ligand treatment in breast epithelial cells. This MUC1-mediated protection against erbB1 degradation can increase total cellular pools of erbB1 over time. Biotinylation of surface proteins demonstrates that cell-surface associated erbB1 receptor is protected by MUC1 against ligand-induced degradation, although this is accompanied by an increase in erbB1 internalization. The MUC1-mediated protection against degradation occurs with a decrease in EGF-stimulated ubiquitination of erbB1, and an increase in erbB1 recycling. These data indicate that MUC1 expression is a potent regulator of erbB1 receptor stability upon activation and may promote transformation through the inhibition of erbB1 degradation.
• Lopez, J. I., Camenisch, T. D., Stevens, M. V., Sands, B. J., McDonald, J., & Schroeder, J. A. (2005). CD44 attenuates metastatic invasion during breast cancer progression. Cancer research, 65(15), 6755-63.
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  Metastatic invasion is the primary cause of breast cancer mortality, and adhesion receptors, such as CD44, are believed to be critical in this process. Historically, primary breast tumor epithelium has been investigated in isolation from other tissue components, leading to the common interpretation that CD44 and its primary ligand, hyaluronan, promote invasion. Here, we provide in vivo evidence showing CD44 antagonism to breast cancer metastasis. In a mouse model of spontaneously metastasizing breast cancer (MMTV-PyV mT), we found that loss of CD44 promotes metastasis to the lung. Localization studies, in combination with a novel hyaluronan synthase-GFP transgenic mouse, show a restricted pattern of expression for CD44 and hyaluronan. Whereas CD44 is expressed in tumor epithelium, hyaluronan synthase expression is restricted to stromal-associated cells. This distinct CD44 and hyaluronan pattern of distribution suggests a role for epithelial-stromal interaction in CD44 function. To define the relevance of this spatial regulation, we developed an in vitro invasion assay to emulate invasion into the extracellular matrix. Invasion of CD44-positive tumor cells was inhibited in hyaluronan-containing matrices, whereas blocking CD44-hyaluronan association increased invasion. Collectively, these data show that during breast cancer progression, hyaluronan-CD44 dynamics occurring through epithelial-stromal interactions are protective against metastasis.
• Schroeder, J. A., Camenisch, T. D., Lopez, J. I., Stevens, M. V., Sands, B. J., & McDonald, J. (2005). CD44 Attenuates Metastatic Invasion during Breast Cancer Progression. Cancer Research, 65(15), 6755-6763. doi:10.1158/0008-5472.can-05-0863
• Hattrup, C. L., Fernandez-Rodriguez, J., Schroeder, J. A., Hansson, G. C., & Gendler, S. J. (2004). MUC1 can interact with adenomatous polyposis coli in breast cancer. Biochemical and Biophysical Research Communications, 316(2), 364-369.
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  PMID: 15020226;Abstract: The MUC1 tumor antigen is overexpressed on most breast tumors and metastases. It interacts with signaling proteins such as the ErbB kinases and β-catenin, and is involved in mammary gland oncogenesis and tumor progression. Herein, we report a novel interaction between MUC1 and adenomatous polyposis coli (APC), a tumor suppressor involved in downregulating β-catenin signaling. Initially identified in colorectal cancer, APC is also downregulated in breast tumors and presumably involved in mammary carcinogenesis. MUC1 and APC co-immunoprecipitate from the ZR-75-1 human breast carcinoma cell line and co-localize in mouse mammary glands and tumors. These studies also indicate that the association of MUC1 and APC may be increased by epidermal growth factor stimulation. Intriguingly, the co-immunoprecipitation of MUC1 and APC increases in human breast tumors and metastases as compared to adjacent normal tissues, indicating that this association may play a role in the formation and progression of breast tumors. © 2004 Elsevier Inc. All rights reserved.
• Schroeder, J. A., Masri, A. A., Adriance, M. C., Tessier, J. C., Kotlarczyk, K. L., Thompson, M. C., & Gendler, S. J. (2004). MUC1 overexpression results in mammary gland tumorigenesis and prolonged alveolar differentiation. Oncogene, 23(34), 5739-5747.
