Panoply™ Human CD44 Knockdown Stable Cell Line

Cluster of differentiation 44 (CD44) is a transmembrane glycoprotein that has been identified as a cancer stem cell marker in a variety of cancer cells. Although many studies have focused on CD44 as a cancer stem cell marker, its effects on cancer cell metabolism remain unclear. To explore the role of CD44 on cancer cell metabolism, researchers established CD44 knockdown cells in human breast cancer cells. Silencing of CD44 reduced the glycolytic phenotype of cancer cells, affecting glucose uptake, ATP production, and lactate production. Knockdown of CD44-induced lactate dehydrogenase (LDH) isozymes resulted in downregulation of LDHA and upregulation of LDHB, leading to the shift of LDH1, indicating that the regulation of LDH isozymes is important for cancer metabolism. CD44 silencing reduced the expression of glycolysis-related proteins, including hypoxia-inducible factor 1α (HIF-1α) and LDHA. These effects were due to decreased c-Src and Akt activities in CD44 knockdown cells, leading to upregulation of liver kinase B1 (LKB1)/AMP-activated protein kinase (AMPK) α activity. Finally, induction of LKB1/AMPKα activity inhibited the expression of HIF-1α and its target gene LDHA. Conversely, the expression of LDHB was inhibited by HIF-1α. Together, these results suggest that in human breast cancer cells, the metabolic shift induced by CD44 silencing is mediated by the regulation of c-Src/Akt/LKB1/AMPKα/HIF-1α signaling.

Given that CD44 knockdown can regulate the expression of metabolism-related genes, the researchers studied the effects of CD44 knockdown on glucose uptake, cellular ATP production, lactate production, LDH isozyme expression patterns, and endogenous cellular oxygen consumption rates. Compared with control shRNA cells, the glucose uptake capacity, cellular ATP levels, and lactate production of CD44 knockdown cells were downregulated (Figure 1A-C). To further confirm the effect of CD44 knockdown on the expression pattern of LDH isozymes, the researchers performed non-denaturing in-gel experiments. The results showed that control shRNA cells mainly contained LDHA-rich LDH5 and LDH4 isozymes. However, CD44 knockdown effectively shifted the LDH isozymes toward the LDHB-rich LDH2 isozymes (Figure 1D). Compared with control shRNA cells, the endogenous cellular oxygen consumption rate of CD44 knockdown cells was increased (Figure 1E). These results suggest that the ablation of CD44 undergoes metabolic shift toward mitochondrial respiration.

Figure 1. CD44 ablation suppresses glucose uptake, ATP production, and lactate production and induces an LDH isoenzyme shift in human breast cancer cells. (Nam K S, Oh S, Shin I., 2016)