Trastuzumab aggravates radiation induced cardiotoxicity in mice.

Some breast cancer patients with overexpression of human epidermal growth factor receptor 2 need both chest radiotherapy and targeted therapy with trastuzumab (TRZ). The cardiotoxicity associated with combined treatment potentially restricts the clinical benefits of antitumor therapy. There is no consensus on whether and how chest radiotherapy can be given in concurrent with TRZ at present, considering the cardiotoxicity. This study intends to establish an in vitro and in vivo heart injury model by irradiation and TRZ, analyze whether there is a synergistic effect in heart, and to explore the molecular changes. First, an in vitro irradiation model of H9C2 cardiomyocytes was established. The effects of TRZ and radiation on cardiomyocyte injury were observed by cell flow cytometry, CCK-8 test, Western blot, γ-H2AX fluorescence focus formation and cell Reactive Oxygen Species (ROS) content test. Second, the mouse heart injury model was set up by X-ray cardiac irradiation combined with TRZ. Six months later, the cardiac function was analyzed by small animal ultrasound and 18FDG-micro PET/CT. The morphological changes of heart tissue were assessed by histological section. We found that concurrent TRZ aggravates the injury effect of irradiation on cardiomyocytes in vitro. The influence of TRZ might be consequence of inhibiting Akt phosphorylation, promoting the excessive accumulation of ROS in cells and promoting intracellular DNA damage. In animal experiments, the dysfunction of diastolic and myocardial ischemia of mouse heart was observed by echocardiography and 18FDG-micro PET/CT, respectively; myocardial fibrosis and cardiomyocyte apoptosis were also observed. Therefore, our in vitro and in vivo experiments have revealed that TRZ combined irradiation caused more cardiotoxicity than irradiation or TRZ alone. These results suggested that the concurrent management of TRZ and radiotherapy should be carefully made in clinical practice, and more attention is needed on cardiac safety.

RNF19A-mediated ubiquitination of BARD1 prevents BRCA1/BARD1-dependent homologous recombination.

BRCA1-BARD1 heterodimers act in multiple steps during homologous recombination (HR) to ensure the prompt repair of DNA double strand breaks. Dysfunction of the BRCA1 pathway enhances the therapeutic efficiency of poly-(ADP-ribose) polymerase inhibitors (PARPi) in cancers, but the molecular mechanisms underlying this sensitization to PARPi are not fully understood. Here, we show that cancer cell sensitivity to PARPi is promoted by the ring between ring fingers (RBR) protein RNF19A. We demonstrate that RNF19A suppresses HR by ubiquitinating BARD1, which leads to dissociation of BRCA1-BARD1 complex and exposure of a nuclear export sequence in BARD1 that is otherwise masked by BRCA1, resulting in the export of BARD1 to the cytoplasm. We provide evidence that high RNF19A expression in breast cancer compromises HR and increases sensitivity to PARPi. We propose that RNF19A modulates the cancer cell response to PARPi by negatively regulating the BRCA1-BARD1 complex and inhibiting HR-mediated DNA repair.

Asparaginyl endopeptidase (AEP) regulates myocardial apoptosis in response to radiation exposure via alterations in NRF2 activation.

Radiation-induced heart disease (RIHD) leads to myocardial dysfunction and metabolic abnormalities in patients treated with thoracic irradiation which restricts the long-term survival benefits of radiotherapy. There is no specific or effective manner of intervention currently available. Asparaginyl endopeptidase (AEP) plays a pivotal role in the maintenance of cellular functions through regulating proteolytic cleavage as peptidase enzyme. We aimed to investigate the role of unique cardiac AEP in cardiac function by modulating key signaling elements in the myocardium. The murine heart was exposed to a single dose of 14 Gy radiation. Cellular signaling and apoptosis was analyzed in human and rat cardiomyocytes treated with various doses of radiation, we observed expression of AEP was increased by immunohistochemical staining in murine heart exposed to radiation. The AEP production along with its increased level of mRNA expression was associated with increased doses of radiation (0, 2, 5, 10 Gy) in cardiomyocytes. The myocardial cells transfected with AEP overexpression showed overall cellular viability enhancement, DNA damage inhibition, the foci formation of γ-H2AX suppressed and DNA repair enhancement significantly after radiation exposure. Small interfering RNA-mediated AEP knockdown was with reduced cardiomyocyte viability, elevated apoptotic rate, increased γ-H2AX foci formation and inhibited DNA repair as well after irradiation. After radiation exposure of 10 Gy, the expression of AEP increased in P53 overexpressing cardiomyocytes and decreased in the P53 knockdown cells, indicates that radiation-induced expression of AEP might be regulated by P53. Moreover, treatments with either AEP overexpression or knockdown showed enhanced NRF2 activity in the nuclear or suppressed NRF2 expression in the cytoplasm of myocardial cells after irradiation, respectively, defined a possible regulatory effect of AEP associated with diminished NRF2 translocation and activation by radiation exposure, including impair myocardium and myocardial apoptosis. These findings suggest that increased levels of AEP in failing myocardium after irradiation is mediated by P53 and regulate a novel pathway that involves NRF2 activation. AEP is essential for maintaining cellular redox homeostasis of cardiac function.

