48 results on '"Mark Duquette"'
Search Results
2. 897-F Wnt inhibition improves efficacy of immune checkpoint blockade in glioblastoma
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Dai Fukumura, Rakesh K Jain, Peigen Huang, Lance L Munn, Patrik Andersson, Zohreh Amoozgar, Somin Lee, Meenal Datta, Sylvie Roberge, Mark Duquette, Neal I Lindeman, Shanmugarajan Krishnan, Sonu Subudhi, Ashwin Srinivasan Kumar, Jessica Posada, and Pinji Lei
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Neoplasms. Tumors. Oncology. Including cancer and carcinogens ,RC254-282 - Published
- 2023
- Full Text
- View/download PDF
3. Anti-VEGF therapy improves EGFR-vIII-CAR-T cell delivery and efficacy in syngeneic glioblastoma models in mice
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Dai Fukumura, Rakesh K Jain, Jun Ren, Xinyue Dong, Zohreh Amoozgar, Somin Lee, Meenal Datta, Sylvie Roberge, and Mark Duquette
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Neoplasms. Tumors. Oncology. Including cancer and carcinogens ,RC254-282 - Abstract
Chimeric antigen receptor (CAR)-T cells have revolutionized the treatment of multiple types of hematological malignancies, but have shown limited efficacy in patients with glioblastoma (GBM) or other solid tumors. This may be largely due to the immunosuppressive tumor microenvironment (TME) that compromises CAR-T cells’ delivery and antitumor activity. We previously showed that blocking vascular endothelial growth factor (VEGF) signaling can normalize tumor vessels in murine and human tumors, including GBM, breast, liver, and rectal carcinomas. Moreover, we demonstrated that vascular normalization can improve the delivery of CD8+ T cells and the efficacy of immunotherapy in breast cancer models in mice. In fact, the US FDA (Food and drug administration) has approved seven different combinations of anti-VEGF drugs and immune checkpoint blockers for liver, kidney, lung and endometrial cancers in the past 3 years. Here, we tested the hypothesis that anti-VEGF therapy can improve the delivery and efficacy of CAR-T cells in immunocompetent mice bearing orthotopic GBM tumors. We engineered two syngeneic mouse GBM cell lines (CT2A and GSC005) to express EGFRvIII—one of the most common neoantigens in human GBM—and CAR T cells to recognize EGFRvIII. We found that treatment with the anti-mouse VEGF antibody (B20) improved CAR-T cell infiltration and distribution throughout the GBM TME, delayed tumor growth, and prolonged survival of GBM-bearing mice compared with EGFRvIII-CAR-T cell therapy alone. Our findings provide compelling data and a rationale for clinical evaluation of anti-VEGF agents with CAR T cells for GBM patients.
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- 2023
- Full Text
- View/download PDF
4. Corrigendum: Thrombospondin-1 Silencing Down-Regulates Integrin Expression Levels in Human Anaplastic Thyroid Cancer Cells With BRAFV600E: New Insights in the Host Tissue Adaptation and Homeostasis of Tumor Microenvironment
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Mark Duquette, Peter M. Sadow, Jack Lawler, and Carmelo Nucera
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BRAFV600E ,integrins ,thyroid cancer ,microenvironment ,extracellular matrix ,TSP-1 ,Diseases of the endocrine glands. Clinical endocrinology ,RC648-665 - Published
- 2020
- Full Text
- View/download PDF
5. Data from A Double Hit to Kill Tumor and Endothelial Cells by TRAIL and Antiangiogenic 3TSR
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Roya Khosravi-Far, Jack Lawler, Nordine Benhaga, Xuefeng Zhang, Mark Duquette, Robin Humphreys, Min Ye, Taiguang Jin, Sareh Parangi, Keli Song, and Bin Ren
- Abstract
As tumor development relies on a coordination of angiogenesis and tumor growth, an efficient antitumor strategy should target both the tumor and its associated vessels. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in a tumor-selective manner. Additionally, thrombospondin-1, a naturally occurring inhibitor of angiogenesis, and a recombinant protein containing functional domains of thrombospondin-1, 3TSR, have been shown to be necessary and sufficient to inhibit tumor angiogenesis. Here, we show that a combination of a TRAIL receptor 2 agonist antibody, Lexatumumab, and 3TSR results in a significantly enhanced and durable tumor inhibition. We further observed that 3TSR induces apoptosis in primary endothelial cells by up-regulating the expression of TRAIL receptors 1 and 2 in a CD36 and Jun NH2-terminal kinase-dependent manner leading to the activation of both intrinsic and extrinsic apoptotic machineries. The modulation of these pathways is critical for 3TSR-induced apoptosis as disrupting either via specific inhibitors reduced apoptosis. Moreover, 3TSR attenuates the Akt survival pathway. These studies indicate that 3TSR plays a critical role in regulating the proapoptotic signaling pathways that control growth and death in endothelial cells and that a combination of TRAIL and 3TSR acts as a double hit against tumor and tumor-associated vessels. [Cancer Res 2009;69(9):3856–65]
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- 2023
6. Supplementary Figure 1 from A Double Hit to Kill Tumor and Endothelial Cells by TRAIL and Antiangiogenic 3TSR
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Roya Khosravi-Far, Jack Lawler, Nordine Benhaga, Xuefeng Zhang, Mark Duquette, Robin Humphreys, Min Ye, Taiguang Jin, Sareh Parangi, Keli Song, and Bin Ren
- Abstract
Supplementary Figure 1 from A Double Hit to Kill Tumor and Endothelial Cells by TRAIL and Antiangiogenic 3TSR
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- 2023
7. Supplementary Figure Legends 1-3 from A Double Hit to Kill Tumor and Endothelial Cells by TRAIL and Antiangiogenic 3TSR
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Roya Khosravi-Far, Jack Lawler, Nordine Benhaga, Xuefeng Zhang, Mark Duquette, Robin Humphreys, Min Ye, Taiguang Jin, Sareh Parangi, Keli Song, and Bin Ren
- Abstract
Supplementary Figure Legends 1-3 from A Double Hit to Kill Tumor and Endothelial Cells by TRAIL and Antiangiogenic 3TSR
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- 2023
8. 911 Anti-VEGF therapy improves EGFR-vIII-CAR-T cell delivery and efficacy in syngeneic glioblastoma models in mice
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Xinyue Dong, Jun Ren, Zohreh Amoozgar, Meenal Datta, Somin Lee, Sylvie Roberge, Mark Duquette, Dai Fukumura, and Rakesh Jain
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- 2022
9. Losartan controls immune checkpoint blocker-induced edema and improves survival in glioblastoma
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Meenal Datta, Sampurna Chatterjee, Elizabeth M. Perez, Simon Gritsch, Sylvie Roberge, Mark Duquette, Ivy X. Chen, Kamila Naxerova, Ashwin S. Kumar, Mitrajit Ghosh, Kyrre E. Emblem, Mei R. Ng, William W. Ho, Pragya Kumar, Shanmugarajan Krishnan, Xinyue Dong, Maria C. Speranza, Martha R. Neagu, David A. Reardon, Arlene H. Sharpe, Gordon J. Freeman, Mario L. Suvà, Lei Xu, and Rakesh K. Jain
- Abstract
Immune checkpoint blockers (ICBs) have failed in all Phase III glioblastoma trials. Here, we found that ICBs induce cerebral edema in some patients and mice with glioblastoma. Through single-cell RNA sequencing, intravital imaging, and T cell blocking studies in mice, we demonstrated that this edema results from an inflammatory response following anti-PD1 antibody treatment that disrupts the blood-tumor-barrier. Used in lieu of immunosuppressive corticosteroids, the angiotensin receptor blocker losartan prevented this ICB-induced edema and reprogrammed the tumor microenvironment, curing 20% of mice which increased to 40% in combination with standard of care treatment. Using a bihemispheric tumor model, we identified a “hot” tumor immune signature prior to losartan+anti-PD1 therapy that predicted long-term survival. Our findings provide the rationale and associated biomarkers to test losartan with ICBs in glioblastoma patients.One-Sentence SummaryLosartan prevents immunotherapy-associated edema and enhances the outcome of immunotherapy in glioblastoma.
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- 2022
10. Fatty acid synthesis is required for breast cancer brain metastasis
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Jiang Chen, Anna M. Westermark, Lewis C. Cantley, Rakesh K. Jain, Brendan Prideaux, John M. Asara, Clary B. Clish, David P. Kodack, Neal I. Lindeman, Christopher W. Ng, Gino B. Ferraro, Ivy X. Chen, Costas A. Lyssiotis, Keene L. Abbott, Ahmed Ali, Jens Nielsen, Zohreh Amoozgar, Dai Fukumura, Raphael Ferreira, Dan G. Duda, Mark Duquette, David E. Housman, Jessica M. Possada, Véronique Dartois, Kamila Naxerova, Shawn M. Davidson, Xin Jin, Matthew G. Vander Heiden, Sylvie Roberge, Todd R. Golub, Amy Deik, Christopher R. Chin, Divya Bezwada, Elena F. Brachtel, Alba Luengo, and Landry Blanc
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Cancer Research ,Systemic disease ,Breast Neoplasms ,chemistry.chemical_compound ,Breast cancer ,Tumor Microenvironment ,medicine ,Humans ,skin and connective tissue diseases ,Fatty acid synthesis ,biology ,Brain Neoplasms ,business.industry ,Fatty Acids ,Metabolism ,medicine.disease ,Phenotype ,Metastatic breast cancer ,Fatty acid synthase ,Oncology ,chemistry ,Cancer research ,biology.protein ,Female ,Fatty Acid Synthases ,business ,Brain metastasis - Abstract
Brain metastases are refractory to therapies that control systemic disease in patients with human epidermal growth factor receptor 2 (HER2+) breast cancer, and the brain microenvironment contributes to this therapy resistance. Nutrient availability can vary across tissues, therefore metabolic adaptations required for brain metastatic breast cancer growth may introduce liabilities that can be exploited for therapy. Here, we assessed how metabolism differs between breast tumors in brain versus extracranial sites and found that fatty acid synthesis is elevated in breast tumors growing in brain. We determine that this phenotype is an adaptation to decreased lipid availability in brain relative to other tissues, resulting in a site-specific dependency on fatty acid synthesis for breast tumors growing at this site. Genetic or pharmacological inhibition of fatty acid synthase (FASN) reduces HER2+ breast tumor growth in the brain, demonstrating that differences in nutrient availability across metastatic sites can result in targetable metabolic dependencies.
- Published
- 2021
11. Anti-VEGF therapy improves EGFR-vIII-CAR-T cell delivery and efficacy in syngeneic glioblastoma models in mice
- Author
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Xinyue Dong, Jun Ren, Zohreh Amoozgar, Somin Lee, Meenal Datta, Sylvie Roberge, Mark Duquette, Dai Fukumura, and Rakesh K Jain
- Subjects
Pharmacology ,Cancer Research ,Oncology ,Immunology ,Molecular Medicine ,Immunology and Allergy - Abstract
Chimeric antigen receptor (CAR)-T cells have revolutionized the treatment of multiple types of hematological malignancies, but have shown limited efficacy in patients with glioblastoma (GBM) or other solid tumors. This may be largely due to the immunosuppressive tumor microenvironment (TME) that compromises CAR-T cells’ delivery and antitumor activity. We previously showed that blocking vascular endothelial growth factor (VEGF) signaling can normalize tumor vessels in murine and human tumors, including GBM, breast, liver, and rectal carcinomas. Moreover, we demonstrated that vascular normalization can improve the delivery of CD8+ T cells and the efficacy of immunotherapy in breast cancer models in mice. In fact, the US FDA (Food and drug administration) has approved seven different combinations of anti-VEGF drugs and immune checkpoint blockers for liver, kidney, lung and endometrial cancers in the past 3 years. Here, we tested the hypothesis that anti-VEGF therapy can improve the delivery and efficacy of CAR-T cells in immunocompetent mice bearing orthotopic GBM tumors. We engineered two syngeneic mouse GBM cell lines (CT2A and GSC005) to express EGFRvIII—one of the most common neoantigens in human GBM—and CAR T cells to recognize EGFRvIII. We found that treatment with the anti-mouse VEGF antibody (B20) improved CAR-T cell infiltration and distribution throughout the GBM TME, delayed tumor growth, and prolonged survival of GBM-bearing mice compared with EGFRvIII-CAR-T cell therapy alone. Our findings provide compelling data and a rationale for clinical evaluation of anti-VEGF agents with CAR T cells for GBM patients.
