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Molecular characterization of a novel cancer cell line established from human carcinoma in pleomorphic adenoma
M Fujioka1, K Shimada, S Kitazawa
1Second Department of Pathology, Kobe University School of Medicine, Japan.
Abstract:
A cultured cell line (CaPA-4), derived from an undifferentiated carcinoma in a pleomorphic adenoma of the submandibular gland, was established through xenografted tumors in nude mice. Geneticin treatment eliminated surrounding mouse fibroblasts and yielded enriched tumor cells at an early stage of cell passage. In vitro, the line grew in a cobblestone pattern, revealing its epithelial origin. Chromosomal analysis by Giemsabanding confirmed its human origin, while electron microscopic examination showed its squamous-cell characteristics. CaPA-4 cells stained positive for the c-myc and Ha-ras antibodies. Molecular analysis showed over-expression of both c-myc and Ha-ras mRNA, with point mutation of p53 at codon 248 and of Ha-ras at codon 61. Amplification and rearrangement of the Ha-ras gene were observed; however, no loss of heterozygosity (LOH) of the p53 gene was detected by Southern blotting. This sequence of cancer-related gene activation may represent the malignant transformation from benign pleomorphic adenoma. This report describes the establishment and molecular characterization of this novel cell line from carcinoma in pleomorphic adenoma exhibiting squamous-cell differentiation. This could represent a useful model for investigating the cause of malignant transformation from human salivary-gland mixed tumors.
Insights
A novel human cell line, CaPA-4, was established from submandibular gland carcinoma in pleomorphic adenoma. This cell line exhibits squamous cell characteristics and genetic alterations, offering a model for studying malignant transformation.
Area of Science:
- Oncology
- Cell Biology
- Genetics
Background:
- Pleomorphic adenomas are common benign salivary gland tumors.
- Malignant transformation of pleomorphic adenoma to carcinoma is rare but significant.
- Establishing cell lines is crucial for studying tumor biology and transformation mechanisms.
Purpose of the Study:
- To establish and characterize a novel human cell line (CaPA-4) from a carcinoma ex pleomorphic adenoma.
- To investigate the molecular alterations associated with malignant transformation in this cell line.
- To provide a valuable in vitro model for salivary gland tumor research.
Main Methods:
- Xenografting in nude mice for tumor establishment.
- Geneticin treatment for selective enrichment of human tumor cells.
- In vitro cell culture and characterization (morphology, epithelial origin).
- Cytogenetic analysis (Giemsa banding) for human origin confirmation.
- Electron microscopy for squamous cell differentiation assessment.
- Immunohistochemistry for c-myc and Ha-ras expression.
- Molecular analyses including mRNA expression, gene mutation (p53, Ha-ras), gene amplification/rearrangement, and Southern blotting for loss of heterozygosity (LOH).
Main Results:
- The CaPA-4 cell line was successfully established and maintained, showing epithelial (cobblestone) morphology.
- Confirmed human origin and squamous cell characteristics.
- Demonstrated overexpression of c-myc and Ha-ras mRNA.
- Identified point mutations in p53 (codon 248) and Ha-ras (codon 61).
- Observed amplification and rearrangement of the Ha-ras gene.
- No loss of heterozygosity (LOH) of the p53 gene was detected.
Conclusions:
- The CaPA-4 cell line represents a unique model of carcinoma ex pleomorphic adenoma with squamous differentiation.
- The observed genetic alterations (c-myc, Ha-ras, p53 mutations) may play a role in malignant transformation.
- This cell line provides a platform for investigating the molecular pathways driving malignant transformation in human salivary gland tumors.