High-throughput screening of small-molecules libraries identified antibacterials against clinically relevant multidrug-resistant A. baumannii and K. pneumoniae.

BACKGROUND: The current pipeline for new antibiotics fails to fully address the significant threat posed by drug-resistant Gram-negative bacteria that have been identified by the World Health Organization (WHO) as a global health priority. New antibacterials acting through novel mechanisms of action are urgently needed. We aimed to identify new chemical entities (NCEs) with activity against Klebsiella pneumoniae and Acinetobacter baumannii that could be developed into a new treatment for drug-resistant infections. METHODS: We developed a high-throughput phenotypic screen and selection cascade for generation of hit compounds active against multidrug-resistant (MDR) strains of K. pneumoniae and A. baumannii. We screened compound libraries selected from the proprietary collections of three pharmaceutical companies that had exited antibacterial drug discovery but continued to accumulate new compounds to their collection. Compounds from two out of three libraries were selected using "eNTRy rules" criteria associated with increased likelihood of intracellular accumulation in Escherichia coli. FINDINGS: We identified 72 compounds with confirmed activity against K. pneumoniae and/or drug-resistant A. baumannii. Two new chemical series with activity against XDR A. baumannii were identified meeting our criteria of potency (EC50 ≤50 µM) and absence of cytotoxicity (HepG2 CC50 ≥100 µM and red blood cell lysis HC50 ≥100 µM). The activity of close analogues of the two chemical series was also determined against A. baumannii clinical isolates. INTERPRETATION: This work provides proof of principle for the screening strategy developed to identify NCEs with antibacterial activity against multidrug-resistant critical priority pathogens such as K. pneumoniae and A. baumannii. The screening and hit selection cascade established here provide an excellent foundation for further screening of new compound libraries to identify high quality starting points for new antibacterial lead generation projects. FUNDING: BMBF and GARDP.

Aging exacerbates murine lung ischemia-reperfusion injury by excessive inflammation and impaired tissue repair response.

Donor shortage is a major problem in lung transplantation (LTx), and the use of lungs from elderly donors is one of the possible solutions in a rapidly aging population. However, the utilization of organs from donors aged >65 years has remained infrequent and may be related to a poor outcome. To investigate the molecular events in grafts from elderly donors early after LTx, the left lungs of young and old mice were subjected to 1 hour of ischemia and subsequent reperfusion. The left lungs were collected at 1 hour, 1 day, and 3 days after reperfusion and subjected to wet-to-dry weight ratio measurement, histological analysis, and molecular biological analysis, including RNA sequencing. The lungs in old mice exhibited more severe and prolonged pulmonary edema than those in young mice after ischemia reperfusion, which was accompanied by upregulation of the genes associated with inflammation and impaired expression of cell cycle-related genes. Apoptotic cells increased and proliferating type 2 alveolar epithelial cells decreased in the lungs of old mice compared with young mice. These factors could become conceptual targets for developing interventions to ameliorate lung ischemia-reperfusion injury after LTx from elderly donors, which may serve to expand the old donor pool.

Murine Intrapulmonary Tracheal Transplantation: A Model for Investigating Obliterative Airway Disease After Lung Transplantation.

Murine intrapulmonary tracheal transplantation (IPTT) is used as a model of obliterative airway disease (OAD) following lung transplantation. Initially reported by our team, this model has gained use in the study of OAD due to its high technical reproducibility and suitability for investigating immunological behaviors and therapeutic interventions. In the IPTT model, a rodent tracheal graft is directly inserted into the recipient's lung through the pleura. This model is distinct from the heterotopic tracheal transplantation (HTT) model, wherein grafts are transplanted into subcutaneous or omental sites, and from the orthotopic tracheal transplantation (OTT) model in which the donor trachea replaces the recipient's trachea. Successful implementation of the IPTT model requires advanced anesthetic and surgical skills. Anesthetic skills include endotracheal intubation of the recipient, setting appropriate ventilatory parameters, and appropriately timed extubation after recovery from anesthesia. Surgical skills are essential for precise graft placement within the lung and for ensuring effective sealing of the visceral pleura to prevent air leakage and bleeding. In general, the learning process takes approximately 2 months. In contrast to the HTT and OTT models, in the IPTT model, the allograft airway develops airway obliteration in the relevant lung microenvironment. This allows investigators to study lung-specific immunological and angiogenic processes involved in airway obliteration after lung transplantation. Furthermore, this model is also unique in that it exhibits tertiary lymphoid organs (TLOs), which are also seen in human lung allografts. TLOs are comprised of T and B cell populations and characterized by the presence of high endothelial venules that direct immune cell recruitment; therefore, they are likely to play a crucial role in graft acceptance and rejection. We conclude that the IPTT model is a useful tool for studying intrapulmonary immune and profibrotic pathways involved in the development of airway obliteration in the lung transplant allograft.

