Browsing by Author "Yeh, Elizabeth S."

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    Correction: HUNK phosphorylates EGFR to regulate breast cancer metastasis
    (Springer Nature, 2021-05) Williams, Carly B.; Phelps-Polirer, Kendall; Dingle, Ivan P.; Williams, Christina J.; Rhett, Matthew J.; Eblen, Scott T.; Armeson, Kent; Hill, Elizabeth G.; Yeh, Elizabeth S.; Pharmacology and Toxicology, School of Medicine
    Erratum for 10.1038/s41388-019-1046-5
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    HUNK as a key regulator of tumor-associated macrophages in triple negative breast cancer
    (Taylor & Francis, 2024-06-05) Ramos Solis, Nicole; Cannon, Anthony; Dilday, Tinslee; Abt, Melissa; Oblak, Adrian L.; Soloff, Adam C.; Kaplan, Mark H.; Yeh, Elizabeth S.; Pharmacology and Toxicology, School of Medicine
    Triple-negative breast cancer (TNBC) lacks the expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). TNBC tumors are not sensitive to endocrine therapy, and standardized TNBC treatment regimens are lacking. TNBC is a more immunogenic subtype of breast cancer, making it more responsive to immunotherapy intervention. Tumor-associated macrophages (TAMs) constitute one of the most abundant immune cell populations in TNBC tumors and contribute to cancer metastasis. This study examines the role of the protein kinase HUNK in tumor immunity. Gene expression analysis using NanoString's nCounter PanCancer Immune Profiling panel identified that targeting HUNK is associated with changes in the IL-4/IL-4 R cytokine signaling pathway. Experimental analysis shows that HUNK kinase activity regulates IL-4 production in mammary tumor cells, and this regulation is dependent on STAT3. In addition, HUNK-dependent regulation of IL-4 secreted from tumor cells induces polarization of macrophages into an M2-like phenotype associated with TAMs. In return, IL-4 induces cancer metastasis and macrophages to produce epidermal growth factor. These findings delineate a paracrine signaling exchange between tumor cells and TAMs regulated by HUNK and dependent on IL-4/IL-4 R. This highlights the potential of HUNK as a target for reducing TNBC metastasis through modulation of the TAM population.
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    HUNK Gene Alterations in Breast Cancer
    (MDPI, 2022-11-29) Ramos-Solis, Nicole; Dilday, Tinslee; Kritikos, Alex E.; Yeh, Elizabeth S.; Pharmacology and Toxicology, School of Medicine
    Hormonally upregulated neu-associated kinase (HUNK) is a serine/threonine (S/T) protein kinase related to the adenosine monophosphate-activated protein kinase (AMPK) family of kinases. HUNK was originally discovered using a screen to identify kinases expressed in the mouse mammary gland. Therefore, the majority of studies to date have been carried out in models specific to this tissue, and the kinase was named to reflect its mammary gland-specific physiology and pathology. Prior studies show a clear pathogenic role for HUNK in breast cancer. HUNK is upregulated in response to oncogenic HER2/neu and Akt, and there is strong evidence that HUNK is critical for the survival of breast cancer cells. Further evidence shows that inhibiting HUNK using a variety of breast cancer models, including those that are resistant, inhibits tumorigenesis and metastasis. However, HUNK alterations are infrequent. Here, the incidence and consequence of HUNK alterations in breast cancer is reviewed using data mined from the online database cBioPortal and considered in relation to prior research studies.
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    HUNK phosphorylates EGFR to regulate breast cancer metastasis
    (Springer Nature, 2019-10-09) Williams, Carly B.; Phelps-Polirer, Kendall; Dingle, Ivan P.; Williams, Christina J.; Rhett, Matthew J.; Eblen, Scott T.; Armeson, Kent; Hill, Elizabeth G.; Yeh, Elizabeth S.; Pharmacology and Toxicology, School of Medicine
    Epidermal growth factor receptor (EGFR) is commonly over-expressed in metastatic breast cancer yet metastatic breast cancer is generally resistant to anti-EGFR therapies, and the mechanism for resistance to EGFR inhibitors in this setting is not fully understood. Hormonally up-regulated neu-associated kinase (HUNK) kinase is up-regulated in aggressive breast cancers and is thought to play a role in breast cancer metastasis. However, no studies have been conducted to examine a relationship between EGFR and HUNK in breast cancer metastasis. We performed a kinase substrate screen and identified that EGFR is phosphorylated by HUNK. Our studies show that HUNK phosphorylates EGFR at T654, enhancing receptor stability and downstream signaling. We found that increased phosphorylation of T654 EGFR correlates with increased epithelial to mesenchymal, migration and invasion, and metastasis. In addition, we found that HUNK expression correlates with overall survival and distant metastasis free survival. This study shows that HUNK directly phosphorylates EGFR at T654 to promote metastasis and is the first study to show that the phosphorylation of this site in EGFR regulates metastasis.
