Effect of molecular imaging on validation of developed anti-hVEGFR2 therapeutic antibody.

Vascular endothelial growth factor receptor type 2 (VEGFR2)-targeted tumor treatment is an antiangiogenic therapeutic strategy. The human sodium iodide symporter (hNIS) gene is a useful reporter gene for tumor imaging and radiotherapy. In this study, we investigated the evaluation of therapeutic efficacy in hNIS gene-transfected tumor xenografts using a gamma imaging system after treatment with an anti-VEGFR2 antibody. Human breast cancer MDA-MB-231 cells transfected with the hNIS gene were injected subcutaneously into the right flanks of BALB/c nude mice. Therapy was initiated when the tumor volume reached approximately 130-180 mm(3). The animals were intravenously injected with 50, 100, or 150 µg of antibody every 3 days for 16 days. Gamma imaging was performed 1 and 2 weeks after the first injection to monitor the effects of tumor therapy. Mice were sacrificed 2 weeks after the first injection of antibody and the tumors were removed for CD31 staining and reverse transcription-polymerase chain reaction (RT-PCR) assay. All groups of mice that were treated with anti-hVEGFR2 antibody showed markedly reduced tumor growth compared to control mice. In vivo gamma imaging results showed that, at 1 week after the first injection of the anti-hVEGFR2 antibody, (125)I uptake of a tumor treated with 150 µg of antibody was 24.5% lower than that in the controls. At 2 weeks, (125)I uptake in the tumor treated with 150 µg of antibody was as low as 44.3% of that in the controls. CD31 staining and RT-PCR assays showed that blood vessel formation and expression of the hNIS gene were reduced with increased treatment doses. This study demonstrated the feasibility of molecular imaging and the therapeutic efficacy of developing therapeutic antibody anti-hVEGFR2 using a gamma imaging system in hNIS gene-transfected tumor xenograft mice.

Human peroxiredoxin 1 and 2 are not duplicate proteins: the unique presence of CYS83 in Prx1 underscores the structural and functional differences between Prx1 and Prx2.

Human peroxiredoxins 1 and 2, also known as Prx1 and Prx2, are more than 90% homologous in their amino acid sequences. Prx1 and Prx2 are elevated in various cancers and are shown to influence diverse cellular processes. Although their growth regulatory role has traditionally been attributed to the peroxidase activity, the physiological significance of this function is unclear because the proteins are highly susceptible to inactivation by H(2)O(2). A chaperone activity appears to emerge when their peroxidase activity is lost. Structural studies suggest that they may form a homodimer or doughnut-shaped homodecamer. However, little information is available whether human Prx1 and Prx2 are duplicative in structure and function. We noted that Prx1 contains a cysteine (Cys(83)) at the putative dimer-dimer interface, which is absent in Prx2. We studied the role of Cys(83) in regulating the peroxidase and chaperone activities of Prx1, because the redox status of Cys(83) might influence the oligomeric structure and consequently the functions of Prx1. We show that Prx1 is more efficient as a molecular chaperone, whereas Prx2 is better suited as a peroxidase enzyme. Substituting Cys(83) with Ser(83) (Prx1C83S) results in dramatic changes in the structural and functional characteristics of Prx1 in a direction similar to those of Prx2. Here we also report the first crystal structure of human Prx1 and the presence of the Cys(83)-Cys(83) bond at the dimer-dimer interface of decameric Prx1. These findings are consistent with the hypothesis that human Prx1 and Prx2 possess unique functions and regulatory mechanisms and that Cys(83) bestows a distinctive identity to Prx1.

Prx1 suppresses radiation-induced c-Jun NH2-terminal kinase signaling in lung cancer cells through interaction with the glutathione S-transferase Pi/c-Jun NH2-terminal kinase complex.

Radiotherapy is one of the major treatment modalities for lung cancer. Cell killing by ionizing radiation is mediated primarily through the reactive oxygen species (ROS) and ROS-driven oxidative stress. Prx1, a peroxiredoxin family member, was shown to be frequently elevated in lung cancer cells and tissues. Although the antioxidant function of Prx1 is expected to affect the radiotherapy response of lung cancer, the physiologic significance of its peroxidase activity in irradiated cells is unclear because the catalytic Cys52 is easily inactivated by ROS due to its overoxidation to sulfinic or sulfonic acid. In this study, we investigated the role of Prx1 in radiation sensitivity of human lung cancer cells, with special emphasis on the redox status of the catalytic Cys52. We found that overexpression of Prx1 enhances the clonogenic survival of irradiated cells and suppresses ionizing radiation-induced c-Jun NH2-terminal kinase (JNK) activation and apoptosis. The peroxidase activity of Prx1, however, is not essential for inhibiting JNK activation. The latter effect is mediated through its association with the glutathione S-transferase pi (GSTpi)-JNK complex, thereby preventing JNK release from the complex. Reduced JNK activation is observed when the peroxidase activity of Prx1 is compromised by Cys52 overoxidation or in the presence of the Cys52 to Ser52 mutant (Prx1C52S) lacking peroxidase activity. We show that both Prx1 and Prx1C52S interact with the GSTpi-JNK complex and suppress the release of JNK from the complex. Our study provides new insight into the antiapoptotic function of Prx1 in modulating radiosensitivity and provides the impetus to monitor the influence of Prx1 levels in the management of lung cancer.