Long noncoding RNA HOXC-AS3 indicates a poor prognosis and regulates tumorigenesis by binding to YBX1 in breast cancer.

Multiple studies have highlighted the importance of long noncoding RNAs in tumorigenesis. However, the molecular mechanisms underlying the role of lncRNAs in breast cancer are not well understood. Recently, the lncRNA HOXC-AS3 has drawn significant attention due to its regulatory effects on the tumorigenesis of human cancers. However, the potential molecular mechanisms whereby it mediates breast cancer progression remain unknown. Based on public breast cancer expression data and using bioinformatics methods, we discovered significantly upregulated expression levels of HOXC-AS3 in diseased tissues. We verified this result in breast cancer samples and found that the expression of HOXC-AS3 was well correlated with the prognosis of breast cancer. In vitro and in vivo experimental evidence suggests that HOXC-AS3 has the potential to regulate tumorigenesis. Further, mechanistic studies demonstrated the potential of HOXC-AS3 in the transcriptional activation of TK1 via its binding to YBX1. Furthermore, the silencing of TK1 reversed HOXC-AS3-mediated increase in breast cancer cell growth and migration. In conclusion, these results indicated the potential value of HOXC-AS as a prognostic biological marker for breast cancer, and possibly, as a therapeutic target.

Administration of trastuzumab with heart irradiation induced acute cardiotoxicity in mice.

Cardiac toxicity is one of the major advese effect associated with thoracic irradiation. Breast cancer patients with human epidermal factor receptor-2 (Her-2) overexpression could be indicated for both radiation and anti-Her2 target therapy. We aimed to investigate the early detection of radiation and Trastuzumab (TRZ) induced acute cardiotoxicity in mice. In the present study, the heart of animal was subjected to irradiation (IR, 14 Gy/1 Fx), TRZ was intraperitonealy (i.p.) administrated to mice in 2 weeks (6 fractions). The IR plus TRZ group received heart IR after TRZ. We found that body weight of mouse in treatment groups reduced significantly as compared with that of mouse in control group (P<0.05). At day 21, the diastolic function of mice decreased significantly in IR plus TRZ group compared with control group measured by E/E' parameter using echocardiography (57.72 vs 40.82, P<0.05). The left ventricular posterior wall (LVPW) and interventricular septum (IVS) were also increased significantly in diastolic phase at day 21 in the combined group compared with TRZ alone (LVPW: 0.95 mm vs 0.70 mm, P<0.05; IVS: 0.94 mm vs 0.65 mm P<0.05). Moreover, hematoxylin and eosin (HE) staining of cardiac tissue showed that the arrangement of myocardial cell was disordered in the combined group with vacuolar and adipocyte changes, as well as the loose of structure of myocardial cells and the pyknosis of the nucleus. Moderate damage was observed in irradiation-treated group and TRZ-treated group. The expressions of γ-H2AX, vascular cell adhesion molecule-1 (VCAM-1) and von Willebrand Factor (vWF) were remarkedly appeared in co-treatment group. Heart irradiation combined with TRZ treatment simultaneously might cause acute cardiac toxicity in terms of the parameter of E/E', LVPW and IVS. Our results suggest that the diastolic function could detect the early stage of acute cardiotoxicity in heart exposed to irradiation and TRZ co-treatment in mice. The DNA injury and microangiopathy might involve in cardiac injury that aggravated by radiation and Trastuzumab treatments.

