Chaperone-Targeting Cytotoxin and Endoplasmic Reticulum Stress-Inducing Drug Synergize to Kill Cancer Cells

Abstract

Diverse physiological and therapeutic insults that increase the amount of unfolded or misfolded proteins in the endoplasmic reticulum (ER) induce the unfolded protein response, an evolutionarily conserved protective mechanism that manages ER stress. Glucose-regulated protein 78/immunoglobulin heavy-chain binding protein (GRP78/BiP) is an ER-resident protein that plays a central role in the ER stress response and is the only known substrate of the proteolytic A subunit (SubA) of a novel bacterial AB(5) toxin. Here, we report that an engineered fusion protein, epidermal growth factor (EGF)-SubA, combining EGF and SubA, is highly toxic to growing and confluent epidermal growth factor receptor-expressing cancer cells, and its cytotoxicity is mediated by a remarkably rapid cleavage of GRP78/BiP. Systemic delivery of EGF-SubA results in a significant inhibition of human breast and prostate tumor xenografts in mouse models. Furthermore, EGF-SubA dramatically increases the sensitivity of cancer cells to the ER stress-inducing drug thapsigargin, and vice versa, demonstrating the first example of mechanism-based synergism in the action of a cytotoxin and an ER-targeting drug.

Inducible knockout of GRP78/BiP in the hematopoietic system suppresses Pten-null leukemogenesis and AKT oncogenic signaling

“…Therapeutics targeting GRP78 have recently emerged and have shown anti-tumor activity in solid tumors. 16,[51][52][53][54] Whether these and other agents capable of suppressing GRP78 expression or activity can be applied for suppression of leukemogenesis warrants vigorous investigation.…”

Targeting the Unfolded Protein Response in Cancer Therapy

“…We found that EGF-SubA rapidly accumulates in cancer cells via EGFR-mediated endocytosis and is cytotoxic to both growing and confluent EGFR-expressing cells in the picomolar concentration range, and that this toxicity is mediated by cleavage of GRP78 (Backer et al, 2009). In this respect, despite different routes of intracellular delivery, EGF-SubA is similar to the bacterial SubAB holotoxin (Paton et al, 2006).…”

“…Viability of control untreated cells was defined as 100%. For these cell lines, the IC 50 values differ 20-fold, 2 pM for PC3 cells and 40 pM for MDA231luc cells, despite similar levels of EGFR and GRP78 in these cells (Backer et al, 2009). …”

“…We cloned cDNA encoding an EGFR ligand EGF in-frame with cDNA encoding SubA, and expressed the resulting immunotoxin, named EGFSubA, with functionally active EGF and SubA moieties (Backer et al, 2009). We found that EGF-SubA rapidly accumulates in cancer cells via EGFR-mediated endocytosis and is cytotoxic to both growing and confluent EGFR-expressing cells in the picomolar concentration range, and that this toxicity is mediated by cleavage of GRP78 (Backer et al, 2009).…”

“…Notice that usually there are two distinct GRP78 responses to EGF-SubA, cleavage of GRP78 and upregulation of GRP78 expression (Backer et al, 2009). Typically, EGF-SubA-induced cleavage reflects the general susceptibility of cells to the immunotoxin.…”