- Published
- 2023
12. TMIC-61. LOSARTAN PREVENTS IMMUNOTHERAPY-ASSOCIATED EDEMA AND ENHANCES SURVIVAL IN GLIOBLASTOMA
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Meenal Datta, Sampurna Chatterjee, Elizabeth Perez, Simon Gritsch, Sylvie Roberge, Mark Duquette, Ivy Chen, Kamila Naxerova, Ashwin Kumar, Mitrajit Ghosh, Kyrre Emblem, Rosa Ng, William Ho, Pragya Kumar, Shanmugarajan Krishnan, Xinyue Dong, Maria Speranza, Martha Neagu, David A Reardon, Arlene Sharpe, Gordon Freeman, Mario Suva, Lei Xu, and Rakesh Jain
- Subjects
Cancer Research ,Oncology ,Neurology (clinical) - Abstract
Immune checkpoint blockers (ICBs) have revolutionized the treatment of some solid cancers but have failed to benefit the majority of glioblastoma (GBM) patients. Two reasons underlying limited ICB benefit are: 1) immune-related adverse events, and 2) resistance conferred by the tumor microenvironment. Here, we show that ICBs induce cerebral edema in patients and GBM mouse models. This edema results from an inflammatory response to ICB treatment that disrupts the blood-tumor-barrier, as confirmed by intravital imaging, mechanistic blocking studies, and single-cell RNA sequencing. Losartan – a commonly prescribed antihypertensive agent – controls ICB-induced edema, reprograms the immunosuppressive tumor microenvironment, and improves survival under ICB therapy. In combination with a standard of care regimen in mice mimicking clinical treatment (surgical resection, chemoradiation), losartan increases the percent of long-term surviving (cured) mice under ICB therapy from 16% to 43%. Finally, a bihemispheric “resect-and-response” model to establish predictive biomarkers from the tumor microenvironment reveals that cured mice have an immunostimulatory (“hot”) immune tumor compartment prior to therapy. These results provide the basis for clinical testing of adding to losartan to ICB treatment for GBM patients.
- Published
- 2022
13. DDRE-07. FATTY ACID SYNTHESIS IS REQUIRED FOR BREAST CANCER BRAIN METASTASIS
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Lewis C. Cantley, Shawn M. Davidson, Landry Blanc, Gino B. Ferraro, Costas A. Lyssiotis, Xin Jin, Ahmed Ali, John M. Asara, Divya Bezwada, Clary B. Clish, Véronique Dartois, Rakesh K. Jain, Sylvie Roberge, Kamila Naxerova, Christopher R. Chin, Jiang Chen, Brendan Prideaux, David P. Kodack, Dai Fukumura, Anna M. Westermark, Todd R. Golub, Alba Luengo, Keene L. Abbott, Christopher W. Ng, Amy Deik, Matthew G. Vander Heiden, Raphael Ferreira, Mark Duquette, Dan G. Duda, Jessica M. Possada, Ivy X. Chen, and Zohreh Amoozgar
- Subjects
Systemic disease ,Fatty acid biosynthesis ,Metabolic Drug Targets, Resistance ,Metabolism ,Biology ,medicine.disease ,Phenotype ,Supplement Abstracts ,chemistry.chemical_compound ,Fatty acid synthase ,Breast cancer ,chemistry ,medicine ,biology.protein ,Cancer research ,AcademicSubjects/MED00300 ,AcademicSubjects/MED00310 ,skin and connective tissue diseases ,Fatty acid synthesis ,Brain metastasis - Abstract
Brain metastases are refractory to therapies that otherwise control systemic disease in patients with human epidermal growth factor receptor 2 (HER2+) breast cancer, and the unique brain microenvironment contributes to this therapy resistance. Nutrient availability can vary across tissues, therefore metabolic adaptations required for breast cancer growth in the brain microenvironment may also introduce liabilities that can be exploited for therapy. Here, we assessed how metabolism differs between breast tumors growing in the brain versus extracranial sites and found that fatty acid synthesis is elevated in breast tumors growing in the brain. We determine that this phenotype is an adaptation to decreased lipid availability in the brain relative to other tissues, which results in a site-specific dependency on fatty acid synthesis for breast tumors growing at this site. Genetic or pharmacological inhibition of fatty acid synthase (FASN) reduces HER2+ breast tumor growth in the brain, demonstrating that differences in nutrient availability across metastatic sites can result in targetable metabolic dependencies.
- Published
- 2021
14. Author Correction: Fatty acid synthesis is required for breast cancer brain metastasis
- Author
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Rakesh K. Jain, Neal I. Lindeman, Raphael Ferreira, Christopher R. Chin, Clary B. Clish, John M. Asara, Gino B. Ferraro, Keene L. Abbott, Costas A. Lyssiotis, Jessica M. Posada, Alba Luengo, Landry Blanc, Amy Deik, Mark Duquette, Véronique Dartois, Kamila Naxerova, Divya Bezwada, Brendan Prideaux, Shawn M. Davidson, Todd R. Golub, Matthew G. Vander Heiden, David P. Kodack, Ahmed Ali, Dai Fukumura, Elena F. Brachtel, Ivy X. Chen, Xin Jin, Anna M. Westermark, Christopher W. Ng, Dan G. Duda, Zohreh Amoozgar, Sylvie Roberge, Lewis C. Cantley, Jiang Chen, David E. Housman, and Jens Nielsen
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Cancer Research ,business.industry ,Cancer ,medicine.disease ,Metastasis ,chemistry.chemical_compound ,Breast cancer ,Oncology ,chemistry ,medicine ,Cancer research ,business ,Fatty acid synthesis ,Brain metastasis - Published
- 2021
15. Abstract 90: Fatty acid synthesis is required for breast cancer brain metastasis
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Dai Fukumura, Rakesh K. Jain, Anna M. Westermark, Alba Luengo, Ahmed Ali, Raphael Ferreira, Keene L. Abbott, Matthew G. Vander Heiden, Véronique Dartois, Kamila Naxerova, Todd R. Golub, Sylvie Roberge, Landry Blanc, Dan G. Duda, Gino B. Ferraro, Jessica M. Possada, Jiang Chen, Ivy X. Chen, Zohreh Amoozgar, Costas A. Lyssiotis, Amy Deik, Clary B. Clish, Shawn M. Davidson, Christopher W. Ng, Xin Jin, Mark Duquette, Christopher R. Chin, Divya Bezwada, Brendan Prideaux, and David P. Kodack
- Subjects
Cancer Research ,biology ,business.industry ,Cancer ,medicine.disease ,chemistry.chemical_compound ,Fatty acid synthase ,Breast cancer ,Oncology ,chemistry ,Cancer research ,biology.protein ,Medicine ,In patient ,Treatment resistance ,business ,Human Epidermal Growth Factor Receptor 2 ,Fatty acid synthesis ,Brain metastasis - Abstract
Brain metastases are refractory to therapies that otherwise control systemic disease in patients with human epidermal growth factor receptor 2 (HER2+) breast cancer, and the unique brain microenvironment contributes to this therapy resistance. Nutrient availability can vary across tissues, therefore metabolic adaptations required for breast cancer growth in the brain microenvironment may also introduce liabilities that can be exploited for therapy. Here, we assessed how metabolism differs between breast tumors growing in the brain versus extracranial sites and found that fatty acid synthesis is elevated in breast tumors growing in the brain. We determine that this phenotype is an adaptation to decreased lipid availability in the brain relative to other tissues, which results in a site-specific dependency on fatty acid synthesis for breast tumors growing at this site. Genetic or pharmacological inhibition of fatty acid synthase (FASN) reduces HER2+ breast tumor growth in the brain, demonstrating that differences in nutrient availability across metastatic sites can result in targetable metabolic dependencies. Citation Format: Gino B. Ferraro, Ahmed Ali, Alba Luengo, David P. Kodack, Amy Deik, Keene L. Abbott, Divya Bezwada, Landry Blanc, Brendan Prideaux, Xin Jin, Jessica M. Possada, Jiang Chen, Christopher R. Chin, Zohreh Amoozgar, Raphael Ferreira, Ivy Chen, Kamila Naxerova, Christopher Ng, Anna M. Westermark, Mark Duquette, Sylvie Roberge, Costas A. Lyssiotis, Dan G. Duda, Todd R. Golub, Shawn M. Davidson, Dai Fukumura, Véronique A. Dartois, Clary B. Clish, Matthew G. Vander Heiden, Rakesh K. Jain. Fatty acid synthesis is required for breast cancer brain metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 90.
- Published
- 2021
16. Metastasis-associated MCL1 and P16 copy number alterations dictate resistance to vemurafenib in a BRAFV600E patient-derived papillary thyroid carcinoma preclinical model
- Author
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G. Mike Makrigiorgos, Vania Nosé, Michelle Vinco, Thomas J. Giordano, Pier Paolo Pandolfi, Roderick T. Bronson, Chen Song, Paul Van Hummelen, Peter M. Sadow, Junichi Kurebayashi, Andrew H. Fischer, Zeus A. Antonello, Mark Duquette, Jennifer N. Sims, Carmelo Nucera, Elena Castellanos-Rizaldos, Amjad Husain, and Dora Dias-Santagata
- Subjects
Pathology ,Indoles ,endocrine system diseases ,medicine.medical_treatment ,Gene Dosage ,Apoptosis ,Mice, SCID ,Targeted therapy ,Metastasis ,Mice ,0302 clinical medicine ,Mice, Inbred NOD ,CDKN2A ,MCL1 ,Neoplasm Metastasis ,Vemurafenib ,Oligonucleotide Array Sequence Analysis ,Sulfonamides ,0303 health sciences ,Neovascularization, Pathologic ,vemurafenib resisatnce ,Immunohistochemistry ,BRAFV600E papillary thyroid cancer pre-clinical model ,3. Good health ,Oncology ,MCL1 and P16/CDKN2A somatic copy number ,Thyroid Cancer, Papillary ,030220 oncology & carcinogenesis ,Heterografts ,medicine.drug ,Proto-Oncogene Proteins B-raf ,medicine.medical_specialty ,Cell Survival ,Blotting, Western ,Antineoplastic Agents ,Enzyme-Linked Immunosorbent Assay ,Transfection ,Thyroid carcinoma ,03 medical and health sciences ,In Situ Nick-End Labeling ,medicine ,Carcinoma ,Animals ,Humans ,Thyroid Neoplasms ,neoplasms ,030304 developmental biology ,business.industry ,Genes, p16 ,medicine.disease ,microenvironment ,Carcinoma, Papillary ,digestive system diseases ,Disease Models, Animal ,Drug Resistance, Neoplasm ,Mutation ,Cancer research ,Myeloid Cell Leukemia Sequence 1 Protein ,business ,V600E ,Priority Research Paper - Abstract
BRAF(V600E) mutation exerts an essential oncogenic function in many tumors, including papillary thyroid carcinoma (PTC). Although BRAF(V600E) inhibitors are available, lack of response has been frequently observed. To study the mechanism underlying intrinsic resistance to the mutant BRAF(V600E) selective inhibitor vemurafenib, we established short-term primary cell cultures of human metastatic/recurrent BRAF(V600E)-PTC, intrathyroidal BRAF(V600E)-PTC, and normal thyroid (NT). We also generated an early intervention model of human BRAF(V600E)-PTC orthotopic mouse. We find that metastatic BRAF(V600E)-PTC cells elicit paracrine-signaling which trigger migration of pericytes, blood endothelial cells and lymphatic endothelial cells as compared to BRAF(WT)-PTC cells, and show a higher rate of invasion. We further show that vemurafenib therapy significantly suppresses these aberrant functions in non-metastatic BRAF(V600E)-PTC cells but lesser in metastatic BRAF(V600E)-PTC cells as compared to vehicle treatment. These results concur with similar folds of down-regulation of tumor microenvironment-associated pro-metastatic molecules, with no effects in BRAF(WT)-PTC and NT cells. Our early intervention preclinical trial shows that vemurafenib delays tumor growth in the orthotopic BRAF(WT/V600E)-PTC mice. Importantly, we identify high copy number gain of MCL1 (chromosome 1q) and loss of CDKN2A (P16, chromosome 9p) in metastatic BRAF(V600E)-PTC cells which are associated with resistance to vemurafenib treatment. Critically, we demonstrate that combined vemurafenib therapy with BCL2/MCL1 inhibitor increases metastatic BRAF(V600E)-PTC cell death and ameliorates response to vemurafenib treatment as compared to single agent treatment. In conclusion, short-term PTC and NT cultures offer a predictive model for evaluating therapeutic response in patients with PTC. Our PTC pre-clinical model suggests that combined targeted therapy might be an important therapeutic strategy for metastatic and refractory BRAF(V600E)-positive PTC.
