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Article ; Online: Netrin-1 Stimulates Migration of Neogenin Expressing Aggressive Melanoma Cells.

Untiveros, Gustavo / Raskind, Aleksandr / Linares, Laura / Dotti, Alessandro / Strizzi, Luigi

International journal of molecular sciences

2022 Volume 23, Issue 21

Abstract: Netrin-1 is a neural guidance factor that regulates migration and positioning of neural crest-derived cells during embryonic development. Depending on the type of Netrin-1 receptor expression, cells are either attracted or repulsed by Netrin-1. Postnatal ...

Abstract	Netrin-1 is a neural guidance factor that regulates migration and positioning of neural crest-derived cells during embryonic development. Depending on the type of Netrin-1 receptor expression, cells are either attracted or repulsed by Netrin-1. Postnatal expression of Netrin-1 is detected in brain, colon, liver, and kidney, which are common sites of cancer metastasis, including melanoma. Thus, understanding the dynamics between Netrin-1 and its receptors could explain the attraction of melanoma towards these Netrin-1-expressing tissues. Here, we investigate whether the Netrin-1-attractive receptor Neogenin can affect migration of melanoma cells towards a Netrin-1 source. Results from Western blot (WB) analysis show higher expression of Neogenin in aggressive compared to non-aggressive melanoma cells. Cell migration experiments show increased migration of Neogenin-expressing aggressive melanoma cells towards exogenous, soluble recombinant human Netrin-1 and towards a Netrin-1-expressing cell line. Furthermore, WB reveals ERK1/2 activation and increased N-cadherin expression in Neogenin-expressing aggressive melanoma cells treated with rhNetrin-1. Moreover, treatment with anti-Neogenin blocking antibody caused decreased migration towards Netrin-1-expressing cells and reduced ERK1/2 activity in Neogenin-expressing aggressive melanoma cells. These results suggest Neogenin may play a role during migration of melanoma cells towards Netrin-1 via ERK1/2 signaling.
MeSH term(s)	Humans ; Cell Line ; Cell Movement/physiology ; Melanoma/genetics ; Nerve Growth Factors/metabolism ; Netrin-1 ; Transcription Factors
Chemical Substances	Nerve Growth Factors ; Netrin-1 (158651-98-0) ; Transcription Factors
Language	English
Publishing date	2022-10-22
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms232112751
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Article ; Online: Netrin-1 Stimulates Migration of Neogenin Expressing Aggressive Melanoma Cells

Gustavo Untiveros / Aleksandr Raskind / Laura Linares / Alessandro Dotti / Luigi Strizzi

International Journal of Molecular Sciences, Vol 23, Iss 12751, p

2022 Volume 12751

Abstract	Netrin-1 is a neural guidance factor that regulates migration and positioning of neural crest-derived cells during embryonic development. Depending on the type of Netrin-1 receptor expression, cells are either attracted or repulsed by Netrin-1. Postnatal expression of Netrin-1 is detected in brain, colon, liver, and kidney, which are common sites of cancer metastasis, including melanoma. Thus, understanding the dynamics between Netrin-1 and its receptors could explain the attraction of melanoma towards these Netrin-1-expressing tissues. Here, we investigate whether the Netrin-1-attractive receptor Neogenin can affect migration of melanoma cells towards a Netrin-1 source. Results from Western blot (WB) analysis show higher expression of Neogenin in aggressive compared to non-aggressive melanoma cells. Cell migration experiments show increased migration of Neogenin-expressing aggressive melanoma cells towards exogenous, soluble recombinant human Netrin-1 and towards a Netrin-1-expressing cell line. Furthermore, WB reveals ERK1/2 activation and increased N-cadherin expression in Neogenin-expressing aggressive melanoma cells treated with rhNetrin-1. Moreover, treatment with anti-Neogenin blocking antibody caused decreased migration towards Netrin-1-expressing cells and reduced ERK1/2 activity in Neogenin-expressing aggressive melanoma cells. These results suggest Neogenin may play a role during migration of melanoma cells towards Netrin-1 via ERK1/2 signaling.
Keywords	Netrin-1 ; Neogenin ; migration ; aggressive melanoma cells ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Subject code	570 ; 612
Language	English
Publishing date	2022-10-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
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Article ; Online: Normal Skin Cells Increase Aggressiveness of Cutaneous Melanoma by Promoting Epithelial-to-Mesenchymal Transition via Nodal and Wnt Activity.

