Impaired DNA damage response--an Achilles' heel sensitizing cancer to chemotherapy and radiotherapy

Abstract

Despite the progress in targeting particular molecular abnormalities specific to different cancers (targeted therapy), chemo- and radiotherapies are still the most effective of all anticancer modalities. Induction of DNA damage and inhibition of cell proliferation are the objects of most chemotherapeutic agents and radiation. Their effectiveness was initially thought to be due to the high rate of proliferation of cancer cells. However, normal cell proliferation rate in some tissues often exceeds that of curable tumors. Most tumors have impaired DNA damage response (DDR) and the evidence is forthcoming that this confers sensitivity to chemo- or radiotherapy. DDR is a complex set of events which elicits a plethora of molecular interactions engaging signaling pathways designed to: (a) halt cell cycle progression and division to prevent transfer of DNA damage to progeny cells; (b) increase the accessibility of the damaged sites to the DNA repair machinery; (c) engage DNA repair mechanisms and (d) activate the apoptotic pathway when DNA cannot be successfully repaired. A defective DDR makes cancer cells unable to effectively stop cell cycle progression, engage in DNA repair and/or trigger the apoptotic program when treated with DNA damaging drugs. With continued exposure to the drug, such cells accumulate DNA damage which leads to their reproductive death that may have features of cell senescence. Cancers with nonfunctional BRCA1 and BRCA2 are particularly sensitive to combined treatment with DNA damaging drugs and inhibitors of poly(ADP-ribose) polymerase. Antitumor strategies are being designed to treat cancers having particular defects in their DDR, concurrent with protecting normal cells.

Fig. 1. The model of the ATM signaling pathway in response to induction of DSBs, as proposed by Kitagawa et al., (2004)

Induction of DSB causes unwinding and relaxation of torsional stress of the DNA double helix, which leads to local decondensation of chromatin and recruitment of the MRE11, RAD50 and NBS1 proteins (MRN complex), as well as BRCA1 to the DSB site. This event activates ATM by triggering its Ser139 auto-phosphorylation which causes dissociation of the ATM dimer into two enzymatically active monomers. Activated ATM is then recruited to the site of DSB at which point it phosphorylates several substrates including NBS1, BRCA1 and SMC1. Phosphorylation of NBS1 targets ATM to Chk1, and is also essential for targeting ATM to phosphorylate Chk2. Phosphorylation of SMC1 activates the S-phase checkpoint while BRCA1 phosphorylation engages this protein in DSB repair. ATM also phosphorylates other substrates such as E2F1, Chk1, p53, Mdm2, Chk2, and H2AX. Activated p53 induces transcription of p21 and/or Bax p53 genes whose products arrest cells in G₁ or promote apoptosis, respectively.

Fig. 2. Activation of Chk2 and Chk2 major substrates

Chk2 is activated either by ATM (in response to DNA damage) or by ATR (in response to replication stress). The activation is initiated by Thr68 phosphorylation which leads to dimerization of Chk2. The dimerization facilitates further intermolecular trans-phosphorylations at Thr383, Thr387 and Ser516. The dimer then dissociates into monomers. Both multi-phosphorylated dimers and monomers of Chk2 are enzymatically active and able to phosphorylate several substrates. Phosphorylation of the Cdc25C and Cdc25A phosphatases by Chk2 opens a binding site for 14-3-3 proteins, sequestrates them within the complex with 14-3-3 and thereby prevents their translocation into the nucleus and dephosphorylation of inhibitory phosphorylations at Thr14 and Tyr15 on cyclin/CDK complexes, which stalls cell cycle progression at the G₂ to M and G₁ to S transitions, respectively (Ahn et al., 2004). Phosphorylation of Cdc25 phosphatases also accelerates their proteasomal degradation (Boutros et al., 2006). Cell arrest in G₁ can also be mediated by Chk2 through phosphorylation of p53 which may lead to upregulation of the cdk2 inhibitor p21^WAF1. Phosphorylation of p53 may also result in upregulation of Bax, which promotes apoptosis. Induction of apoptosis is additionally promoted by phosphorylation of PML and E2F-1, while phosphorylation of BRCA1 (mediated by both ATM and Chk2) engages the DNA repair pathway.