Macomb Community College

Adjunct Professor

Arts and Sciences Department
Macomb Community College South Campus

• Knowledgeable curriculum designer and developer using backward design and learning outcomes for assessments and activities in an Introductory lecture and laboratory Biology course.
• Strong leadership and technological skills resulting in being chosen as the first adjunct to teach Anatomy and Physiology using the Willey-Plus publisher web-based learning platform.

Marygrove College

Adjunct Professor

Science and Math Department
Marygrove College

• Strong team player and communicator working with other adjunct and full-time faculty leading to uniformity among assessments in 2 sections of an Introductory Biology course.
• Excellent resource and time management skills responsible for the development and organization of all sections of Introductory Biology laboratories resulting in newer inquiry-based lab exercises.
• Strong leadership skills resulting in being chosen as first adjunct to co-teach Introduction to Organic and Biochemistry, focusing on the Biochemistry portion of the course.
• Competent technological skills utilizing online materials from McGraw Hill Connect websites for Biology and Biochemistry resulting in decreased assessment development and grading time.

National Science Foundation (NSF)

Postdoctoral Teaching Fellow and Intermittent Lecturer

NSF FIRST IV Fellowship, Co-Principle Investigators of the NSF grant: Diane Ebert-May, Ph.D. (Michigan State University, East Lansing, MI) &Terry Derting, Ph.D. (Murray State University, Murray, KY
• Strong development of professional skills after being chosen to participate in a competitive program for postdoctoral fellows interested in STEM education reform.
• Attended two, all expenses paid summer workshops, designed and taught Human Genetics for the Molecular Cellular and Developmental Biology Department, University of Michigan, Ann Arbor, MI (Summer 2011), and received feedback on teaching experience.

Oakland University

Visiting Assistant Professor

Interdisciplinary Health
Oakland University School of Health Sciences

• High level educator with diverse teaching and curriculum development on the prevention, management, and treatment of diseases resulting in the training of Health Science students.
• Excellent interdisciplinary collaboration skills demonstrated working in cross functional teams to design assessments and train 6 undergraduate student graders to maximize grading efficiency.
• Expanded professional awareness and development in evidence-based learning in STEM fields including: flipping the classroom, adult learning theory, active and problem-based learning resulting in a learner-centered approach to teaching.
• Strong project management skills in community research demonstrated by planning 5 individual and one large community class project for the Parks and Recreation Department of Pontiac, MI.
• Passionate presentation and communication skills demonstrated at oral sessions at local and national conferences including awarding of a travel grant to a Lilly teaching conference ($1,200).

University of Michigan - Research and Sponsored Projects

Project Manager and Medical Writer Experience

Postdoctoral Fellow
University of Michigan Comprehensive Cancer Center
Principle Investigator: Max Wicha, M.D.
Projects: Activation and Tumorigenesis of Notch Stem Cell Pathway in Breast Cancer Cells and Identification of Cancer Stem Cell Biomarkers in Normal Mammary Tissue

• Excellent teamwork and project management skills working on several multifaceted research projects resulting in two first author journal publications, 3 contributing author publications, and
National Cancer Institute (NCI) Research Training (T32) Fellowship, $42,000/year.
o Member of the Stand Up to Cancer Dream Team resulting in the sharing of $73 million dollars towards new treatments from lab to cancer patient.
• Relationship builder with Merck to conduct preclinical tumor imaging studies using cell lines and NOD/SCID mice for drug development of a Gamma Secretase Inhibitor (GSI) against the Notch pathway leading to establishment of new therapeutic combinations against breast cancer.
• High-level technical and problem-solving skills demonstrated in the process of generating cells from reduction mammoplasties resulting in the development of complex multi-color flow cytometry biomarker protocols.
• Strong leadership skills demonstrated in the teaching of daily research activities, project development, and writing of fellowship applications leading to specialized training of undergraduate students, visiting medical fellows, researchers, and pathologists.
• Knowledge of current industry trends and experience communicating complex topics to diverse audiences resulting in presentations at international meetings like AACR.

Turning Point

Education and counseling for victims of rape and violence.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Mol Cell

Abstract Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Mol Cell

Abstract Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

SOCS3-mediated regulation of inflammatory cytokines in PTEN and p53 inactivated triple negative breast cancer model.