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    HUNK Phosphorylates Rubicon to Support Autophagy
    (MDPI, 2019-11-19) Zambrano, Joelle N.; Eblen, Scott T.; Abt, Melissa; Rhett, J. Matthew; Muise-Helmericks, Robin; Yeh, Elizabeth S.; Pharmacology and Toxicology, School of Medicine
    Background: Autophagy is a catabolic cellular recycling pathway that is essential for maintaining intracellular homeostasis. Autophagosome formation is achieved via the coordination of the Beclin-1 protein complex. Rubicon is a Beclin-1 associated protein that suppresses autophagy by impairing the activity of the class III PI3K, Vps34. However, very little is known about the molecular mechanisms that regulate Rubicon function. Methods: In this study, co-immunoprecipitation and kinase assays were used to investigate the ability of Hormonally Upregulated Neu-associated Kinase (HUNK) to bind to and phosphorylate Rubicon. LC3B was monitored by immunofluorescence and immunoblotting to determine whether phosphorylation of Rubicon by HUNK controls the autophagy suppressive function of Rubicon. Results: Findings from this study identify Rubicon as a novel substrate of HUNK and show that phosphorylation of Rubicon inhibits its function, promoting autophagy.
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    Identification and characterization of a potent and selective HUNK inhibitor for treatment of HER2+ breast cancer
    (Elsevier, 2024) Dilday, Tinslee; Abt, Melissa; Ramos-Solís, Nicole; Dayal, Neetu; Larocque, Elizabeth; Oblak, Adrian L.; Sintim, Herman O.; Yeh, Elizabeth S.; Pharmacology and Toxicology, School of Medicine
    Human epidermal growth factor receptor 2 (HER2)-targeted agents have proven to be effective, however, the development of resistance to these agents has become an obstacle in treating HER2+ breast cancer. Evidence implicates HUNK as an anti-cancer target for primary and resistant HER2+ breast cancers. In this study, a selective inhibitor of HUNK is characterized alongside a phosphorylation event in a downstream substrate of HUNK as a marker for HUNK activity in HER2+ breast cancer. Rubicon has been established as a substrate of HUNK that is phosphorylated at serine (S) 92. Findings indicate that HUNK-mediated phosphorylation of Rubicon at S92 promotes both autophagy and tumorigenesis in HER2/neu+ breast cancer. HUNK inhibition prevents Rubicon S92 phosphorylation in HER2/neu+ breast cancer models and inhibits tumorigenesis. This study characterizes a downstream phosphorylation event as a measure of HUNK activity and identifies a selective HUNK inhibitor that has meaningful efficacy toward HER2+ breast cancer.
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    Isoforms of Neuropilin-2 Denote Unique Tumor-Associated Macrophages in Breast Cancer
    (Frontiers Media, 2022-04-27) Dhupar, Rajeev; Jones, Katherine E.; Powers, Amy A.; Eisenberg, Seth H.; Ding, Kai; Chen, Fangyuan; Nasarre, Cecile; Cen, Zhanpeng; Gong, Yi-Nan; LaRue, Amanda C.; Yeh, Elizabeth S.; Luketich, James D.; Lee, Adrian V.; Oesterreich, Steffi; Lotze, Michael T.; Gemmill, Robert M.; Soloff, Adam C.; Pharmacology and Toxicology, School of Medicine
    Tumor-associated macrophages (TAMs) exert profound influence over breast cancer progression, promoting immunosuppression, angiogenesis, and metastasis. Neuropilin-2 (NRP2), consisting of the NRP2a and NRP2b isoforms, is a co-receptor for heparin-binding growth factors including VEGF-C and Class 3 Semaphorins. Selective upregulation in response to environmental stimuli and independent signaling pathways endow the NRP2 isoforms with unique functionality, with NRP2b promoting increased Akt signaling via receptor tyrosine kinases including VEGFRs, MET, and PDGFR. Although NRP2 has been shown to regulate macrophage/TAM biology, the role of the individual NRP2a/NRP2b isoforms in TAMs has yet to be evaluated. Using transcriptional profiling and spectral flow cytometry, we show that NRP2 isoform expression was significantly higher in TAMs from murine mammary tumors. NRP2a/NRP2b levels in human breast cancer metastasis were dependent upon the anatomic location of the tumor and significantly correlated with TAM infiltration in both primary and metastatic breast cancers. We define distinct phenotypes of NRP2 isoform-expressing TAMs in mouse models of breast cancer and within malignant pleural effusions from breast cancer patients which were exclusive of neuropilin-1 expression. Genetic depletion of either NRP2 isoform in macrophages resulted in a dramatic reduction of LPS-induced IL-10 production, defects in phagosomal processing of apoptotic breast cancer cells, and increase in cancer cell migration following co-culture. By contrast, depletion of NRP2b, but not NRP2a, inhibited production of IL-6. These results suggest that NRP2 isoforms regulate both shared and unique functionality in macrophages and are associated with distinct TAM subsets in breast cancer.
