Gemcitabine And Its Effect On Cell Cycle
Gemcitabine is a chemotherapeutic medication used to treat various cancers such as pancreatic, breast, ovarian, bladder, and lung cancers. It was discovered in the early1980s during a pancreatic cancer trial when the researchers discovered that the patients who were injected with Gemcitabine had a longer survival rate, it was then FDA approved in 1996. The main mechanism of action of Gemcitabine is incorporating Gemcitabine phosphate into DNA to induce apoptosis.
Once the Gemcitabine phosphate enters the DNA, the single subsequent nucleotide is formed which then blocks the DNA polymerase from processing. This allows the drug to escape from masked chain termination and begins apoptosis to slow down the rate of the dividing cells. The first experiment tested this cell cycle out by combining the effect of 125I and Gemcitabine on PANC-1 cells to distinguish apoptosis and cell cycle arrest. This experiment was aimed to study the combined effects of these two drugs on pancreatic carcinoma cells to see the underlying outcome of the combined drugs. This experiment started out by incubating the pancreatic cells at 37 degrees Celsius with 5% CO2 in DMEM, and 10% calf serum. They were then placed in in vitro irradiation and exposed to antibodies shortly after. After counting the cells, the media was replaced with CCK-8 reagent and incubated for 3 hours and scanned with a microplate reader. After counting the colonies, the cells were then introduced to the 125I and the gemcitabine and assessed with a flow cytometer. The cells were then observed for the morphological changes with a fluorescence microscope followed by a western blot. The results stated that the gemcitabine increased the radiation induced apoptosis and arrested the G1 expression. The antibody ratios were also lower when the combination of 125I and gemcitabine was induced rather than only 1 being induced. This concludes that the combination of both gemcitabine and 125I may conduct to increased numbers of apoptosis.
The next experiment tested if a pharmacodynamic model of cell cycle apoptosis is due to gemcitabine exposure on pancreatic adenocarcinoma cells. The method started out by obtaining three pancreatic adenocarcinoma cell line which were AsPC-1, BxPC-3, and MiaPaca-2). These cell lines were then exposed to different concentrations of gemcitabine for 8 days. The main purpose is to observe if the drug can induce a cycle phase transition in the G1, S, and G2 phase causing apoptosis. The results showed that low concentration of gemcitabine caused lower apoptotic rates within the S phase and higher rates in the G1 and G2 phases, and higher concentration of gemcitabine caused a high death rate in all stages of the cell cycle. This concludes that the pharmacodynamic model insinuates apoptosis efficiency within the pancreatic cell line when introduced to high concentrations of gemcitabine.
The third experiment tested the effects of honokiol, an active constituent of the grandiflora herb and gemcitabine on 2 pancreatic cancer cell lines PANC-1 and Miapaca. The cells were infused with 10% penicillin and streptomycin in DMEM and incubated at 37 degrees Celsius in 5%CO2. The cells were then induced with the gemcitabine alone, honokiol herb alone, and both together, all in equal concentrations. The effects were then determined by flow cytometry and PI staining followed by a western blot analysis. The results stated that the apoptotic rate was higher when the herb and gemcitabine were both induced in the pancreatic cell line rather than them both being induced alone. This concludes that both of these can reduce cell cycle arrest within the G1 phase of the cell cycle and alter the expression of it allowing for apoptosis to occur.
The fourth experiment tested the effects on mangiferin against the gemcitabine resistant pancreatic cancer line MiaPaca-2. The experiment started after the cells were incubated and induced 100 microliters of mangiferin and acridine orange and ethidium bromide staining for 12 hours. The cells were then washed with phosphate buffer saline and induced with PI staining and analyzed through flow cytometry followed by a western blot. The results stated that the mangiferin allowed growth of the pancreatic cancer cells and induced apoptosis. It also showed that the mangiferin stimulated autophagy in the cells which prevents the invasion of potential cancer cells. This concludes that mangiferin could be used as a chemotherapeutic treatment for pancreatic cancer.
The most important concepts I learned about this drug is that it can treat various different types of cancers, most commonly pancreatic cancer. I learned that it is also important to administer this drug in a certain amount of concentration in order for it to succeed efficiently. This drug can also cause cell death and cell arrest within different stages of the cell cycle. It also words even better when paired with certain herbs or other chemotherapeutic medications.