Oncogene

Abstract Somatic mutations or deletions of TP53 and PTEN in ductal carcinoma in situ lesions have been implicated in progression to invasive ductal carcinomas. A recent molecular and mutational analysis of breast cancers revealed that inactivation of tumor suppressors, p53 and PTEN, are strongly associated with triple negative breast cancer. In addition, these tumor suppressors have important roles in regulating self-renewal in normal and malignant stem cells. To investigate their role in breast carcinogenesis, we knocked down these genes in human mammary cells and in non-transformed MCF10A cells. p53 and PTEN knockdown synergized to activate pro-inflammatory interleukin-6 (IL6)/Stat3/nuclear factor κB signaling. This resulted in generation of highly metastatic epithelial-to-mesenchymal transition-like cancer stem cells resulting in tumors whose gene expression profile mimicked that found in basal/claudin-low molecular subtype within the triple negative breast tumors. Constitutive activation of this loop in transformed cells was dependent on proteolytic degradation of suppressor of cytokine signaling 3 (SOCS3) resulting in low levels of this protein in basal/claudin-low cell lines and primary tumors. In non-transformed cells, transient activation of the IL6 inflammatory loop induced SOCS3 expression leading to pathway inactivation. In transformed cells, enforced expression of SOCS3 or interfering with IL6 pathway via IL6R blockade inhibited tumor growth and metastasis in mouse xenograft models. Furthermore, circulating tumor cells were significantly reduced in tumor-bearing animals when treated with anti-IL6R antibodies. These studies uncover important connections between inflammation and carcinogenesis and suggest that blocking pro-inflammatory cytokines may be utilized as an attractive strategy to target triple negative breast tumors, which currently lacks molecularly targeted therapies.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Mol Cell

Abstract Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

SOCS3-mediated regulation of inflammatory cytokines in PTEN and p53 inactivated triple negative breast cancer model.

Oncogene

Abstract Somatic mutations or deletions of TP53 and PTEN in ductal carcinoma in situ lesions have been implicated in progression to invasive ductal carcinomas. A recent molecular and mutational analysis of breast cancers revealed that inactivation of tumor suppressors, p53 and PTEN, are strongly associated with triple negative breast cancer. In addition, these tumor suppressors have important roles in regulating self-renewal in normal and malignant stem cells. To investigate their role in breast carcinogenesis, we knocked down these genes in human mammary cells and in non-transformed MCF10A cells. p53 and PTEN knockdown synergized to activate pro-inflammatory interleukin-6 (IL6)/Stat3/nuclear factor κB signaling. This resulted in generation of highly metastatic epithelial-to-mesenchymal transition-like cancer stem cells resulting in tumors whose gene expression profile mimicked that found in basal/claudin-low molecular subtype within the triple negative breast tumors. Constitutive activation of this loop in transformed cells was dependent on proteolytic degradation of suppressor of cytokine signaling 3 (SOCS3) resulting in low levels of this protein in basal/claudin-low cell lines and primary tumors. In non-transformed cells, transient activation of the IL6 inflammatory loop induced SOCS3 expression leading to pathway inactivation. In transformed cells, enforced expression of SOCS3 or interfering with IL6 pathway via IL6R blockade inhibited tumor growth and metastasis in mouse xenograft models. Furthermore, circulating tumor cells were significantly reduced in tumor-bearing animals when treated with anti-IL6R antibodies. These studies uncover important connections between inflammation and carcinogenesis and suggest that blocking pro-inflammatory cytokines may be utilized as an attractive strategy to target triple negative breast tumors, which currently lacks molecularly targeted therapies.

