ABSTRACT Background Resolvin E1 (RvE1) derived from the ω-3 polyunsaturated fatty acid eicosapentaenoic acid is known to be a potent pro-resolving lipid mediator that prevents chronic inflammation ...and osteoclastogenesis. We investigated the inhibitory effects of RvE1 on osteoclastogenesis and bone resorption to clarify its therapeutic potential for rheumatoid arthritis (RA). Methods Receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation was assessed with tartrate-resistant acid phosphatase staining. RANKL-induced bone resorption was assessed by the measurement of pit formation using calcium phosphate-labeled fluorescent polyanionic molecules in RAW264.7 cells as osteoclast precursors. The effects of RvE1 on the RANKL-induced mRNA expression of osteoclast- specific genes and transcriptional factors such as c-fos and nuclear factor of activated T cells c1 (NFATc1) in RAW264.7 cells were measured by quantitative real-time PCR. The distribution of NFATc1 induced by RANKL was evaluated by immunofluorescence staining in RAW264.7 cells. To analyze the mechanism of the inhibitory effect of RvE1 on osteoclastogenesis, we measured IL-17-induced RANKL mRNA expression in MC3T3-E1 osteoblast cells treated with RvE1 using quantitative real-time PCR and determined the level of prostaglandin E2 (PGE2) production by enzyme-linked immunosorbent assay. Results RvE1 significantly suppressed RANKL-induced osteoclast differentiation and bone resorption. RvE1 inhibited the RANKL-induced mRNA expression of osteoclast-specific genes along with the transcription factors NFATc1 and c-fos. Moreover, NFATc1 translocation from the cytoplasm to the nucleus of RAW264.7 cells was suppressed following RvE1 treatment. RvE1 also inhibited IL-17-induced RANKL mRNA expression and PGE2 production in MC3T3-E1 cells. Conclusion RvE1 inhibited osteoclastogenesis and bone resorption by suppressing RANKL-induced NFATc1 and c-fos expression in osteoclasts and IL-17-induced RANKL expression through the autocrine action of PGE2 in osteoblasts. Our data suggest RvE1 as a new therapeutic target of RA.
Epidermal growth factor receptor (EGFR) is commonly overexpressed in lung cancer. Cetuximab is a chimeric mouse-human antibody targeted against EGFR. Compared with its inhibitory properties, its ...immunologic mechanisms have not been well studied. In this study, we investigated the antibody-dependent cellular cytotoxicity (ADCC) activity of cetuximab against lung cancer cell lines.
We studied the correlation between EGFR expression in lung cancer cell lines and the ADCC activity of cetuximab as well as the influence of interleukin-2 and chemotherapy on the ADCC activity. EGFR expression was measured by a quantitative flow cytometric analysis and immunohistochemistry. The ADCC activity was assessed by a 4-h (51)Cr release assay. Peripheral blood mononuclear cells, purified T cells, natural killer (NK) cells, and monocytes from healthy donors or lung cancer patients were used as effector cells.
Fresh peripheral blood mononuclear cells exhibited cetuximab-mediated ADCC activity against lung cancer cell lines at a low concentration of cetuximab (0.25 microg/mL). A logarithmic correlation was observed between the number of EGFRs and ADCC activity. Even low EGFR expression, which was weakly detectable by immunohistochemistry, was sufficient for maximum ADCC activity, and further increases in EGFR expression on the target cells had no further effect on the ADCC activity. In addition, ADCC activity was enhanced by interleukin-2 mainly through activation of NK cells and was less susceptible to immunosuppression by chemotherapy than NK activity in lung cancer patients.
These observations suggest the importance of ADCC activity as an immunologic mechanism of cetuximab in biological therapy for lung cancer patients.
Background
Adenomyoepithelioma (AME) of the breast is a very rare tumor and is generally considered to be benign. However, some show malignant transformation, which results in local recurrences or ...distant metastases. The morphological features of AME that might predict malignant potential have not been elucidated. Moreover, there is also no established multidisciplinary treatment for malignant AME aside from complete excision at an early stage.
