[4D electromagnetic navigation bronchoscopy for the diagnosis of peripheral pulmonary nodules - An overview and preliminary clinical results]. / 4D-Elektromagnetische Navigationsbronchoskopie zur Diagnostik peripherer Rundherde.

BACKGROUND: The diagnostic of peripheral pulmonary nodules (PPN) is a particular challenge in interventional bronchology, which is why navigation systems such as electromagnetic navigation (ENB) are increasingly being used. The 4D-ENB represents the most current development of the ENB. It utilizes inspiratory and expiratory CT scans for mapping and thus helps compensate for respiratory movements-induced CT-to-body divergence. The aim of this work was to present the first clinical data and experiences using the 4D-ENB method for diagnosis of PPNs. METHODS: We retrospectively describe the results of the first nine consecutive patient cases diagnosed at Klinikum Braunschweig using 4D-ENB in a unimodal diagnostic procedure. RESULTS: Of the first 9 PPNs examined by 4D-ENB, navigation and puncture of the lesion was successful in 8 patients (89%). Diagnostic biopsy was could be carried out in six out of nine patients (67%). There were no significant procedure-related complications. CONCLUSION: Our preliminary data suggest that 4D-ENB is a promising new alternative for the diagnosis of PPNs. To further improve diagnostic yield, 4D-END, which lacks real-time visualization, should be embedded in a multimodal diagnostic procedure with rEBUS and/or fluoroscopy.

Group 2 Innate Lymphoid Cells (ILC2) Suppress Beneficial Type 1 Immune Responses During Pulmonary Cryptococcosis.

Cryptococcus neoformans is an opportunistic fungal pathogen preferentially causing disease in immunocompromised individuals such as organ-transplant-recipients, patients receiving immunosuppressive medications or, in particular, individuals suffering from HIV infection. Numerous studies clearly indicated that the control of C. neoformans infections is strongly dependent on a prototypic type 1 immune response and classical macrophage activation, whereas type 2-biased immunity and alternative activation of macrophages has been rather implicated in disease progression and detrimental outcomes. However, little is known about regulatory pathways modulating and balancing immune responses during early phases of pulmonary cryptococcosis. Here, we analyzed the role of group 2 innate lymphoid cells (ILC2s) for the control of C. neoformans infection. Using an intranasal infection model with a highly virulent C. neoformans strain, we found that ILC2 numbers were strongly increased in C. neoformans-infected lungs along with induction of a type 2 response. Mice lacking ILC2s due to conditional deficiency of the transcription factor RAR-related orphan receptor alpha (Rora) displayed a massive downregulation of features of type 2 immunity as reflected by reduced levels of the type 2 signature cytokines IL-4, IL-5, and IL-13 at 14 days post-infection. Moreover, ILC2 deficiency was accompanied with increased type 1 immunity and classical macrophage activation, while the pulmonary numbers of eosinophils and alternatively activated macrophages were reduced in these mice. Importantly, this shift in pulmonary macrophage polarization in ILC2-deficient mice correlated with improved fungal control and prolonged survival of infected mice. Conversely, adoptive transfer of ILC2s was associated with a type 2 bias associated with less efficient anti-fungal immunity in lungs of recipient mice. Collectively, our date indicate a non-redundant role of ILC2 in orchestrating myeloid anti-cryptococcal immune responses toward a disease exacerbating phenotype.

Emergent Geometric Frustration and Flat Band in Moiré Bilayer Graphene.

So far the physics of moiré graphene bilayers at large, incommensurate rotation angles has been considered uninteresting. It has been held that the interlayer coupling in such structures is weak and the system can be thought of as a pair of decoupled single graphene sheets to a good approximation. Here, we demonstrate that for large rotation angles near commensurate ones, the interlayer coupling, far from being weak, is able to completely localize electrons to within a large scale, geometrically frustrated network of topologically protected modes. The emergent geometric frustration of the system gives rise to completely flat bands, with strong correlation physics as a result. All of this arises although in the lattice structure no large scale pattern appears to the unguided eye. Sufficiently close to commensuration the low-energy physics of this remarkable system has an exact solution.

A CCL1/CCR8-dependent feed-forward mechanism drives ILC2 functions in type 2-mediated inflammation.

