Panoply™ Human PDK1 Knockdown Stable Cell Line

As a key enzyme in glycolysis, the role of pyruvate dehydrogenase kinase 1 (PDK1) in the development and progression of colorectal cancer (CRC) remains unclear. Here, researchers found that high expression of PDK1 is associated with poor prognosis in colorectal cancer patients, and that PDK1 knockdown significantly reduces liver metastasis of colorectal cancer in nude mice and immunocompetent BALB/c mice. When combined with the STAT3-p-Y705 inhibitor cryptotanshinone (CPT), liver metastasis was further inhibited. PDK1 knockdown significantly increased reactive oxygen species levels under anoikis conditions and led to increased anoikis, but the effect of PDK1 knockdown on anoikis was attenuated when combined with CPT. Based on this difference, the researchers further examined the adhesion ability of colorectal cancer cells to the extracellular matrix protein fibronectin. The results showed that the adhesion ability of colorectal cancer cells to fibronectin was significantly reduced after PDK1 knockdown combined with CPT treatment. These results suggest that inhibiting PDK1 can reduce the number of surviving colorectal cancer cells in the bloodstream by upregulating anoikis, while inhibiting STAT3-p-Y705 can prevent cancer cells from colonizing the pre-metastatic microenvironment in the liver, ultimately reducing liver metastasis.

Here, EdU assay was used to determine whether blocking PDK1 could inhibit the proliferation of colorectal cancer cells. Flow cytometry results showed that the proliferation ability of PDK1-knockdown HCT116 cells was reduced by approximately 20% compared to the control group (Figure 1A, B). Similarly, the proliferation ability of PDK1-knockdown SW480 cells was reduced by approximately 20%. Colony formation assays further confirmed the crucial role of PDK1 in tumorigenesis, with a significant reduction in the number of colonies formed by PDK1-knockdown HCT116 cells (Figure 1C, D). It is generally believed that STAT3 promotes tumorigenesis by regulating the expression of various target genes, including PDK1. To assess the role of endogenous PDK1 in the STAT3 signaling pathway, researchers treated HCT116 cells with or without PDK1 knockdown with a p-STAT3-Y705 inhibitor (cryptotanshinone, CPT). Western blot results showed that the level of p-STAT3-Y705 was significantly inhibited in PDK1-knockdown cells compared to the control group and the random sequence control group (Figure 1E). In particular, combined treatment with PDK1 knockdown and CPT completely inhibited p-STAT3-Y705 (Figure 1E). To determine whether STAT3 directly interacts with PDK1 in colorectal cancer cells, researchers performed co-immunoprecipitation (Co-IP) experiments on HCT116 and SW480 cells using an anti-PDK1 antibody. The results confirmed the direct interaction between PDK1 and STAT3 in both colorectal cancer cell lines (Figure 1F).

Figure 1. The direct interaction between PDK1 and p-STAT3 may contribute to CRC proliferation. (Qin W, et al., 2019)