TIMP-1 via TWIST1 Induces EMT Phenotypes in Human Breast Epithelial Cells

Mol Cancer Res

Abstract Tissue inhibitor of metalloproteinase-1 (TIMP-1) regulates intracellular signaling networks for inhibition of apoptosis. Tetraspanin (CD63), a cell surface binding partner for TIMP-1, was previously shown to regulate integrin-mediated survival pathways in the human breast epithelial cell line MCF10A. In the current study, we show that TIMP-1 expression induces phenotypic changes in cell morphology, cell adhesion, cytoskeletal remodeling, and motility, indicative of an epithelial-mesenchymal transition (EMT). This is evidenced by loss of the epithelial cell adhesion molecule E-cadherin with an increase in the mesenchymal markers vimentin, N-cadherin, and fibronectin. Signaling through TIMP-1, but not TIMP-2, induces the expression of TWIST1, an important EMT transcription factor known to suppress E-cadherin transcription, in a CD63-dependent manner. RNAi-mediated knockdown of TWIST1 rescued E-cadherin expression in TIMP-1-overexpressing cells, demonstrating a functional significance of TWIST1 in TIMP-1-mediated EMT. Furthermore, analysis of TIMP-1 structural mutants reveals that TIMP-1 interactions with CD63 that activate cell survival signaling and EMT do not require the matrix metalloproteinase (MMP)-inhibitory domain of TIMP-1. Taken together, these data demonstrate that TIMP-1 binding to CD63 activates intracellular signal transduction pathways, resulting in EMT-like changes in breast epithelial cells, independent of its MMP-inhibitory function. IMPLICATIONS: TIMP-1's function as an endogenous inhibitor of MMP or as a "cytokine-like" signaling molecule may be a critical determinant for tumor cell behavior.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Mol Cell

Abstract Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

SOCS3-mediated regulation of inflammatory cytokines in PTEN and p53 inactivated triple negative breast cancer model.

Oncogene

Abstract Somatic mutations or deletions of TP53 and PTEN in ductal carcinoma in situ lesions have been implicated in progression to invasive ductal carcinomas. A recent molecular and mutational analysis of breast cancers revealed that inactivation of tumor suppressors, p53 and PTEN, are strongly associated with triple negative breast cancer. In addition, these tumor suppressors have important roles in regulating self-renewal in normal and malignant stem cells. To investigate their role in breast carcinogenesis, we knocked down these genes in human mammary cells and in non-transformed MCF10A cells. p53 and PTEN knockdown synergized to activate pro-inflammatory interleukin-6 (IL6)/Stat3/nuclear factor κB signaling. This resulted in generation of highly metastatic epithelial-to-mesenchymal transition-like cancer stem cells resulting in tumors whose gene expression profile mimicked that found in basal/claudin-low molecular subtype within the triple negative breast tumors. Constitutive activation of this loop in transformed cells was dependent on proteolytic degradation of suppressor of cytokine signaling 3 (SOCS3) resulting in low levels of this protein in basal/claudin-low cell lines and primary tumors. In non-transformed cells, transient activation of the IL6 inflammatory loop induced SOCS3 expression leading to pathway inactivation. In transformed cells, enforced expression of SOCS3 or interfering with IL6 pathway via IL6R blockade inhibited tumor growth and metastasis in mouse xenograft models. Furthermore, circulating tumor cells were significantly reduced in tumor-bearing animals when treated with anti-IL6R antibodies. These studies uncover important connections between inflammation and carcinogenesis and suggest that blocking pro-inflammatory cytokines may be utilized as an attractive strategy to target triple negative breast tumors, which currently lacks molecularly targeted therapies.

TIMP-1 via TWIST1 Induces EMT Phenotypes in Human Breast Epithelial Cells

Mol Cancer Res

Abstract Tissue inhibitor of metalloproteinase-1 (TIMP-1) regulates intracellular signaling networks for inhibition of apoptosis. Tetraspanin (CD63), a cell surface binding partner for TIMP-1, was previously shown to regulate integrin-mediated survival pathways in the human breast epithelial cell line MCF10A. In the current study, we show that TIMP-1 expression induces phenotypic changes in cell morphology, cell adhesion, cytoskeletal remodeling, and motility, indicative of an epithelial-mesenchymal transition (EMT). This is evidenced by loss of the epithelial cell adhesion molecule E-cadherin with an increase in the mesenchymal markers vimentin, N-cadherin, and fibronectin. Signaling through TIMP-1, but not TIMP-2, induces the expression of TWIST1, an important EMT transcription factor known to suppress E-cadherin transcription, in a CD63-dependent manner. RNAi-mediated knockdown of TWIST1 rescued E-cadherin expression in TIMP-1-overexpressing cells, demonstrating a functional significance of TWIST1 in TIMP-1-mediated EMT. Furthermore, analysis of TIMP-1 structural mutants reveals that TIMP-1 interactions with CD63 that activate cell survival signaling and EMT do not require the matrix metalloproteinase (MMP)-inhibitory domain of TIMP-1. Taken together, these data demonstrate that TIMP-1 binding to CD63 activates intracellular signal transduction pathways, resulting in EMT-like changes in breast epithelial cells, independent of its MMP-inhibitory function. IMPLICATIONS: TIMP-1's function as an endogenous inhibitor of MMP or as a "cytokine-like" signaling molecule may be a critical determinant for tumor cell behavior.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Mol Cell

Abstract Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

SOCS3-mediated regulation of inflammatory cytokines in PTEN and p53 inactivated triple negative breast cancer model.

