Targeting nonhomologous end-joining through epidermal growth factor receptor inhibition: rationale and strategies for radiosensitization

Abstract

DNA double-strand breaks (DSBs) are the most lethal type of DNA damage induced by ionizing radiation or chemotherapeutic drugs used to eradicate cancer cells. The ability of cancer cells to effectively repair DSBs significantly influences the outcome of therapeutic regimens. Therefore, a new and important area of clinical cancer research is the development of DNA repair inhibitors that can be used as radio- or chemosensitizers. Nonhomologous end joining (NHEJ) is the predominant pathway for the repair of radiation-induced DSBs. A series of recent reports indicates that the epidermal growth factor receptor (EGFR) or its downstream components may modulate NHEJ through direct interaction with the DNA repair enzyme, DNA-dependent protein kinase. Because EGFR is overexpressed or activated in many cancers, these findings provide a compelling rationale for combining radiotherapy with therapies that block EGFR or its downstream signaling components. In this review, we delineate how these novel connections between a cell-surface receptor (EGFR) and a predominantly nuclear event (NHEJ) provide vulnerable nodes that can be selectively targeted to improve cancer therapy.

Figure 1. Simplified schematic depicting steps in the non-homologous end-joining (NHEJ) pathway of DNA double-strand break (DSB) repair

DSBs induced by ionizing radiation (IR) or chemotherapeutic agents are recognized by the Ku70/80 heterodimer which then recruits DNA-PKcs (DNA-dependent protein kinase, catalytic subunit) resulting in activation of the latter. Activated DNA-PKcs phosphorylates itself and other proteins involved in repair or damage signaling. Following DNA-end processing by nucleases (such as Artemis and WRN) and DNA polymerases (such as pol λ and pol μ), the ends are ligated together by a complex of XRCC4 (X-ray repair complementing defective repair in Chinese-hamster cells 4), DNA ligase IV, and XLF (XRCC4-like factor, also called Cernunnos). The exact choreography of events at a break and the precise nature of the protein sub-complexes that participate in the repair process may be flexible. As the kinase activity of DNA-PKcs is essential for NHEJ, small molecule inhibitors that bind to and block the kinase domain of DNA-PKcs are very potent radiosensitizers.

Figure 2. Simplified schematic depicting connections between epidermal growth factor receptor (EGFR) signaling and non-homologous end-joining (NHEJ)

Membrane-bound wild type EGFR is activated after binding its ligands: epidermal growth factor (EGF) or transforming growth factor-α (TGF-α). The three domains of EGFR are highlighted with brackets: i (ligand-binding domain), ii (transmembrane domain), and iii (tyrosine kinase domain or TKD). Activated EGFR signals through the following pathways: 1) phosphatidylinositol 3-kinase (PI3K)-Akt-1 pathway, 2) Ras/RAF/mitogen-activated protein kinase (MAPK)/extracellular signal regulated (ERK) pathway, and 3) signal transducer and activation of transcription (STAT) pathway (only the PI3K-Akt-1 pathway is shown for simplicity). The two commonest categories of mutations observed in EGFR are: 1) Somatic “gain-of-function” mutations in the TKD, seen prominently in lung cancers (not shown), and 2) deletion mutations in the extracellular domain, seen commonly in brain cancers (EGFRvIII). EGFRvIII is constitutively active and signals preferentially via the PI3K-Akt-1 pathway. In this pathway, activated phosphatidylinositol kinase (PI3K) phosphorylates phosphatidylinositol-4,5-biphosphate (PIP2) to generate phosphatidylinositol-3,4,5-triphosphate (PIP3). PIP3 anchors Akt-1 to the plasma membrane, where it is phosphorylated by mTOR complex 2 (mTORC2) and 3-phosphoinositide-dependent kinase 1 (PDK1). Activated Akt-1 phosphorylates a plethora of downstream targets resulting in enhanced proliferation and decreased cell death. The PTEN tumor suppressor reverses PIP3 back to PIP2, negatively regulating PI3K-Akt-1 signaling. In response to ionizing radiation (IR), wild type EGFR can translocate into the nucleus and interact with DNA-PKcs (DNA-dependent protein kinase, catalytic subunit) stimulating its DNA repair activity (I). However, EGFR with somatic mutations in the TKD is defective in nuclear translocation and has a radiosensitizing effect (not shown). PI3K-Akt-1 signaling triggered by EGFR or EGFRvIII can also promote DSB repair (II). In response to IR, Akt-1 translocates into the nucleus and interacts with DNA-PKcs. DNA-PKcs phosphorylates Akt-1 in response to IR promoting survival (curved arrow) and we speculate that reciprocal phosphorylation of DNA-PKcs by Akt-1 (?) might promote DSB repair by NHEJ. As EGFR signaling promotes NHEJ, inhibitors of EGFR and Akt-1 can serve as effective radiosensitizers.