- Published
- 2015
17. Abstract B06: The angiotensin receptor blocker and partial PPARγ agonist telmisartan inhibits the growth of pancreatic ductal adenocarcinoma
- Author
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Yves Boucher, Shiwei Han, Andrew S. Liss, Mark Duquette, Hao Liu, Daniel H. Schanne, Rakesh K. Jain, Matthew G. Vander Heiden, Alba Luengo, and Jelena Grahovac
- Subjects
Cancer Research ,Angiotensin receptor ,Angiotensin II receptor type 1 ,Chemistry ,Autophagy ,medicine.disease ,Angiotensin II ,Oncology ,Tumor progression ,Pancreatic cancer ,medicine ,Cancer research ,Telmisartan ,Receptor ,medicine.drug - Abstract
Pancreatic ductal adenocarcinoma (PDAC) is highly resistant to chemotherapy, partly due to the presence of a dense-fibrotic stroma and adaptive metabolism. Telmisartan is an angiotensin II type receptor 1 (AT1) antagonist with partial peroxisome proliferator-activated receptor gamma (PPARγ) agonistic activity used for treatment of hypertension. The aim of this study was to determine the effects of telmisartan on the viability of PDAC cells and tumor progression. In panels of 4 murine and 8 human PDAC cells, the telmisartan IC50 was lower in cells with a low steady-state expression of PPARγ and a mesenchymal cell morphology. In contrast, losartan—a selective AT1 inhibitor—did not affect the viability of PDAC cells. The siRNA knockdown of PPARγ enhanced the sensitivity of telmisartan and stimulated epithelial-mesenchymal transition, which was accompanied by an increase in Wnt signaling. PPARγ regulates glucose metabolism and autophagy. We thus assessed effects of telmisartan on bioenergetics and autophagy of PDAC cells. In PPARγ-knockdown and -scrambled cells telmisartan significantly reduced glucose uptake, without affecting ATP production, but increased respiratory capacity, which can maintain the production of ATP during hypoglycemia. Immunoblotting revealed that PPARγ knockdown compared to scramble cells had increased levels of phosphorylated-AMP-activated protein kinase (p-AMPK) and increased expression of LC3A/B—structural proteins of autophagosomal membranes—which implies higher levels of autophagy. We also compared effects of telmisartan treatment on LC3A/B expression to well-established autophagy modulators, chloroquine and verapamil. Under nutrient-rich conditions and as expected, chloroquine and verapamil treatment induced LC3A/B accumulation, consistent with active autophagic flux in these cells. Telmisartan treatment decreased the levels of LC3A/B in both scramble and PPARγ knockdown cells and decreased the formation of LC3A/B positive granules in other PDAC cell lines. Telmisartan can also induce the accumulation of the signal adaptor protein p62 (SQSTM1), even in the presence of verapamil, which is also consistent with autophagy inhibition. Telmisartan did not prevent the accumulation LC3A/B in the presence of chloroquine, implying that telmisartan acts after the autophagosome-lysosome fusion step. To assess the effects of telmisartan in vivo, we used an orthotopic PDAC model. Telmisartan monotherapy inhibited the growth of primary tumors, decreased the incidence of liver metastasis, and significantly improved the survival of mice. Hence, telmisartan can reduce autophagy and the viability of PDAC cells, and PDAC progression. Because telmisartan is an FDA-approved drug, our findings provide the scientific rationale for testing its efficacy in the prevention of PDAC progression. Citation Format: Jelena Grahovac, Shiwei Han, Hao Liu, Mark Duquette, Alba Luengo, Daniel Schanne, Andrew S. Liss, Matthew G. Vander Heiden, Rakesh K. Jain, Yves Boucher. The angiotensin receptor blocker and partial PPARγ agonist telmisartan inhibits the growth of pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr B06.
- Published
- 2019
18. Members of the thrombospondin gene family bind stromal interaction molecule 1 and regulate calcium channel activity
- Author
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Mark Duquette, Jack Lawler, Monica J. S. Nadler, Trevor J. Shuttleworth, Dayne Okuhara, and Jill L. Thompson
- Subjects
inorganic chemicals ,Patch-Clamp Techniques ,Blotting, Western ,chemistry.chemical_element ,Biology ,Calcium ,Mass Spectrometry ,Article ,Thrombospondin 1 ,Mice ,Animals ,Humans ,Immunoprecipitation ,Arachidonate-regulated calcium channel ,Thrombospondins ,Molecular Biology ,Calcium signaling ,Thrombospondin ,Voltage-dependent calcium channel ,HEK 293 cells ,Stromal interaction molecule 1 ,Membrane Proteins ,STIM1 ,Recombinant Proteins ,Neoplasm Proteins ,Cell biology ,HEK293 Cells ,Biochemistry ,Membrane protein ,chemistry ,Calcium Channels - Abstract
The thrombospondins (TSPs) are a family of matricellular proteins that regulate cellular phenotype through interactions with a myriad of other proteins and proteoglycans. We have identified a novel interaction of the members of the TSP gene family with stromal interaction molecule 1 (STIM1). This association is robust since it is preserved in Triton X-100, can be detected with multiple anti-TSP-1 and anti-STIM1 antibodies, and is detected in a wide range of cell types. We have also found that STIM1 co-immunoprecipitates with TSP-4 and cartilage oligomeric matrix protein (COMP), and that a recombinant version of the N-terminal domain of STIM1 binds to the signature domain of TSP-1 and COMP. The association of the TSPs with STIM1 is observed in both the presence and absence of calcium indicating that the calcium-dependent conformation of the signature domain of TSPs is not required for binding. Thus, this interaction could occur in the ER under conditions of normal or low calcium concentration. Furthermore, we observed that the expression of COMP in HEK 293 cells decreases STIM1-mediated calcium release activated calcium (CRAC) channel currents and increases arachidonic acid calcium (ARC) channel currents. These data indicate that the TSPs regulate STIM1 function and participate in the reciprocal regulation of two channels that mediate calcium entry into the cell.
- Published
- 2014
- Full Text
- View/download PDF
19. Abstract LB-057: Hypoxia regulation of antigen presentation machinery expression in breast cancer
- Author
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Elena F. Brachtel, Mark Badeaux, Kamila Naxerova, Soldano Ferrone, Gino B. Ferraro, Rakesh K. Jain, Divya Bezwada, Shan M. Chin, Mei Rosa Ng, Francesco Sabbatino, and Mark Duquette
- Subjects
Cancer Research ,Breast cancer ,Oncology ,business.industry ,Antigen presentation ,Cancer research ,Medicine ,Hypoxia (medical) ,medicine.symptom ,business ,medicine.disease - Abstract
Antigen presentation plays a major role in tumor cell recognition and targeting by immune cells, and is critical to the success of many cancer immunotherapies. How the abnormal tumor microenvironment affects tumor cell antigen presentation is unclear. Hypoxia is a prevalent feature of the tumor microenvironment. Here, we showed that the expression of major histocompatibility complex class I (MHCI) is associated with regions of hypoxia in human breast tumors. The association between hypoxia and MHCI is independent of the breast tumor hormone receptor and HER2 expression status. In vitro studies revealed that hypoxia directly regulates the expression levels of MHCI along with other components of the antigen presentation machinery. Multiple kinase regulators of MHCI expression are responsive to hypoxia. These results suggest that hypoxia effects on cancer cell antigen presentation may be a potential mechanism of tumor immune evasion and treatment resistance. Citation Format: Mei Rosa Ng, Francesco Sabbatino, Mark Duquette, Kamila Naxerova, Mark Badeaux, Gino B. Ferraro, Shan M. Chin, Divya Bezwada, Elena F. Brachtel, Soldano Ferrone, Rakesh K. Jain. Hypoxia regulation of antigen presentation machinery expression in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-057.
- Published
- 2018
20. A Double Hit to Kill Tumor and Endothelial Cells by TRAIL and Antiangiogenic 3TSR
- Author
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Jack Lawler, Xuefeng Zhang, Nordine Benhaga, Sareh Parangi, Min Ye, Robin Humphreys, Roya Khosravi-Far, Keli Song, Tai-Guang Jin, Mark Duquette, and Bin Ren
- Subjects
Cancer Research ,MAP Kinase Kinase 4 ,Lexatumumab ,Angiogenesis ,Mice, Nude ,Apoptosis ,Biology ,Vascular endothelial growth inhibitor ,Caspase 8 ,Article ,TNF-Related Apoptosis-Inducing Ligand ,Mice ,Antineoplastic Combined Chemotherapy Protocols ,medicine ,Animals ,Humans ,Protein kinase B ,Cells, Cultured ,Neovascularization, Pathologic ,Caspase 3 ,Antibodies, Monoclonal ,Endothelial Cells ,HCT116 Cells ,Xenograft Model Antitumor Assays ,Peptide Fragments ,Recombinant Proteins ,Cell biology ,Enzyme Activation ,Endothelial stem cell ,Receptors, TNF-Related Apoptosis-Inducing Ligand ,Oncology ,Colonic Neoplasms ,Female ,Tumor necrosis factor alpha ,Signal transduction ,Thrombospondins ,medicine.drug - Abstract
As tumor development relies on a coordination of angiogenesis and tumor growth, an efficient antitumor strategy should target both the tumor and its associated vessels. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in a tumor-selective manner. Additionally, thrombospondin-1, a naturally occurring inhibitor of angiogenesis, and a recombinant protein containing functional domains of thrombospondin-1, 3TSR, have been shown to be necessary and sufficient to inhibit tumor angiogenesis. Here, we show that a combination of a TRAIL receptor 2 agonist antibody, Lexatumumab, and 3TSR results in a significantly enhanced and durable tumor inhibition. We further observed that 3TSR induces apoptosis in primary endothelial cells by up-regulating the expression of TRAIL receptors 1 and 2 in a CD36 and Jun NH2-terminal kinase-dependent manner leading to the activation of both intrinsic and extrinsic apoptotic machineries. The modulation of these pathways is critical for 3TSR-induced apoptosis as disrupting either via specific inhibitors reduced apoptosis. Moreover, 3TSR attenuates the Akt survival pathway. These studies indicate that 3TSR plays a critical role in regulating the proapoptotic signaling pathways that control growth and death in endothelial cells and that a combination of TRAIL and 3TSR acts as a double hit against tumor and tumor-associated vessels. [Cancer Res 2009;69(9):3856–65]
- Published
- 2009
21. The crystal structure of the signature domain of cartilage oligomeric matrix protein: implications for collagen, glycosaminoglycan and integrin binding
- Author
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Jack Lawler, Kemin Tan, Andrzej Joachimiak, and Mark Duquette
- Subjects
Models, Molecular ,Integrins ,DNA, Complementary ,Static Electricity ,Integrin ,Plasma protein binding ,Cartilage Oligomeric Matrix Protein ,In Vitro Techniques ,Crystallography, X-Ray ,Ligands ,Biochemistry ,Research Communications ,Multiple epiphyseal dysplasia ,Thrombospondin 1 ,Genetics ,medicine ,Humans ,Matrilin Proteins ,Cysteine ,Binding site ,Thrombospondins ,Molecular Biology ,Glycoproteins ,Glycosaminoglycans ,Integrin binding ,Cartilage oligomeric matrix protein ,Extracellular Matrix Proteins ,Binding Sites ,Base Sequence ,biology ,Chemistry ,Cartilage ,medicine.disease ,Recombinant Proteins ,Protein Structure, Tertiary ,medicine.anatomical_structure ,Mutation ,Biophysics ,biology.protein ,Collagen ,Oligopeptides ,Protein Binding ,Biotechnology - Abstract
Cartilage oligomeric matrix protein (COMP), or thrombospondin-5 (TSP-5), is a secreted glycoprotein that is important for growth plate organization and function. Mutations in COMP cause two skeletal dysplasias, pseudoachondroplasia (PSACH) and multiple epiphyseal dysplasia (EDM1). In this study, we determined the structure of a recombinant protein that contains the last epidermal growth factor repeat, the type 3 repeats and the C-terminal domain (CTD) of COMP to 3.15-Å resolution limit by X-ray crystallography. The CTD is a β-sandwich that is composed of 15 antiparallel β-strands, and the type 3 repeats are a contiguous series of calcium binding sites that associate with the CTD at multiple points. The crystal packing reveals an exposed potential metal-ion-dependent adhesion site (MIDAS) on one edge of the β-sandwich that is common to all TSPs and may serve as a binding site for collagens and other ligands. Disease-causing mutations in COMP disrupt calcium binding, disulfide bond formation, intramolecular interactions, or sites for potential ligand binding. The structure presented here and its unique molecular packing in the crystal identify potential interactive sites for glycosaminoglycans, integrins, and collagens, which are key to cartilage structure and function.—Tan, K., Duquette, M., Joachimiak, A., Lawler, J. The crystal structure of the signature domain of cartilage oligomeric matrix protein: implications for collagen, glycosaminoglycan and integrin binding.