Group 2 innate lymphoid cells (ILC2s) possess indispensable roles during type 2-mediated inflammatory diseases. Although their physiological and detrimental immune functions seem to depend on the anatomical compartment they reside, their tissue tropism and the molecular and immunological processes regulating the self-renewal of the local pool of ILC2s in the context of inflammation or infection are incompletely understood. Here, we analyzed the role of the CC-chemokine receptor CCR8 for the biological functions of ILC2s. In vitro and in vivo experiments indicated that CCR8 is in comparison to the related molecule CCR4 less important for migration of these cells. However, we found that activated mouse and human ILC2s produce the CCR8 ligand CCL1 and are a major source of CCL1 in vivo. CCL1 signaling to ILC2s regulates their proliferation and supports their capacity to protect against helminthic infections. In summary, we identify a novel chemokine receptor-dependent mechanism by which ILC2s are regulated during type 2 responses.

Soluble CD83 Triggers Resolution of Arthritis and Sustained Inflammation Control in IDO Dependent Manner.

Interference with autoimmune-mediated cytokine production is a key yet poorly developed approach to treat autoimmune and inflammatory diseases such as rheumatoid arthritis. Herein, we show that soluble CD83 (sCD83) enhances the resolution of autoimmune antigen-induced arthritis (AIA) by strongly reducing the expression levels of cytokines such as IL-17A, IFNγ, IL-6, and TNFα within the joints. Noteworthy, also the expression of RANKL, osteoclast differentiation, and joint destruction was significantly inhibited by sCD83. In addition, osteoclasts which were cultured in the presence of synovial T cells, derived from sCD83 treated AIA mice, showed a strongly reduced number of multinuclear large osteoclasts compared to mock controls. Enhanced resolution of arthritis by sCD83 was mechanistically based on IDO, since inhibition of IDO by 1-methyltryptophan completely abrogated sCD83 effects on AIA. Blocking experiments, using anti-TGF-ß antibodies further revealed that also TGF-ß is mechanistically involved in the sCD83 induced reduction of bone destruction and cartilage damage as well as enhanced resolution of inflammation. Resolution of arthritis was associated with increased numbers of regulatory T cells, which are induced in a sCD83-IDO-TGF-ß dependent manner. Taken together, sCD83 represents an interesting approach for downregulating cytokine production, inducing regulatory T cells and inducing resolution of autoimmune arthritis.

Flupirtine and retigabine as templates for ligand-based drug design of KV7.2/3 activators.

Drug induced liver injury (DILI) and tissue discoloration led to the recent discontinuation of the therapeutic use of the closely related drugs flupirtine and retigabine, respectively. Experience gained with these drugs strongly suggests that heterotetramer, voltage-gated potassium channels 2 and 3 (KV7.2/3) are valid targets for effective treatment of pain and epilepsy. Because the adverse effects are not related to the mechanism of action, it appears promising to investigate chemical modifications of these clinically validated, drug-like leads. In the present retro-metabolic drug design study, a series of 43 compounds were synthesized and characterized with regard to KV7.2/3 opening activity and efficacy. The most active compound 22d displays excellent potency (EC50 = 4 nM) and efficacy (154%) as a KV7.2/3 opener. Limited aqueous solubility hampered toxicity testing at concentrations higher than 63 µM, but this concentration was nontoxic to two hepatocellular cell lines (HEP-G2 and TAMH) in culture. The slightly less active but more soluble compound 25b (EC50 = 11 nM, efficacy 111%) showed an improved toxicity/activity ratio compared to flupirtine by three orders of magnitude and represents an attractive lead structure for the development of safer analgesics and antiepileptics.

Sulfide Analogues of Flupirtine and Retigabine with Nanomolar KV 7.2/KV 7.3 Channel Opening Activity.

The potassium channel openers flupirtine and retigabine have proven to be valuable analgesics or antiepileptics. Their recent withdrawal due to occasional hepatotoxicity and tissue discoloration, respectively, leaves a therapeutic niche unfilled. Metabolic oxidation of both drugs gives rise to the formation of electrophilic quinones. These elusive, highly reactive metabolites may induce liver injury in the case of flupirtine and blue tissue discoloration after prolonged intake of retigabine. We examined which structural features can be altered to avoid the detrimental oxidation of the aromatic ring and shift oxidation toward the formation of more benign metabolites. Structure-activity relationship studies were performed to evaluate the KV 7.2/3 channel opening activity of 45 derivatives. Sulfide analogues were identified that are devoid of the risk of quinone formation, but possess potent KV 7.2/3 opening activity. For example, flupirtine analogue 3-(3,5-difluorophenyl)-N-(6-(isobutylthio)-2-(pyrrolidin-1-yl)pyridin-3-yl)propanamide (48) has 100-fold enhanced activity (EC50 =1.4 nm), a vastly improved toxicity/activity ratio, and the same efficacy as retigabine in vitro.

