Complex Entry Research Articles

A Single Center Analysis of Advanced Non-small Cell Lung Cancer Patients Treated with Immunotherapy in Real-world Practice

背景与目的近些年，多项临床试验显示免疫检查点抑制剂（immunocheckpoint inhibitor, ICI）为晚期非小细胞肺癌（non-small cell lung cancer, NSCLC）患者带来生存获益，但临床试验有着严格而复杂的纳入与排除标准，其结果不能完全反映真实世界的实际情况。本研究拟探讨真实世界中免疫治疗的临床疗效和安全性以及可能相关的预后因素。方法回顾性分析2017年1月-2019年7月在北京胸科医院接受免疫治疗的晚期NSCLC患者，收集患者基本临床资料、治疗疗效、无进展生存期（progression-free survival, PFS）和药物不良反应等资料，探讨临床疗效、不良反应及可能相关的预后因素。结果研究共纳入34例患者，中位PFS为5.66个月（95%CI：4.48个月-6.84个月），1级-2级不良反应和3级-4级不良反应发生率分别为61.71%（22/34）和14.71%（5/34），共有3例（8.82%）患者出现致死性免疫相关不良反应（immune-related adverse event, irAE），其中2例为免疫相关肺炎，1例为免疫相关心肌炎。单因素分析显示肿瘤-淋巴结-转移（tumor-node-metastasis, TNM）分期、转移部位与中位PFS相关（P < 0.05），多因素分析显示存在肺外转移（OR=6.42, P=0.029）、胸膜转移（OR=14.14, P=0.006）为患者PFS的独立预后因素。结论真实世界中免疫治疗对晚期NSCLC患者具有良好的疗效，但其严重irAE的发生率也较高。存在肺外转移、胸膜转移是接受免疫治疗的晚期NSCLC患者的不良预后因素。

A Hyperstabilizing Mutation in the Base of the Ebola Virus Glycoprotein Acts at Multiple Steps To Abrogate Viral Entry

Ebola virus (EBOV) causes highly lethal disease outbreaks against which no FDA-approved countermeasures are available. Although many host factors exploited by EBOV for cell entry have been identified, including host cell surface phosphatidylserine receptors, endosomal cysteine proteases, and the lysosomal cholesterol trafficking protein NPC1, key questions remain. Specifically, late entry steps culminating in viral membrane fusion remain enigmatic. Here, we investigated a set of glycoprotein (GP) mutants previously hypothesized to be entry defective and identified one mutation, R64A, that abolished infection with no apparent impact on GP expression, folding, or viral incorporation. R64A profoundly thermostabilized EBOV GP and rendered it highly resistant to proteolysis in vitro Forward-genetics and cell entry studies strongly suggested that R64A's effects on GP thermostability and proteolysis arrest viral entry at least at two distinct steps: the first upstream of NPC1 binding and the second at a late entry step downstream of fusion activation. Concordantly, toremifene, a small-molecule entry inhibitor previously shown to bind and destabilize GP, may selectively enhance the infectivity of viral particles bearing GP(R64A) at subinhibitory concentrations. R64A provides a valuable tool to further define the interplay between GP stability, proteolysis, and viral membrane fusion; to explore the rational design of stability-modulating antivirals; and to spur the development of next-generation Ebola virus vaccines with improved stability.IMPORTANCE Ebola virus is a medically relevant virus responsible for outbreaks of severe disease in western and central Africa, with mortality rates reaching as high as 90%. Despite considerable effort, there are currently no FDA-approved therapeutics or targeted interventions available, highlighting the need of development in this area. Host-cell invasion represents an attractive target for antivirals, and several drug candidates have been identified; however, our limited understanding of the complex viral entry process challenges the development of such entry-targeting drugs. Here, we report on a glycoprotein mutation that abrogates viral entry and provides insights into the final steps of this process. In addition, the hyperstabilized phenotype of this mutant makes it useful as a tool in the discovery and design of stability-modulating antivirals and next-generation vaccines against Ebola virus.

An exploration of conditions proposed to trigger the Ebola virus glycoprotein for fusion.

