Panoply™ Human IGF2R Knockdown Stable Cell Line

Cervical cancer is one of the most common gynecological malignancies worldwide. However, the survival rate for patients with advanced and recurrent cervical cancer remains low. The multifunctional protein insulin-like growth factor 2 receptor (IGF2R) has multiple ligands, including IGF-2 and mannose-6-phosphate (M6P)-labeled proteins. Given its antagonistic effect on IGF1R signaling, IGF2R is currently considered a tumor suppressor gene, but its significance as an M6P receptor is unclear. Based on transcriptome analysis of TCGA and GEO open datasets, researchers found that IGF2R is upregulated in cervical cancer and is associated with poor prognosis. Several experiments using cervical cancer cell lines showed that IGF2R depletion induces apoptosis, reduces cell viability, and increases cell sensitivity to certain anticancer drugs (such as cisplatin). Unlike its weak effect on IGF1R signaling, IGF2R depletion disrupts the transport of M6P-labeled cathepsins from the Golgi apparatus to lysosomes, leading to decreased lysosomal activity, accompanied by their abnormal accumulation and autophagy and mitophagy dysfunction, resulting in the accumulation of misfolded proteins and the production of reactive oxygen species. Therefore, IGF2R plays an oncogenic role in cervical cancer by transporting M6P-labeled cargo and can serve as a predictive biomarker for prognostic classification.

IGF2R binds to M6P-tagged proteins and transports them from the trans-Golgi network (TGN) to lysosomes via the vesicular transport system. Here, researchers verified the function of lysosomes in IGF2R knockdown cells, as lysosomes are considered the organelles most affected by insufficient delivery of M6P-tagged lysosomal enzymes. Accumulation of acidic organelles and LAMP1-positive organelles over time was observed in IGF2R knockdown cells (Figure 1b), indicating that this receptor maintains lysosomal homeostasis. Increased lysosomal membrane permeability (LMP) leads to the release of lysosomal enzymes into the cytoplasm, thereby inducing apoptosis; however, IGF2R knockdown did not induce LMP. Instead, IGF2R deficiency reduced the activity of lysosomal enzymes (Figure 1c). Lysosomes are closely related to various types of autophagy (e.g., macroautophagy and mitophagy). Here, IGF2R knockdown promoted the formation of autophagosomes (Figure 1d). Considering that IGF2R knockdown did not enhance bafilomycin A-induced LC3-II conversion (confirmed by densitometric analysis of three independent experiments), this suggests that IGF2R knockdown has an inhibitory effect on autophagy (Figure 1e). Consequently, accumulation of polyubiquitinated proteins was observed in IGF2R knockdown cells (Figure 1f). Furthermore, IGF2R knockdown significantly reduced mitochondrial membrane potential, indicating the accumulation of damaged mitochondria in the cells. As a result, ROS production increased in IGF2R knockdown cells (Figure 1g). Immunocytochemical analysis showed a decrease in mitophagy in IGF2R knockdown cells (Figure 1h), indicating that IGF2R is crucial for autophagy and mitophagy.

Figure 1. IGF2R maintains autophagy and mitophagy through lysosome homeostasis. (Takeda T, et al., 2019)