- Published
- 2009
22. The Crystal Structure of the Heparin-Binding Reelin-N Domain of F-Spondin
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Kemin Tan, Mark Duquette, Jia-huai Wang, Jin-huan Liu, and Jack Lawler
- Subjects
EGF-like domain ,Surface Properties ,Cell Adhesion Molecules, Neuronal ,Dimer ,Molecular Sequence Data ,Protein domain ,Protein Data Bank (RCSB PDB) ,Nerve Tissue Proteins ,Plasma protein binding ,Biology ,Crystallography, X-Ray ,Protein Structure, Secondary ,Article ,chemistry.chemical_compound ,Structural Biology ,Humans ,Amino Acid Sequence ,Binding site ,Molecular Biology ,Protein secondary structure ,Extracellular Matrix Proteins ,Binding Sites ,Heparin ,Serine Endopeptidases ,Protein Structure, Tertiary ,Reelin Protein ,Crystallography ,nervous system ,chemistry ,Biophysics ,Dimerization ,Sequence Alignment ,Protein Binding ,Binding domain - Abstract
The extracellular matrix protein F-spondin mediates axon guidance during neuronal development. Its N-terminal domain, termed the reelin-N domain, is conserved in F-spondins, reelins, and other extracellular matrix proteins. In this study, a recombinant human reelin-N domain has been expressed, purified, and shown to bind heparin. The crystal structure of the reelin-N domain resolved to 2.0 A reveals a variant immunoglobulin-like fold and potential heparin-binding sites. Substantial conformational variations even in secondary structure are observed between the two chemically identical reelin-N domains in one crystallographic asymmetric unit. The variations may result from extensive, highly specific interactions across the interface of the two reelin-N domains. The calculated values of buried surface area and the interface's shape complementarity are consistent with the formation of a weak dimer. The homophilic asymmetric dimer can potentially offer advantages in binding to ligands such as glycosaminoglycans, which may, in turn, bridge the two reelin-N domains and stabilize the dimer.
- Published
- 2008
23. The effect of thrombospondin-1 on breast cancer metastasis
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Raymond Washington, Jack Lawler, Shideh Kazerounian, Mark Duquette, Karen O. Yee, and Caitlin M. Connolly
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Genetically modified mouse ,endocrine system ,Cancer Research ,Pathology ,medicine.medical_specialty ,Angiogenesis ,Breast Neoplasms ,Mice, Transgenic ,Biology ,Article ,Metastasis ,Thrombospondin 1 ,Mice ,Cell Movement ,medicine ,Animals ,Neoplasm Metastasis ,Mammary tumor ,Angiogenesis Modulating Agents ,virus diseases ,Cancer ,Cell migration ,medicine.disease ,Primary tumor ,Disease Models, Animal ,Oncology ,Tumor progression ,Disease Progression ,Female - Abstract
Thrombospondin-1 (TSP-1) has been proposed to have both pro-metastatic and anti-metastatic properties. To elucidate its role in breast cancer metastasis, we compared tumor progression in the polyomavirus middle T antigen (Pyt) transgenic mouse and the TSP-1-null Pyt transgenic mouse. We characterized the tumors in these mice at 45, 60 and 90 days of age. Tumor size, areas of necrosis, macrophage infiltration, levels of active and total TGF-beta, vessel morphology, and lung and blood metastasis were measured in these mice. Mammary tumors were larger in the TSP-1-null mouse, and vessels were larger, but fewer in number in these tumors. The level of total TGF-beta was significantly higher in the Pyt tumors at 90 days of age. Importantly, significantly fewer metastases were observed in the lungs of the TSP-1-null/Pyt mouse. Primary Pyt tumor cells were more migratory than TSP-1-null Pyt tumor cells on collagen. Treatment of Pyt mice with recombinant proteins that contain the type-1 repeats of TSP-1 resulted in decreased primary tumor growth and metastasis. Sequences that are involved in CD36 binding and those required for TGF-beta activation mediated the inhibition of primary tumor growth. Thus, TSP-1 in the mammary tumor microenvironment inhibits angiogenesis and tumor growth, but promotes metastasis to the lung in the Pyt transgenic mouse. The ability of TSP-1 to support metastasis correlates with its ability to promote tumor cell migration.
- Published
- 2008
24. Continuous administration of the three thrombospondin-1 type 1 repeats recombinant protein improves the potency of therapy in an orthotopic human pancreatic cancer model
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Mark Duquette, Xuefeng Zhang, Sareh Parangi, Caitlin M. Connolly, and Jack Lawler
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Cancer Research ,Angiogenesis ,Angiogenesis Inhibitors ,Pharmacology ,Article ,law.invention ,Thrombospondin 1 ,Mice ,Text mining ,In vivo ,law ,Pancreatic cancer ,medicine ,Animals ,Humans ,Potency ,Bovine serum albumin ,Neovascularization, Pathologic ,biology ,business.industry ,medicine.disease ,Xenograft Model Antitumor Assays ,Recombinant Proteins ,Pancreatic Neoplasms ,Oncology ,Immunology ,biology.protein ,Recombinant DNA ,business - Abstract
Thrombospondin-1 is one of most important natural angiogenic inhibitors. The three thrombospondin-1 type 1 repeats (3TSR), an anti-angiogenic domain of thrombospondin-1, is a promising novel agent for anti-angiogenic treatment. In the present study, we showed 3TSR was biologically stable at least for 7 days in mini-osmotic pumps in vivo, and continuous administration of 3TSR decreased the dosage and improved the potency of therapy in an orthotopic pancreatic cancer model. By using different dosage and delivery routes, we proved that the anti-tumor efficacy of 3TSR was correlated with its anti-angiogenic efficacy. 3TSR treatment also decreased tumor vessel patency and blood flow. The results indicate the advantage of continuous administration of angiogenic inhibitors and provide rationale for using such delivery methods for cancer treatment.
- Published
- 2007
25. The Structures of the Thrombospondin-1 N-Terminal Domain and Its Complex with a Synthetic Pentameric Heparin
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Rongguang Zhang, Kemin Tan, Andrzej Joachimiak, Mark Duquette, Jia-huai Wang, Jack Lawler, and Jin-huan Liu
- Subjects
Integrin ,Molecular Sequence Data ,Antiparallel (biochemistry) ,Crystallography, X-Ray ,Protein Structure, Secondary ,Article ,Thrombospondin 1 ,03 medical and health sciences ,Polysaccharides ,Structural Biology ,Concanavalin A ,Humans ,Amino Acid Sequence ,Binding site ,Cell adhesion ,Peptide sequence ,Molecular Biology ,030304 developmental biology ,0303 health sciences ,Binding Sites ,biology ,Chemistry ,030302 biochemistry & molecular biology ,Peptide Fragments ,Protein Structure, Tertiary ,Biochemistry ,Fondaparinux ,G-domain ,Structural Homology, Protein ,biology.protein ,Biophysics ,Crystallization ,Linker ,Sequence Alignment - Abstract
Summary The N-terminal domain of thrombospondin-1 (TSPN-1) mediates the protein's interaction with (1) glycosaminoglycans, calreticulin, and integrins during cellular adhesion, (2) low-density lipoprotein receptor-related protein during uptake and clearance, and (3) fibrinogen during platelet aggregation. The crystal structure of TSPN-1 to 1.8 A resolution is a β sandwich with 13 antiparallel β strands and 1 irregular strand-like segment. Unique structural features of the N- and C-terminal regions, and the disulfide bond location, distinguish TSPN-1 from the laminin G domain and other concanavalin A-like lectins/glucanases superfamily members. The crystal structure of the complex of TSPN-1 with heparin indicates that residues R29, R42, and R77 in an extensive positively charged patch at the bottom of the domain specifically associate with the sulfate groups of heparin. The TSPN-1 structure and identified adjacent linker region provide a structural framework for the analysis of the TSPN domain of various molecules, including TSPs, NELLs, many collagens, TSPEAR, and kielin.
- Published
- 2006
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26. Combined therapy with thrombospondin-1 type I repeats (3TSR) and chemotherapy induces regression and significantly improves survival in a preclinical model of advanced stage epithelial ovarian cancer
- Author
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Jim Petrik, S. Russell, Ronny Drapkin, Mark Duquette, Jack Lawler, and Joyce Ying Liu
- Subjects
CD36 Antigens ,Pathology ,medicine.medical_specialty ,Maximum Tolerated Dose ,Cell Survival ,CD36 ,medicine.medical_treatment ,Angiogenesis Inhibitors ,Antineoplastic Agents ,Apoptosis ,Biochemistry ,Cell Line ,Thrombospondin 1 ,Mice ,Research Communication ,In vivo ,Cell Line, Tumor ,Genetics ,medicine ,Animals ,Humans ,Hypoxia ,Molecular Biology ,Survival rate ,Cell Proliferation ,Ovarian Neoplasms ,Chemotherapy ,biology ,Cluster of differentiation ,business.industry ,Protein Tyrosine Phosphatase, Non-Receptor Type 6 ,Metronomic Chemotherapy ,Combined Modality Therapy ,Mice, Inbred C57BL ,Receptors, Vascular Endothelial Growth Factor ,Treatment Outcome ,biology.protein ,Cancer research ,Female ,business ,Neoplasm Transplantation ,Biotechnology - Abstract
Most women are diagnosed with epithelial ovarian cancer (EOC) at advanced stage, where therapies have limited effectiveness and the long-term survival rate is low. We evaluated the effects of combined antiangiogenic and chemotherapy treatments on advanced stage EOC. Treatment of EOC cells with a recombinant version of the thrombospondin-1 type I repeats (3TSR) induced more apoptotic cell death (36.5 ± 9.6%) in vitro compared to untreated controls (4.1 ± 1.4). In vivo, tumors were induced in an orthotopic, syngeneic mouse model of advanced stage EOC. Mice were treated with 3TSR (4 mg/kg per day) alone or in combination with chemotherapy drugs delivered with maximum tolerated dose or metronomic scheduling. Pretreatment with 3TSR induced tumor regression, normalized tumor vasculature, and improved uptake of chemotherapy drugs. Combination 3TSR and metronomic chemotherapy induced the greatest tumor regression (6.2-fold reduction in size compared to PBS-treated controls) and highest survival when treatment was initiated at advanced stage. 3TSR binding to its receptor, CD36 (cluster of differentiation 36), increased binding of CD36 and SHP-1, which significantly inhibited phosphorylation of the VEGF receptor. In this study, we describe a novel treatment approach and mechanism of action with 3TSR and chemotherapy that induces regression of advanced stage EOC and significantly improves survival.-Russell, S., Duquette, M., Liu, J., Drapkin, R., Lawler, J., Petrik, J. Combined therapy with thrombospondin-1 type I repeats (3TSR) and chemotherapy induces regression and significantly improves survival in a preclinical model of advanced stage epithelial ovarian cancer.
