Cyclophilin inhibitors restrict Middle East respiratory syndrome coronavirus via interferon-λ in vitro and in mice

While severe coronavirus infections, including Middle East respiratory syndrome coronavirus (MERS-CoV), cause lung injury with high mortality rates, protective treatment strategies are not approved for clinical use. We elucidated the molecular mechanisms by which the cyclophilin inhibitors cyclosporin A (CsA) and alisporivir (ALV) restrict MERS-CoV to validate their suitability as readily available therapy in MERS-CoV infection. Calu-3 cells and primary human alveolar epithelial cells (hAECs) were infected with MERS-CoV and treated with CsA or ALV or inhibitors targeting cyclophilin inhibitor-regulated molecules including calcineurin, nuclear factor of activated T-cells (NFATs) or mitogen-activated protein kinases. Novel CsA-induced pathways were identified by RNA sequencing and manipulated by gene knockdown or neutralising antibodies. Viral replication was quantified by quantitative real-time PCR and 50% tissue culture infective dose. Data were validated in a murine MERS-CoV infection model. Both CsA and ALV reduced MERS-CoV titres and viral RNA replication in Calu-3 cells and hAECs, improving epithelial integrity. While neither calcineurin nor NFAT inhibition reduced MERS-CoV propagation, blockade of c-Jun N-terminal kinase diminished infectious viral particle release but not RNA accumulation. Importantly, CsA induced interferon regulatory factor 1 (IRF1), a pronounced type III interferon (IFNλ) response and expression of antiviral genes. Downregulation of IRF1 or IFNλ increased MERS-CoV propagation in the presence of CsA. Importantly, oral application of CsA reduced MERS-CoV replication in vivo, correlating with elevated lung IFNλ levels and improved outcome. We provide evidence that cyclophilin inhibitors efficiently decrease MERS-CoV replication in vitro and in vivo via upregulation of inflammatory antiviral cell responses, in particular IFNλ. CsA might therefore represent a promising candidate for treating MERS-CoV infection

Video-Assisted Mediastinoscopic Lymphadenectomy (VAMLA)

Background:The development of a two-bladed spreadable videomediastinoscope in 1992 allowed increased exposure and bimanual dissection of mediastinal structures. Concurrent with technical progress in mediastinoscopy, neoadjuvant treatment of stage III lung cancer was introduced, and accuracy of pretreatment mediastinal staging became a topic at issue. In this setting, development of a videomediastinoscopic technique for complete mediastinal lymphadenectomy was the obvious thing to do.Methods:Video-assisted mediastinoscopic lymphadenectomy (VAMLA) dissection is guided by anatomical landmarks, very similar to open lymphadenectomy. It includes en bloc resection of the right and central compartments and dissection and lymphadenectomy of the left-sided compartments. In a preliminary case-control study of 40 patients, VAMLA technique was standardized and evaluated against open lymphadenectomy. A second study investigated 130 patients with resectable lung cancer and radiographically normal mediastinum who underwent VAMLA and consecutive lung resection with mediastinal reexploration.Results:VAMLA harvested significantly more nodes than open lymphadenectomy. With a mean duration of 54 minutes and a complication rate of 4.6%, VAMLA appeared applicable to clinical routine. We noted a sensitivity of 93.8%, a specificity of 100%, and a false-negative rate of 0.9%.Conclusions:In our experience, VAMLA is a feasible method of mediastinal staging. Its accuracy and radicality can equal open lymphadenectomy. However, VAMLA is minimally invasive and therefore pretherapeutically available. Its advantages might be of interest with neoadjuvant strategies, trials, involved field radiation, video-assisted thoroscopic lobectomy, and left-sided tumors