Glycyrrhizin mitigates radiation-induced acute lung injury by inhibiting the HMGB1/TLR4 signalling pathway.

Radiation-induced lung injury (RILI) is the major complication of thoracic radiation therapy, and no effective treatment is available. This study explored the role of high-mobility group box 1 (HMGB1) in acute RILI and the therapeutic effect of glycyrrhizin, an inhibitor of HMGB1, on RILI. C57BL/6 mice received a 20 Gy dose of X-ray radiation to the whole thorax with or without administration of glycyrrhizin. Severe lung inflammation was present 12 weeks after irradiation, although only a mild change was noted at 2 weeks and could be alleviated by administration of glycyrrhizin. Glycyrrhizin decreased the plasma concentrations of HMGB1 and sRAGE as well as TNF-α, IL-1ß and IL-6 levels in the bronchoalveolar lavage fluid (BALF). The expression of RAGE was decreased while that of TLR4 was significantly increased at 12 weeks, but not 2 weeks, after irradiation in mouse lung tissue. In vitro, the expression of TLR4 increased in RAW 264.7 cells after conditioning with the supernatant from the irradiated MLE-12 cells containing HMGB1 but showed no change when conditioned medium without HMGB1 was used. However, conditioned culture had no effect on RAGE expression in RAW 264.7 cells. Glycyrrhizin also inhibited the related downstream transcription factors of HMGB/TLR4, such as NF-κB, JNK and ERK1/2, in lung tissue and RAW 264.7 cells when TLR4 was activated. In conclusion, the HMGB1/TLR4 pathway mediates RILI and can be mitigated by glycyrrhizin.

Pituitary adenylate cyclase-activating polypeptide ameliorates radiation-induced cardiac injury.

Radiation-induced heart disease (RIHD) is a common sequelae of thoracic irradiation. Currently, there is no effective prevention and treatment strategy. Oxidative stress is associated with the development of RIHD. Pituitary adenylate cyclase-activating polypeptide 38 (PACAP38) has been defined as the multipotent properties of cytoprotective effect on its anti-apoptotic and antioxidant activities. Here, we set to investigate whether PACAP38 plays a role in attenuating RIHD. We established radiation-related cardiac injury models using 6MV X-ray based on H9C2 cardiomyocytes and male C57/BL6 mice which were pre-treated with PACAP38 prior to radiation exposure. PACAP38 protected mice from radiation-induced histological damage including myocardial apoptosis and fibrosis. Also, cell viability and colony-forming efficiency were enhanced and intracellular ROS generation was reduced in PACAP38 treated H9C2 cardiomyocytes exposed to radiation. Moreover, PACAP38 suppressed myocardial apoptosis and G2/M arrest through blunting the radiation-induced down-regulation of Bcl-2, CyclinB1 and CDC2, and inhibiting the up-regulation of Bax. Furthermore, irradiation resulted in activating of NRF2 and HO-1 expressions were further enhanced by PACAP38 in H9C2 cells and the protective effect of PACAP38 was partially blocked by NRF2 siRNA silencing. In summary, PACAP38 has the potential to effectively protect against acute radiation-induced cardiac injury and its cardioprotective effect involves upregulation of NRF2/HO-1-dependent signaling activation.

Intraoperative radiotherapy for the treatment of thyroid cancer: a pilot study.

We preliminarily evaluated the clinical feasibility and efficacy of intraoperative radiotherapy in patients with thyroid carcinoma. Nine thyroid cancer patients received intraoperative radiotherapy using an Intrabeam system. The dose was 3-4 Gy and the irradiation time ranged from 1 min 32 s to 7 min 33s. One case was a primary thyroid carcinoma, while the other cases were recurrent disease. Adverse effects, recurrence and survival were analyzed. In one patient, poorly differentiated thyroid carcinoma recurred 5 months after treatment, one patient developed a postoperative tracheal skin fistula, and one patient developed a wound infection. Because the affected areas were treated with both surgical resection and then radiotherapy, it is difficult to know which of those led to the adverse effects. Nonetheless, our results indicate that intraoperative radiotherapy can relieve the symptoms associated with thyroid cancer and improve the quality of life for these patients. It thus appears feasible to treat thyroid cancer patients with intraoperative radiotherapy.