Ebolaviruses continue to inflict horrific disease and instill fear. The 2013–2016 outbreak in Western Africa caused unfathomable morbidity and mortality (over 11,000 deaths), and the second largest outbreak is on-going in the Democratic Republic of the Congo. The first stage of an Ebolavirus infection is entry, culminating in delivery of the viral genome into the cytoplasm to initiate replication. Among enveloped viruses, Ebolaviruses use a complex entry pathway: they bind to attachment factors on cell surfaces, are engulfed by macropinocytosis, and traffic through the endosomal system. En route, the receptor binding subunit of the glycoprotein (GP) is reduced from ~130 to ~19 kDa by cathepsins. This event allows cleaved GP (GPcl) to bind to Niemann-Pick C1 (NPC1), its endosomal receptor. The virus then fuses with a late endosomal membrane, but how this occurs remains a subject of debate. An early, but standing, observation is that entry of particles bearing GPcl is inhibited by agents that raise endosomal pH or inhibit cysteine proteases, suggesting the need for an additional factor(s). Yet, some have concluded that NPC1 is sufficient to trigger the fusion activity of GPcl. Here, we re-examined this question using sensitive cell-cell and pseudovirus-cell fusion assays. We did not observe detectable GPcl-mediated fusion with NPC1 or its GPcl binding domain at any pH tested, while robust fusion was consistently observed with GP from lymphocytic choriomeningitis virus at low pH. Addition of proposed fusion-enhancing factors—cations (Ca++ and K+), a reducing agent, the anionic lipid Bis(Monoacylglycero)Phosphate, and a mixture of cathepsins B and L—did not induce detectable fusion. Our findings are in line with the earlier proposal that an additional factor is required to trigger the full fusion activity of GPcl after binding to NPC1. We discuss caveats to our study and what the missing factor(s) might be.

Biosynthetically Inspired Transformation of Iboga to Monomeric Post-iboga Alkaloids

Summary Organisms have evolved to produce various natural products from a common precursor as a means of maximizing the number of secondary metabolites, thus acquiring selectional advantages. By adopting nature's divergent biosynthetic strategy, we transformed (+)-catharanthine to various iboga and post-iboga alkaloids, including (−)-conodusine A, (+)-conodusines B and C, (−)-voatinggine, (−)-tabertinggine, and (+)-dippinine B. Well-orchestrated oxidations and reorganizations of catharanthine derivatives enabled chemical access to these natural products.

Lighting Environment in the Outdoor Space of Apartment Complexes

This study aims to identify the current state of night outdoor-lighting environments for apartment complexes in Korea in order to provide ways to improve that environment. Field studies of two apartment complexes were conducted to measure physical environments and analyze the psychological assessment results. The physical environments were measured with horizontal illumination, and the psychological assessments were carried out with a survey of students using the semantic differential method (SD method). The study was conducted from November 2015 to January 2016. The study results were as follows. The mean illuminance at the complex entry, square, pedestrian paths, exercise facility, and children’s playground did not meet the standard illuminance of the KS illuminance criteria. The assessment result on location and sense of security revealed that the complex entry and building entry were safe, which was a positive assessment; however, the pedestrian path, square facility, and childrens playground were felt to be insecure, which was a negative assessment. The assessment of sense of security according to gender showed that female students felt more insecure than male students. The factor analysis result showed that four factors were derived from each apartment: security, activity, openness, and glare for the S apartment complex, and brightness, security, activity, and openness for the I apartment complex.

Evaluating the Paper-to-Screen Translation of Participant-Aided Sociograms with High-Risk Participants.

While much social network data exists online, key network metrics for high-risk populations must still be captured through self-report. This practice has suffered from numerous limitations in workflow and response burden. However, advances in technology, network drawing libraries and databases are making interactive network drawing increasingly feasible. We describe the translation of an analog-based technique for capturing personal networks into a digital framework termed netCanvas that addresses many existing shortcomings such as: 1) complex data entry; 2) extensive interviewer intervention and field setup; 3) difficulties in data reuse; and 4) a lack of dynamic visualizations. We test this implementation within a health behavior study of a high-risk and difficult-to-reach population. We provide a within-subjects comparison between paper and touchscreens. We assert that touchscreen-based social network capture is now a viable alternative for highly sensitive data and social network data entry tasks.