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  PMID: 15221004;Abstract: MUC1 is a transmembrane mucin that was initially cloned from malignant mammary epithelial cells as a tumor antigen. More than 90% of human breast carcinomas overexpress MUC1. Numerous studies have demonstrated an interaction between MUC1 and other oncogenic proteins such as β-catenin, erbB receptors and c-Src, but a functional role for MUC1 in transformation has not been identified. We previously reported the development of transgenic mice that overexpress human MUC1 in the mouse mammary gland (MMTV-MUC1). Analysis of these transgenic mice at an early age demonstrated the ability of MUC1 to potentiate EGF-dependent activation of MAP kinase signaling pathways in the lactating mammary gland. We now report that multiparous MMTV-MUC1 transgenic mice stochastically develop unifocal mammary gland carcinomas late in life. Molecular analysis of these tumors shows a tumor-specific coimmunoprecipitation between MUC1 and β-catenin. Examination of the contralateral glands in MMTV-MUC1 transgenics demonstrates that the development of frank carcinomas is accompanied by a failure of multiparous glands to undergo postlactational involution. Furthermore, uniparous MMTV-MUC1 transgenic mice display decreased postlactational apoptosis, elevated whey acidic protein expression and aberrant pErk2 activation. These findings are the first to determine that MUC1 overexpression promotes in vivo transformation of the mammary gland.
• Schroeder, J. A., Adriance, M. C., Thompson, M. C., Camenisch, T. D., & Gendler, S. J. (2003). MUC1 alters β-catenin-dependent tumor formation and promotes cellular invasion. Oncogene, 22(9), 1324-1332.
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  PMID: 12618757;Abstract: MUC1 is aberrantly expressed in greater than 90% of all breast carcinomas, yet its function as a tumor antigen is not fully understood. Recently, studies have shown that MUC1 interacts with β-catenin, erbB receptors, src, GSK-3β and protein kinase Cδ, possibly in a complex that promotes the disassembly of adherens junctions and the invasion of cells. Here we show that the deletion of Muc1 expression from MMTV-Wnt-1 transgenic mice results in a significant increase in the time to mammary gland tumor onset. Analysis of MMTV-Wnt-1 tumors on a wild-type Muc1 background shows a tumor-specific complex formation between Muc1 and β-catenin that can be observed in both the membrane and the cytoplasm of transformed epithelium. Analysis of primary human adenocarcinomas revealed that this MUC1/β-catenin interaction occurs in both primary and metastatic tumors, but is dramatically increased in metastatic lesions. Addition of MUC1-cytoplasmic domain peptides to the invasive MDA-MB-468 and MDA-MB-231 cell lines increases their invasive capability, and these peptides colocalize with both β-catenin and the focal adhesion protein vinculin, primarily at sites of membrane invasion into a collagen matrix. These data indicate a potential mechanism for MUC1 promotion of invasive tumorigenesis in the breast through the modulation of β-catenin localization and subsequent cytoskeletal dynamics.
• Schroeder, J. A., Jackson, L. F., Lee, D. C., & Camenisch, T. D. (2003). Form and function of developing heart valves: coordination by extracellular matrix and growth factor signaling. Journal of molecular medicine (Berlin, Germany), 81(7), 392-403.
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  It is becoming clear that converging pathways coordinate early heart valve development and remodeling into functional valve leaflets. The integration of these pathways begins with macro and molecular interactions outside the cell in the extracellular matrix separating the myocardial and endocardial tissue components of the rudimentary heart. Such interactions regulate events at the cell surface through receptors, proteases, and other membrane molecules which in turn transduce signals into the cell. These signals trigger intracellular cascades that transduce cellular responses through both transcription factor and cofactor activation mediating gene induction or suppression. Chamber septation and valve formation occur from these coordinated molecular events within the endocardial cushions to sustain unidirectional blood flow and embryo viability. This review discusses the emerging connection between extracellular matrix and growth factor receptor signaling during endocardial cushion morphogenesis by highlighting the extracellular component, hyaluronan, and erbB receptor functions during early valve development.