Innate lymphoid cells in intestinal cancer development.

Colorectal cancer (CRC) is a highly prominent cause of cancer-related deaths worldwide. Although the functions of immune cells in the colorectal tumor microenvironment are complex and heterogeneous, dysregulated changes in the composition and activation state of immune cells are believed to represent key events supporting the establishment of pro- or anti-tumorigenic immune states. Recently, innate lymphoid cells (ILCs) emerged as central innate immune mediators during both gastrointestinal homeostasis and inflammatory pathologies. Hence, ILCs might also represent promising targets in the context of cancer therapy and are increasingly recognized as innate immune cells with potent immunomodulatory properties. In this review, we summarize the pleiotropic roles of the different ILC subsets for intestinal homeostasis and discuss the recent evidence on their potential involvement in the development and growth of intestinal cancers.

Flupirtine Analogues: Explorative Synthesis and Influence of Chemical Structure on KV7.2/KV7.3 Channel Opening Activity.

Neuronal voltage-gated potassium channels KV7.2/KV7.3 are sensitive to small-molecule drugs such as flupirtine, even though physiological response occurs in the absence of ligands. Clinically, prolonged use of flupirtine as a pain medication is associated with rare cases of drug-induced liver injury. Thus, safety concerns prevent a broader use of this non-opioid and non-steroidal analgesic in therapeutic areas with unmet medical needs such as hyperactive bladder or neonatal seizures. With the goal of studying influences of chemical structure on activity and toxicity of flupirtine, we explored modifications of the benzylamino bridge and the substitution pattern in both rings of flupirtine. Among twelve derivatives, four novel thioether derivatives showed the desired activity in cellular assays and may serve as leads for safer KV channel openers.

Group 2 Innate Lymphoid Cells Attenuate Inflammatory Arthritis and Protect from Bone Destruction in Mice.

Group 2 innate lymphoid cells (ILC2s) were detected in the peripheral blood and the joints of rheumatoid arthritis (RA) patients, serum-induced arthritis (SIA), and collagen-induced arthritis (CIA) using flow cytometry. Circulating ILC2s were significantly increased in RA patients compared with healthy controls and inversely correlated with disease activity. Induction of arthritis in mice led to a fast increase in ILC2 number. To elucidate the role of ILC2 in arthritis, loss- and gain-of-function mouse models for ILC2 were subjected to arthritis. Reduction of ILC2 numbers in RORαcre/GATA3fl/fl and Tie2cre/RORαfl/fl mice significantly exacerbated arthritis. Increasing ILC2 numbers in mice by IL-25/IL-33 mini-circles or IL-2/IL-2 antibody complex and the adoptive transfer of wild-type (WT) ILC2s significantly attenuated arthritis by affecting the initiation phase. In addition, adoptive transfer of IL-4/13-competent WT but not IL-4/13-/- ILC2s and decreased cytokine secretion by macrophages. These data show that ILC2s have immune-regulatory functions in arthritis.

ILC2s in infectious diseases and organ-specific fibrosis.

Type 2 immune responses evolved to provide host protection against parasitic infections and to support the repair of infection-induced tissue injury. However, persistent chronic organ damage can result in dysregulated production of critical type 2 cytokines supporting tissue remodeling and fibrosis development. Recently, group 2 innate lymphoid cells (ILC2s) were newly described as central innate mediators of type 2 responses. In particular, by secretion of the cytokines IL-5, IL-9, and IL-13 and the growth factor amphiregulin in response to the release of tissue-derived alarmins, ILC2s have been shown to substantially contribute to both the dismissal of metazoan parasites and the repair of infection-dependent or sterile tissue damage. Conversely, cytokine production by ILC2s emerged as a driving force for tissue remodeling and excessive fibrosis in several organ systems including the lung, liver, and skin. In this review, we discuss how ILC2s are specifically implicated in the body's immune response to different pathogenic infections and how dysregulated ILC2s may promote organ-specific fibrosis.

Chemically induced mouse models of acute and chronic intestinal inflammation.