Perovskite-related structures of Ba2YAlO5 and the ß and α phases of Ba6Y2Al4O15 containing AlO4 tetrahedra.

Single crystals of Ba2YAlO5 and of the α and ß phases of Ba6Y2Al4O15 suitable for X-ray structure analysis were obtained via grain growth of polycrystalline samples prepared by solid-state reactions. Ba2YAlO5 was found to have a monoclinic crystal structure, with lattice parameters a = 7.2333 (7), b = 6.0254 (5), c = 7.4294 (7) Šand ß = 117.249 (3)°, and to belong to the space group P21/m, while α-Ba6Y2Al4O15 was determined to be monoclinic, with a = 5.9019 (2), b = 7.8744 (3), c = 9.6538 (3) Šand ß = 107.7940 (10)°, and the space group Pm, and ß-Ba6Y2Al4O15 was found to be monoclinic, with a = 7.8310 (2), b = 5.8990 (2), c = 18.3344 (6) Šand ß = 91.6065 (11)°, and the space group P2/c. In each of these compounds, BO6 octahedra in ABO3 perovskite-type structures were replaced by AlO4 tetrahedra and YO6 octahedra. Polycrystalline samples in which some Y atoms were replaced with Eu exhibited orange-red luminescence in the range 580-730 nm in response to exposure to radiation having a wavelength of approximately 250 nm.

Impact of intraoperative use of venovenous extracorporeal membrane oxygenation on the status of von Willebrand factor large multimers during single lung transplantation.

Background: von Willebrand factors (vWFs), hemostatic factors, are produced as large multimers and are shear stress-dependently cleaved to become the appropriate size. A reduction in vWF large multimers develops in various conditions including the use of extracorporeal life support, which can cause excessive-high shear stress in the blood flow and result in hemostatic disorders. The objective of this prospective study was to investigate the impact of venovenous extracorporeal membrane oxygenation (VV ECMO) use on the status of vWF large multimers and hemostatic disorders during single lung transplantation (SLT). Methods: We prospectively enrolled 12 patients who underwent SLT at our center. Among them, seven patients were supported by VV ECMO intraoperatively (ECMO group) and the remaining five patients underwent SLT without ECMO support (control group). The vWF large multimer index (%) was defined as the ratio of the large multimer proportion in total vWF (vWF large multimer ratio) derived from a patient's plasma to that from standard human plasma. Results: The vWF large multimer index at the end of the surgery was significantly lower in the ECMO group than in the control group (112.6% vs. 75.8%, respectively; P<0.05). The intraoperative blood loss and the amounts of intraoperative transfusion products in the ECMO group tended to be greater than those in the control group; however, the differences were not significant. Conclusions: During SLT, the use of VV ECMO caused a decrease in the vWF large multimer index. The short duration of time of VV ECMO use in our study did not significantly affect the intra- and postoperative outcomes including blood loss, blood transfusion, and re-exploration thoracotomy for bleeding. Nevertheless, to comprehensively evaluate the actual influence of this decrease in the vWF large multimer index on intra- and postoperative outcomes, a multicenter larger-scale study is warranted.

Bilateral lung transplant with pulmonary artery reconstruction using donor aorta for pulmonary hypertension with a giant pulmonary arterial aneurysm.