Untiveros, Gustavo / Dezi, Lindsay / Gillette, Megan / Sidor, Julia / Strizzi, Luigi

International journal of molecular sciences

2021 Volume 22, Issue 21

Abstract: Melanoma is a lethal form of skin cancer triggered by genetic and environmental factors. Excision of early-stage, poorly aggressive melanoma often leads to a successful outcome; however, left undiagnosed these lesions can progress to metastatic disease. ... ...

Abstract	Melanoma is a lethal form of skin cancer triggered by genetic and environmental factors. Excision of early-stage, poorly aggressive melanoma often leads to a successful outcome; however, left undiagnosed these lesions can progress to metastatic disease. This research investigates whether the exposure of poorly aggressive melanoma to certain normal skin cells can explain how non-metastatic melanoma becomes more aggressive while still confined to the skin. To this end, we used a serial co-culture approach to sequentially expose cells from two different, poorly aggressive human melanoma cell lines against normal cells of the skin beginning with normal melanocytes, then epidermal keratinocytes, and finally dermal fibroblasts. Protein extraction of melanoma cells occurred at each step of the co-culture sequence for western blot (WB) analysis. In addition, morphological and functional changes were assessed to detect differences between the serially co-cultured melanoma cells and non-co-cultured cells. Results show that the co-cultured melanoma cells assumed a more mesenchymal morphology and displayed a significant increase in proliferation and invasiveness compared to control or reference cells. WB analysis of protein from the co-cultured melanoma cells showed increased expression of Snail and decreased levels of E-cadherin suggesting that epithelial-to-mesenchymal transition (EMT) is occurring in these co-cultured cells. Additional WB analysis showed increased levels of Nodal protein and signaling and signs of increased Wnt activity in the co-cultured melanoma cells compared to reference cells. These data suggest that interaction between poorly aggressive melanoma cells with normal cells of the skin may regulate the transition from localized, poorly aggressive melanoma to invasive, metastatic disease via Nodal and/or Wnt induced EMT.
MeSH term(s)	Cell Line ; Cell Line, Tumor ; Coculture Techniques ; Epithelial-Mesenchymal Transition ; Humans ; Melanoma/metabolism ; Melanoma/pathology ; Nodal Protein/metabolism ; Skin/cytology ; Skin/metabolism ; Skin/pathology ; Skin Neoplasms/metabolism ; Skin Neoplasms/pathology ; Wnt Signaling Pathway ; Melanoma, Cutaneous Malignant
Chemical Substances	NODAL protein, human ; Nodal Protein
Language	English
Publishing date	2021-10-29
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms222111719
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Article ; Online: Role of Presenilin-1 in Aggressive Human Melanoma.

Sidor, Julia / Gillette, Megan / Dezi, Lindsay Ann / Untiveros, Gustavo / Strizzi, Luigi

International journal of molecular sciences

2022 Volume 23, Issue 9

Abstract: Presenilin-1 (PS-1), a component of the gamma (γ)-secretase catalytic complex, has been implicated in Alzheimer's disease (AD) and in tumorigenesis. Interestingly, AD risk is inversely related to melanoma, suggesting that AD-related factors, such as PS-1, ...