Case presentation
A 64-year-old female diagnosed with AME of the left breast underwent lumpectomy. The surgical margins were negative. Six months after the operation, however, malignant AME recurred locally in the left breast. MRI showed multiple masses, which invaded the skin. A left mastectomy with axillary lymph node dissection was performed. Additional areas of AME were found in about one third of the entire breast. Eight months after the mastectomy, lung metastases were detected. She underwent chemotherapy with fluorouracil, epirubicin, and cyclophosphamide (FEC) for 9 cycles with little response. Lung metastasectomy was performed. Nine months after lung metastasectomy, the metastases were widespread to the brain, heart, and kidney; she subsequently died 2 months later.
Conclusions
Malignant AME has various morphological features, and in this report, we characterize new findings from both imaging and pathology/autopsy. Malignant potency is related to the tumor size, tumor appearance, and mitoses, even if only a few. Given that ductal spread is one of the morphological features of malignant AME, it is of paramount importance to assess the surgical margins.
Purpose
To elucidate whether adjuvant taxane monotherapy is a feasible and tolerable for postoperative breast cancer patients, we evaluated the severity of chemotherapy-induced peripheral neuropathy ...(CIPN) and the relative tolerability of regimens by health-related quality of life (HRQOL) assessment in node-positive breast cancer patients treated with taxane-containing regimens.
Methods
We evaluated CIPN and HRQOL in the first 300 patients enrolled in a larger (1,060 total) multicenter phase III trial randomized to one of four adjuvant regimens: (1) anthracycline–cyclophosphamide followed by paclitaxel (ACP), (2) AC followed by docetaxel (ACD), (3) paclitaxel alone (PTX), or (4) docetaxel alone (DTX). CIPN was assessed by the Patient Neurotoxicity Questionnaire (PNQ) and the National Cancer Institute Common Toxicity Criteria, and HRQOL by Functional Assessment of Cancer Therapy-General (FACT-G). CIPN and HRQOL scores were compared between ACP and ACD vs. PTX and DTX, and ACP and PTX vs. ACD and DTX.
Results
PNQ sensory scores were significantly higher in patients treated with taxane monotherapy compared to treatment with AC followed by taxane (
P
= .003). No significant differences in PNQ sensory scores were observed between the ACP and PTX vs. ACD and DTX regimens (
P
= .669). Regardless of taxane regimen, PNQ severity scores for CIPN appear to be largely reversible within 1 year of adjuvant treatment. No significant difference in FACT-G scores was observed between any regimens during the study treatments.
Conclusions
Patient-reported CIPN was significantly more severe with single-agent adjuvant taxane compared to AC followed by taxane treatment; however, the HRQOL findings support that single-agent taxane treatment is tolerable.
ABSTRACT Background DNA double-strand breaks (DSBs) are the most cytotoxic form of DNA damage and are induced by ionizing radiation and specific chemotherapeutic agents, such as topoisomerase ...inhibitors. Cancer cells acquire resistance to such therapies by repairing DNA DSBs. A major pathway for the repair of DNA DSBs is non-homologous end-joining (NHEJ), which requires DNA-dependent protein kinase (DNA-PK) activity. In this study, we investigated the effect of NU7441, a synthetic small-molecule compound, as a specific inhibitor of DNA-PK on the chemosensitization of non-small cell lung carcinoma (NSCLC) A549 cells. Methods The combined effects of chemotherapeutic agents and NU7441 were evaluated by isobologram analysis using Cell Counting Kit-8. DNA DSBs were assessed by immunofluorescence assay. Apoptosis was examined by flow cytometry using an Annexin V apoptosis kit. Activation of DNA-PK was assayed by western blotting. Results The combination of NU7441 and topoisomerase inhibitors such as amrubicin and irinotecan had a synergistic effect on cell proliferation in A549 cells. NU7441 increased 53BP1 foci and apoptosis induced by topoisomerase inhibitors and decreased phospho-DNA-dependent protein kinase, catalytic subunit (pDNA-PKcs) (S2056) protein expression caused by topoisomerase inhibitors. Interestingly, mitotic inhibitors such as pacritaxel did not cause the pDNA-PKcs (S2056) protein expression and the combination of NU7441 and pacritaxel had an only additive effect. Conclusion NU7441 inhibited the growth of NSCLC cells and enhanced the chemosensitization to topoisomerase inhibitors by blocking DNA repair. A combination of NU7441 and topoisomerase inhibitor may be a promising treatment for NSCLC.
