4E-BP1 Is a Tumor Suppressor Protein Reactivated by mTOR Inhibition in Head and Neck Cancer

Abstract

Aberrant activation of the PI3K-mTOR signaling pathway occurs in >80% of head and neck squamous cell carcinomas (HNSCC), and overreliance on this signaling circuit may in turn represent a cancer-specific vulnerability that can be exploited therapeutically. mTOR inhibitors (mTORi) promote tumor regression in genetically defined and chemically induced HNSCC animal models, and encouraging results have been recently reported. However, the mTOR-regulated targets contributing to the clinical response have not yet been identified. Here, we focused on EIF4E-BP1 (4E-BP1), a direct target of mTOR that serves as key effector for protein synthesis. A systematic analysis of genomic alterations in the PIK3CA-mTOR pathway in HNSCC revealed that 4E-BP1 is rarely mutated, but at least one 4E-BP1 gene copy is lost in over 35% of the patients with HNSCC, correlating with decreased 4E-BP1 protein expression. 4E-BP1 gene copy number loss correlated with poor disease-free and overall survival. Aligned with a tumor-suppressive role, 4e-bp1/2 knockout mice formed larger and more lesions in models of HNSCC carcinogenesis. mTORi treatment or conditional expression of a mutant 4E-BP1 that cannot be phosphorylated by mTOR was sufficient to disrupt the translation-initiation complex and prevent tumor growth. Furthermore, CRISPR/Cas9-targeted 4E-BP1 HNSCC cells resulted in reduced sensitivity to mTORi in vitro and in vivo. Overall, these findings indicate that in HNSCC, mTOR persistently restrains 4E-BP1 via phosphorylation and that mTORi can restore the tumor-suppressive function of 4E-BP1. Our findings also support 4E-BP1 expression and phosphorylation status as a mechanistic biomarker of mTORi sensitivity in patients with HNSCC. Significance: These findings suggest that EIF4E-BP1 acts as a tumor suppressor in HNSCC and that 4E-BP1 dephosphorylation mediates the therapeutic response to mTORi, providing a mechanistic biomarker for future precision oncology trials.

Metformin Inhibits Progression of Head and Neck Squamous Cell Carcinoma by Acting Directly on Carcinoma-Initiating Cells

“…Remarkably, we found that metformin represses the expression of HNSCC stem cell programs, and causes the loss of expression of markers associated with cancer stem cells and promotes HNSCC terminal differentiation. These findings can be explained by our recent observations that 4E-BP1 can suppress the translation of stemness-related genes in HNSCC when reactivated upon mTOR inhibition (15). This is relevant considering that metformin caused the association of dephosphorylated 4E-BP1 with eIF4E, and disruption of the association between eIF4E and eIF4G.…”

“…To begin gaining a mechanistic understanding of metformin actions in HNSCC cells, we first ask if mTOR inhibition and p4E-BP1 reduction results in the reactivation of 4E-BP1, based on our recent findings that 4E-BP1 may represent a candidate tumor suppressor protein in HNSCC, which is reactivated by mTOR blockade (15). Using 7mGTP pull down and eIF4G coimmunoprecipitation assays, we confirmed that the effect of metformin on p4E-BP1 reduction resulted in the increased association of eIF4E with de-phosphorylated 4E-BP1 (de p4E-BP) and a concomitant decreased association of eIF4E with eIF4G ( Fig.…”

Re-expression of DIRAS3 and p53 induces apoptosis and impaired autophagy in head and neck squamous cell carcinoma

“…eIF4E recognizes the 5′-cap structure of mRNAs, delivering these mRNAs to the eIF4F complex for translation initiation. However, 4E-BP1 could bind eIF4E, thereby preventing eIF4F complex assembly and translation initiation [33,34]. The repression of eIF4E function preferentially and disproportionately reduces the expression of numerous growth and survival factors critical for malignancy [35].…”

Mechanism underlying the regulation of sortilin expression and its trafficking function

“…Eukaryotic translation initiation factor 4E-BP1 is a highly conserved eukaryotic initiation factor-4E (eIF-4E) inhibitory protein that regulates translation initiation by binding with eIF-4E (Wang et al, 2019). Low levels of 4E-BP1 phosphorylation inhibit the binding of eIF-4E to the cap structure of 7-methylguanosine at the 5′terminus of a target mRNA via eIF-4E-binding, which prevents the initiation of translation (Choi et al, 2019;Woodcock et al, 2019).…”