Inflammatory bowel diseases (IBDs) result in diarrhea and abdominal pain with further potential complications such as tissue fibrosis and stenosis. Animal models help in understanding the immunopathogenesis of IBDs and in the design of novel therapeutic concepts. Here we present an updated version of a protocol we published in 2007 for key models of acute and chronic forms of colitis induced by 2,4,6-trinitro-benzene sulfonic acid (TNBS), oxazolone and dextran sulfate sodium (DSS). This protocol update describes an adaptation of the existing protocol that modifies the technique. This protocol has been used to generate improved mouse models that better reflect the nature of IBDs in humans. In TNBS and oxazolone colitis models, topical administration of hapten reagents results in T-cell-mediated immunity against haptenized proteins and luminal antigens. By contrast, to generate DSS colitis models, mice orally receive DSS, causing death of epithelial cells, compromising barrier function and causing subsequent inflammation. The analysis of the acute colitis models can be performed within 1-2 weeks, whereas that of the chronic models may take 2-4 months. The strengths of the acute models are that they are based on the analysis of short-lasting barrier alterations, innate immune effects and flares. The advantages of the chronic models are that they may offer better insight into adaptive immunity and complications such as neoplasia and tissue fibrosis. The protocol requires basic skills in laboratory animal research.

Programming of Intestinal Epithelial Differentiation by IL-33 Derived from Pericryptal Fibroblasts in Response to Systemic Infection.

The intestinal epithelium constitutes an efficient barrier against the microbial flora. Here, we demonstrate an unexpected function of IL-33 as a regulator of epithelial barrier functions. Mice lacking IL-33 showed decreased Paneth cell numbers and lethal systemic infection in response to Salmonella typhimurium. IL-33 was produced upon microbial challenge by a distinct population of pericryptal fibroblasts neighboring the intestinal stem cell niche. IL-33 programmed the differentiation of epithelial progenitors toward secretory IEC including Paneth and goblet cells. Finally, IL-33 suppressed Notch signaling in epithelial cells and induced expression of transcription factors governing differentiation into secretory IEC. In summary, we demonstrate that gut pericryptal fibroblasts release IL-33 to translate bacterial infection into an epithelial response to promote antimicrobial defense.

3H-1,3-Azaphospholo[4,5-b]pyridines--novel heterocyclic P,N-bridging or hybrid ligands: synthesis and first d(8)-transition metal complexes.

The first 3H-1,3-azaphospholo-pyridines 2a-c were synthesized as racemic mixtures in modest to medium yield by the reaction of N-(2-chloropyrid-3-yl)-trimethylacetimidoyl chloride 1 with RPLi2 (R = Ph, n-Bu, i-Bu), generated from RPH2 and BuLi in THF at -70 °C, and studied with respect to their suitability as ligands (L) in transition metal complexes. Reactions of 2a with group 6 metal(0) pentacarbonyls led to P-coordinated LM(CO)5 complexes 3a-5a (Cr, Mo, W) and the reaction of 2c with (norbornadiene)Mo(CO)4 surprisingly to 4c. [Rh(1,5-COD)Cl]2 and 2a,b, in metal/ligand ratio 1 : 1, furnished LRh(1,5-COD)Cl complexes 6a,b with P-coordination, 6b accompanied by a minor contamination by the bis-coordinated L[Rh(COD)Cl]2 complex 7b. Reactions of 2a,b with [(allyl)PdCl]2 proceeded in THF with dismutation of N-coordinated (allyl)PdCl and formed with 2a a labile crude product [(2a){(allyl)PdCl}1.2(PdCl2)0.8]·C4H8O, with the composition close to L[Pd(allyl)Cl]PdCl2 THF (8a·THF), which converted during crystallization to 9a, whereas 2b directly formed the N,N'-PdCl2-bridged bis[LPd(allyl)chloride] complex 9b. Conversion of 2b with equimolar amounts of Pd(CH3CN)2Cl2 in THF, or Na2PdCl4 in methanol, gave rise to the dimeric P,N-bridging complex 10b. Crystal structure analyses of 6a (rac), 9b·2CDCl3 (meso), 10b·4.5THF and 10b·2D6-acetone (rac) provided detailed structural information. 10b, but more efficiently complexes formed in situ from 2a,b and Pd2(DBA)3 or Pd(OAc)2, catalysed the arylamination of 2-bromopyridine with 2,4,6-trimethylaniline.

The ring opening reaction of 1,3-dithiol-2-one systems is fully reversible.

The deprotection of a common precursor moiety in dithiolene chemistry was discovered to be fully reversible, which, besides being relevant for researchers working in very different fields with these non-innocent ligand systems, may even have an impact on CO2 housekeeping, as the deprotected ligand acts as an efficient trap.