- Published
- 2014
27. Thrombospondin-1 Modulates Actin Filament Remodeling and Cell Motility in Mouse Mammary Tumor cells in Vitro
- Author
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Cameron A. Duquette, Jack Lawler, Shideh Kazerounian, Dorothy Ndishabandi, Mark Duquette, Ghita El-Moatassim Billah, and Millys A. Reyes
- Subjects
0303 health sciences ,Tumor microenvironment ,Mammary tumor ,endocrine system ,Stromal cell ,Angiogenesis ,virus diseases ,Biology ,medicine.disease ,Article ,3. Good health ,Metastasis ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,immune system diseases ,030220 oncology & carcinogenesis ,Cancer cell ,Thrombospondin 1 ,Cancer research ,medicine ,Actin filament polymerization ,030304 developmental biology - Abstract
It is well established that the secretion of thrombospondin-1 (TSP-1) by activated stromal cells and its accumulation in the tumor microenvironment during dysplasia inhibits primary tumor growth through inhibition of angiogenesis. This inhibitory function of TSP-1 is actuated either by inhibiting MMP9 activation and the release of VEGF from extracellular matrix or by an interaction with CD36 on the surface of endothelial cells resulting in an increase in apoptosis. In contrast, several published articles have also shown that as tumor cells become more invasive and enter the early stage of carcinoma, they up-regulate TSP-1 expression, which may promote invasion and migration. In our in vivo studies using the polyoma middle T antigen (PyT) transgenic mouse model of breast cancer, we observed that the absence of TSP-1 significantly increased the growth of primary tumors, but delayed metastasis to the lungs. In this study, we propose a mechanism for the promigratory function of TSP-1 in mouse mammary tumor cells in vitro. We demonstrate the correlations between expression of TSP-1 and its receptor integrin α3β1, which is considered a promigratory protein in cancer cells. In addition we propose that binding of TSP-1 to integrin α3β1 is important for mediating actin filament polymerization and therefore, cell motility. These findings can help explain the dual functionality of TSP-1 in cancer progression.
- Published
- 2014
28. Thrombospondin-1 Gene Expression Affects Survival and Tumor Spectrum of p53-Deficient Mice
- Author
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Jack Lawler, Richard O. Hynes, Mark Duquette, Wei-Min Miao, Roderick T. Bronson, and Noel P. Bouck
- Subjects
Male ,Teratocarcinoma ,Stromal cell ,Genotype ,Tumor suppressor gene ,Gene Expression ,Loss of Heterozygosity ,Biology ,Pathology and Forensic Medicine ,Thrombospondin 1 ,Mice ,Testicular Neoplasms ,In vivo ,medicine ,Animals ,Melanoma ,Mice, Knockout ,Cell growth ,Neoplasms, Experimental ,medicine.disease ,Mice, Inbred C57BL ,Tumor progression ,Apoptosis ,Cancer research ,Female ,Tumor Suppressor Protein p53 ,Neoplasm Transplantation ,Regular Articles - Abstract
In vitro and in vivo data indicate that thrombospondin-1 (TSP1) inhibits tumor progression in several ways including direct effects on cellular growth and apoptosis in the stromal compartment. To evaluate the importance of TSP1 for the progression of naturally arising tumors in vivo, we have crossed TSP1-deficient mice with p53-deficient mice. In p53-null mice, the absence of TSP1 decreases survival from 160 +/- 52 days to 149 +/- 42 days. A log-rank test comparing survival curves for these two populations yields a two-sided P value of 0.0272. For mice that are heterozygous for the p53-null allele, survival is 500 +/- 103 days in the presence of TSP1 expression, and 426 +/- 125 days in its absence (P = 0.0058). Whereas TSP1 expression did not cause a measurable change in the incidence of the majority of tumor types, a statistically significant (Por = 0.05) decrease in the incidence of osteosarcomas is observed in the absence of TSP1. To determine more directly if host TSP1 inhibits tumor growth, B16F10 melanoma and F9 testicular teratocarcinoma cells have been implanted in C57BL/6J and 129Sv TSP1-null mice, respectively. The B16F10 tumors grow approximately twice as fast in the TSP1-null background and exhibit an increase in vascular density, a decrease in the rate of tumor cell apoptosis, and an increase in the rate of tumor cell proliferation. Increased tumor growth is also observed in the absence of TSP1 on the 129Sv genetic background. These data indicate that endogenous host TSP1 functions as a modifier or landscaper gene to suppress tumor growth.
- Published
- 2001
29. Thrombospondin-1 is required for normal murine pulmonary homeostasis and its absence causes pneumonia
- Author
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Elizabeth George, Jack Lawler, Helen Rayburn, Mark Duquette, Valérie Thibert, Richard O. Hynes, and Mary E. Sunday
- Subjects
Pathology ,Platelet Aggregation ,Neutrophils ,Restriction Mapping ,Monocytes ,Thrombospondin 1 ,Extracellular matrix ,Leukocyte Count ,Mice ,Homeostasis ,Lung ,Cells, Cultured ,Mice, Knockout ,Recombination, Genetic ,biology ,Thrombin ,virus diseases ,Diffuse alveolar hemorrhage ,General Medicine ,medicine.anatomical_structure ,Lordosis ,Collagen ,Research Article ,Blood Platelets ,endocrine system ,medicine.medical_specialty ,Genetic Vectors ,Hemorrhage ,Spleen ,Hemosiderin ,Transfection ,Congenital Abnormalities ,Ribonucleases ,medicine ,Animals ,Thrombospondins ,Thrombospondin ,Hyperplasia ,Macrophages ,Proteins ,Epithelial Cells ,DNA ,Pneumonia ,Elastin ,Eosinophils ,Radiography ,biology.protein - Abstract
The thrombospondins are a family of extracellular calcium-binding proteins that modulate cellular phenotype. Thrombospondin-1 (TSP-1) reportedly regulates cellular attachment, proliferation, migration, and differentiation in vitro. To explore its function in vivo, we have disrupted the TSP-1 gene by homologous recombination in the mouse genome. Platelets from these mice are completely deficient in TSP-1 protein; however, thrombin-induced platelet aggregation is not diminished. TSP-1-deficient mice display a mild and variable lordotic curvature of the spine that is apparent from birth. These mice also display an increase in the number of circulating white blood cells, with monocytes and eosinophils having the largest percent increases. The brain, heart, kidney, spleen, stomach, intestines, aorta, and liver of TSP-1-deficient mice showed no major abnormalities. However, consistent with high levels of expression of TSP-1 in lung, we observe abnormalities in the lungs of mice that lack the protein. Although normal at birth, histopathological analysis of lungs from 4-wk-old TSP-1-deficient mice reveals extensive acute and organizing pneumonia, with neutrophils and macrophages. The macrophages stain for hemosiderin, indicating that diffuse alveolar hemorrhage is occurring. At later times, the number of neutrophils decreases and a striking increase in the number of hemosiderin-containing macrophages is observed associated with multiple-lineage epithelial hyperplasia and the deposition of collagen and elastin. A thickening and ruffling of the epithelium of the airways results from increasing cell proliferation in TSP-1-deficient mice. These results indicate that TSP-1 is involved in normal lung homeostasis.
- Published
- 1998
30. Thrombospondin-1 Silencing Down-Regulates Integrin Expression Levels in Human Anaplastic Thyroid Cancer Cells with BRAFV600E: New Insights in the Host Tissue Adaptation and Homeostasis of Tumor Microenvironment
- Author
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Carmelo Nucera, Peter M. Sadow, Jack Lawler, and Mark Duquette
- Subjects
Integrins ,endocrine system ,Pathology ,medicine.medical_specialty ,Stromal cell ,Endocrinology, Diabetes and Metabolism ,Cell ,Cell morphology ,lcsh:Diseases of the endocrine glands. Clinical endocrinology ,03 medical and health sciences ,Endocrinology ,BRAFV600E ,0302 clinical medicine ,immune system diseases ,thyroid cancer ,medicine ,TSP-1 ,Gene silencing ,Anaplastic thyroid cancer ,Thyroid cancer ,Original Research ,030304 developmental biology ,0303 health sciences ,Tumor microenvironment ,lcsh:RC648-665 ,business.industry ,Correction ,virus diseases ,Cancer ,medicine.disease ,microenvironment ,Extracellular Matrix ,3. Good health ,medicine.anatomical_structure ,030220 oncology & carcinogenesis ,Cancer research ,business - Abstract
Background and Rationale: Anaplastic thyroid cancer (ATC) is characterized by pleomorphic cells, has a poor prognosis, is highly devastating disease, and is not curable. No reliable biomarkers of metastatic potential, helpful for early diagnosis of ATC and therapeutic response have been found yet. Thrombospondin-1 (TSP-1) plays a fundamental role in cancer progression by regulating cell stromal cross-talk in the tumor microenvironment. Goals: Our goal was to understand whether TSP-1 could affect protein levels of its integrin receptors (e.g., ITGα3, α6, and β1) and cell morphology in BRAFV600E-ATC cells in vitro and in vivo. Experimental Design: Anaplastic thyroid cancer-derived cell cultures and western blotting were used to assess integrin protein expression upon TSP-1 silencing. Immunohistochemistry was performed on orthotopic primary human ATC and metastatic ATC in lung tissue to compare TSP-1 and integrin protein expression levels. Results: TSP-1 knock-down down-regulates ITGα3, α6, and β1 in BRAFV600E-human ATC cells. BRAFV600E-ATC cells with TSP-1 knock-down were rounded compared to control cells, which displayed a spread morphology. TSP-1 knock-down also reduced TSP-1, ITGα3, α6, and β1 protein expression levels in vivo in the ATC microenvironment, which is enriched in stromal and inflammatory cells. Conclusion: TSP-1 silencing causes changes in ITG levels and ATC cell morphology. The assessment of TSP-1 and ITG levels might contribute to earlier metastatic potential of BRAFV600E-positive aggressive thyroid cancers, and allow improved patient selection for clinical trials.
- Published
- 2013
31. Characterization of Human Thrombospondin-4
- Author
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Mark Duquette, Jack Lawler, Laura H. Derick, and Katherine McHenry
- Subjects
Pentameric protein ,Molecular Sequence Data ,chemistry.chemical_element ,Calcium ,Biology ,Biochemistry ,law.invention ,Mice ,Xenopus laevis ,Affinity chromatography ,law ,Thrombospondin 4 ,Calcium-binding protein ,Complementary DNA ,Animals ,Humans ,Trypsin ,Amino Acid Sequence ,Cloning, Molecular ,education ,Thrombospondins ,Molecular Biology ,education.field_of_study ,Membrane Glycoproteins ,Base Sequence ,Hydrolysis ,Calcium-Binding Proteins ,3T3 Cells ,Cell Biology ,Molecular biology ,Recombinant Proteins ,Microscopy, Electron ,chemistry ,Recombinant DNA - Abstract
The thrombospondins are a family of extracellular calcium binding proteins that are involved in cell proliferation, adhesion, and migration. We have sequenced full-length human thrombospondin-4 and characterized the recombinant protein. In contrast to Xenopus laevis thrombospondin-4, the human protein contains an RGD cell binding sequence in the third type 3 repeat. Transfection of mouse NIH3T3 fibroblasts or C2C12 myoblasts with a full-length human thrombospondin-4 cDNA results in the expression of a polypeptide with a reduced molecular weight of 140,000. In the absence of reducing agent, the expressed protein has an apparent molecular weight of 550,000. Recombinant thrombospondin-4 has been purified from the culture supernatant by heparin-Sepharose and anti-thrombospondin-4 antibody-Affi-gel affinity chromatography. Electron microscopy indicates that thrombospondin-4 is composed of five subunits with globular domains at each end. The observation of a calcium-dependent change in the electron microscopic appearance of thrombospondin-4 is consistent with limited tryptic digestion data that indicate that thrombospondin-4 is resistant to digestion in the presence of calcium. These data indicate that thrombospondin-4 is a pentameric protein that binds to heparin and calcium.
