Engineered ubiquitin ligase PTB-U-box targets insulin/insulin-like growth factor receptor for degradation and coordinately inhibits cancer malignancy

Abstract

The type 1 insulin-like growth factor receptor (IGF-1R) is a promising target for cancer therapy with antibodies and small molecule tyrosine kinase inhibitors (TKIs) which have been actively tested clinically. Evidences have demonstrated that insulin receptor (IR), which is implicated in tumorigenesis, conveys resistance to IGF-1R targeted therapy. This provided the compelling rationale for co-targeting IGF-1R and IR. Herein we have developed an approach to simultaneously down-regulate IGF-1R and IR in protein levels. By generating and screening several engineered ubiquitin ligases, we have identified that, PTB-U-box, which is composed of an IGF-1R/IR-binding domain and a functional E3 ubiquitin ligase domain, binds activated IGF-1R/IR and targets their ubiquitination and degradation. When ectopically expressed in HepG2 and HeLa cells, PTB-U-box inhibits cell proliferation and invasion, increases chemo-sensitivity, as well as interrupts glucose metabolism. Finally, intratumoral injection of adenovirus carrying PTB-U-box dramatically retards the growth of HepG2 xenograft. Therefore, well-designed engineered ubiquitin ligase represents an effective therapeutic strategy for the treatment of the cancers with co-expressed IGF-1R/IR.

Edgeworthia gardneri (Wall.) Meisn. Water Extract Ameliorates Palmitate Induced Insulin Resistance by Regulating IRS1/GSK3β/FoxO1 Signaling Pathway in Human HepG2 Hepatocytes

“…The activation of Akt induced by insulin accelerates the translocation of GLUT4 vesicles to the plasma membrane, which results in enhanced glucose uptake in the adipose and muscle cells (Gonzalez and McGraw, 2006). Notably, it has been reported that GLUT4 translocation into plasma membrane promotes hepatic glucose uptake, and insulin treatment promoted the translocation of GLUT4 in HepG2 cells (Wang et al, 2014). Our results showed that GLUT4 migration from cytosol to membrane induced by insulin was prevented by PA treatment.…”

The chimeric ubiquitin ligase SH2-U-box inhibits the growth of imatinib-sensitive and resistant CML by targeting the native and T315I-mutant BCR-ABL

“…Another approach is to create a non-endogenous, chimeric E3 ubiquitin ligase that can specifically target the proteins of interest, by replacing the “recognizing and binding” region of an endogenous E3 by an adaptor molecule or directly fusing an E3 “catalytic” domain with an interacting domain of a specific adaptor. With these chimeric E3s, we and others successfully rewired the oncoproteins such as Myc 49 , KRAS 50 , HER2 32 42 , EGFR 33 42 , and IR/IGF-1R 34 to ubiquitin-proteasome degradation route, and as a result, the oncogenes-related malignant behaviors of tumor cells as well as in vivo tumor growth were remarkably inhibited. These promising results prompted us to test whether such a protein knockout strategy could be applied to targeted degradation of the wild type and mutant BCR-ABL in CML.…”

“…The U-box protein CHIP (carboxy terminus of Hsc70 interacting protein) can bind to the molecular chaperone Hsc70 and Hsp90 and mediates ubiquitination and subsequent degradation of their client proteins 31 . Previously, we successfully generated several chimeric ubiquitin ligases by fusing RING domain of Cbl or U-box domain of CHIP with a protein-protein interaction domain to specifically target the oncoproteins such as HER2 32 , EGFR 33 and IR/IGF-1R 34 , and we also demonstrated that they could effectively inhibited these oncoproteins-related tumors.…”

Ligand-Mediated Endocytosis and Trafficking of the Insulin-Like Growth Factor Receptor I and Insulin Receptor Modulate Receptor Function

“…More recent data have shown that an engineered ubiquitin ligase PTBU-box can promote the ubiquitination and degradation of IGF-IR and IR, and thus effectively inhibit in vitro and in vivo tumorigenesis of liver cancer HepG2 and cervical cancer HeLa cells that over-express IGF-IR and IR ( 39 ).…”

“…Because different ubiquitin ligase proteins have been implicated in mediating the ubiquitination of the IGF-IR ( 28 , 33 , 37 , 39 ) we can speculate that different complexes may have different abilities in promoting either polyubiquitination or multiubiquitination of the receptor depending on either cell background or tumor model thus differentially affecting receptor sorting, stability, and biological activity (Figure 1 ).…”