A Criterion for Essential Self-Adjointness of a Symmetric Operator Defined by Some Infinite Hermitian Matrix with Unbounded Entries

We shall consider a double infinite, Hermitian, complex entry matrix \({A=[a_{x,y}]_{x,y\in\mathbb{Z}}}\). In the present note we give a criterion, expressed in terms of the entries of the matrix, for the corresponding symmetric operator defined on compactly supported sequences, to be essentially self-adjoint in the space \({\ell_2(\mathbb{Z})}\). Roughly speaking, assuming that x denotes the row number, we require that: (1) there exist \({\gamma\in[0,1)}\) and n > 0 for which the entries that are at distance larger than \({n(|x|^2+1)^{\gamma/2}}\) from the diagonal vanish and (2) the \({\ell^1}\) norm of the xth row grows slower that \({|x|^{\gamma-1}}\), as \({|x|\to+\infty}\).

Translocation of botulinum neurotoxin serotype A and associated proteins across the intestinal epithelia.

Botulinum neurotoxins (BoNTs) are some of the most poisonous natural toxins. Botulinum neurotoxins associate with neurotoxin-associated proteins (NAPs) forming large complexes that are protected from the harsh environment of the gastrointestinal tract. However, it is still unclear how BoNT complexes as large as 900 kDa traverse the epithelial barrier and what role NAPs play in toxin translocation. In this study, we examined the transit of BoNT serotype A (BoNT/A) holotoxin, complex and recombinantly purified NAP complex through cultured and polarized Caco-2 cells and, for the first time, in the small mouse intestine. Botulinum neurotoxin serotype A and NAPs in the toxin complex were detectable inside intestinal cells beginning at 2 h post intoxication. Appearance of the BoNT/A holotoxin signal was slower, with detection starting at 4–6 h. This indicated that the holotoxin alone was sufficient for entry but the presence of NAPs enhanced the rate of entry. Botulinum neurotoxin serotype A detection peaked at approximately 6 and 8 h for complex and holotoxin, respectively, and thereafter began to disperse with some toxin remaining in the epithelia after 24 h. Purified HA complexes alone were also internalized and followed a similar time course to that of BoNT/A complex internalization. However, recombinant HA complexes did not enhance BoNT/A holotoxin entry in the absence of a physical link with BoNT/A. We propose a model for BoNT/A toxin complex translocation whereby toxin complex entry is facilitated by NAPs in a receptor-mediated mechanism. Understanding the intestinal uptake of BoNT complexes will aid the development of new measures to prevent or treat oral intoxications.

Biological responses to nanoscale particles.

Biological responses to nanoscale particles.

Occipital Condyle-C1 Complex Screw for Fixation of Basilar Invagination Patients with Atlas Assimilation.

To investigate whether C1-occipital condyle complex (CC complex) screws can be safely and rapidly placed without guidance in basilar invagination patients with atlas assimilation. The occipital-cervical (OC) junction was fixed posteriorly in 8 basilar invagination patients with atlas assimilation using polyaxial titanium screws inserted unicortically into the CC complex and C2 pedicles and subsequent fixation to a 3-mm rod. Anatomic landmarks were used during the drilling. The screw angles and lengths of the CC complex were analyzed. The width, length, and height values of the left-side CC complex were 7.96±2.23, 16.06±2.73, and 13.76±2.06 mm; those of the right-side CC complex were 7.84±1.38, 16.66±2.58, and 12.81±2.62 mm. The CC complex entry point was at the central point of posterior surface of the CC complex. The angle used for medialization was 10-15°, which was also the maximal superior screw angulation in the sagittal plane. The screw length required for unicortical purchase was 16-22 mm. The screw was not misplaced or poorly positioned, and no neurovascular complications associated with screw insertion were detected. In patients with atlas assimilation, CC complex screws can be placed safely. The CC complex screws can be safely inserted assisted by microscope without image guidance.

