Panoply™ Human CDA Knockdown Stable Cell Line

Cytidine deaminase (CDA) plays a role in the pyrimidine rescue pathway of DNA and RNA synthesis and has been shown to protect cancer cells from damage by deoxycytidine chemotherapy drugs. Here, researchers observed that CDA is overexpressed in tissues of pancreatic adenocarcinoma patients at baseline and is crucial for the growth of experimental tumors. Mechanistic studies revealed that CDA localizes at replication forks, increasing replication rate, improving replication fork restart efficiency, reducing endogenous replication stress, decreasing DNA breaks, and regulating genetic stability during DNA replication. In a cellular pancreatic cancer model, high CDA expression is associated with resistance to DNA-damaging drugs. Silencing CDA in in vitro patient-derived primary cultured cells and in vivo orthotopic xenograft tumors increases replication stress and makes pancreatic adenocarcinoma cells more sensitive to oxaliplatin. This study elucidates the role of CDA in pancreatic adenocarcinoma and explores how this tumor type modulates replication stress. The results suggest that CDA expression may predict treatment efficacy, and that targeting CDA induces intolerable levels of replication stress in cancer cells, especially when used in combination with DNA-targeting therapies.

Here, researchers found that the reduction in DNA strand length caused by CDA deletion was comparable to that caused by the DNA replication inhibitor aphidicolin (Figure 1A and B), indicating that the replication fork speed was affected. Simultaneously, CDA inhibition altered the cell cycle progression of cancer cells, leading to a decrease in G1 phase cells and an increase in S and G2-M phase cells (Figure 1C). Researchers rescued CDA deletion by supplementing with uridine. The results showed that, compared to control cells, CDA deletion not only restored cell cycle progression (Figure 1D) but also promoted tumor cell proliferation. Transcriptome analysis revealed a significant enrichment of ATR response signatures to replication stress in CDA knockdown cells compared to control cells (Figure 1E). Compared to control cells, the activation of CHK1 effector kinase was significantly enhanced in CDA-knockdown MIA PaCa-2 (Figure 1F), Capan-1, and BxPC3 cells, further supporting these findings. Next, researchers examined the number of γH2AX foci in S phase cells, a classic marker of DNA breakage and replication stress. As shown in Figure 1G and H, CDA overexpression in MIA PaCa-2 cells significantly reduced the number of γH2AX foci in EdU-positive cells compared to control cells. This effect was entirely dependent on CDA deaminase activity, as expression of catalytically inactivated CDA mutants had no effect on the number of γH2AX foci (Figure 1E and F). Conversely, CDA silencing in BxPC3 cells led to the accumulation of γH2AX foci in S-phase cells (Figures 1I and J).

Figure 1. CDA controls replication stress levels in PDAC cells. (Lumeau A, et al., 2024)