OBJECTIVES: Standard bilateral lung transplantation (BLT) is not feasible for patients with pulmonary arterial hypertension (PAH) complicated with a giant pulmonary arterial aneurysm (PAA). This study aimed to describe the outcomes of BLT with pulmonary artery reconstruction (PAR) using donor aorta for such patients. METHODS: This is a retrospective single-centre study reviewing PAH patients with a PAA who received BLT with PAR using donor aorta from January 2010 through December 2020. We compared the characteristics and short- and long-term outcomes of recipients receiving PAR (PAR group) with those who had no PAA and received standard BLT (non-PAR group). RESULTS: Nineteen adult PAH patients underwent cadaveric lung transplantation during the study period. Among them, 5 patients with a giant PAA (median pulmonary artery trunk diameter, 69.9 mm) underwent BLT with PAR using donor aorta and the others received standard BLT. Although the operation time tended to be longer in the PAR group compared with the non-PAR group (1239 vs 958 mins, P = 0.087), 90-day mortality (PAR group: 0% vs non-PAR group: 14.3%, P > 0.99), and 5-year survival rate (PAR group: 100% vs non-PAR group: 85.7%, P = 0.74) was comparable between the groups. No dilatation, constriction or infection of the aortic grafts were recorded during the study period with a median follow-up time of 94 months in the PAR group. CONCLUSIONS: Lung transplantation with PAR using donor aorta is a valid surgical option for PAH patients complicated with a giant PAA.

Honeycomb lung is a major risk factor for preoperative radiological tumor size underestimation in patients with primary lung cancer.

Background: Lung cancer frequently occurs in lungs with background idiopathic interstitial pneumonias (IIPs). Limited resection is often selected to treat lung cancer in patients with IIPs in whom respiratory function is already compromised. However, accurate surgical margins are essential for curative resection; underestimating these margins is a risk for residual lung cancer after surgery. We aimed to investigate the findings of lung fields adjacent to cancer segments affect the estimation of tumor size on computed tomography compared with the pathological specimen. Methods: This analytical observational study retrospectively investigated 896 patients with lung cancer operated on at Fujita Health University from January 2015 to June 2020. The definition of underestimation was a ≥10 mm difference between the radiological and pathological maximum sizes of the tumor. Results: The lung tumors were in 15 honeycomb, 30 reticulated, 207 emphysematous, and 628 normal lungs. The ratio of underestimation in honeycomb lungs was 33.3% compared to 7.4% without honeycombing (P=0.004). Multivariate analysis showed that honeycombing was a significant risk factor for tumor size underestimation. A Bland-Altman plot represented wide 95% limits of agreement, -40.8 to 70.2 mm, between the pathological and radiological maximum tumor sizes in honeycomb lungs.

The phase angle as a predictor of postoperative complications in patients undergoing lung cancer surgery.

PURPOSE: The phase angle (PhA), calculated by bioelectrical impedance analysis, is used as a nutritional risk indicator. A low preoperative PhA has been reported as a marker of postoperative complications in patients with cancer; however, the relationship between the PhA and postoperative complications in patients with lung cancer remains unknown. We conducted this study to assess the predictive ability of the preoperative PhA for postoperative complications in patients undergoing surgery for primary lung cancer. METHODS: We reviewed the data on 240 patients who underwent surgery for primary lung cancer at our institution between August, 2019 and August, 2021. RESULTS: The PhA value in this study was 4.7 ± 0.7°. According to the Clavien-Dindo classification, grade ≥ II postoperative complications occurred in 53 patients (22.0%). Based on the multivariate logistic analysis, only the PhA (odds ratio, 0.51, 95% confidence interval, 0.29-0.90, p = 0.018) was an independent predictor of Clavien-Dindo grade ≥ II postoperative complications. CONCLUSIONS: The PhA may be a valuable marker for predicting the risk of postoperative complications following lung cancer surgery.

Tracheobronchial reconstruction by inverted Barclay's method for tracheobronchial injury in an 8-year-old girl: a case report.