• Camenisch, T. D., Schroeder, J. A., Bradley, J., Klewer, S. E., & McDonald, J. A. (2002). Heart-valve mesenchyme formation is dependent on hyaluronan-augmented activation of ErbB2-ErbB3 receptors. Nature medicine, 8(8), 850-5.
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  Heart septation and valve malformations constitute the most common anatomical birth defects. These structures arise from the endocardial cushions within the atrioventricular canal (AVC) through dynamic interactions between cushion cells and the extracellular matrix (termed cardiac jelly). Transformation of endothelial cells to mesenchymal cells is essential for the proper development of the AVC and subsequent septation and valve formation. Atrioventricular septal defects can result from incomplete endocardial cushion morphogenesis. We show that hyaluronan-deficient AVC explants from Has2(-/-) embryos, which normally lack mesenchyme formation, are rescued by heregulin treatment, which restores phosphorylation of ErbB2 and ErbB3. These events were blocked using a soluble ErbB3 molecule, as well as with an inhibitor of ErbB2, herstatin. We show further that ErbB3 is activated during hyaluronan treatment of Has2(-/-) explants. These data provide a link between extracellular matrix-hyaluronan and ErbB receptor activation during development of early heart-valve and septal mesenchyme.
• Schroeder, J. A., Adriance, M. C., McConnell, E. J., Thompson, M. C., Pockaj, B., & Gendler, S. J. (2002). ErbB-beta-catenin complexes are associated with human infiltrating ductal breast and murine mammary tumor virus (MMTV)-Wnt-1 and MMTV-c-Neu transgenic carcinomas. The Journal of biological chemistry, 277(25), 22692-8.
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  Simultaneous deregulation of both Wnt and ErbB growth factors has previously been shown to result in the cooperative induction of mammary gland tumors. Using the murine mammary tumor virus (MMTV)-Wnt-1 transgenic model of mammary carcinoma, we have identified an unvarying association between beta-catenin and epidermal growth factor receptor/c-Neu (ErbB1/ErbB2) heterodimers in mammary gland tumors, indicating a requirement for ErbB signaling in Wnt-mediated tumorigenesis. Expansion of these observations to a second transgenic model, MMTV-c-Neu, demonstrated similar tumor-specific interactions, including an ErbB1 ligand-inducible phosphorylation of both beta-catenin and c-Neu. Direct relevance of these findings to human breast cancer was established upon examination of a set of human infiltrating ductal breast adenocarcinoma and lymph node metastasis tissues taken at surgery. These data revealed increased levels of beta-catenin in tumors and metastases versus normal breast as well as an association between beta-catenin and c-Neu that measurably occurs only in neoplasia, most strongly in metastatic lesions. These studies have identified a seemingly indispensable interaction between beta-catenin and epidermal growth factor receptor/c-Neu heterodimers in Wnt-1-mediated breast tumorigenesis that may indicate a fundamental signaling event in human metastatic progression.
• Schroeder, J. A., Thompson, M. C., Gardner, M. M., & Gendler, S. J. (2001). Transgenic MUC1 interacts with epidermal growth factor receptor and correlates with mitogen-activated protein kinase activation in the mouse mammary gland. The Journal of biological chemistry, 276(16), 13057-64.
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  MUC1 is a large (>400 kDa), heavily glycosylated transmembrane protein that is aberrantly expressed on greater than 90% of human breast carcinomas and subsequent metastases. The precise function of MUC1 overexpression in tumorigenesis is unknown, although various domains of MUC1 have been implicated in cell adhesion, cell signaling, and immunoregulation. Stimulation of the MDA-MB-468 breast cancer line as well as mouse mammary glands with epidermal growth factor results in the co-immunoprecipitation of MUC1 with a tyrosine-phosphorylated protein of approximately 180 kDa. We have generated transgenic lines overexpressing full-length (MMF), cytoplasmic tail deleted (DeltaCT), or tandem repeat deleted (DeltaTR)-human MUC1 under the control of the mouse mammary tumor virus promoter to further examine the role of MUC1 in signaling and tumorigenesis. Immunoprecipitation experiments revealed that full-length transgenic MUC1 physically associates with all four erbB receptors, and co-localizes with erbB1 in the lactating gland. Furthermore, we detected a sharp increase in ERK1/2 activation in MUC1 transgenic mammary glands compared with Muc1 null and wild-type animals. These results point to a novel function of increased MUC1 expression, potentiation of erbB signaling through the activation of mitogenic MAP kinase pathways.
