EMT-associated factors promote invasive properties of uveal melanoma cells

Laura Asnaghi, Gülçin Gezgin, Arushi Tripathy, James T. Handa, Shannath L. Merbs, Pieter A. van der Velden, Martine J. Jager, J. William Harbour, Charles G. Eberhart

Research output: Contribution to journalArticle

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Abstract

Purpose: Transcription factors regulating the epithelial-to-mesenchymal transition (EMT) program contribute to carcinogenesis and metastasis in many tumors, including cutaneous melanoma. However, little is known about the role of EMT factors in the growth and metastatic dissemination of uveal melanoma cells. Here, we analyzed the expression and functions of the EMT factors ZEB1, Twist1, and Snail1 in uveal melanoma cell lines and primary tumors. Methods: ZEB1, Twist1, and Snail1 mRNA levels were measured using qPCR in five uveal melanoma cell lines and in 30 primary tumors. Gene expression was used to determine class 1 and class 2 signatures in the primary tumors. Short hairpin RNA was used to downregulate the expressions of the EMT factors; then, growth and transwell invasion assays were performed. Results: ZEB1, Twist1, and Snail1 were expressed in all five uveal melanoma lines, with ZEB1 having the highest protein levels. ZEB1 mRNA was significantly elevated in highly metastatic class 2 primary tumors for which survival data were not available, whereas a high gene expression of Twist1 was associated with a worse prognosis in a separate tumor cohort analyzed by expression profiling. The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50% reduction in invasion, but only suppressed growth in OMM1 cells. Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50%. The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50%, but did not interfere with growth. Conclusions: The downregulation of ZEB1, Twist1, and Snail1 reduces the invasive properties of uveal melanoma cells, and the elevated mRNA levels of ZEB1 and Twist1 are associated with a more aggressive clinical phenotype in uveal melanoma samples. Therefore, these factors could represent new therapeutic targets in patients with ocular melanoma.

Original languageEnglish (US)
Pages (from-to)919-929
Number of pages11
JournalMolecular Vision
Volume21
StatePublished - Aug 25 2015

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Epithelial-Mesenchymal Transition
Down-Regulation
Neoplasms
Growth
Messenger RNA
Melanoma
Gene Expression
Cell Line
Tumor Cell Line
Small Interfering RNA
Uveal melanoma
Intercellular Signaling Peptides and Proteins
Carcinogenesis
Transcription Factors
Neoplasm Metastasis
Phenotype
Skin
Survival
Proteins

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Asnaghi, L., Gezgin, G., Tripathy, A., Handa, J. T., Merbs, S. L., van der Velden, P. A., ... Eberhart, C. G. (2015). EMT-associated factors promote invasive properties of uveal melanoma cells. Molecular Vision, 21, 919-929.

EMT-associated factors promote invasive properties of uveal melanoma cells. / Asnaghi, Laura; Gezgin, Gülçin; Tripathy, Arushi; Handa, James T.; Merbs, Shannath L.; van der Velden, Pieter A.; Jager, Martine J.; William Harbour, J.; Eberhart, Charles G.

In: Molecular Vision, Vol. 21, 25.08.2015, p. 919-929.

Research output: Contribution to journalArticle

Asnaghi, L, Gezgin, G, Tripathy, A, Handa, JT, Merbs, SL, van der Velden, PA, Jager, MJ, William Harbour, J & Eberhart, CG 2015, 'EMT-associated factors promote invasive properties of uveal melanoma cells', Molecular Vision, vol. 21, pp. 919-929.
Asnaghi L, Gezgin G, Tripathy A, Handa JT, Merbs SL, van der Velden PA et al. EMT-associated factors promote invasive properties of uveal melanoma cells. Molecular Vision. 2015 Aug 25;21:919-929.
Asnaghi, Laura ; Gezgin, Gülçin ; Tripathy, Arushi ; Handa, James T. ; Merbs, Shannath L. ; van der Velden, Pieter A. ; Jager, Martine J. ; William Harbour, J. ; Eberhart, Charles G. / EMT-associated factors promote invasive properties of uveal melanoma cells. In: Molecular Vision. 2015 ; Vol. 21. pp. 919-929.
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abstract = "Purpose: Transcription factors regulating the epithelial-to-mesenchymal transition (EMT) program contribute to carcinogenesis and metastasis in many tumors, including cutaneous melanoma. However, little is known about the role of EMT factors in the growth and metastatic dissemination of uveal melanoma cells. Here, we analyzed the expression and functions of the EMT factors ZEB1, Twist1, and Snail1 in uveal melanoma cell lines and primary tumors. Methods: ZEB1, Twist1, and Snail1 mRNA levels were measured using qPCR in five uveal melanoma cell lines and in 30 primary tumors. Gene expression was used to determine class 1 and class 2 signatures in the primary tumors. Short hairpin RNA was used to downregulate the expressions of the EMT factors; then, growth and transwell invasion assays were performed. Results: ZEB1, Twist1, and Snail1 were expressed in all five uveal melanoma lines, with ZEB1 having the highest protein levels. ZEB1 mRNA was significantly elevated in highly metastatic class 2 primary tumors for which survival data were not available, whereas a high gene expression of Twist1 was associated with a worse prognosis in a separate tumor cohort analyzed by expression profiling. The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50{\%} reduction in invasion, but only suppressed growth in OMM1 cells. Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50{\%}. The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50{\%}, but did not interfere with growth. Conclusions: The downregulation of ZEB1, Twist1, and Snail1 reduces the invasive properties of uveal melanoma cells, and the elevated mRNA levels of ZEB1 and Twist1 are associated with a more aggressive clinical phenotype in uveal melanoma samples. Therefore, these factors could represent new therapeutic targets in patients with ocular melanoma.",
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AU - Asnaghi, Laura