Oncogene

Abstract Somatic mutations or deletions of TP53 and PTEN in ductal carcinoma in situ lesions have been implicated in progression to invasive ductal carcinomas. A recent molecular and mutational analysis of breast cancers revealed that inactivation of tumor suppressors, p53 and PTEN, are strongly associated with triple negative breast cancer. In addition, these tumor suppressors have important roles in regulating self-renewal in normal and malignant stem cells. To investigate their role in breast carcinogenesis, we knocked down these genes in human mammary cells and in non-transformed MCF10A cells. p53 and PTEN knockdown synergized to activate pro-inflammatory interleukin-6 (IL6)/Stat3/nuclear factor κB signaling. This resulted in generation of highly metastatic epithelial-to-mesenchymal transition-like cancer stem cells resulting in tumors whose gene expression profile mimicked that found in basal/claudin-low molecular subtype within the triple negative breast tumors. Constitutive activation of this loop in transformed cells was dependent on proteolytic degradation of suppressor of cytokine signaling 3 (SOCS3) resulting in low levels of this protein in basal/claudin-low cell lines and primary tumors. In non-transformed cells, transient activation of the IL6 inflammatory loop induced SOCS3 expression leading to pathway inactivation. In transformed cells, enforced expression of SOCS3 or interfering with IL6 pathway via IL6R blockade inhibited tumor growth and metastasis in mouse xenograft models. Furthermore, circulating tumor cells were significantly reduced in tumor-bearing animals when treated with anti-IL6R antibodies. These studies uncover important connections between inflammation and carcinogenesis and suggest that blocking pro-inflammatory cytokines may be utilized as an attractive strategy to target triple negative breast tumors, which currently lacks molecularly targeted therapies.

TIMP-1 via TWIST1 Induces EMT Phenotypes in Human Breast Epithelial Cells

Mol Cancer Res

Abstract Tissue inhibitor of metalloproteinase-1 (TIMP-1) regulates intracellular signaling networks for inhibition of apoptosis. Tetraspanin (CD63), a cell surface binding partner for TIMP-1, was previously shown to regulate integrin-mediated survival pathways in the human breast epithelial cell line MCF10A. In the current study, we show that TIMP-1 expression induces phenotypic changes in cell morphology, cell adhesion, cytoskeletal remodeling, and motility, indicative of an epithelial-mesenchymal transition (EMT). This is evidenced by loss of the epithelial cell adhesion molecule E-cadherin with an increase in the mesenchymal markers vimentin, N-cadherin, and fibronectin. Signaling through TIMP-1, but not TIMP-2, induces the expression of TWIST1, an important EMT transcription factor known to suppress E-cadherin transcription, in a CD63-dependent manner. RNAi-mediated knockdown of TWIST1 rescued E-cadherin expression in TIMP-1-overexpressing cells, demonstrating a functional significance of TWIST1 in TIMP-1-mediated EMT. Furthermore, analysis of TIMP-1 structural mutants reveals that TIMP-1 interactions with CD63 that activate cell survival signaling and EMT do not require the matrix metalloproteinase (MMP)-inhibitory domain of TIMP-1. Taken together, these data demonstrate that TIMP-1 binding to CD63 activates intracellular signal transduction pathways, resulting in EMT-like changes in breast epithelial cells, independent of its MMP-inhibitory function. IMPLICATIONS: TIMP-1's function as an endogenous inhibitor of MMP or as a "cytokine-like" signaling molecule may be a critical determinant for tumor cell behavior.

TIMP-1 induces an EMT-like phenotypic conversion in MDCK cells independent of its MMP-inhibitory domain.