• Schroeder, J. A., Troyer, K. L., & Lee, D. C. (2000). Cooperative induction of mammary tumorigenesis by TGFα and Wnts. Oncogene, 19(28), 3193-3199.
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  PMID: 10918574;Abstract: We previously reported that multiparous WAP-TGFα transgenic mice develop mammary gland carcinomas with complete incidence. TGFα-induced tumors appear stochastically and with relatively long latency, indicating an additional requirement for other genetic alterations. To identify genes that cooperate with TGFα in mammary tumorigenesis, we used a retroviral insertion approach featuring a cloned and infectious hybrid MMTV (C3H/Mtv-1). Tumor latency was decreased approximately 30% in MMTV-infected WAP-TGFα transgenic animals compared to non-infected transgenic controls, and > 30% of the corresponding tumors displayed evidence of integrated C3H/Mtv-1 DNA. PCR-based analyses of DNAs from two virus-infected, transgenic tumors revealed integration of hybrid MMTV in 3' untranslated exons of the Wnt-1 or Wnt-3 oncogenes. Moreover, Northern blots confirmed dramatic induction of Wnt-1 or Wnt-3 transcripts in the respective tumors, indicating that MMTV integration resulted in activated expression of these genes. Semiquantitative RT-PCR analyses showed that overexpression of Wnt-1 or Wnt-3 was a common occurrence in MMTV-infected WAP-TGFα tumors, and some non-infected WAP-TGFα tumors also showed evidence of elevated Wnt-3 transcripts. Collectively, these results reveal cooperative induction of mammary gland tumorigenesis by simultaneous deregulation of EGF-like (TGFα) and Wnt growth factors.
• Schroeder, J. A., & Lee, D. C. (1998). Dynamic expression and activation of ERBB receptors in the developing mouse mammary gland. Cell Growth and Differentiation, 9(6), 451-464.
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  PMID: 9663464;Abstract: The complex system of ERBB receptors and ligands is implicated in growth and differentiation of the mammary gland. However, it has not been comprehensively examined in this dynamic tissue. Combined RNA and protein analyses of glands in different stages from virgin to involution revealed differential expression of the four ERBB receptors, as well as distinctive patterns of ERBB ligand expression that suggested specialized function. ERBB localization was linked to mammary gland function. Thus, in the virgin gland, ERBB1 and ERBB2 were colocalized to all major cell types during ductal morphogenesis but differentially localized in the mature gland. All four ERBB receptors were restricted to epithelia in the differentiated gland. Analyses of ERBB tyrosine phosphorylation provided strong evidence of interaction between the four receptors in this physiological context. Thus, exogenous EGF induced stage-dependent transphosphorylation of ERBB2-4 as well as ERBB1, whereas endogenous phosphorylation of all four receptors peaked in late pregnancy and lactation.
• Schroeder, J. A., & Lee, D. C. (1997). Transgenic mice reveal roles for TGFα and EGF receptor in mammary gland development and neoplasia. Journal of Mammary Gland Biology and Neoplasia, 2(2), 119-129.
