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Development and Validation of an Ultrasensitive Single Molecule Array Digital Enzyme-linked Immunosorbent Assay for Human Interferon-α
Published on: June 14, 2018
Identification of proteins regulated by interferon-alpha in resistant and sensitive malignant melanoma cell lines
Rachel A Craven1, Anthea J Stanley, Sarah Hanrahan
1Cancer Research UK Clinical Centre, St. James's University Hospital, Leeds, UK.
Abstract:
Treatment of patients with malignant melanoma with interferon-alpha achieves a response in a small but significant subset of patients. Currently, although much is known about interferon biology, little is known about either the particular mechanisms of interferon-alpha activity that are crucial for response or why only some patients respond to interferon-alpha therapy. Two melanoma cell lines (MeWo and MM418) that are known to differ in their response to the antiproliferative activity of interferon-alpha, have been used as a model system to investigate interferon-alpha action. Using a proteomics approach based on two-dimensional polyacrylamide gel electrophoresis and mass spectrometry, several proteins induced in response to interferon-alpha have been identified. These include a number of gene products previously known to be type I interferon responsive (tryptophanyl tRNA synthetase, leucine aminopeptidase, ubiquitin cross-reactive protein, gelsolin, FUSE binding protein 2 and hPNPase) as well as a number of proteins not previously reported to be induced by type I interferon (cathepsin B, proteasomal activator 28alpha and alpha-SNAP). Although the proteins upregulated by interferon-alpha were common between the cell lines when examined at the level of Western blotting, the disparity in the basal level of cathepsin B was striking, raising the possibility that the higher level in MM418 may contribute to the sensitivity of this cell line to interferon-alpha treatment.
Insights
Interferon-alpha treats melanoma by upregulating proteins. Differences in basal cathepsin B levels may explain why some melanoma patients respond to this therapy.
Area of Science:
- Oncology
- Immunology
- Proteomics
Background:
- Malignant melanoma treatment with interferon-alpha yields variable patient responses.
- Understanding interferon-alpha's precise mechanisms and patient-specific responses remains limited.
Purpose of the Study:
- To investigate the molecular mechanisms of interferon-alpha action in melanoma.
- To identify proteins differentially regulated by interferon-alpha in melanoma cell lines with varying sensitivities.
Main Methods:
- Utilized a proteomics approach involving two-dimensional polyacrylamide gel electrophoresis and mass spectrometry.
- Employed two melanoma cell lines (MeWo and MM418) with known differences in antiproliferative response to interferon-alpha.
- Validated protein expression using Western blotting.
Main Results:
- Identified several interferon-alpha-induced proteins, including known type I interferon-responsive genes and novel proteins like cathepsin B, proteasomal activator 28alpha, and alpha-SNAP.
- Observed common upregulated proteins between cell lines via Western blotting.
- Noted a striking disparity in basal cathepsin B levels between the two cell lines, with MM418 exhibiting higher levels.
Conclusions:
- Proteomics identified novel interferon-alpha-responsive proteins in melanoma cells.
- Differential basal expression of cathepsin B may contribute to varying sensitivities to interferon-alpha therapy in melanoma.
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