PLoS One

Abstract Matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) regulate epithelial-mesenchymal transition (EMT) critical for the development of epithelial organs as well as cancer cell invasion. TIMP-1 is frequently overexpressed in several types of human cancers and serves as a prognostic marker. The present study investigates the roles of TIMP-1 on the EMT process and formation of the lumen-like structure in a 3D Matrigel culture of MDCK cells. We show that TIMP-1 overexpression effectively prevents cell polarization and acinar-like structure formation. TIMP-1 induces expression of the developmental EMT transcription factors such as SLUG, TWIST, ZEB1 and ZEB2, leading to downregulation of epithelial marker and upregulation of mesenchymal markers. Importantly, TIMP-1's ability to induce the EMT-like process is independent of its MMP-inhibitory domain. To our surprise, TIMP-1 induces migratory and invasive properties in MDCK cells. Here, we present a novel finding that TIMP-1 signaling upregulates MT1-MMP and MMP-2 expression, and potentiates MT1-MMP activation of pro-MMP-2, contributing to tumor cell invasion. In spite of the fact that TIMP-1, as opposed to TIMP-2, does not interact with and inhibit MT1-MMP, TIMP-1 may act as a key regulator of MT1-MMP/MMP-2 axis. Collectively, our findings suggest a model in which TIMP-1 functions as a signaling molecule and also as an endogenous inhibitor of MMPs. This concept represents a paradigm shift in the current view of TIMP-1/MT1-MMP interactions and functions during cancer development/progression.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Mol Cell

Abstract Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

SOCS3-mediated regulation of inflammatory cytokines in PTEN and p53 inactivated triple negative breast cancer model.

Oncogene

Abstract Somatic mutations or deletions of TP53 and PTEN in ductal carcinoma in situ lesions have been implicated in progression to invasive ductal carcinomas. A recent molecular and mutational analysis of breast cancers revealed that inactivation of tumor suppressors, p53 and PTEN, are strongly associated with triple negative breast cancer. In addition, these tumor suppressors have important roles in regulating self-renewal in normal and malignant stem cells. To investigate their role in breast carcinogenesis, we knocked down these genes in human mammary cells and in non-transformed MCF10A cells. p53 and PTEN knockdown synergized to activate pro-inflammatory interleukin-6 (IL6)/Stat3/nuclear factor κB signaling. This resulted in generation of highly metastatic epithelial-to-mesenchymal transition-like cancer stem cells resulting in tumors whose gene expression profile mimicked that found in basal/claudin-low molecular subtype within the triple negative breast tumors. Constitutive activation of this loop in transformed cells was dependent on proteolytic degradation of suppressor of cytokine signaling 3 (SOCS3) resulting in low levels of this protein in basal/claudin-low cell lines and primary tumors. In non-transformed cells, transient activation of the IL6 inflammatory loop induced SOCS3 expression leading to pathway inactivation. In transformed cells, enforced expression of SOCS3 or interfering with IL6 pathway via IL6R blockade inhibited tumor growth and metastasis in mouse xenograft models. Furthermore, circulating tumor cells were significantly reduced in tumor-bearing animals when treated with anti-IL6R antibodies. These studies uncover important connections between inflammation and carcinogenesis and suggest that blocking pro-inflammatory cytokines may be utilized as an attractive strategy to target triple negative breast tumors, which currently lacks molecularly targeted therapies.

TIMP-1 via TWIST1 Induces EMT Phenotypes in Human Breast Epithelial Cells

Mol Cancer Res

Abstract Tissue inhibitor of metalloproteinase-1 (TIMP-1) regulates intracellular signaling networks for inhibition of apoptosis. Tetraspanin (CD63), a cell surface binding partner for TIMP-1, was previously shown to regulate integrin-mediated survival pathways in the human breast epithelial cell line MCF10A. In the current study, we show that TIMP-1 expression induces phenotypic changes in cell morphology, cell adhesion, cytoskeletal remodeling, and motility, indicative of an epithelial-mesenchymal transition (EMT). This is evidenced by loss of the epithelial cell adhesion molecule E-cadherin with an increase in the mesenchymal markers vimentin, N-cadherin, and fibronectin. Signaling through TIMP-1, but not TIMP-2, induces the expression of TWIST1, an important EMT transcription factor known to suppress E-cadherin transcription, in a CD63-dependent manner. RNAi-mediated knockdown of TWIST1 rescued E-cadherin expression in TIMP-1-overexpressing cells, demonstrating a functional significance of TWIST1 in TIMP-1-mediated EMT. Furthermore, analysis of TIMP-1 structural mutants reveals that TIMP-1 interactions with CD63 that activate cell survival signaling and EMT do not require the matrix metalloproteinase (MMP)-inhibitory domain of TIMP-1. Taken together, these data demonstrate that TIMP-1 binding to CD63 activates intracellular signal transduction pathways, resulting in EMT-like changes in breast epithelial cells, independent of its MMP-inhibitory function. IMPLICATIONS: TIMP-1's function as an endogenous inhibitor of MMP or as a "cytokine-like" signaling molecule may be a critical determinant for tumor cell behavior.

