Lung cancer is the leading cause of cancer-related deaths worldwide. alimta is a new-generation antifolate drug, approved for the treatment of lung cancer. this drug inhibits several folate-dependent enzymes, leading to a global reduction in dna synthasis. dna is not naked in the cell nucleus and forms a chromatin complex in association with histone. chromatin non-histone proteins ,hmgb1, is the fundemental protein in chromatin structure and participates in metabolism and translocates into extracellular space acting as a cytokine in inflammation, angiogenesis, and late apoptosis in tumor cells. in the present study, we have investigated the mechanism of alimta action through hmgb1.
Mtt, sds-page, western blot, fluorescence staining, diphenylamine method
The results showed that alimta decreased viability of a549 cells in a dose- and time-dependent manner. the content of non-histone protein hmgb1 decreased as drug concentration increased but hmgb1 release into extracellular space was not detected in the cells exposed to alimta for 48 h. apoptosis was studied in a549 cells incubated with different concentrations of alimta. acridine orange/ ethidium bromide and hoechst staining of drug treated cells revealed morphological changes such as chromatin condensation and nuclear fragmentation indicating occurrence of apoptosis. dna fragmentation by diphenylamine method showed that alimta induces dna fragmentation in a dose dependent manner.
from the results, it is concluded that alimta induces apoptosis in a549 cells independent of hmgb1 release. it is suggested that possibly alimta binds to hmgb1 and by compaction of chromation precedes the cells in to apoptosis.