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Gene amplification-associated cytogenetic aberrations and protein changes in vincristine-resistant Chinese hamster,
Abstract:
We carried out cytogenetic studies of four Chinese hamster, mouse, and human cell lines selected for high levels of resistance (500- to 4,000-fold) to vincristine (VCR) by a multistep selection procedure. All cells examined contained gene amplification-associated metaphase chromosome abnormalities, either homogeneously staining regions (HSRs), abnormally banding regions (ABRs), or double-minute chromosomes (DMs); control actinomycin D- and daunorubicin-resistant hamster lines did not exhibit this type of chromosomal abnormality. VCR-resistant Chinese hamster sublines exhibited both increased synthesis of the protein V19 (Mr 19,000; pl = 5.7) and increased concentrations of V19 polysomal mRNA. When VCR-resistant cells were grown in drug-free medium, level of resistance, synthesis of V19, and amount of V19 mRNA declined in parallel with mean length of the HSR or mean number of DMs per cell. Cross-resistance studies indicate that VCR-resistant cells have increased resistance both to antimitotic agents and to a wide variety of agents unrelated to VCR in chemical structure and/or mechanism of action. Our studies of tubulin synthesis in Chinese hamster cells indicate no overproduction of tubulin or presence of a mutant tubulin species. Comparison with antifolate-resistant Chinese hamster cells known to contain amplified dihydrofolate reductase genes localized to HSRs or ABRs strongly suggests that the HSRs, ABRs, or DMs of the Vinca alkaloid-resistant sublines likewise represent cytological manifestations of specifically amplified genes, possibly encoding V19, involved in development of resistance to VCR.
Insights
High resistance to vincristine (VCR) in cancer cells is linked to gene amplification, causing chromosome abnormalities like HSRs and DMs. This resistance involves increased V19 protein and mRNA, which decrease when drug pressure is removed.
Area of Science:
- Cytogenetics
- Molecular Biology
- Cancer Research
Background:
- Multidrug resistance (MDR) is a significant challenge in cancer chemotherapy.
- Vincristine (VCR), a Vinca alkaloid, is a widely used antineoplastic agent.
- Understanding the genetic basis of VCR resistance is crucial for improving treatment efficacy.
Purpose of the Study:
- To investigate the cytogenetic and molecular mechanisms underlying high-level VCR resistance in mammalian cell lines.
- To identify potential genetic alterations associated with VCR resistance.
- To explore the relationship between drug resistance, gene expression, and chromosomal abnormalities.
Main Methods:
- Cytogenetic analysis (metaphase chromosome banding) of VCR-resistant cell lines.
- Biochemical assays to measure protein synthesis (V19) and mRNA levels.
- Drug resistance profiling through cross-resistance studies.
- Comparison with known gene amplification models (e.g., dihydrofolate reductase).
Main Results:
- VCR-resistant cell lines exhibited gene amplification-associated chromosomal abnormalities (HSRs, ABRs, DMs).
- Increased synthesis of a 19 kDa protein (V19) and its corresponding mRNA was observed in VCR-resistant cells.
- Resistance levels and V19 expression decreased in parallel with chromosomal abnormality reduction upon drug withdrawal.
- Cross-resistance studies indicated broad resistance to various cytotoxic agents, not limited to VCR.
Conclusions:
- High-level VCR resistance in mammalian cells is associated with specific gene amplification events manifesting as chromosomal abnormalities.
- The protein V19 and its mRNA are likely involved in the development of VCR resistance.
- These findings suggest that HSRs, ABRs, or DMs represent cytological evidence of amplified genes contributing to Vinca alkaloid resistance.