Abstract	Presenilin-1 (PS-1), a component of the gamma (γ)-secretase catalytic complex, has been implicated in Alzheimer's disease (AD) and in tumorigenesis. Interestingly, AD risk is inversely related to melanoma, suggesting that AD-related factors, such as PS-1, may affect melanomagenesis. PS-1 has been shown to reduce Wnt activity by promoting degradation of beta-catenin (β-catenin), an important Wnt signaling partner. Since Wnt is known to enhance progression of different cancers, including melanoma, we hypothesized that PS-1 could affect Wnt-associated melanoma aggressiveness. Western blot results showed that aggressive melanoma cells expressed significantly lower levels of both PS-1 and phosphorylated-β-catenin (P-β-catenin) than nonaggressive melanoma cells. Immunohistochemistry of human melanoma samples showed significantly reduced staining for PS-1 in advanced stage melanoma compared with early stage melanoma. Furthermore, γ-secretase inhibitor (GSI) treatment of aggressive melanoma cells was followed by significant increases in PS-1 and P-β-catenin levels, suggesting impaired Wnt signaling activity as PS-1 expression increased. Finally, a significant reduction in cell migration was associated with the higher levels of PS-1 and P-β-catenin in the GSI-treated aggressive melanoma cells. We demonstrate for the first time that PS-1 levels can be used to assess melanoma aggressiveness and suggest that by enhancing PS-1 expression, Wnt-dependent melanoma progression may be reduced.
MeSH term(s)	Alzheimer Disease ; Amyloid Precursor Protein Secretases/metabolism ; Humans ; Melanoma/genetics ; Melanoma/metabolism ; Presenilin-1/genetics ; Presenilin-1/metabolism ; Wnt Signaling Pathway ; beta Catenin/metabolism
Chemical Substances	Presenilin-1 ; beta Catenin ; Amyloid Precursor Protein Secretases (EC 3.4.-)
Language	English
Publishing date	2022-04-28
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms23094904
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Article ; Online: Normal Skin Cells Increase Aggressiveness of Cutaneous Melanoma by Promoting Epithelial-to-Mesenchymal Transition via Nodal and Wnt Activity

Gustavo Untiveros / Lindsay Dezi / Megan Gillette / Julia Sidor / Luigi Strizzi

International Journal of Molecular Sciences, Vol 22, Iss 11719, p

2021 Volume 11719

Abstract	Melanoma is a lethal form of skin cancer triggered by genetic and environmental factors. Excision of early-stage, poorly aggressive melanoma often leads to a successful outcome; however, left undiagnosed these lesions can progress to metastatic disease. This research investigates whether the exposure of poorly aggressive melanoma to certain normal skin cells can explain how non-metastatic melanoma becomes more aggressive while still confined to the skin. To this end, we used a serial co-culture approach to sequentially expose cells from two different, poorly aggressive human melanoma cell lines against normal cells of the skin beginning with normal melanocytes, then epidermal keratinocytes, and finally dermal fibroblasts. Protein extraction of melanoma cells occurred at each step of the co-culture sequence for western blot (WB) analysis. In addition, morphological and functional changes were assessed to detect differences between the serially co-cultured melanoma cells and non-co-cultured cells. Results show that the co-cultured melanoma cells assumed a more mesenchymal morphology and displayed a significant increase in proliferation and invasiveness compared to control or reference cells. WB analysis of protein from the co-cultured melanoma cells showed increased expression of Snail and decreased levels of E-cadherin suggesting that epithelial-to-mesenchymal transition (EMT) is occurring in these co-cultured cells. Additional WB analysis showed increased levels of Nodal protein and signaling and signs of increased Wnt activity in the co-cultured melanoma cells compared to reference cells. These data suggest that interaction between poorly aggressive melanoma cells with normal cells of the skin may regulate the transition from localized, poorly aggressive melanoma to invasive, metastatic disease via Nodal and/or Wnt induced EMT.
Keywords	melanoma ; normal skin cells ; co-culture ; epithelial-to-mesenchymal transition ; Nodal ; Wnt ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Subject code	570
Language	English
Publishing date	2021-10-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Role of Presenilin-1 in Aggressive Human Melanoma

