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The progression in the mouse skin carcinogenesis model correlates with ERK1/2 signaling
Kostas D Katsanakis1, Vassilis Gorgoulis, Athanasios G Papavassiliou
1Institute of Biological Research and Biotechnology, National Hellenic Research Foundation, Athens, Greece.
Background:
The ras family of proto-oncogenes encodes for small GTPases that play critical roles in cell-cycle progression and cellular transformation. ERK1/2 MAP kinases are major ras effectors. Tumors in chemically treated mouse skin contain mutations in the Ha-ras proto- oncogene. Amplification and mutation of Ha-ras has been shown to correlate with malignant progression of these tumors. Cell lines isolated from mouse skin tumors represent the stages of tumor development, such as the PDV:PDVC57 cell line pair and B9 squamous carcinoma and A5 spindle cells. PDVC57 cells were selected from PDV cells, which were transformed with dimethyl-benzanthracene (DMBA) in vitro and then transplanted in adult syngeneic mice. The PDV:PDVC57 pair contains ratio of normal:mutant Ha-ras 2:1 and 1:2, respectively. This genetic alteration correlates with more advanced tumorigenic characteristics of PDVC57 compared to PDV. The squamous carcinoma B9 cell clone was isolated from the same primary tumor as A5 spindle cell line. The mutant Ha-ras allele, also present in B9, is amplified and overexpressed in A5 cells. Therefore these cell line pairs represent an in vivo model for studies of Ha-ras and ERK1/2 signaling in mouse tumorigenesis.
Materials And Methods:
The ERK1/2 status in the above mouse cell lines was examined by using various molecular techniques. For the study of the tumorigenic properties and the role of the ras/MEK/ERK1/2 pathway in the cell lines mentioned, phenotypic characteristics, colony formation assay, anchorage-independent growth, and gelatin zymography were assessed, after or without treatment with the MEK inhibitor, PD98059.
Results:
ERK1/2 phosphorylation was found to be increased in PDVC57 when compared to PDV. This also applies to A5 spindle carcinoma cells when compared to squamous carcinoma and papilloma cells. The above finding was reproduced when transfecting human activated Ha-ras allele into PDV, thus demonstrating that Ha-ras enhances ERK1/2 signaling. To further test whether ERK1/2 activation was required for growth we used the MEK-1 inhibitor, PD98059. The latter inhibited cell proliferation and anchorage-independent growth of squamous and spindle cells. In addition, PD98059 treatment partially reverted the spindle morphology of A5 cells.
Conclusions:
These data suggest, for the first time, that oncogenicity and the degree of progression in the mouse skin carcinogenesis model correlates with ERK1/2 signaling.
Insights
Mouse skin cancer progression correlates with increased ERK1/2 signaling, driven by Ha-ras mutations. Inhibiting this pathway reduces tumor cell growth and alters morphology, highlighting its role in tumorigenesis.
Area of Science:
- Oncology
- Molecular Biology
- Cancer Research
Background:
- Ras proto-oncogenes regulate cell cycle and transformation; ERK1/2 MAP kinases are key downstream effectors.
- Mouse skin tumors exhibit Ha-ras mutations, correlating with malignant progression.
- Established cell line pairs (PDV:PDVC57, B9:A5) model different stages of mouse skin carcinogenesis and Ha-ras alterations.
Purpose of the Study:
- To investigate the role of Ha-ras and ERK1/2 signaling in mouse skin tumorigenesis.
- To analyze ERK1/2 activation status in different stages of tumor development using specific cell lines.
- To determine the impact of MEK inhibition on the tumorigenic properties of these cell lines.
Main Methods:
- Examined ERK1/2 phosphorylation status using molecular techniques.
- Assessed phenotypic characteristics, colony formation, anchorage-independent growth, and gelatin zymography.
- Utilized the MEK inhibitor PD98059 to evaluate pathway dependency.
Main Results:
- Increased ERK1/2 phosphorylation observed in PDVC57 and A5 cells compared to their less malignant counterparts.
- Ha-ras activation was shown to enhance ERK1/2 signaling.
- PD98059 treatment inhibited cell proliferation and anchorage-independent growth, and partially reverted A5 cell spindle morphology.
Conclusions:
- Oncogenicity and tumor progression in mouse skin carcinogenesis correlate with ERK1/2 signaling.
- The ras/MEK/ERK1/2 pathway is crucial for the growth and malignant characteristics of these tumor cells.
- Targeting ERK1/2 signaling presents a potential therapeutic strategy for skin cancer.