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  PMID: 10882298;Abstract: Transforming growth factor-alpha (TGFα)4 and/or the EGF receptor (EGFR) are frequently overexpressed by human and rodent breast tumors, as well as tumor-derived cell lines. Additionally, various observations suggest a role for TGFα and the EGFR signaling system in normal mouse mammary gland development. Recently, several laboratories have established TGFα transgenic mice with which to study the role of this growth factor in normal and neoplastic mammary biology. Examination of these mice revealed that overexpression of TGFα has profound consequences for this tissue. Most strikingly, transgenic mice expressing TGFα under the control of tissue-specific and nonspecific promoters stochastically developed focal mammary tumors with an incidence and latency that was markedly affected by pregnancy. Most TGFα-induced tumors were well-differentiated adenomas/adenocarcinomas, although some were undifferentiated and locally invasive. Distant metastases were only occasionally observed. Administration of the genotoxic carcinogen, 7,12-dimethylbenzanthracene (DMBA), dramatically accelerated mammary tumorigenesis induced by the TGFα transgene, raising the possibility that TGFα acts as a promoter in this tissue. Mice harboring dual transgenes encoding TGFα and either wild-type ERBB2 or c-myc displayed markedly accelerated tumorigenesis compared to mice carrying any of the single transgenes alone, indicative of potent cooperativity. Moreover, tumorigenesis in the bitransgenic mice was less dependent on pregnancy, and tumors were generally more malignant in appearance. Finally, TGFα also affected mammary gland dynamics. TGFα transgenic mice consistently displayed precocious alveolar development, were variably impaired with respect to lactation, and showed markedly reduced postlactional involution. As a result, the glands of multiparous females accumulated hyperplastic lesions that generally resembled milk-producing alveoli. Limited data support the hypothesis that these lesions were precursors to TGFα-induced tumors. In summary, these various findings underscore the potential importance of TGFα for cellular differentiation and transformation in the mammary gland. They also establish TGFα transgenic mice as a powerful model with which to study the role of EGFR signaling molecules in this dynamic tissue. © 1997 Plenum Publishing Corporation.
• Berkowitz, E. A., Seroogy, K. B., Schroeder, J. A., Russell, W. E., Evans, E. P., Riedel, R. F., Phillips, H. K., Harrison, C. A., Lee, D. C., & Luetteke, N. C. (1996). Characterization of the mouse transforming growth factor gene: Its expression during eyelid development and in waved 1 tissues. Cell Growth and Differentiation, 7(9), 1271-1282.
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  PMID: 8877107;Abstract: The spontaneous mouse waved 1 (wa1) mutation is allelic with the transforming growth factor α (TGF-α) gene and produces phenotypes similar to those of TGF-α knockout mice. Here, we show that TGF-α mRNA and protein levels are measurable in wa1 tissues but reduced 5- to 30-fold relative to wild type. Because the wa1-coding sequence is identical to that of the normal mRNA, wa1 is not a null mutation. Nuclear run-on analyses revealed decreased transcription of the TGF-α gene in wa1 tissues, but the sequence of a 3.2- kb 5' flanking fragment containing the promoter was unaltered. Moreover, pulsed field gel electrophoresis analysis did not reveal alterations within 750 kb upstream or 350 kb downstream of the gene, and chromosome 6 was karyotypically normal. Hence, we speculate that the wa1 mutation may be subtle and/or reside at a greater distance from the TGF-α gene. TGF-α deficiency elicits a spectrum of variably penetrant eye anomalies in wa1 and knockout mice that are associated with open eyes at birth. We found that late-gestation wa1 and TGF-α-null embryos display a significant delay in eyelid closure, although the eyes of most embryos fuse prior to birth. In situ hybridization localized TGF-α expression to the advancing margins of the eyelid epithelium and epidermal growth factor receptor expression throughout the eyelid and corneal epithelia. These results suggest that eye problems observed in TGF-α-deficient adult mice arise from premature exposure and trauma to open eyes during or following parturition.
• Sandgren, E. P., Schroeder, J. A., Qui, T. H., Palmiter, R. D., Brinster, R. L., & Lee, D. C. (1995). Inhibition of mammary gland involution is associated with transforming growth factor α but not c-myc-induced tumorigenesis in transgenic mice. Cancer Research, 55(17), 3915-3927.