Resources-for-infrastructure (R4I) swaps

Purpose – This paper aims to understand the role resources-for-infrastructure (R4I) swaps play in internationalisation strategies, thereby contributing to a modern theory of the multinational enterprises (MNEs) based on experiences of rising power firms. Since 2004, the Chinese Government; state-owned policy banks; and oil, mining and construction corporations have used a relatively unique form of internationalisation through complex, large-scale R4I swaps in Africa. Design/methodology/approach – This paper uses a resource bundling perspective and political economy lens to analyse complex entry decisions and success, as well as the failure of R4I swaps. The paper is based on a comparative analysis of published case studies of R4I swaps in seven African countries complemented by field research by the first author. Findings – The findings show that, under very specific circumstances, R4I swaps can be considered as a successful internationalisation strategy. R4I swaps enable Chinese MNEs to build and maintain relationships with non-market elites that control access to natural resources and infrastructure contracts. Research limitations/implications – The sample of cases, although representing all relevant R4I-swaps, is too small to come for more quantitative conclusions on success/failure factors. Practical implications – R4I swaps are a very unlikely model for Western MNEs, as they lack the necessary country-specific competitive advantages and institutional mechanisms. Originality/value – To the authors’ knowledge, this is the first comprehensive study of all relevant Chinese R4I swaps in Africa and contains original data from fieldwork in Ghana and D.R. Congo.

Morphing: The Linkage of Inward Private Equity and Outward Ventures

Private equity is impacting the global economy and competitive landscape of multinational enterprises (MNEs) worldwide. In this pioneering study, we find an important new pattern for global interfirm connections: an inward private equity investment is inductive for emerging MNEs to conduct an outward venture. This inward-outward linkage implies that emerging MNEs are more likely to undergo a series of organizational changes after receiving private equity investments. These changes include restructuring their boards of directors, rebuilding their top management teams, reconfiguring their corporate resources, reframing the industry structure, and altering competitive dynamics. These organizational changes induce emerging MNEs to modify their strategies such as speeding up the internationalization process, locating additional outward ventures in advanced economies, and choosing more complex entry modes, particularly cross-border mergers and acquisitions (M&As). We describe this type of evolving internationalization of emerging MNEs as a “morphing” process with in-depth case studies on emerging Chinese MNEs including Lenovo, Zoomlion, Geely, Alibaba.com, Huawei, NVC Lighting, and BYD. © 2014 Wiley Periodicals, Inc.

New Trends in Foreign Market Entry Mode Choices: The Case of Italian Mid-Sized Companies

As Italian SMEs increasingly expand their markets overseas, the internationalization processes evolve towards more complex market entry modes based on equity forms of international expansion and investment such as JVs and WOSs. Research on international high-control entry mode has tended to concentrate on large firms. In order to test the choice between equity and non-equity entry modes to the specific category of the mid-sized companies, the paper provides two main contributions: 1) it proposes an integration of three perspectives, transaction cost, institutional and resource-based theory; 2) it empirically verifies the applicability to the medium sized firms of the proposed framework.

Immunity protein release from a cell‐bound nuclease colicin complex requires global conformational rearrangement

Nuclease colicins bind their target receptor BtuB in the outer membrane of sensitive Escherichia coli cells in the form of a high-affinity complex with their cognate immunity proteins. The release of the immunity protein from the colicin complex is a prerequisite for cell entry of the colicin and occurs via a process that is still relatively poorly understood. We have previously shown that an energy input in the form of the cytoplasmic membrane proton motive force is required to promote immunity protein (Im9) release from the colicin E9/Im9 complex and colicin cell entry. We report here that engineering rigidity in the structured part of the colicin translocation domain via the introduction of disulfide bonds prevents immunity protein release from the colicin complex. Reduction of the disulfide bond by the addition of DTT leads to immunity protein release and resumption of activity. Similarly, the introduction of a disulfide bond in the DNase domain previously shown to abolish channel formation in planar bilayers also prevented immunity protein release. Importantly, all disulfide bonds, in the translocation as well as the DNase domain, also abolished the biological activity of the Im9-free colicin E9, the reduction of which led to a resumption of activity. Our results show, for the first time, that conformational flexibility in the structured translocation and DNase domains of a nuclease colicin is essential for immunity protein release, providing further evidence for the hypothesis that global structural rearrangement of the colicin molecule is required for disassembly of this high-affinity toxin-immunity protein complex prior to outer membrane translocation.