TIMP-1 induces an EMT-like phenotypic conversion in MDCK cells independent of its MMP-inhibitory domain.

PLoS One

Abstract Matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) regulate epithelial-mesenchymal transition (EMT) critical for the development of epithelial organs as well as cancer cell invasion. TIMP-1 is frequently overexpressed in several types of human cancers and serves as a prognostic marker. The present study investigates the roles of TIMP-1 on the EMT process and formation of the lumen-like structure in a 3D Matrigel culture of MDCK cells. We show that TIMP-1 overexpression effectively prevents cell polarization and acinar-like structure formation. TIMP-1 induces expression of the developmental EMT transcription factors such as SLUG, TWIST, ZEB1 and ZEB2, leading to downregulation of epithelial marker and upregulation of mesenchymal markers. Importantly, TIMP-1's ability to induce the EMT-like process is independent of its MMP-inhibitory domain. To our surprise, TIMP-1 induces migratory and invasive properties in MDCK cells. Here, we present a novel finding that TIMP-1 signaling upregulates MT1-MMP and MMP-2 expression, and potentiates MT1-MMP activation of pro-MMP-2, contributing to tumor cell invasion. In spite of the fact that TIMP-1, as opposed to TIMP-2, does not interact with and inhibit MT1-MMP, TIMP-1 may act as a key regulator of MT1-MMP/MMP-2 axis. Collectively, our findings suggest a model in which TIMP-1 functions as a signaling molecule and also as an endogenous inhibitor of MMPs. This concept represents a paradigm shift in the current view of TIMP-1/MT1-MMP interactions and functions during cancer development/progression.

Breast cancer stem cells transition between epithelial and mesenchymal states reflective of their normal counterparts.

Stem Cell Reports.

Abstract Previous studies have suggested that breast cancer stem cells (BCSCs) mediate metastasis, are resistant to radiation and chemotherapy, and contribute to relapse. Although several BCSC markers have been described, it is unclear whether these markers identify the same or independent BCSCs. Here, we show that BCSCs exist in distinct mesenchymal-like (epithelial-mesenchymal transition [EMT]) and epithelial-like (mesenchymal-epithelial transition [MET]) states. Mesenchymal-like BCSCs characterized as CD24(-)CD44(+) are primarily quiescent and localized at the tumor invasive front, whereas epithelial-like BCSCs express aldehyde dehydrogenase (ALDH), are proliferative, and are located more centrally. The gene-expression profiles of mesenchymal-like and epithelial-like BCSCs are remarkably similar across different molecular subtypes of breast cancer, and resemble those of distinct basal and luminal stem cells found in the normal breast. We propose that the plasticity of BCSCs that allows them to transition between EMT- and MET-like states endows these cells with the capacity for tissue invasion, dissemination, and growth at metastatic sites.

Therapeutic targeting of the Notch pathway in breast cancer stem cells.

American Association of Cancer Research Annual Meeting, Orlando, FL

Poster Presentation at 2 Conferences/Meetings: Cancer Research Symposium Poster, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI. Dec 17, 2010 AACR Annual Meeting April 2-6, 2011

Mapping Out Cellular Respiration

Oakland University Center for Teaching and Learning (CETL) Instructional Fair,

Poster presentation and demonstration of concept mapping for Nutrient Metabolism,

Notch reporter activity in breast cancer cell lines identifies a subset of cells with stem cell activity.