BACKGROUND: Tracheobronchial injury in children is rare but can be highly fatal in severe cases. Therefore, prompt diagnosis and treatment are required. The appropriate treatment method depends on the extent and severity of the injury. CASE PRESENTATION: An 8-year-old girl fell from the fifth floor and was transported to a local hospital. She had a tracheobronchial injury, went into cardiopulmonary arrest during transportation to our hospital. She was revived with cardiopulmonary resuscitation, and veno-arterial extracorporeal membrane oxygenation (VA-ECMO) was commenced. Subsequently, we performed tracheobronchial reconstruction by inverted Barclay's method for tracheobronchial injury. She was switched from VA-ECMO to venovenous (VV)-ECMO 4 days postoperatively, and VV-ECMO was eventually discontinued 27 days after the surgery. The patient was awake and weaned off the ventilator on postoperative day 58. She was discharged 97 days after the surgery. CONCLUSIONS: Tracheobronchial reconstruction by inverted Barclay's method is the preferred surgical technique when other reconstruction techniques are expected to cause excessive tension on the anastomosis of the right main bronchus.

Malignant solitary fibrous tumors of the left atrial endocardium.

Solitary fibrous tumors are typically benign and usually develop in the pleura. We herein report the first case of a solitary fibrous tumor that was pathologically malignant and developed in the left atrial endocardium. A 24-year-old woman underwent resection of a malignant solitary fibrous tumor in her right forearm at another hospital. Computed tomography demonstrated a mass in her right pleura 2 months after the surgery. She was referred to our hospital, and a tumor in her left atrium was subsequently found. She underwent resection of these tumors, and pathological examination showed that they were both malignant solitary fibrous tumors.

Effect of CTLA4-Ig on Obliterative Bronchiolitis in a Mouse Intrapulmonary Tracheal Transplantation Model.

OBJECTIVES: One of the serious problems after lung transplantation is chronic lung allograft dysfunction (CLAD). Most CLAD patients pathologically characterized by obliterative bronchiolitis (OB). Cytotoxic T-lymphocyte-associated antigen 4 (CTLA4)-Ig is a combination protein of the Fc fragment of human IgG1 linked to the extracellular domain of CTLA4. The aim of the study was to examine the effect of CTLA4-Ig therapy on OB using a mouse intrapulmonary tracheal transplantation (IPTT) model. METHODS: IPTT was performed between BALB/c (donor) and C57BL/6 (recipient) mice. Abatacept, which is a commercially available form of CTLA4-Ig, was intraperitoneally injected in recipient mice immediately after surgery, on days 7, 14, and 21. The mice in the control group received human IgG. RESULTS: We performed semi-quantitative analysis of graft luminal obliteration at post-transplant day 28. We calculated the obliteration ratio of the lumen of the transplanted trachea in each case. The obliteration ratio was significantly lower in the CTLA4-Ig group than that in the control group (91.2 ± 2.1% vs. 47.8 ± 7.9%, p = 0.0008). Immunofluorescent staining revealed significantly decreased lymphoid neogenesis in the lung. CONCLUSIONS: CTLA4-Ig therapy attenuated tracheal obliteration with fibrous tissue in the mouse IPTT model. The attenuation of fibrous obliteration was correlated with the inhibition of lymphoid neogenesis.

Targeting stanniocalcin-1-expressing tumor cells elicits efficient antitumor effects in a mouse model of human lung cancer.

Lung cancer is the most common cause of cancer-related death in developed countries; therefore, the generation of effective targeted therapeutic regimens is essential. Recently, gene therapy approaches toward malignant cells have emerged as attractive molecular therapeutics. Previous studies have indicated that stanniocalcin-1 (STC-1), a hormone involved in calcium and phosphate homeostasis, positively regulates proliferation, apoptosis resistance, and glucose metabolism in lung cancer cell lines. In this study, we investigated if targeting STC-1 in tumor cells could be a promising strategy for lung cancer gene therapy. We confirmed that STC-1 levels in peripheral blood were higher in lung cancer patients than in healthy donors and that STC-1 expression was observed in five out of eight lung cancer cell lines. A vector expressing a suicide gene, uracil phosphoribosyltransferase (UPRT), under the control of the STC-1 promoter, was constructed (pPSTC-1 -UPRT) and transfected into three STC-1-positive cell lines, PC-9, A549, and H1299. When stably transfected, we observed significant cell growth inhibition using 5-fluorouracil (5-FU) treatment. Furthermore, growth of the STC-1-negative lung cancer cell line, LK-2 was significantly arrested when combined with STC-1-positive cells transfected with pPSTC-1 -UPRT. We believe that conferring cytotoxicity in STC-1-positive lung cancer cells using a suicide gene may be a useful therapeutic strategy for lung cancer.