- Published
- 1995
32. Abstract A12: Microenvironmental Interleukin-6 induced activation of STAT3 signaling in medulloblastoma: Implications for molecular pathology and therapy
- Author
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Trupti Vardam, Tai Hato, Sylvie Roberge, Vasileios Askoxylakis, Dai Fukumura, Sampurna Chatterjee, Rakesh K. Jain, Shuji Kitahara, Shuang Yan, Mark Duquette, and Lei Xu
- Subjects
Medulloblastoma ,Cancer Research ,Molecular pathology ,Biology ,medicine.disease ,medicine.disease_cause ,Pediatric cancer ,Metastasis ,Oncology ,medicine ,Cancer research ,STAT protein ,biology.protein ,Carcinogenesis ,STAT3 ,Janus kinase - Abstract
Aberrant activation of Janus kinase (JAK) / signal transducer and activator of transcription (STAT) 3 pathway has been implicated in tumor initiation, progression and metastasis which make it a potential therapeutic target in human malignancies. In vitro STAT3 inhibition exhibited anti-tumor effect in medulloblastoma (MB) cell lines; however, little has been reported about the mechanisms of STAT3 activation in medulloblastoma (MB). We found that STAT3 was constitutively activated in both medulloblastoma patient samples and 3 mice models that were clinically classified as SHH subgroup, Group 3 and Group 4 in pediatric MB patients. By using PCR arrays we investigated the early expression profile of local cytokines in host cerebellum during MB pathogenesis and found increased IL17B, IL6 and TNFα in the microenvironment of MB mice models. In vitro cytokine stimulation assay indicated that IL6 was the major activator of STAT3 in D283 MB cells. Confocal immunofluorescence staining of different cell components in the cerebellum in D283 xenografts showed that the endothelial cells and Purkinje cells were the two major sources of IL6 secretion. Our data provides molecular and cellular clue for the activation of STAT3 signaling during MB tumorigenesis and might help to develop more effective therapeutic targets for STAT3 blockade in MB. Citation Format: Shuang Yan, Sampurna Chatterjee, Trupti Vardam, Shuji Kitahara, Tai Hato, Sylvie Roberge, Vasileios Askoxylakis, Mark Duquette, Dai Fukumura, Lei Xu, Rakesh Jain. Microenvironmental Interleukin-6 induced activation of STAT3 signaling in medulloblastoma: Implications for molecular pathology and therapy. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Pediatric Cancer Research: From Mechanisms and Models to Treatment and Survivorship; 2015 Nov 9-12; Fort Lauderdale, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(5 Suppl):Abstract nr A12.
- Published
- 2016
33. Thrombospondin‐1 binding protein on the surface of Trypanosoma cruzi enhances cellular infection
- Author
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Maria F. Lima, Tatiana C. Cardenas, Fernando Villalta, Mark Duquette, Pius N. Nde, Siddharth Pratap, Candice A. Johnson, and Jack Lawler
- Subjects
biology ,Chemistry ,Binding protein ,Thrombospondin 1 ,Genetics ,Trypanosoma cruzi ,biology.organism_classification ,Molecular Biology ,Biochemistry ,Biotechnology ,Cell biology - Published
- 2012
34. Priming of the vascularendothelial growth factor signaling pathway by thrombospondin-1, CD36, and spleentyrosine kinase
- Author
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Isaac Rabinovitz, Millys A. Reyes, Carole A. Perruzzi, Luca Primo, Shideh Kazerounian, Federico Bussolino, Mark Duquette, Keli Song, James T. Lawler, Roya Khosravi-Far, and Jack Lawler
- Subjects
CD36 Antigens ,Vascular Endothelial Growth Factor A ,Umbilical Veins ,Angiogenesis ,Immunology ,Syk ,Neovascularization, Physiologic ,Biology ,Biochemistry ,environment and public health ,Thrombospondin 1 ,chemistry.chemical_compound ,angiogenesis ,Vascular Biology ,Cell Movement ,parasitic diseases ,Humans ,Syk Kinase ,Phosphorylation ,Cells, Cultured ,cell adhesion ,Intracellular Signaling Peptides and Proteins ,Endothelial Cells ,hemic and immune systems ,Cell Biology ,Hematology ,Tyrosine-Protein Kinase SYK ,Protein-Tyrosine Kinases ,Vascular Endothelial Growth Factor Receptor-2 ,Up-Regulation ,Vascular endothelial growth factor ,Vascular endothelial growth factor A ,enzymes and coenzymes (carbohydrates) ,chemistry ,Cancer research ,Signal transduction ,biological phenomena, cell phenomena, and immunity ,Signal Transduction - Abstract
CD36 plays a critical role in the inhibition of angiogenesis through binding to the type 1 repeats of thrombospondin-1 (TSP-1) and activating Fyn tyrosine kinase and MAPK pathways. Here, we reveal a novel association of CD36 with VEGFR-2 and spleen tyrosine kinase (Syk). We also address the correlation between the expression of CD36 and Syk by demonstrating that overexpression of CD36 in HUVECs up-regulates endogenous Syk expression. We also define a new role for TSP-1 and CD36 in the activation of the VEGFR-2 signaling pathway that requires Syk. Our findings also identify a role for Syk as a stimulator of VEGF-A–induced angiogenesis by increasing phosphorylation of Y1175 in VEGFR-2, which is a major tyrosine for promoting VEGF-A–induced endothelial cell migration. Together, these studies introduce a new signaling pathway for TSP-1, CD36, and Syk, and address the role of these proteins in regulating the angiogenic switch.
- Published
- 2011
35. Identification and characterization of thrombospondin-4, a new member of the thrombospondin gene family
- Author
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Jack Lawler, Douglas W. DeSimone, Charles A. Whittaker, Katherine McHenry, Josephine C. Adams, and Mark Duquette
- Subjects
Molecular Sequence Data ,Restriction Mapping ,Xenopus ,Gene Expression ,Platelet Membrane Glycoproteins ,Biology ,Polymerase Chain Reaction ,Xenopus laevis ,Thrombospondin 4 ,Gene expression ,Thrombospondin 1 ,Gene family ,Animals ,Humans ,Northern blot ,Amino Acid Sequence ,RNA, Messenger ,Cloning, Molecular ,education ,Gene ,Repetitive Sequences, Nucleic Acid ,Thrombospondin ,education.field_of_study ,Base Sequence ,Cell Biology ,Articles ,biology.organism_classification ,Molecular biology ,Genes ,Oligodeoxyribonucleotides ,Multigene Family ,Thrombospondins ,Sequence Alignment - Abstract
A new member of the thrombospondin gene family, designated thrombospondin-4, has been identified in the Xenopus laevis genome. The predicted amino acid sequence indicates that the protein is similar to the other members of this gene family in the structure of the type 3 repeats and the COOH-terminal domain. Thrombospondin-4 contains four type 2 repeats and lacks the type 1 repeats that are found in thrombospondin-1 and 2. The amino-terminal domain of thrombospondin-4 has no significant homology with the other members of the thrombospondin gene family or with other proteins in the database. RNAse protection analysis establishes that the initial expression of Xenopus thrombospondin-4 is observed during neurulation. Levels of mRNA expression increase twofold during tailbud stages but decrease by the feeding tadpole stage. The size of the thrombospondin-4 message is 3.3 Kb and 3.4 Kb in the frog and human, respectively. Northern blot analysis of human tissues reveals high levels of thrombospondin-4 expression in heart and skeletal muscle, low levels in brain, lung and pancreas and undetectable levels in the placenta, liver and kidney. These data establish the existence of a new member of the thrombospondin gene family that may participate in the genesis and function of cardiac and skeletal muscle.
- Published
- 1993
36. Thrombospondin-1 modulates vascular endothelial growth factor activity at the receptor level
- Author
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Shideh Kazerounian, Carole A. Perruzzi, Mark Duquette, Jack Lawler, Sareh Parangi, Janice A. Nagy, Harold F. Dvorak, and Xuefeng Zhang
- Subjects
Vascular Endothelial Growth Factor A ,Angiogenesis ,Vascular permeability ,Biology ,Vascular endothelial growth inhibitor ,Biochemistry ,Research Communications ,Capillary Permeability ,Thrombospondin 1 ,chemistry.chemical_compound ,Mice ,Genetics ,Animals ,Humans ,Molecular Biology ,S1PR1 ,Integrin beta1 ,Cell Membrane ,Vascular Endothelial Growth Factor Receptor-2 ,Mice, Mutant Strains ,Cell biology ,Vascular endothelial growth factor ,Vascular endothelial growth factor B ,Vascular endothelial growth factor A ,chemistry ,Vascular endothelial growth factor C ,Immunology ,Receptors, Complement 3b ,Endothelium, Vascular ,Biotechnology - Abstract
Vascular endothelial growth factor (VEGF) is a well-established stimulator of vascular permeability and angiogenesis, whereas thrombospondin-1 (TSP-1) is a potent angiogenic inhibitor. In this study, we have found that the TSP-1 receptors CD36 and β1 integrin associate with the VEGF receptor 2 (VEGFR2). The coclustering of receptors that regulate angiogenesis may provide the endothelial cell with a platform for integration of positive and negative signals in the plane of the membrane. Thus, this complex may represent a molecular switch that regulates angiogenesis and determines endothelial cell behavior. In this context, physiological levels of TSP-1 appear to support VEGFR2 function on both the cellular and tissue level, because phosphorylation of VEGFR2 and vascular permeability in response to VEGF are decreased in TSP-1-null mice and isolated endothelial cells. A therapeutic agent based on the antiangiogenic domain of TSP-1, designated 3TSR (for three TSP-1 type 1 repeats), has significant antiangiogenic and antitumor efficacy. Systemic treatment of wild-type mice with 3TSR significantly decreased VEGF-induced permeability. Consistent with this result, VEGF-stimulated phosphorylation of VEGFR2 was also significantly decreased in lung extracts from 3TSR-treated mice. Moreover, 3TSR significantly decreased VEGF-stimulated VEGFR2 phosphorylation in human dermal microvascular endothelial cells in culture. Taken together, the results indicate that TSP-1 and 3TSR modulate the function of VEGFR2.—Zhang, X., Kazerounian, S., Duquette, M., Perruzzi, C., Nagy, J. A., Dvorak, H. J., Parangi, S., and Lawler, J. Thrombospondin-1 modulates vascular endothelial growth factor activity at the receptor level.
- Published
- 2009
37. Characterization of the murine thrombospondin gene
- Author
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Jack Lawler, Debra J. Gilbert, Mark Duquette, Neal G. Copeland, Paula Ferro, and Nancy A. Jenkins
- Subjects
Untranslated region ,Molecular Sequence Data ,CAAT box ,Platelet Membrane Glycoproteins ,Biology ,Mice ,Exon ,Lectins ,Sequence Homology, Nucleic Acid ,Consensus Sequence ,Genetics ,Consensus sequence ,Animals ,Humans ,Coding region ,Amino Acid Sequence ,Cloning, Molecular ,Promoter Regions, Genetic ,Gene ,Crosses, Genetic ,Repetitive Sequences, Nucleic Acid ,Regulation of gene expression ,Thrombospondin ,Base Sequence ,Exons ,Molecular biology ,Mice, Inbred C57BL ,Gene Expression Regulation ,Thrombospondins - Abstract
Thrombospondin is an adhesive glycoprotein that supports cell attachment, spreading, and migration. The murine thrombospondin gene is approximately 18 kb in length and includes 22 exons. Interspecific backcross analysis using progeny derived from matings of (C57BL/6J x Mus spretus) F1 x C57BL/6J mice indicates that the thrombospondin gene is tightly linked to the Fshb, Actcl, Ltk, and B2M loci on murine chromosome 2. The sequence of the murine gene is very similar to that of the human gene in (1) regions of the promoter, (2) the coding region, and (3) the 3'-untranslated region. The predicted amino acid sequence of the mature murine thrombospondin subunit is 95.1% identical to that of the human. The sequences of these two species are most similar at the regions containing the type 1, 2, and 3 repeats as well as the COOH-terminal globular domain. The thrombospondin promoter is similar to the 5' flanking region of some housekeeping and growth control genes in that it contains multiple GC-rich regions and lacks a CAAT box. The presence of various consensus sequences suggests that thrombospondin gene expression is regulated by cAMP, cytokines, and steroid hormones.
- Published
- 1991
38. Heparin-induced cis- and trans-dimerization modes of the thrombospondin-1 N-terminal domain
- Author
-
Jack Lawler, Kumaran Shanmugasundaram, John T. Gallagher, Andrzej Joachimiak, Jia-huai Wang, Mark Duquette, Kemin Tan, Jin-huan Liu, and Alan C. Rigby
- Subjects
Models, Molecular ,Stereochemistry ,Protein Conformation ,Dimer ,Crystallography, X-Ray ,Biochemistry ,Article ,Glycosaminoglycan ,Extracellular matrix ,Cell membrane ,Thrombospondin 1 ,chemistry.chemical_compound ,Protein structure ,medicine ,Humans ,Molecular Biology ,Heparin ,Cell Biology ,Recombinant Proteins ,medicine.anatomical_structure ,chemistry ,Chromatography, Gel ,Crystallization ,Dimerization ,Cis–trans isomerism ,medicine.drug - Abstract
Through its interactions with proteins and proteoglycans, thrombospondin-1 (TSP-1) functions at the interface of the cell membrane and the extracellular matrix to regulate matrix structure and cellular phenotype. We have previously determined the structure of the high affinity heparin-binding domain of TSP-1, designated TSPN-1, in association with the synthetic heparin, Arixtra. To establish that the binding of TSPN-1 to Arixtra is representative of the association with naturally occurring heparins, we have determined the structures of TSPN-1 in complex with heparin oligosaccharides containing eight (dp8) and ten (dp10) subunits, by x-ray crystallography. We have found that dp8 and dp10 bind to TSPN-1 in a manner similar to Arixtra and that dp8 and dp10 induce the formation of trans and cis TSPN-1 dimers, respectively. In silico docking calculations partnered with our crystal structures support the importance of arginine residues in positions 29, 42, and 77 in binding sulfate groups of the dp8 and dp10 forms of heparin. The ability of several TSPN-1 domains to bind to glycosaminoglycans simultaneously probably increases the affinity of binding through multivalent interactions. The formation of cis and trans dimers of the TSPN-1 domain with relatively short segments of heparin further enhances the ability of TSP-1 to participate in high affinity binding to glycosaminoglycans. Dimer formation may also involve TSPN-1 domains from two separate TSP-1 molecules. This association would enable glycosaminoglycans to cluster TSP-1.