Julia Sidor / Megan Gillette / Lindsay Ann Dezi / Gustavo Untiveros / Luigi Strizzi

International Journal of Molecular Sciences, Vol 23, Iss 4904, p

2022 Volume 4904

Abstract: Presenilin-1 (PS-1), a component of the gamma (γ)-secretase catalytic complex, has been implicated in Alzheimer’s disease (AD) and in tumorigenesis. Interestingly, AD risk is inversely related to melanoma, suggesting that AD-related factors, such as PS-1, ...

Abstract	Presenilin-1 (PS-1), a component of the gamma (γ)-secretase catalytic complex, has been implicated in Alzheimer’s disease (AD) and in tumorigenesis. Interestingly, AD risk is inversely related to melanoma, suggesting that AD-related factors, such as PS-1, may affect melanomagenesis. PS-1 has been shown to reduce Wnt activity by promoting degradation of beta-catenin (β-catenin), an important Wnt signaling partner. Since Wnt is known to enhance progression of different cancers, including melanoma, we hypothesized that PS-1 could affect Wnt-associated melanoma aggressiveness. Western blot results showed that aggressive melanoma cells expressed significantly lower levels of both PS-1 and phosphorylated-β-catenin (P-β-catenin) than nonaggressive melanoma cells. Immunohistochemistry of human melanoma samples showed significantly reduced staining for PS-1 in advanced stage melanoma compared with early stage melanoma. Furthermore, γ-secretase inhibitor (GSI) treatment of aggressive melanoma cells was followed by significant increases in PS-1 and P-β-catenin levels, suggesting impaired Wnt signaling activity as PS-1 expression increased. Finally, a significant reduction in cell migration was associated with the higher levels of PS-1 and P-β-catenin in the GSI-treated aggressive melanoma cells. We demonstrate for the first time that PS-1 levels can be used to assess melanoma aggressiveness and suggest that by enhancing PS-1 expression, Wnt-dependent melanoma progression may be reduced
Keywords	presenilin-1 ; melanoma ; biomarker ; Wnt signaling ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Subject code	570
Language	English
Publishing date	2022-04-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: The in vivo biocompatibility of novel tannic acid-collagen type I injectable bead scaffold material for breast reconstruction post-lumpectomy.

Baldwin, Andrew / Uy, Lisa / Frank-Kamenetskii, Anastasia / Strizzi, Luigi / Booth, Brian W

Journal of biomaterials applications

2020 Volume 34, Issue 9, Page(s) 1315–1329

Language	English
Publishing date	2020-01-05
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	639283-0
ISSN	1530-8022 ; 0885-3282
ISSN (online)	1530-8022
ISSN	0885-3282
DOI	10.1177/0885328219899238
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Article ; Online: The role of Nodal and Cripto-1 in human oral squamous cell carcinoma.

Daraghma, Hussein / Untiveros, Gustavo / Raskind, Aleksandr / Iaccarino, Emanuela / Sandomenico, Annamaria / Ruvo, Menotti / Arnouk, Hilal / Ciancio, Mae J / Cuevas-Nunez, Maria / Strizzi, Luigi

Oral diseases

2020 Volume 27, Issue 5, Page(s) 1137–1147

Abstract: Oral squamous cell carcinoma (OSCC) is a common epithelial malignancy of the oral cavity. Nodal and Cripto-1 (CR-1) are important developmental morphogens expressed in several adult cancers and are associated with disease progression. Whether Nodal and ... ...