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    Mapping the Anti-Cancer Activity of α-Connexin Carboxyl-Terminal (aCT1) Peptide in Resistant HER2+ Breast Cancer
    (MDPI, 2024-01-19) Baker, Kimberly M.; Abt, Melissa; Doud, Emma H.; Oblak, Adrian L.; Yeh, Elizabeth S.; Pharmacology and Toxicology, School of Medicine
    Connexin 43 (Cx43) is a protein encoded by the GJA1 gene and is a component of cell membrane structures called gap junctions, which facilitate intercellular communication. Prior evidence indicates that elevated GJA1 expression in the HER2-positive (HER2+) subtype of breast cancer is associated with poor prognosis. Prior evidence also suggests that HER2+ breast cancers that have become refractory to HER2-targeted agents have a loss of Cx43 gap junction intercellular communication (GJIC). In this study, a Cx43-targeted agent called alpha-connexin carboxyl-terminal peptide (aCT1) is examined to determine whether GJIC can be rescued in refractory HER2+ breast cancer cells. A proposed mechanism of action for aCT1 is binding to the tight junction protein Zonal Occludens-1 (ZO-1). However, the true scope of activity for aCT1 has not been explored. In this study, mass spectrometry proteomic analysis is used to determine the breadth of aCT1-interacting proteins. The NanoString nCounter Breast Cancer 360 panel is also used to examine the effect of aCT1 on cancer signaling in HER2+ breast cancer cells. Findings from this study show a dynamic range of binding partners for aCT1, many of which regulate gene expression and RNA biology. nCounter analysis shows that a number of pathways are significantly impacted by aCT1, including upregulation of apoptotic factors, leading to the prediction and demonstration that aCT1 can boost the cell death effects of cisplatin and lapatinib in HER2+ breast cancer cells that have become resistant to HER2-targeted agents.
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    Mass Spectrometry-Based Glycoproteomic Workflows for Cancer Biomarker Discovery
    (Sage, 2023) Doud, Emma H.; Yeh, Elizabeth S.; Biochemistry and Molecular Biology, School of Medicine
    Glycosylation has a clear role in cancer initiation and progression, with numerous studies identifying distinct glycan features or specific glycoproteoforms associated with cancer. Common findings include that aggressive cancers tend to have higher expression levels of enzymes that regulate glycosylation as well as glycoproteins with greater levels of complexity, increased branching, and enhanced chain length1. Research in cancer glycoproteomics over the last 50-plus years has mainly focused on technology development used to observe global changes in glycosylation. Efforts have also been made to connect glycans to their protein carriers as well as to delineate the role of these modifications in intracellular signaling and subsequent cell function. This review discusses currently available techniques utilizing mass spectrometry-based technologies used to study glycosylation and highlights areas for future advancement.
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    NF-κB Signaling Is Regulated by Fucosylation in Metastatic Breast Cancer Cells
    (MDPI, 2020-12-12) Doud, Emma H.; Shetty, Trupti; Abt, Melissa; Mosley, Amber L.; Corson, Timothy W.; Mehta, Anand; Yeh, Elizabeth S.; Biochemistry and Molecular Biology, School of Medicine
    A growing body of evidence indicates that the levels of fucosylation correlate with breast cancer progression and contribute to metastatic disease. However, very little is known about the signaling and functional outcomes that are driven by fucosylation. We performed a global proteomic analysis of 4T1 metastatic mammary tumor cells in the presence and absence of a fucosylation inhibitor, 2-fluorofucose (2FF). Of significant interest, pathway analysis based on our results revealed a reduction in the NF-κB and TNF signaling pathways, which regulate the inflammatory response. NF-κB is a transcription factor that is pro-tumorigenic and a prime target in human cancer. We validated our results, confirming that treatment of 4T1 cells with 2FF led to a decrease in NF-κB activity through increased IκBα. Based on these observations, we conclude that fucosylation is an important post-translational modification that governs breast cancer cell signaling.