- Published
- 2007
39. Antiangiogenic treatment with three thrombospondin-1 type 1 repeats versus gemcitabine in an orthotopic human pancreatic cancer model
- Author
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Eric Galardi, Jack Lawler, Sareh Parangi, Mark Duquette, and Xuefeng Zhang
- Subjects
Cancer Research ,Pancreatic disease ,Time Factors ,Angiogenesis Inhibitors ,Apoptosis ,Mice, SCID ,Deoxycytidine ,Thrombospondin 1 ,Mice ,Medicine ,Hypoxia ,Neovascularization, Pathologic ,Flow Cytometry ,Immunohistochemistry ,Recombinant Proteins ,Platelet Endothelial Cell Adhesion Molecule-1 ,medicine.anatomical_structure ,Oncology ,Tumor necrosis factor alpha ,Female ,Pancreas ,medicine.drug ,medicine.medical_specialty ,Antimetabolites, Antineoplastic ,medicine.drug_class ,Molecular Sequence Data ,Antimetabolite ,Necrosis ,Internal medicine ,Pancreatic cancer ,Cell Line, Tumor ,In Situ Nick-End Labeling ,Animals ,Humans ,Amino Acid Sequence ,Cell Proliferation ,business.industry ,Microcirculation ,Endothelial Cells ,medicine.disease ,Gemcitabine ,Protein Structure, Tertiary ,Pancreatic Neoplasms ,Endocrinology ,Microscopy, Fluorescence ,Cancer cell ,Cancer research ,business ,Neoplasm Transplantation - Abstract
Purpose: In this study, we investigated the antitumor efficacy of thrombospondin-1 three type 1 repeats (3TSR), the antiangiogenic domain of thrombospondin-1, in comparison and in combination with gemcitabine, in an orthotopic pancreatic cancer model. Experimental Design: Human pancreatic cancer cells were injected into the pancreas of severe combined immunodeficient mice. The animals were treated with 3TSR, gemcitabine, 3TSR plus gemcitabine, or vehicle for 3 weeks. Subsequently, the effects of 3TSR and/or gemcitabine on tumor growth, tumor necrosis, microvessel density, cancer cell proliferation, apoptosis, and endothelial cell apoptosis were analyzed. Results: After 3 weeks of treatment, 3TSR reduced tumor volume by 65%, and gemcitabine by 84%. Tumor volume was not statistically different between gemcitabine group and combinatorial treatment group. Extensive necrotic areas were observed in tumors from 3TSR-treated mice, whereas tumors from gemcitabine and combinatorially treated mice were less necrotic than control tumors. 3TSR reduced tumor microvessel density and increased tumor blood vessel endothelial cell apoptosis. In contrast, gemcitabine induced apoptosis and inhibited proliferation of cancer cells. Conclusion: 3TSR, the antiangiogenic domain of thrombospondin-1, showed comparable antitumor efficacy to gemcitabine in a human pancreatic cancer orthotopic mouse model. No synergistic effect was found when the two drugs were combined and possible reasons are discussed in detail. A delicate balance between normalization and excessive regression of tumor vasculature is important when initiating alternative combinatorial regimens for treatment of patients with pancreatic cancer.
- Published
- 2005
40. Antiangiogenic treatment with the three thrombospondin-1 type 1 repeats recombinant protein in an orthotopic human pancreatic cancer model
- Author
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Xuefeng Zhang, Marko Delic, Sareh Parangi, Mark Duquette, Jack Lawler, and Eric Galardi
- Subjects
Cancer Research ,Pathology ,medicine.medical_specialty ,Pancreatic disease ,Angiogenesis ,Angiogenesis Inhibitors ,Apoptosis ,Mice, SCID ,Neovascularization ,Thrombospondin 1 ,Mice ,Pancreatic cancer ,medicine ,In Situ Nick-End Labeling ,Tumor Cells, Cultured ,Animals ,Humans ,Microvessel ,Cell Proliferation ,Repetitive Sequences, Nucleic Acid ,Neovascularization, Pathologic ,business.industry ,Endothelial Cells ,medicine.disease ,Xenograft Model Antitumor Assays ,Recombinant Proteins ,Angiogenesis inhibitor ,Pancreatic Neoplasms ,medicine.anatomical_structure ,Treatment Outcome ,Oncology ,Cancer research ,Female ,medicine.symptom ,business ,Pancreas - Abstract
Purpose: This study investigates the antiangiogenesis and antitumor efficacy of a recombinant protein composed of the three type 1 repeats (3TSR) of thrombospondin-1 in an orthotopic human pancreatic cancer model and provides useful preclinical data for pancreatic cancer treatment. Experimental Design: Human pancreatic cancer cells (AsPC-1) were injected into the pancreas of severe combined immunodeficient mice. The animals were treated with 3TSR (3 mg per kg per day) or PBS for 3 weeks. Subsequently, the effects of 3TSR on tumor growth, microvessel density, cancer cell proliferation, apoptosis, and endothelial cell apoptosis were analyzed. The in vitro effects of 3TSR on human pancreatic cancer cells were also studied. Results: 3TSR treatment significantly reduced angiogenesis and tumor growth of orthotopic pancreatic cancer. 3TSR-treated mice had a 69% reduction in tumor volume (316.6 ± 79.3 versus 1,012.2 ± 364.5 mm3; P = 0.0001), and a significant increase in tumor necrotic area. After 3TSR treatment, both the vessel number and average microvessel size were significantly decreased, and microvessel density was decreased from 8.0% to 3.7% (P < 0.0001). The apoptotic rate of tumoral endothelial cells in 3TSR-treated tumors increased to 14.7% comparing to 4.2% in control tumors (P < 0.0001). 3TSR showed no direct effects on pancreatic cancer cell proliferation or apoptosis either in vivo or in vitro. Conclusion: 3TSR, a domain of a natural occurring angiogenesis inhibitor, showed potent therapeutic effect in pancreatic cancer by inhibiting tumor angiogenesis and may prove to be a promising agent for clinical pancreatic cancer treatment.
- Published
- 2005
41. Purification and Analysis of Thrombospondin‐1
- Author
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Jack Lawler, Anna Ludlow, Mark Duquette, and Karen O. Yee
- Subjects
Blood Platelets ,endocrine system ,Cell signaling ,biology ,Angiogenesis ,Molecular Sequence Data ,Matricellular protein ,virus diseases ,Cell migration ,Cell Biology ,Transforming growth factor beta ,Adhesion ,Peptide Fragments ,Cell biology ,Thrombospondin 1 ,immune system diseases ,Methods ,biology.protein ,Humans ,Tumor growth ,Amino Acid Sequence ,Peptide Hydrolases - Abstract
Thromboapondin 1 (TSP-1) is a trimeric matricellular protein that is expressed by many cells. It contains several different domains that allow it to participate in cell adhesion, cell migration, and cell signaling. Recently TSP-1 has been shown to activate transforming growth factor beta (TGF-beta) and to inhibit both angiogenesis and tumor growth. This unit contains protocols for the purification of TSP-1 from platelet-rich plasma and the purification of TSP-1 proteolytic fragments.
- Published
- 2003
42. Crystal structure of the TSP-1 type 1 repeats: a novel layered fold and its biological implication
- Author
-
Yicheng Dong, Jia-huai Wang, Kemin Tan, Jack Lawler, Jin-huan Liu, Rongguang Zhang, Andrzej Joachimiak, and Mark Duquette
- Subjects
endocrine system ,Protein Folding ,Stereochemistry ,Molecular Sequence Data ,Crystal structure ,Biology ,thrombospondin ,TSR domain ,X-ray structure ,angiogenesis ,GAG binding ,Antiparallel (biochemistry) ,Crystallography, X-Ray ,Article ,Thrombospondin 1 ,03 medical and health sciences ,Structure-Activity Relationship ,0302 clinical medicine ,Structure–activity relationship ,Animals ,Humans ,Amino Acid Sequence ,Binding site ,Cell adhesion ,Peptide sequence ,030304 developmental biology ,0303 health sciences ,Glycosaminoglycan binding ,Binding Sites ,Comment ,Cell Biology ,3. Good health ,Protein Structure, Tertiary ,Biochemistry ,030220 oncology & carcinogenesis ,Protein folding ,Extracellular Space ,Signal Transduction - Abstract
In this issue, Tan et al. (2002) report the first high resolution (1.9 A) structural data for thrombospondin (TSP)-1, a large multifunctional protein that regulates cell adhesion, angiogenesis, cell proliferation and survival, TGFbeta activation, and protease function (for review see Chen et al., 2000). Because TSP-1 has multiple binding partners and many functions, precise structural information is crucial to understanding its biology. The structure now reported, derived from crystals of the second and third type I repeats of TSP-1 is of particular interest because of the specific functions attributed to these repeats and because domains homologous to the repeats appear in many other proteins in nature. The novel layered fold motif described brings great insight into how the complicated functions of TSP-1 and related molecules are affected.
- Published
- 2002
43. Comparison of antiangiogenic therapy with thrombospondin type I repeats and gemcitabine in an orthotopic model of pancreatic cancer in SCID mice
- Author
-
Xuefeng Zhang, Eric Galardi, Sareh Parangi, Mark Duquette, Jack Lawler, and Xue Zhang
- Subjects
Thrombospondin ,medicine.medical_specialty ,Tumor size ,business.industry ,Antiangiogenic therapy ,Urology ,Scid mice ,medicine.disease ,Gemcitabine ,law.invention ,law ,Pancreatic cancer cell ,Pancreatic cancer ,Immunology ,medicine ,Recombinant DNA ,Surgery ,business ,medicine.drug - Abstract
Introduction: Pancreatic cancer is a devastating disease. Efficacy of type I repeats of thrombospondin (TSP-1) in mice with orthotopic pancreatic cancer was tested and compared to gemcitabine. Methods: SCID mice treated with an intrapancreatic injection of a human pancreatic cancer cell line (ASPC-1) developed large orthotopic tumors with local invasion, peritoneal and and mesenteric implants. Recombinant Thrombospondin Type I repeats (TSR), one of the antiangiogenic domains of TSP, was cloned and purified. One week after tumor implantation, mice were treated with 3TSR (n = 9, 0.3 mg/kg/day), gemcitabine (n = 6, 15 mg/kg, twice weekly) or the combination of the two drugs (n = 11) for 3 weeks. Subsequently, mouse weight, tumor size, number of metasteses were compared. Tumor volume was calculated using the formula 0.52 (width) 2 (length). Student's t test (Mann-Whitney) was used for statistical comparison. Results: Tumor bearing mice treated with TSR had a 65% reduction in tumor volume (371 ± 97 vs 1064 ± 375 mm 3 , p 3 ) and 79% with gemcitabine plus TSR treatment (219.39 ± 78.37 mm 3 ) p 0.05 when compared to eachother. Mean vessel density was lowest in the TSR treated animals (3.74 ± 1.2) and highest in the control group (7.96 ± 6.7), p Conclusions: Antiangiogenic therapy using TSR reduces tumor volume and vascular density in mice with orthotopic pancreatic tumors and can be used safely alone or in combination with gemcitabine.