Abstract	Oral squamous cell carcinoma (OSCC) is a common epithelial malignancy of the oral cavity. Nodal and Cripto-1 (CR-1) are important developmental morphogens expressed in several adult cancers and are associated with disease progression. Whether Nodal and CR-1 are simultaneously expressed in the same tumor and how this affects cancer biology are unclear. We investigate the expression and potential role of both Nodal and CR-1 in human OSCC. Immunohistochemistry results show that Nodal and CR-1 are both expressed in the same human OSCC sample and that intensity of Nodal staining is correlated with advanced-stage disease. However, this was not observed with CR-1 staining. Western blot analysis of lysates from two human OSCC line experiments shows expression of CR-1 and Nodal, and their respective signaling molecules, Src and ERK1/2. Treatment of SCC25 and SCC15 cells with both Nodal and CR-1 inhibitors simultaneously resulted in reduced cell viability and reduced levels of P-Src and P-ERK1/2. Further investigation showed that the combination treatment with both Nodal and CR-1 inhibitors was capable of reducing invasiveness of SCC25 cells. Our results show a possible role for Nodal/CR-1 function during progression of human OSCC and that targeting both proteins simultaneously may have therapeutic potential.
MeSH term(s)	Adult ; Carcinoma, Squamous Cell ; Cell Line, Tumor ; Head and Neck Neoplasms ; Humans ; Mouth Neoplasms ; Squamous Cell Carcinoma of Head and Neck
Language	English
Publishing date	2020-09-25
Publishing country	Denmark
Document type	Journal Article
ZDB-ID	1290529-x
ISSN	1601-0825 ; 1354-523X
ISSN (online)	1601-0825
ISSN	1354-523X
DOI	10.1111/odi.13640
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Article ; Online: An innovative strategy to investigate microbial protein modifications in a reliable fast and sensitive way: A therapy oriented proof of concept based on UV-C irradiation of SARS-CoV-2 spike protein.

Strizzi, Sergio / Bernardo, Letizia / D'Ursi, Pasqualina / Urbinati, Chiara / Bianco, Andrea / Limanaqi, Fiona / Manconi, Andrea / Milanesi, Maria / Macchi, Alberto / Di Silvestre, Dario / Cavalleri, Adalberto / Pareschi, Giovanni / Rusnati, Marco / Clerici, Mario / Mauri, PierLuigi / Biasin, Mara

Pharmacological research

2023 Volume 194, Page(s) 106862

Abstract: The characterization of modifications of microbial proteins is of primary importance to dissect pathogen lifecycle mechanisms and could be useful in identifying therapeutic targets. Attempts to solve this issue yielded only partial and non-exhaustive ... ...

Abstract	The characterization of modifications of microbial proteins is of primary importance to dissect pathogen lifecycle mechanisms and could be useful in identifying therapeutic targets. Attempts to solve this issue yielded only partial and non-exhaustive results. We developed a multidisciplinary approach by coupling in vitro infection assay, mass spectrometry (MS), protein 3D modelling, and surface plasma resonance (SPR). As a proof of concept, the effect of low UV-C (273 nm) irradiation on SARS-CoV-2 spike (S) protein was investigated. Following UV-C exposure, MS analysis identified, among other modifications, the disruption of a disulphide bond within the conserved S2 subunit of S protein. Computational analyses revealed that this bond breakage associates with an allosteric effect resulting in the generation of a closed conformation with a reduced ability to bind the ACE2 receptor. The UV-C-induced reduced affinity of S protein for ACE2 was further confirmed by SPR analyses and in vitro infection assays. This comprehensive approach pinpoints the S2 domain of S protein as a potential therapeutic target to prevent SARS-CoV-2 infection. Notably, this workflow could be used to screen a wide variety of microbial protein domains, resulting in a precise molecular fingerprint and providing new insights to adequately address future epidemics.
MeSH term(s)	Humans ; COVID-19 ; SARS-CoV-2/metabolism ; Spike Glycoprotein, Coronavirus/metabolism ; Angiotensin-Converting Enzyme 2/metabolism ; Protein Binding
Chemical Substances	spike protein, SARS-CoV-2 ; Spike Glycoprotein, Coronavirus ; Angiotensin-Converting Enzyme 2 (EC 3.4.17.23)
Language	English
Publishing date	2023-07-20
Publishing country	Netherlands
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1003347-6
ISSN	1096-1186 ; 0031-6989 ; 1043-6618
ISSN (online)	1096-1186
ISSN	0031-6989 ; 1043-6618
DOI	10.1016/j.phrs.2023.106862
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Article ; Online: Structure-based design of small bicyclic peptide inhibitors of Cripto-1 activity.