Pneumonia With Scedosporium apiospermum and Lomentospora prolificans in a Patient After Bilateral Lung Transplantation for Pulmonary Hypertension: A Case Report.

Infections caused by the Scedosporium genus have become recognized as a fatal complication after lung transplantation in Europe and Australia, but the reports have been rare from Asian countries including Japan. We present a case of pneumonia caused by a mixed infection of Scedosporium apiospermum (SA) and Lomentospora prolificans (LP) that developed after augmentation of immunosuppression for chronic lung allograft dysfunction (CLAD) after lung transplantation. A 13-year-old man underwent bilateral lung transplantation for pulmonary hypertension. One year after surgery, he was treated with a series of augmented immunosuppressive therapy for severe acute rejection and subsequent CLAD. Three months following the first steroid pulse therapy, his serum ß-D-glucan elevated without any sign of fungal infection by other tests. The serum ß-D-glucan once returned to a normal level by empirical administration of micafungin; however, the patient's condition worsened again by discontinuation of it. He did not recover by restarting micafungin, and computed tomography (CT) scans eventually demonstrated new infiltrates in his lung field 6 weeks after the elevation of serum ß-D-glucan. Microscopic findings of transbronchial lung biopsy specimens showed filamentous fungi, and the culture of bronchoalveolar lavage fluid revealed the growth of SA and LP. Despite subsequent voriconazole administration, he died 14 days after the start of voriconazole. Early and aggressive inspection including bronchoscopy should be performed for the diagnosis of Scedosporium infection in immunocompromised patients, even if CT scans and sputum culture show no evidence of infection.

Novel mouse model for evaluating in vivo efficacy of xCT inhibitor.

xCT, a well-known cystine transporter, is reported to be involved in the proliferation of various cells, such as cancer cells, immune cells, and fibroblasts. xCT inhibitor is expected to be a promising drug for cancer or immune diseases. However, there are little studies reporting that xCT inhibitors improve disease progression in vivo. To invent potent xCT inhibitors in vivo, we established a new in vivo model for assessing efficacy of xCT inhibition. dl-propargylglycine (PPG) was administered intraperitoneally to wild-type C57BL/6J mice. Concentration of cystathionine, another substrate of xCT, in the thymus and spleen was measured by LC-MS/MS. PPG increased cystathionine amounts in the thymus and spleen in a dose- and time-dependent manner. At 7 h after PPG administration, the efficacy of erastin, a representative xCT inhibitor, was clearly shown. We synthesized a new compound, Compound A, which had much higher inhibitory effect on xCT than erastin both in vitro and in vivo. We established a mouse model of PPG-induced cystathionine accumulation for assessing xCT inhibition in vivo. By using this model, we discovered that Compound A was approximately 15 times more effective in vivo than erastin.

Discovery and in vivo effects of novel human natriuretic peptide receptor A (NPR-A) agonists with improved activity for rat NPR-A.

Natriuretic peptide receptor A (NPR-A) agonists were evaluated in vivo by optimizing the structure of quinazoline derivatives to improve agonistic activity for rat NPR-A. A 1,4-Cis-aminocyclohexylurea moiety at 4-position and hydroxy group of d-alaninol at 2-position on the quinazoline ring were found to be important factors in improving rat NPR-A activity. We identified potent quinazoline and pyrido[2,3-d]pyrimidine derivatives against rat NPR-A, with double-digit nanomolar EC50 values. The in vivo results showed that compound 56b administered at 1.0 mg/kg/min significantly increased plasma cGMP concentration and urine volume in rats. We discovered novel potent NPR-A agonists that showed agonistic effects similar to those of atrial natriuretic peptide.

[Second Pneumothorax in an Ehlers-Danlos Syndrome Patient Initially Treated by Bullaectomy Twenty-two Years Ago].