AU - Gezgin, Gülçin

AU - Tripathy, Arushi

AU - Handa, James T.

AU - Merbs, Shannath L.

AU - van der Velden, Pieter A.

AU - Jager, Martine J.

AU - William Harbour, J.

AU - Eberhart, Charles G.

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N2 - Purpose: Transcription factors regulating the epithelial-to-mesenchymal transition (EMT) program contribute to carcinogenesis and metastasis in many tumors, including cutaneous melanoma. However, little is known about the role of EMT factors in the growth and metastatic dissemination of uveal melanoma cells. Here, we analyzed the expression and functions of the EMT factors ZEB1, Twist1, and Snail1 in uveal melanoma cell lines and primary tumors. Methods: ZEB1, Twist1, and Snail1 mRNA levels were measured using qPCR in five uveal melanoma cell lines and in 30 primary tumors. Gene expression was used to determine class 1 and class 2 signatures in the primary tumors. Short hairpin RNA was used to downregulate the expressions of the EMT factors; then, growth and transwell invasion assays were performed. Results: ZEB1, Twist1, and Snail1 were expressed in all five uveal melanoma lines, with ZEB1 having the highest protein levels. ZEB1 mRNA was significantly elevated in highly metastatic class 2 primary tumors for which survival data were not available, whereas a high gene expression of Twist1 was associated with a worse prognosis in a separate tumor cohort analyzed by expression profiling. The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50% reduction in invasion, but only suppressed growth in OMM1 cells. Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50%. The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50%, but did not interfere with growth. Conclusions: The downregulation of ZEB1, Twist1, and Snail1 reduces the invasive properties of uveal melanoma cells, and the elevated mRNA levels of ZEB1 and Twist1 are associated with a more aggressive clinical phenotype in uveal melanoma samples. Therefore, these factors could represent new therapeutic targets in patients with ocular melanoma.

AB - Purpose: Transcription factors regulating the epithelial-to-mesenchymal transition (EMT) program contribute to carcinogenesis and metastasis in many tumors, including cutaneous melanoma. However, little is known about the role of EMT factors in the growth and metastatic dissemination of uveal melanoma cells. Here, we analyzed the expression and functions of the EMT factors ZEB1, Twist1, and Snail1 in uveal melanoma cell lines and primary tumors. Methods: ZEB1, Twist1, and Snail1 mRNA levels were measured using qPCR in five uveal melanoma cell lines and in 30 primary tumors. Gene expression was used to determine class 1 and class 2 signatures in the primary tumors. Short hairpin RNA was used to downregulate the expressions of the EMT factors; then, growth and transwell invasion assays were performed. Results: ZEB1, Twist1, and Snail1 were expressed in all five uveal melanoma lines, with ZEB1 having the highest protein levels. ZEB1 mRNA was significantly elevated in highly metastatic class 2 primary tumors for which survival data were not available, whereas a high gene expression of Twist1 was associated with a worse prognosis in a separate tumor cohort analyzed by expression profiling. The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50% reduction in invasion, but only suppressed growth in OMM1 cells. Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50%. The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50%, but did not interfere with growth. Conclusions: The downregulation of ZEB1, Twist1, and Snail1 reduces the invasive properties of uveal melanoma cells, and the elevated mRNA levels of ZEB1 and Twist1 are associated with a more aggressive clinical phenotype in uveal melanoma samples. Therefore, these factors could represent new therapeutic targets in patients with ocular melanoma.

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