Iaccarino, Emanuela / Calvanese, Luisa / Untiveros, Gustavo / Falcigno, Lucia / D'Auria, Gabriella / Latino, Debora / Sivaccumar, Jwala Priyadarsini / Strizzi, Luigi / Ruvo, Menotti / Sandomenico, Annamaria

The Biochemical journal

2020 Volume 477, Issue 8, Page(s) 1391–1407

Abstract: Bicyclic peptides assembled around small organic scaffolds are gaining an increasing interest as new potent, stable and highly selective therapeutics because of their uncommon ability to specifically recognize protein targets, of their small size that ... ...

Abstract	Bicyclic peptides assembled around small organic scaffolds are gaining an increasing interest as new potent, stable and highly selective therapeutics because of their uncommon ability to specifically recognize protein targets, of their small size that favor tissue penetration and of the versatility and easiness of the synthesis. We have here rationally designed bicyclic peptides assembled around a common tri-bromo-methylbenzene moiety in order to mimic the structure of the CFC domain of the oncogene Cripto-1 and, more specifically, to orient in the most fruitful way the hot spot residues H120 and W123. Through the CFC domain, Cripto-1 binds the ALK4 receptor and other protein partners supporting uncontrolled cell growth and proliferation. Soluble variants of CFC have the potential to inhibit these interactions suppressing the protein activity. A CFC analog named B3 binds ALK4 in vitro with an affinity in the nanomolar range. Structural analyses in solution via NMR and CD show that B3 has rather flexible conformations, like the parent CFC domain. The functional effects of B3 on the Cripto-1-positive NTERA cancer cell line have been evaluated showing that both CFC and B3 are cytotoxic for the cells and block the Cripto-1 intracellular signaling. Altogether, the data suggest that the administration of the soluble CFC and of the structurally related analog has the potential to inhibit tumor growth.
MeSH term(s)	Activin Receptors, Type I/genetics ; Activin Receptors, Type I/metabolism ; Amino Acid Motifs ; Cell Line, Tumor ; Cell Proliferation/drug effects ; Cell Survival/drug effects ; Drug Design ; GPI-Linked Proteins/antagonists & inhibitors ; GPI-Linked Proteins/chemistry ; GPI-Linked Proteins/genetics ; GPI-Linked Proteins/metabolism ; Humans ; Intercellular Signaling Peptides and Proteins/chemistry ; Intercellular Signaling Peptides and Proteins/genetics ; Intercellular Signaling Peptides and Proteins/metabolism ; Magnetic Resonance Spectroscopy ; Neoplasm Proteins/antagonists & inhibitors ; Neoplasm Proteins/chemistry ; Neoplasm Proteins/genetics ; Neoplasm Proteins/metabolism ; Peptides/chemistry ; Peptides/pharmacology
Chemical Substances	GPI-Linked Proteins ; Intercellular Signaling Peptides and Proteins ; Neoplasm Proteins ; Peptides ; TDGF1 protein, human ; ACVR1B protein, human (EC 2.7.11.30) ; Activin Receptors, Type I (EC 2.7.11.30)
Language	English
Publishing date	2020-03-09
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2969-5
ISSN	1470-8728 ; 0006-2936 ; 0306-3275 ; 0264-6021
ISSN (online)	1470-8728
ISSN	0006-2936 ; 0306-3275 ; 0264-6021
DOI	10.1042/BCJ20190953
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