- Published
- 2004
44. Thrombospondin-1 Interacts with Trypanosoma cruzi Surface Calreticulin to Enhance Cellular Infection
- Author
-
Adaeze O. Ikejiani, Aniekanabasi N Udoko, Pius N. Nde, Jack Lawler, Fernando Villalta, Siddharth Pratap, Candice A. Johnson, Tatiana C. Cardenas, Mark Duquette, Maria F. Lima, and Yulia Y Kleshchenko
- Subjects
Proteomics ,lcsh:Medicine ,Pathogenesis ,Plasma protein binding ,Protozoology ,Biochemistry ,Thrombospondin 1 ,Mice ,0302 clinical medicine ,immune system diseases ,Molecular Cell Biology ,Cloning, Molecular ,lcsh:Science ,0303 health sciences ,Multidisciplinary ,biology ,virus diseases ,3. Good health ,Cell biology ,Host-Pathogen Interaction ,Infectious Diseases ,Medicine ,Research Article ,Neglected Tropical Diseases ,Protein Binding ,Chagas disease ,Trypanosoma ,endocrine system ,Immunoprecipitation ,Trypanosoma cruzi ,030231 tropical medicine ,Microbiology ,03 medical and health sciences ,parasitic diseases ,Parasitic Diseases ,medicine ,Animals ,Chagas Disease ,Protein Interactions ,Biology ,030304 developmental biology ,Life Cycle Stages ,Staining and Labeling ,Cell Membrane ,lcsh:R ,Proteins ,medicine.disease ,biology.organism_classification ,Mice, Inbred C57BL ,Membrane protein ,biology.protein ,Parastic Protozoans ,Parasitology ,lcsh:Q ,Calreticulin - Abstract
Trypanosoma cruzi causes Chagas disease, which is a neglected tropical disease that produces severe pathology and mortality. The mechanisms by which the parasite invades cells are not well elucidated. We recently reported that T. cruzi up-regulates the expression of thrombospondin-1 (TSP-1) to enhance the process of cellular invasion. Here we characterize a novel TSP-1 interaction with T. cruzi that enhances cellular infection. We show that labeled TSP-1 interacts specifically with the surface of T. cruzi trypomastigotes. We used TSP-1 to pull down interacting parasite surface proteins that were identified by mass spectrometry. We also show that full length TSP-1 and the N-terminal domain of TSP-1 (NTSP) interact with T. cruzi surface calreticulin (TcCRT) and other surface proteins. Pre-exposure of recombinant NTSP or TSP-1 to T. cruzi significantly enhances cellular infection of wild type mouse embryo fibroblasts (MEF) compared to the C-terminal domain of TSP-1, E3T3C1. In addition, blocking TcCRT with antibodies significantly inhibits the enhancement of cellular infection mediated by the TcCRT-TSP-1 interaction. Taken together, our findings indicate that TSP-1 interacts with TcCRT on the surface of T. cruzi through the NTSP domain and that this interaction enhances cellular infection. Thus surface TcCRT is a virulent factor that enhances the pathogenesis of T. cruzi infection through TSP-1, which is up-regulated by the parasite.
- Published
- 2012
45. The evolution of the thrombospondin gene family
- Author
-
Temple F. Smith, Jack Lawler, Lisa A. Urry, Mark Duquette, and Katherine McHenry
- Subjects
Molecular Sequence Data ,Sequence alignment ,Platelet Membrane Glycoproteins ,Biology ,Mice ,Xenopus laevis ,Species Specificity ,Molecular evolution ,Phylogenetics ,Gene duplication ,Genetics ,Gene family ,Animals ,Humans ,Amino Acid Sequence ,Molecular Biology ,Gene ,Ecology, Evolution, Behavior and Systematics ,Phylogeny ,Thrombospondin ,Phylogenetic tree ,Base Sequence ,Sequence Homology, Amino Acid ,Genes ,Multigene Family ,Thrombospondins ,Chickens ,Sequence Alignment - Abstract
Thrombospondin-1 is an adhesive glycoprotein that is involved in cellular attachment, spreading, migration, and proliferation. To date, four genes have been identified that encode for the members of the thrombospondin gene family. These four genes are homologous to each other in the EGF-like (type 2) repeats, the calcium-binding (type 3) motifs, and the COOH-terminal. The latter has been reported to be a cell-binding domain in thrombospondin-1. Phylogenetic trees have been constructed from the multisequence alignment of thrombospondin sequences from human, mouse, chicken, and frog. Two different algorithms generate comparable results in terms of the topology and the branch lengths. The analysis indicates that an early form of the thrombospondin gene duplicated about 925 million years ago. The gene duplication that produced the thrombospondin-1 and -2 branches of the family is predicted to have occurred 583 million years ago, whereas the gene duplication that produced the thrombospondin-3 and -4 branches of the family is predicted to have occurred 644 million years ago. These results indicate that the members of the thrombospondin gene family have existed throughout the evolution of the animal kingdom and thus probably participate in functions that are common to most of its members.
- Published
- 1993
46. Expression and mutagenesis of thrombospondin
- Author
-
Paula Ferro, Mark Duquette, and Jack Lawler
- Subjects
Protein subunit ,Genetic Vectors ,Molecular Sequence Data ,Gene Expression ,Platelet Membrane Glycoproteins ,Biology ,Transfection ,Biochemistry ,Mice ,Complementary DNA ,Thrombospondin 1 ,Animals ,Humans ,CD36 antigen ,Amino Acid Sequence ,Endothelium ,Site-directed mutagenesis ,Repetitive Sequences, Nucleic Acid ,chemistry.chemical_classification ,Thrombospondin ,Base Sequence ,Heparin ,3T3 Cells ,Molecular biology ,Molecular Weight ,chemistry ,Cell culture ,Mutagenesis, Site-Directed ,Chromosome Deletion ,Glycoprotein ,Carrier Proteins ,Thrombospondins - Abstract
Thrombospondin is a 420,000-dalton adhesive glycoprotein that is composed of three subunits of equivalent molecular weight. When the cDNA for the complete coding region of the human endothelial cell thrombospondin subunit is expressed in mouse NIH 3T3 cells, a 420,000-dalton protein is synthesized and secreted. The expressed protein comigrates with human platelet thrombospondin both in the presence and in the absence of a reducing agent. The expressed protein binds to a monoclonal anti-thrombospondin antibody, heparin, and calcium. In addition to the 420,000-dalton protein, the transfected cell lines also express a variable amount of a 140,000-dalton polypeptide. When the culture supernatants that are produced by cells that are expressing thrombospondin are applied to heparin-Sepharose, the 420,000-dalton and the 140,000-dalton proteins are bound to the column and are eluted with buffer containing 0.55 and 0.3 M NaCl, respectively. The 140,000-dalton protein only binds to heparin-Sepharose in the presence of calcium. Deletion of the region of homology with procollagen results in defective assembly of the trimer. Deletion of the type 1 or type 2 repeats results in decreased stability of the subunit with the predominant polypeptides that are expressed having molecular weights of 127,000 and 130,000, respectively. These polypeptides retain low-affinity heparin-binding activity. High-affinity heparin binding is markedly diminished by mutations in either of two sequence motifs that include clusters of lysines and arginines.(ABSTRACT TRUNCATED AT 250 WORDS)
- Published
- 1992
47. Cloning and sequencing of chicken thrombospondin
- Author
-
Jack Lawler, P Ferro, and Mark Duquette
- Subjects
Molecular Sequence Data ,Restriction Mapping ,Chick Embryo ,Platelet Membrane Glycoproteins ,Biology ,Molecular cloning ,Biochemistry ,Mice ,Molecular evolution ,Cell surface receptor ,Complementary DNA ,Sequence Homology, Nucleic Acid ,Animals ,Humans ,Amino Acid Sequence ,Cloning, Molecular ,Molecular Biology ,Gene Library ,Repetitive Sequences, Nucleic Acid ,Genetics ,chemistry.chemical_classification ,Thrombospondin ,Base Sequence ,Heparin ,Nucleic acid sequence ,Cell Biology ,DNA ,Biological Evolution ,chemistry ,Sequence motif ,Glycoprotein ,Thrombospondins - Abstract
Thrombospondin is a multifunction adhesive protein with the ability to bind proteoglycans, cell surface receptors, other proteins, and calcium ions. Several sequence motifs for some of these interactions have been identified in human thrombospondin. To evaluate the potential functional significance of these sequences and to begin a study of the evolution of thrombospondin, we have isolated and sequenced thrombospondin cDNA clones from a chicken embryo library. Comparison of the chicken and human sequences reveals that the NH2-terminal heparin-binding domains are only 34% identical. By contrast, the type 3 repeats and the COOH-terminal domains are 80 and 82% identical, respectively, when comparing human and chicken sequences. Potential cell recognition sequences of RGD and VTCG are conserved, with the chicken sequence containing an additional copy of the VTCG sequence. Whereas substitutions occur in the two potential heparin-binding motifs that have human counterparts, the chicken sequence contains a third potential heparin-binding motif. The results indicate that the evolutionary constraints on the various types of cell-binding motifs may be quite different.
- Published
- 1991
48. Role of BRAFV600E in the first preclinical model of multifocal infiltrating myopericytoma development and microenvironment
- Author
-
Carmen Priolo, Pier Paolo Pandolfi, Mark Duquette, Seum Chung, Guido Fadda, Peter M. Sadow, Antonina M. De Angelis, Carmelo Nucera, Heinz Kutzner, Oreste Durante, John Parker, Jack Lawler, John G. Clohessy, Simona Nanni, Harold F. Dvorak, Christopher V. Carman, Antonella Farsetti, Alfredo Pontecorvi, Florian A. Karreth, Roberta Martinelli, Thomas Mentzel, Amjad Husain, Celestino Pio Lombardi, Elizabeth P. Henske, Laura E. MacConaill, Christopher D.M. Fletcher, and Emanuele Palescandolo
- Subjects
Cancer Research ,Pathology ,Indoles ,endocrine system diseases ,Angiogenesis ,Myopericytoma ,Angiogenesis Inhibitors ,Mass Spectrometry ,Pathogenesis ,THYROID ,immune system diseases ,Vemurafenib ,Tumor Markers ,Sulfonamides ,Tumor ,Thyroid ,Valine ,medicine.anatomical_structure ,Oncology ,Local ,Tumor Markers, Biological ,Stem cell ,medicine.drug ,Hemangiopericytoma ,Proto-Oncogene Proteins B-raf ,medicine.medical_specialty ,Genotype ,Glutamic Acid ,Biology ,Brief Communication ,BRAF ,Cell Line ,Cell Line, Tumor ,Biomarkers, Tumor ,medicine ,Adjuvant therapy ,Humans ,Thyroid Neoplasms ,neoplasms ,Cell Proliferation ,MYOPERICYTOMA ,Settore MED/06 - ONCOLOGIA MEDICA ,medicine.disease ,Biological ,Xenograft Model Antitumor Assays ,digestive system diseases ,genetic aberration ,BRAF mutation ,Neoplasm Recurrence ,Mutation ,Myopericytoma Development ,Neoplasm Recurrence, Local ,Pericytes ,V600E - Abstract
Myopericytoma (MPC) is a rare tumor with perivascular proliferation of pluripotent stem-cell-like pericytes. Although indolent, MPC may be locally aggressive with recurrent disease. The pathogenesis and diagnostic biomarkers of MPC are poorly understood. We discovered that 15% of benign MPCs (thyroid, skin; 3 of 20 samples) harbored BRAF(WT/V600E); 33.3% (1 of 3 samples) of BRAF(WT/V600E)-MPCs were multifocal/infiltrative/recurrent. Patient-MPC and primary MPC cells harbored BRAF(WT/V600E), were clonal and expressed pericytic-differentiation biomarkers crucial for its microenvironment. BRAF(WT/V600E)-positive thyroid MPC primary cells triggered in vitro (8.8-fold increase) and in vivo (3.6-fold increase) angiogenesis. Anti-BRAF(V600E) therapy with vemurafenib disrupted angiogenic and metabolic properties (~3-fold decrease) with down-regulation (~2.2-fold decrease) of some extracellular-matrix (ECM) factors and ECM-associated long non-coding RNA (LincRNA) expression, with no effects in BRAF(WT)-pericytes. Vemurafenib also inhibited (~3-fold decrease) cell viability in vitro and in BRAF(WT/V600E)-positive thyroid MPC patient-derived xenograft (PDX) mice (n = 5 mice per group). We established the first BRAF(WT/V600E)-dependent thyroid MPC cell culture. Our findings identify BRAF(WT/V600E) as a novel genetic aberration in MPC pathogenesis and MPC-associated biomarkers and imply that anti-BRAF(V600E) agents may be useful adjuvant therapy in BRAF(WT/V600E)-MPC patients. Patients with BRAF(WT/V600E)-MPC should be closely followed because of the risk for multifocality/recurrence.
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