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  PMID: 7641211;Abstract: Deregulated expression of transforming growth factor a (TGF-α) or c-myc has been implicated in the genesis of human breast cancer. To better characterize the role of these molecules in this disease, we generated transgenic mice that express TGF-α or c-myc under control of the mouse whey acidic protein (WAP) promoter. We then compared the resulting mammary gland neoplasia in these mice and in previously described mice expressing a metallothionein-driven TGF-α transgene. Nonvirgin female mice in all transgenic lineages developed mammary tumors with 100% incidence but variable latency. Among TGF-α lines, mean survival time correlated with the level of transgene expression, and the average life spans of high-expressing WAP-TGF- α and WAP-c-myc mice were similarly reduced. The majority of TGF-α-induced tumors were relatively well-differentiated adenomas and adenocarcinomas; in contrast, WAP-c-myc tumors were poorly differentiated, solid carcinomas with a minority of adenocarcinomas. Most TGF-α- and all c-myc-induced tumors were transplantable, but lung metastases were infrequently observed in all transgenic lines. WAP-TGF-α-induced tumors, in marked contrast to those induced by WAP-c-myc, displayed frequent induction of cyclin D1 mRNA, suggesting that expression of this gene may complement that of TGF-α during mammary tumor development. Expression of TGF-α also induced precocious development of pregnant glands and delayed or inhibited mammary involution. As a result, multiparious MT-TGF-α and especially WAP-TGF-α females accumulated large numbers of hyperplastic alveolar nodules that resembled the more differentiated TGF-α-induced tumors. Finally, coexpression of WAP-c- myc and WAP-TGF-α transgenes markedly decreased tumor latency, increased tumor growth, and even induced mammary tumors in virgin female and male mice. These findings provide further evidence for the importance of deregulated TGF-α expression in multistage carcinogenesis, and they suggest that in the mammary gland the mechanism of TGF-α-induced transformation may depend on postlactational survival of differentiated epithelium. They also provide evidence of a potent tumorigenic collaboration between TGF-α and c-myc in mammary epithelium.
• Schluter, S. F., Schroeder, J., Wang, E., & Marchalonis, J. J. (1994). Recognition molecules and immunoglobulin domains in invertebrates. Annals of the New York Academy of Sciences, 712, 74-81.
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  PMID: 8192354;Abstract: We have used specific antibody probes to conserved antigenic motifs to identify and characterize immunoglobulin-related molecules in tunicates and a C-type lectin found in lamprey that is related to molecules found in tunicates and mammals. The tunicate immunoglobulin cross-reactive molecule (μCRM) reacts with antibodies raised to shark IgM heavy chains. Intact tunicate μCRM is a monomer of Ig light-chain-sized subunits and is oligoclonal by IEF. That this molecule is related to Ig is indicated both by immunochemical data and by peptide sequence homologies. The lamprey lectin is a large polymer (> 500,000 kDa) of 35-kDa and 60-kDa subunits. It appears to be related to C-type lectins as shown by peptide sequence homology and the requirement of Ca2+ for activity. Related molecules appear to be present in tunicates and mammals as shown by cross-reactivity of antibodies in Western blots with single bands from hemolymph and T-cell extracts.
• Akins, J. M., Schroeder, J. A., Brower, D. L., & Aposhian, H. V. (1992). Evaluation of Drosophila melanogaster as an alternative animal for studying the neurotoxicity of heavy metals. Biometals, 5(2), 111-120.
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  PMID: 1525477;Abstract: Heavy metals cause irreversible neurobehavioral damage in many developing mammals, but the mechanisms of this damage are unknown. The influence of three heavy metal compounds, triethyllead chloride, lead acetate and cadmium chloride, on lethality, development, behavior and learning was studied using the fruit fly, Drosophila melanogaster. This animal was used because it allows hundreds of subjects to be assayed very easily in individual experiments and because it is a system in which toxicological questions might be answered by using the techniques of modern molecular genetics. When triethyllead chloride, lead acetate or cadmium chloride was placed in the medium, the larval LC50 (± standard error) was found to be 0.090±0.004, 6.60±0.64 and 0.42±0.04 mm, respectively. Each of the tested compounds produced a dose-related delay in development. In particular, they caused a delay in the development of larvae to pupae. When larvae were reared on medium containing triethyllead chloride (0.06 mm), lead acetate (3.07 mm) or cadmium chloride (0.11 mm), phototaxis, locomotion and learning were not inhibited. Since significant neurobehavioral effects were not observed under the experimental conditions used, Drosophila does not appear to be an appropriate animal for the genetic dissection of such effects of heavy metals during development. © 1992 Rapid Communications of Oxford Ltd.
• Schroeder, J. A., Schluter, S. F., & Marchalonis, J. J. (1991). Isolation of a putative protoimmunoglobulin in Pyura hausteria. Developmental and Comparative Immunology, 15(SUPPL. 1), S76.