Molecular Cancer Therapeutics

Developmental pathways such as notch play a pivotal role in tissue-specific stem cell self-renewal as well as in tumor development. however, the role of notch signaling in breast cancer stem cells (csc) remains to be determined. we utilized a lentiviral notch reporter system to identify a subset of cells with a higher notch activity (notch(+)) or reduced activity (notch(-)) in multiple breast cancer cell lines. using in vitro and mouse xenotransplantation assays, we investigated the role of the notch pathway in breast csc regulation. breast cancer cells with increased notch activity displayed increased sphere formation as well as expression of breast csc markers. interestingly notch(+) cells displayed higher notch4 expression in both basal and luminal breast cancer cell lines. moreover, notch(+) cells demonstrated tumor initiation capacity at serial dilutions in mouse xenografts, whereas notch(-) cells failed to generate tumors. γ-secretase inhibitor (gsi), a notch blocker but not a chemotherapeutic agent, effectively targets these notch(+) cells in vitro and in mouse xenografts. furthermore, elevated notch4 and hey1 expression in primary patient samples correlated with poor patient survival. our study revealed a molecular mechanism for the role of notch-mediated regulation of breast cscs and provided a compelling rationale for csc-targeted therapeutics. mol cancer ther; 14(3); 779-87. ©2015 aacr.

Stem cells in normal development and cancer.

Prog Mol Biol Transl Sci.

Abstract In this chapter we provide an overview of stem cells in normal tissues as well as in many different types of cancers. All tissues in the body are derived from organ-specific stem cells that retain the ability to self-renew and differentiate into specific cell types. The cancer stem cell hypothesis suggests that tumors arise from cell populations with dysregulated self-renewal. This may be tissue stem cells or more differentiated cells that acquire self-renewal capabilities. In addition, we outline some useful assays for purification and isolation of cancer stem cells including the dye exclusion side population assay, flow cytometry sorting techniques for identification of putative cancer stem cell markers, tumorspheres assay, aldehyde dehydrogenase activity assay, PKH, and other membrane staining used to label the cancer stem cells, as well as in vivo xenograft transplantation assays. We also examine some of the cell signaling pathways that regulate stem cell self-renewal including the Notch, Hedgehog, HER2/PI3K/Akt/PTEN, and p53 pathways. We also review information demonstrating the involvement of the microenvironment or stem cell niche and its effects on the growth and maintenance of cancer stem cells. Finally, we highlight the therapeutic implications of targeting stem cells by inhibiting these pathways for the treatment and prevention of cancer.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Mol Cell

Abstract Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

SOCS3-mediated regulation of inflammatory cytokines in PTEN and p53 inactivated triple negative breast cancer model.

Oncogene

Abstract Somatic mutations or deletions of TP53 and PTEN in ductal carcinoma in situ lesions have been implicated in progression to invasive ductal carcinomas. A recent molecular and mutational analysis of breast cancers revealed that inactivation of tumor suppressors, p53 and PTEN, are strongly associated with triple negative breast cancer. In addition, these tumor suppressors have important roles in regulating self-renewal in normal and malignant stem cells. To investigate their role in breast carcinogenesis, we knocked down these genes in human mammary cells and in non-transformed MCF10A cells. p53 and PTEN knockdown synergized to activate pro-inflammatory interleukin-6 (IL6)/Stat3/nuclear factor κB signaling. This resulted in generation of highly metastatic epithelial-to-mesenchymal transition-like cancer stem cells resulting in tumors whose gene expression profile mimicked that found in basal/claudin-low molecular subtype within the triple negative breast tumors. Constitutive activation of this loop in transformed cells was dependent on proteolytic degradation of suppressor of cytokine signaling 3 (SOCS3) resulting in low levels of this protein in basal/claudin-low cell lines and primary tumors. In non-transformed cells, transient activation of the IL6 inflammatory loop induced SOCS3 expression leading to pathway inactivation. In transformed cells, enforced expression of SOCS3 or interfering with IL6 pathway via IL6R blockade inhibited tumor growth and metastasis in mouse xenograft models. Furthermore, circulating tumor cells were significantly reduced in tumor-bearing animals when treated with anti-IL6R antibodies. These studies uncover important connections between inflammation and carcinogenesis and suggest that blocking pro-inflammatory cytokines may be utilized as an attractive strategy to target triple negative breast tumors, which currently lacks molecularly targeted therapies.

