The effect of serial passaging on murine PDAC cell line sensitivity to siKRAS treatment

Selective pressures can majorly affect cancer cell lines’ phenotype while undergoing infinite passaging in culture dishes, leading to genetic drift that affects the cell’s characteristics and response to drugs. It is well known that high passage can change cell line behavior to a more aggressive phenotype regarding proliferation rate, invasiveness, and drug resistance, due to positive selection and the gain of novel mutations. As a result, cell lines in constant use can significantly vary from the original cell line, which can dramatically impact experimental results. Here we demonstrate how serial passaging affected the sensitivity of KPC, a murine pancreatic ductal adenocarcinoma (PDAC) cell line that bears a KRAS mutation, to siKRAS therapy. 95% of PDAC patients carry a mutation in KRAS, marking it as a potential therapeutic target. One of the anti-KRAS therapeutic approaches is silencing KRAS by siRNA. We developed a siRNA polymeric nano-carrier (PNC), which can specifically deliver siKRAS to the tumor site. In contrast to the conventional concession, the low passage KPC cell line showed more aggressive characteristics, such as a higher proliferation rate and faster sprouting in a 3D spheroid model, than the higher passaged cell line. Furthermore, the siKRAS-PNC treatment had a minimal effect on the low passage KPC cells but a meaningful impact on the higher passaged KPC cell line, in terms of gene silencing (20% versus 90%, respectively) and proliferation rate (2% versus 50%, respectively). Our findings stress the importance of cell lines’ integrity in producing reliable and reproducible results.