(Ir)regularity of verbs revisited

In three experiments we explored the representation and encoding of verb regularity. Contrasting articulation latencies of present and past regular and irregular forms from paradigms consisting of regular, irregular, or hybrid verbs (regular in present, but irregular in past tense), allowed us to differentiate between affixation and stem selection processes. The analyses revealed that regular verbs in the present and past tense were produced significantly faster than all other forms. Crucially, the naming latencies of hybrid and irregular verbs did not differ from each other in both tenses. We conclude that regularity is not a property of individual verb forms, but generalizes to all forms within a paradigm. In a third, picture word interference experiment, we tested whether regularity, when not bound to individual verb forms, is represented in the form of abstract regularity nodes, as assumed for gender or conjugational class. However, the critical conditions did not exhibit the expected congruency effect. Consequently we conclude that the paradigmatic effects could be explained as a result of the complexity of lexical entries. The retrieval of a complex entry (irregular and hybrid verbs) takes longer than the retrieval of a single stem entry (regular verbs).

Virale Infektion von Hepatozyten

Many viruses infect hepatocytes. On the one hand an understanding of the underlying molecular mechanisms can be used to block infection by pathogenic viruses, on the other hand hepatotropic viruses can be utilized in gene therapy approaches for the directed delivery of genetic material into hepatocytes. The hepatitis C virus (HCV) follows a complex cell entry route utilizing at least four essential cell surface receptors on hepatocytes. Inhibitors of HCV cell entry are in early clinical development and could useful for the prevention of HCV reinfection of the graft after liver transplantation. Although much less is known about the cell entry of hepatitis B virus and hepatitis D virus (HBV; HDV) it can be blocked efficiently by active or passive immunization. Moreover, a highly specific lipopeptide entry inhibitor based on a fragment of the HBV envelope is in clinical development. Finally, approaches are being developed to use hepatotropic viruses to correct genetic defects in hepatocytes. Especially adeno-associated virus based vector systems have recently shown promising results in proof-of-concept studies.

Accurate prediction of pregnancy viability by means of a simple scoring system

What is the performance of a simple scoring system to predict whether women will have an ongoing viable intrauterine pregnancy beyond the first trimester? A simple scoring system using demographic and initial ultrasound variables accurately predicts pregnancy viability beyond the first trimester with an area under the curve (AUC) in a receiver operating characteristic curve of 0.924 [95% confidence interval (CI) 0.900-0.947] on an independent test set. Individual demographic and ultrasound factors, such as maternal age, vaginal bleeding and gestational sac size, are strong predictors of miscarriage. Previous mathematical models have combined individual risk factors with reasonable performance. A simple scoring system derived from a mathematical model that can be easily implemented in clinical practice has not previously been described for the prediction of ongoing viability. This was a prospective observational study in a single early pregnancy assessment centre during a 9-month period. A cohort of 1881 consecutive women undergoing transvaginal ultrasound scan at a gestational age <84 days were included. Women were excluded if the first trimester outcome was not known. Demographic features, symptoms and ultrasound variables were tested for their influence on ongoing viability. Logistic regression was used to determine the influence on first trimester viability from demographics and symptoms alone, ultrasound findings alone and then from all the variables combined. Each model was developed on a training data set, and a simple scoring system was derived from this. This scoring system was tested on an independent test data set. The final outcome based on a total of 1435 participants was an ongoing viable pregnancy in 885 (61.7%) and early pregnancy loss in 550 (38.3%) women. The scoring system using significant demographic variables alone (maternal age and amount of bleeding) to predict ongoing viability gave an AUC of 0.724 (95% CI = 0.692-0.756) in the training set and 0.729 (95% CI = 0.684-0.774) in the test set. The scoring system using significant ultrasound variables alone (mean gestation sac diameter, mean yolk sac diameter and the presence of fetal heart beat) gave an AUC of 0.873 (95% CI = 0.850-0.897) and 0.900 (95% CI = 0.871-0.928) in the training and the test sets, respectively. The final scoring system using demographic and ultrasound variables together gave an AUC of 0.901 (95% CI = 0.881-0.920) and 0.924 (CI = 0.900-0.947) in the training and the test sets, respectively. After defining the cut-off at which the sensitivity is 0.90 on the training set, this model performed with a sensitivity of 0.92, specificity of 0.73, positive predictive value of 84.7% and negative predictive value of 85.4% in the test set. BMI and smoking variables were a potential omission in the data collection and might further improve the model performance if included. A further limitation is the absence of information on either bleeding or pain in 18% of women. Caution should be exercised before implementation of this scoring system prior to further external validation studies This simple scoring system incorporates readily available data that are routinely collected in clinical practice and does not rely on complex data entry. As such it could, unlike most mathematical models, be easily incorporated into normal early pregnancy care, where women may appreciate an individualized calculation of the likelihood of ongoing pregnancy viability. Research by V.V.B. supported by Research Council KUL: GOA MaNet, PFV/10/002 (OPTEC), several PhD/postdoc & fellow grants; IWT: TBM070706-IOTA3, PhD Grants; IBBT; Belgian Federal Science Policy Office: IUAP P7/(DYSCO, `Dynamical systems, control and optimization', 2012-2017). T.B. is supported by the Imperial Healthcare NHS Trust NIHR Biomedical Research Centre. Not applicable.