TIMP-1 via TWIST1 Induces EMT Phenotypes in Human Breast Epithelial Cells

Mol Cancer Res

Abstract Tissue inhibitor of metalloproteinase-1 (TIMP-1) regulates intracellular signaling networks for inhibition of apoptosis. Tetraspanin (CD63), a cell surface binding partner for TIMP-1, was previously shown to regulate integrin-mediated survival pathways in the human breast epithelial cell line MCF10A. In the current study, we show that TIMP-1 expression induces phenotypic changes in cell morphology, cell adhesion, cytoskeletal remodeling, and motility, indicative of an epithelial-mesenchymal transition (EMT). This is evidenced by loss of the epithelial cell adhesion molecule E-cadherin with an increase in the mesenchymal markers vimentin, N-cadherin, and fibronectin. Signaling through TIMP-1, but not TIMP-2, induces the expression of TWIST1, an important EMT transcription factor known to suppress E-cadherin transcription, in a CD63-dependent manner. RNAi-mediated knockdown of TWIST1 rescued E-cadherin expression in TIMP-1-overexpressing cells, demonstrating a functional significance of TWIST1 in TIMP-1-mediated EMT. Furthermore, analysis of TIMP-1 structural mutants reveals that TIMP-1 interactions with CD63 that activate cell survival signaling and EMT do not require the matrix metalloproteinase (MMP)-inhibitory domain of TIMP-1. Taken together, these data demonstrate that TIMP-1 binding to CD63 activates intracellular signal transduction pathways, resulting in EMT-like changes in breast epithelial cells, independent of its MMP-inhibitory function. IMPLICATIONS: TIMP-1's function as an endogenous inhibitor of MMP or as a "cytokine-like" signaling molecule may be a critical determinant for tumor cell behavior.

TIMP-1 induces an EMT-like phenotypic conversion in MDCK cells independent of its MMP-inhibitory domain.

PLoS One

Abstract Matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) regulate epithelial-mesenchymal transition (EMT) critical for the development of epithelial organs as well as cancer cell invasion. TIMP-1 is frequently overexpressed in several types of human cancers and serves as a prognostic marker. The present study investigates the roles of TIMP-1 on the EMT process and formation of the lumen-like structure in a 3D Matrigel culture of MDCK cells. We show that TIMP-1 overexpression effectively prevents cell polarization and acinar-like structure formation. TIMP-1 induces expression of the developmental EMT transcription factors such as SLUG, TWIST, ZEB1 and ZEB2, leading to downregulation of epithelial marker and upregulation of mesenchymal markers. Importantly, TIMP-1's ability to induce the EMT-like process is independent of its MMP-inhibitory domain. To our surprise, TIMP-1 induces migratory and invasive properties in MDCK cells. Here, we present a novel finding that TIMP-1 signaling upregulates MT1-MMP and MMP-2 expression, and potentiates MT1-MMP activation of pro-MMP-2, contributing to tumor cell invasion. In spite of the fact that TIMP-1, as opposed to TIMP-2, does not interact with and inhibit MT1-MMP, TIMP-1 may act as a key regulator of MT1-MMP/MMP-2 axis. Collectively, our findings suggest a model in which TIMP-1 functions as a signaling molecule and also as an endogenous inhibitor of MMPs. This concept represents a paradigm shift in the current view of TIMP-1/MT1-MMP interactions and functions during cancer development/progression.

Breast cancer stem cells transition between epithelial and mesenchymal states reflective of their normal counterparts.

Stem Cell Reports.

Abstract Previous studies have suggested that breast cancer stem cells (BCSCs) mediate metastasis, are resistant to radiation and chemotherapy, and contribute to relapse. Although several BCSC markers have been described, it is unclear whether these markers identify the same or independent BCSCs. Here, we show that BCSCs exist in distinct mesenchymal-like (epithelial-mesenchymal transition [EMT]) and epithelial-like (mesenchymal-epithelial transition [MET]) states. Mesenchymal-like BCSCs characterized as CD24(-)CD44(+) are primarily quiescent and localized at the tumor invasive front, whereas epithelial-like BCSCs express aldehyde dehydrogenase (ALDH), are proliferative, and are located more centrally. The gene-expression profiles of mesenchymal-like and epithelial-like BCSCs are remarkably similar across different molecular subtypes of breast cancer, and resemble those of distinct basal and luminal stem cells found in the normal breast. We propose that the plasticity of BCSCs that allows them to transition between EMT- and MET-like states endows these cells with the capacity for tissue invasion, dissemination, and growth at metastatic sites.

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group

Student Member of Cancer Biology Curriculum Committee

Student Representative for my program

College Diabetes Network

Faculty Representative

American Association of Cancer Research

Associate Member

Member of Women in Cancer Group