Carbon-ion radiotherapy (CIRT) holds promise to treat inoperable locally-advanced non-small cell lung carcinoma (NSCLC), a disease poorly controlled by standard chemoradiotherapy using X-rays. Since ...CIRT is an extremely limited medical resource, selection of NSCLC patients likely to benefit from it is important; however, biological predictors of response to CIRT are ill-defined. The present study investigated the association between the mutational status of EGFR and KRAS, driver genes frequently mutated in NSCLC, and the relative biological effectiveness (RBE) of carbon-ion beams over X-rays. The assessment of 15 NSCLC lines of different EGFR/KRAS mutational status and that of isogenic NSCLC lines expressing wild-type or mutant EGFR revealed that EGFR-mutant NSCLC cells, but not KRAS-mutant cells, show low RBE. This was attributable to (i) the high X-ray sensitivity of EGFR-mutant cells, since EGFR mutation is associated with a defect in non-homologous end joining, a major pathway for DNA double-strand break (DSB) repair, and (ii) the strong cell-killing effect of carbon-ion beams due to poor repair of carbon-ion beam-induced DSBs regardless of EGFR mutation status. These data highlight the potential of EGFR mutation status as a predictor of response to CIRT, i.e., CIRT may show a high therapeutic index in EGFR mutation-negative NSCLC.
EGFR signaling has been implicated in hypoxia-associated resistance to radiation or chemotherapy. Non-small cell lung carcinomas (NSCLC) with activating L858R or ΔE746-E750 EGFR mutations exhibit ...elevated EGFR activity and downstream signaling. Here, relative to wild-type (WT) EGFR, mutant (MT) EGFR expression significantly increases radiosensitivity in hypoxic cells. Gene expression profiling in human bronchial epithelial cells (HBEC) revealed that MT-EGFR expression elevated transcripts related to cell cycle and replication in aerobic and hypoxic conditions and downregulated RAD50, a critical component of nonhomologous end joining and homologous recombination DNA repair pathways. NSCLCs and HBEC with MT-EGFR revealed elevated basal and hypoxia-induced γ-H2AX-associated DNA lesions that were coincident with replication protein A in the S-phase nuclei. DNA fiber analysis showed that, relative to WT-EGFR, MT-EGFR NSCLCs harbored significantly higher levels of stalled replication forks and decreased fork velocities in aerobic and hypoxic conditions. EGFR blockade by cetuximab significantly increased radiosensitivity in hypoxic cells, recapitulating MT-EGFR expression and closely resembling synthetic lethality of PARP inhibition.
This study demonstrates that within an altered DNA damage response of hypoxic NSCLC cells, mutant EGFR expression, or EGFR blockade by cetuximab exerts a synthetic lethality effect and significantly compromises radiation resistance in hypoxic tumor cells.
.