Charge transport through graphene junctions with wetting metal leads.

Graphene is believed to be an excellent candidate material for next-generation electronic devices. However, one needs to take into account the nontrivial effect of metal contacts in order to precisely control the charge injection and extraction processes. We have performed transport calculations for graphene junctions with wetting metal leads (metal leads that bind covalently to graphene) using nonequilibrium Green's functions and density functional theory. Quantitative information is provided on the increased resistance with respect to ideal contacts and on the statistics of current fluctuations. We find that charge transport through the studied two-terminal graphene junction with Ti contacts is pseudo-diffusive up to surprisingly high energies.

A new promising application for highly cytotoxic metal compounds: η6-areneruthenium(II) phosphite complexes for the treatment of alveolar echinococcosis.

Two series of η(6)-areneruthenium(II) phosphite complexes were prepared, characterized, and evaluated in vitro for their toxic potential against Echinococcus multilocularis metacestodes. Neutral complexes of general formula [(η(6)-p-cymene)RuCl(2){P(OR)(3)}] (R = Et, (i)Pr, Ph) with two easily exchangable chloride ligands showed only minor toxicity, whereas the substitution of these moieties against a ß-diketonate (2,2,6,6-tetramethylheptanedionate) ligand led to hydrolytically stable complex salts of type [(η(6)-p-cymene)Ru(ß-diketonate){P(OR)(3)}][BF(4)] (R = Et, (i)Pr, Ph) with comparable in vitro toxicity (50% PGI release at c = 1.4 - 4.7 µM) to the reference drug nitazoxanide (50% PGI release at c = 1.2 µM). In addition, the latter complexes were highly toxic against rat hepatoma cells (IC(50) = 0.40-2.0 µM) and less toxic against human foreskin fibroblasts (IC(50) = 1.1-2.9 µM) and Vero cells (IC(50) = 1.2-8.9 µM). The measured cytotoxicities against mammalian cells are, to the best of our knowledge, among the highest ever observed for ruthenium-based complexes. In conclusion, complex salts of type [(η(6)-p-cymene)Ru(ß-diketonate){P(OR)(3)}][BF(4)] might be interesting candidates for further development toward anthelmintic drugs and/or highly cytotoxic metal compounds.

P=C-N-heterocycles: synthesis of biaryl-type 1,3-benzazaphospholes with ortho-substituted phenyl or 2-heteroaryl groups.

A facile synthesis of functionally substituted 2-(hetero)aryl 1,3-benzazaphospholes via nickel- or palladium-catalyzed phosphonylation of N-acyl-2-bromoanilides 1a-k with triethyl phosphite is presented. Anilidophosphonates 2a-g with naphthoyl-, o-substituted phenyl, furoyl- or thenoyl groups allow direct reductive cyclization with LiAlH(4) to benzazaphospholes 3. The reaction of the o-bromoderivative 2d proceeds with concomitant replacement of bromine by hydrogen, whereas the electron-withdrawing pyridyl group of 2h prevents the synthesis of 3h by this short route. An alternative synthesis of 2-pyridylbenzazaphosphole 3hvia anilidophosphonates succeeded starting from Fmoc-anilinophosphonate 2kvia selective cleavage of the N-protecting group, reduction of the resulting phosphonoaniline to phosphinoaniline and cyclization with pyridine-2-carboxaldehyde via a dihydrobenzazaphosphole 8. N-Substituted pyridylmethylbenzazaphosphole 9 was detected as a side product. The structure elucidation of the new compounds is based on multinuclear NMR data and X-ray crystal structure analyses of a phosphonoanilide, underlining the dominance of N-H···O=P hydrogen bonds over N-H···O=C type hydrogen bonds, of 3h and a supramolecular associate of 3b and its unprecedented air oxidation product 10.

Targeting the epidermal growth factor receptor in non-small cell lung cancer.

The epidermal growth factor receptor (EGFR) has been implicated in a multiplicity of cancer-related signal transduction pathways like cellular proliferation, adhesion, migration, neoangiogenesis and apoptosis inhibition, all of them important features of cancerogenesis and tumour progression. The inhibition of this receptor has been discovered as a suitable pharmaceutical intervention aimed at interrupting tumour activity. In cancer, both monoclonal antibodies and small molecules with anti-tyrosine kinase activity have been assessed in several trials with significant efficacy in clinical applications. The current review focuses in particular on the clinical data of EGFR inhibition in non-small cell lung cancer with emphasis on tyrosine kinase inhibition.