Insertion technique and clinical application of regional method axis pedicle screw

To explore the clinical efficacies and outcomes of regional method axis pedicle screw insertion technique. During the period of April 2004 to June 2010, a total of 23 cases with traumatic instability of upper cervical vertebrae were recruited. There were 19 males and 4 females with a mean age of 45.8 years. They underwent surgical operations after an excellent traction reduction of cervical vertebrae. The entry points were drawn on axial facet joint and all of them distributed in the region of upper inner 1/4 of lower articular process. So the regional method was employed to determine the entry point. All subjects underwent the reconstruction of posterior stability. Axial pedicle screws were inserted by the insertion technique of axial pedicle screw via the "regional method". The entry region was in the upper inner 1/4 area of lower articular process. The entry angle, medial inclination and superior inclination were determined by the direction of inner wall and upper wall of isthmus. Postoperative cervical radiography and CT examination were performed to confirm the screw position. Forty-six axial pedicle screws were implanted. No significant complications occurred. All screws stayed in excellent positions without the invasion of vertebral artery and spinal canal. The "regional method" insertion technique of axial pedicle screw require no memory of complex entry points and entry angle parameters. And there is no need of identifying the anatomical landmarks. Thus this approach is accurate, safe and suitable for most patients.

Asymmetric Synthesis of Allenyl Oxindoles and Spirooxindoles by a Catalytic Enantioselective Saucy–Marbet Claisen Rearrangement

Asymmetric Synthesis of Allenyl Oxindoles and Spirooxindoles by a Catalytic Enantioselective Saucy–Marbet Claisen Rearrangement

Design and Characterization of Stabilized Derivatives of Human CD4D12 and CD4D1

CD4 is present on the surface of T-lymphocytes and is the primary cellular receptor for HIV-1. CD4 consists of a cytoplasmic tail, one transmembrane region, and four extracellular domains, D1-D4. A construct consisting of the first two domains of CD4 (CD4D12) is folded and binds gp120 with similar affinity as soluble 4-domain CD4 (sCD4). However, the first domain alone (CD4D1) was previously shown to be largely unfolded and had 3-fold weaker affinity for gp120 when compared to sCD4 [Sharma, D.; et al. (2005) Biochemistry 44, 16192-16202]. We now report the design and characterization of three single-site mutants of CD4D12 (G6A, L51I, and V86L) and one multisite mutant of CD4D1 (G6A/L51I/L5K/F98T). G6A, L51I, and V86L are cavity-filling mutations while L5K and F98T are surface mutations which were introduced to minimize the aggregation of CD4D1 upon removal of the second domain. Two mutations, G6A and V86L in CD4D12 increased the stability and yield of the protein relative to the wild-type protein. The mutant CD4D1 (CD4D1a) with the 4 mutations was folded and more stable compared to the original CD4D1, but both bound gp120 with comparable affinity. In in vitro neutralization assays, both CD4D1a and G6A-CD4D12 were able to neutralize diverse HIV-1 viruses with similar IC(50)s as 4-domain CD4. These stabilized derivatives of human CD4 can be useful starting points for the design of other more complex viral entry inhibitors.