Proceedings Publications

• Jiang, L., Heimark, R. L., Baygents, J. C., Zohar, Y., Zohar, Y., Zheng, X. J., Schroeder, J. A., Jiang, L., Heimark, R. L., Guzman, R., Cheung, L. S., & Baygents, J. C. (2011). Dynamic states of adhering cancer cells under shear flow in an antibody-functionalized microchannel. In 2011 IEEE 24th International Conference on Micro Electro Mechanical Systems, 849-852.
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  Dynamic states of cancer cells moving under shear flow in an antibody-functionalized microchannel are investigated experimentally and theoretically. A simplified physical model was adopted to analyze the cell motion; it features a rigid sphere, with receptors on its surface, moving above a solid surface with distributed ligands. The cell motion is described by the Langevin equation where the hydrodynamic interactions, gravitational drift force, receptor-ligand binding force, and thermal fluctuations are all taken into account. The receptor-ligand bonds are modeled as Hookean springs. In this study, three dynamic states of cell motion have been identified: (i) free motion, (ii) rolling adhesion, and (iii) firm adhesion depending on the flow shear rate. The numerical simulations allow exploring effects of numerous parameters such as cell-receptor and surface-ligand density.
• Jiang, L., Zohar, Y., Zohar, Y., Zheng, X. J., Schroeder, J. A., Jiang, L., & Cheung, L. S. (2011). A high-performance microsystem for isolating viable circulating tumor cells. In 2011 16th International Solid-State Sensors, Actuators and Microsystems Conference, 226-229.
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  The attachment and detachment of target cancer cells from homogeneous and binary mixtures in antibody-functionalized microchannels have been studied experimentally. Under the same intermediate flow rate, the attachment rate was found to be higher, and detachment flow rate was lower, for cell lines expressing the target receptor at a higher level. For cells that do not express the target receptor, the attachment rate was much lower but did not diminish, due to non-specific binding, and the detachment rate was much higher. The bio-functional microfluidic system performance in selectively isolating target cells from binary mixtures is quantitatively characterized. While the system sensitivity is typically very high, almost 100%, the specificity is lower than 90%. Applying a unique flow scheme of a slow flow rate, for maximum capture of target cells, followed by a faster flow rate, for maximum removal of non-target cells, the specificity is enhanced to levels above 95%, even for mixtures with target cells present at 1∶1,000 relative concentration ratio.

Presentations

• Schroeder, J. A. (2021). Non-Kinase Functions of Receptor Tyrosine Kinases. MCB seminar series. UA.
• Schroeder, J. A., & Liu, R. W. (2020, Aug). Moving Forward, Life after Pandemic. UArizona Virtual Lecture Series. Online: The College of Science and the College of Social and Behavioral Sciences at the University of Arizona.
• Schroeder, J. A. (2018, August). Therapeutic implications of Kinase-independent ERBB functions Breast Cancer. Roche/Ventana Medical Research Forum.
• Schroeder, J. A. (2018, May). Targeting EGFR non-canonical function in inflammatory breast cancer. Mammary Gland Biology and Breast Cancer. Il Ciocco Italy: Gordon conference.
• Schroeder, J. A. (2018, November). Therapeutic implications of Kinase-independent ERBB functions Breast Cancer. Cancer Biology GIDP seminar series. UACC.
• Schroeder, J. A. (2018, spring). Breast cancer research and a novel therapeutic. College of Science Dean's Advisory Board. UA.
• Schroeder, J. A. (2014, spring). Intracellular Peptide Based Breast Cancer Therapeutics. PEGS: Peptide Therapeutics, Conquering Disease. Boston MA: PEGS.
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  Session Chair and speaker
• Schroeder, J. A. (2014, spring). The role of the Epidermal Growth Factor Receptor in Breast Cancer. College of Medicine - Phoenix seminar series. Phoenix AZ: College of Medicine.
• Schroeder, J. A. (2014, spring). The role of the Epidermal Growth Factor Receptor in Breast Cancer. Drug Discovery and Developmental Therapeutics Seminar Series. University of Arizona.