A 2nd pneumothorax developed in a 40-years-old man after his 1st bilateral pneumothoraces were successfully treated by bullaectomies 22 years ago. For the past 10 years, he has suffered major complications such as repeated bleeding into the digestive tract due to vascular-type Ehlers-Danlos syndrome (vEDS). Although preoperative computed tomography demonstrated multiple bullae at the apex of the right lung, any abnormal findings suggesting tissue fragility was not found. At surgery, visceral pleura of the right lung had an almost normal appearance through a thoracoscope. However, we found that gentle touch with soft surgical equipment resulted in serious pleural damage leading to new air leakage. Our case suggests a progressive fragility of the lung in a vEDS patient, and a less invasive pleural covering technique would therefore be preferable when major complications become evident, despite previous successful bullaectomy.

The Effects of a Selective CK2 Inhibitor on Anti-glomerular Basement Membrane Glomerulonephritis in Rats.

Protein kinase CK2 ("casein kinase II") is a protein serine/threonine kinase that plays critical roles in biological processes such as cell growth, cell cycle progression, and apoptosis. So far, we have identified that one catalytic isozyme of CK2, CK2α, is over-expressed in the kidney during the progression of glomerulonephritis (GN). Moreover, we have shown that in vivo inhibition of CK2 by administration of CK2 inhibitors was effective in the treatment of experimental GN. Hence the development of potent CK2 inhibitors should be considered in therapeutic strategies for GN. In the present study we identified compound 13, a pyrazine derivative, as a potent CK2 inhibitor. By performing enzyme kinetics analysis in vitro, we characterized the inhibition of compound 13 toward each CK2 catalytic isozyme. Furthermore, in vivo, we demonstrated that compound 13 is effective in attenuating proteinuria, decreasing the enhanced level of blood urea nitrogen and serum creatinine, and ameliorating glomerular crescent formation in an experimental GN rat model. On the other hand, cellular apoptosis was detected in the rat testis following administration of compound 13. This study provides clues for new strategies for developing applicable compounds into CK2-targeted GN treatments.

Computed tomographic lymphography for sentinel lymph node biopsy in male breast cancer: report of two cases.

Male breast cancer is rare, accounting for less than 1% of breast cancers. Because of its rarity evidence of the usefulness sentinel lymph node biopsy (SLNB) for male breast cancer has not been established. Moreover, a navigation system which can easily determine the incision site of SLNB is needed because a second incision for SLNB is necessary in most cases. We report successful computed tomographic lymphography (CTLG)-guided SLNB in two male breast cancer cases: the first patient was a 79-year-old man and the second was a 64-year-old man. Both had presented with a lump behind the nipple. Clinical diagnoses were early breast carcinoma in both cases. The second patient took tamoxifen 20 mg daily as neoadjuvant endocrine therapy. SLNs were clearly visualized by CTLG, allowing mastectomies with SLNB to be performed. Both SLNB were negative, such that axillary lymph node dissection was not needed. Preoperative CTLG is useful for visualizing lymph flow and detecting SLN in male breast cancer.

Diversity-oriented synthesis of pyrazolo[4,3-b]indoles by gold-catalysed three-component annulation: application to the development of a new class of CK2 inhibitors.

Pyrazolo[4,3-b]indole derivatives have been designed as novel CK2 inhibitor compounds based on the binding mode analysis of a previously reported phenylpyrazole-type CK2 inhibitor. A series of pyrazolo[4,3-b]indoles and related dihydropyrazolo[4,3-b]indoles were efficiently prepared from simple starting materials using a gold-catalysed three-component annulation reaction as a key step. Several of the newly synthesized compounds displayed high levels of inhibitory activity, indicating that the pyrazolo[4,3-b]indole core represents a promising scaffold for the development of potent CK2 inhibitors.

Design and synthesis of a novel class of CK2 inhibitors: application of copper- and gold-catalysed cascade reactions for fused nitrogen heterocycles.

Two classes of fused nitrogen heterocycles were designed as CK2 inhibitor candidates on the basis of previous structure-activity relationship (SAR) studies. Various dipyrrolo[3,2-b:2',3'-e]pyridine and benzo[g]indazole derivatives were prepared using transition-metal-catalysed cascade and/or multicomponent reactions. Biological evaluation of these candidates revealed that benzo[g]indazole is a promising scaffold for potent CK2 inhibitors. The inhibitory activities on cell proliferation of these potent CK2 inhibitors are also presented.