The EGF receptor (EGFR) contributes to tumor radioresistance, in part, through interactions with the catalytic subunit of DNA-dependent protein kinase (DNA-PKc), a key enzyme in the nonhomologous end ...joining DNA repair pathway. We previously showed that EGFR-DNA-PKcs interactions are significantly compromised in the context of activating mutations in EGFR in non-small cell lung carcinoma (NSCLC) and human bronchial epithelial cells. Here, we investigate the reciprocal relationship between phosphorylation status of DNA-PKcs and EGFR-mediated radiation response. The data reveal that both the kinase activity of DNA-PKcs and radiation-induced phosphorylation of DNA-PKcs by the ataxia telangiectasia-mutated (ATM) kinase are critical prerequisites for EGFR-mediated radioresponse. Alanine substitutions at seven key serine/threonine residues in DNA-PKcs or inhibition of DNA-PKcs by NU7441 completely abrogated EGFR-mediated radioresponse and blocked EGFR binding. ATM deficiency or ATM inhibition with KU55933 produced a similar effect. Importantly, alanine substitution at an ATM-dependent DNA-PKcs phosphorylation site, T2609, was sufficient to block binding or radioresponse of EGFR. However, mutation of a DNA-PKcs autophosphorylation site, S2056 had no such effect indicating that DNA-PKcs autophosphorylation is not necessary for EGFR-mediated radioresponse. Our data reveal that in both NSCLCs and human bronchial epithelial cells, activating mutations in EGFR specifically abolished the DNA-PKcs phosphorylation at T2609, but not S2056. Our study underscores the critical importance of a reciprocal relationship between DNA-PKcs phosphorylation and EGFR-mediated radiation response and elucidates mechanisms underlying mutant EGFR-associated radiosensitivity in NSCLCs.
Epidermal growth factor receptor (EGFR) is commonly overexpressed in malignant pleural mesothelioma (MPM). Cetuximab is a chimeric mouse-human antibody targeted against EGFR and induces potent ...antibody-dependent cellular cytotoxicity (ADCC). The action of cetuximab against MPM cells has not been well studied. Therefore, in this study, we investigated the antitumor activity of cetuximab against MPM cell lines, particularly with respect to ADCC activity in vitro and in vivo. EGFR expression of MPM cells was measured by a quantitative flow cytometric analysis and immunohistochemistry. The effect of cetuximab on growth inhibition was assessed using a modified MTT assay. The ADCC activity was measured by a 4-h 51Cr release assay using fresh or IL-2-activated peripheral blood mononuclear cells. In vivo antitumor activity of cetuximab was evaluated using an orthotopic implantation mouse model. Cetuximab-mediated ADCC activity against MPM cells was observed at low concentration (0.25 mg/ml) and was enhanced by IL-2, whereas no direct effect on growth inhibition was detected. A logarithmic correlation was observed between the number of EGFRs on MPM cells and ADCC activity. Low EGFR expression on the MPM cells, which was weakly detectable by immunohistochemistry, was sufficient for maximum ADCC activity. In the mouse model, cetuximab treatment with or without IL-2 significantly inhibited intrathoracic tumor growth and prolonged their survival. Our study shows that cetuximab has potent anti-MPM activity both in vitro and in vivo, mainly through the immunologic mechanism of ADCC. Cetuximab has the potential to be used as a novel therapy for MPM patients.
Oxidative stress plays an important role in the pathogenesis of bronchial asthma. Antioxidant nutrition and supplementation have been used to reduce oxidative stress. However, a clinical trial with ...antioxidant supplementation showed no beneficial effects in patients with asthma. On the other hand, physical activity is related to the prognosis of chronic obstructive pulmonary disease (COPD) and is also related to oxidant status. We investigated the relationships between oxidative stress, serum levels of vitamins, dietary vitamin intake, daily activities, and pulmonary functions in patients with asthma.
Eighteen patients with bronchial asthma were enrolled in this study. Reactive oxidative stress was assessed by measuring organic hydroperoxides (diacron reactive oxygen metabolites: dROM) in sera and by measuring H
O
levels in exhaled breath condensates. The biological antioxidant capacity in serum was evaluated by measuring antioxidant potential capacity against ferric ion. We also assessed pulmonary functions, fraction of exhaled nitric oxide, serum levels of vitamins, dietary vitamin intake, and physical activities.
There were no relationships between the index of oxidative stress (dROM and H
O
in exhaled breathe condensates) and pulmonary functions, serum levels of vitamins, daily vitamin intakes, and activity levels in patients with asthma.
The status of transient oxidative stress may not be related to daily activities, vitamin levels, and pulmonary functions in patients with asthma in a real-life setting. However, our results were obtained in the short-term period from a small number of subjects, so a large longitudinal study is required to ascertain the relationships between oxidative stress, physical activity and vitamin intake in patients with asthma.
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