Dominant negative variants in IKZF2 cause ICHAD syndrome, a new disorder characterised by immunodysregulation, craniofacial anomalies, hearing impairment, athelia and developmental delay.

BACKGROUND: Helios (encoded by IKZF2), a member of the Ikaros family of transcription factors, is a zinc finger protein involved in embryogenesis and immune function. Although predominantly recognised for its role in the development and function of T lymphocytes, particularly the CD4+ regulatory T cells (Tregs), the expression and function of Helios extends beyond the immune system. During embryogenesis, Helios is expressed in a wide range of tissues, making genetic variants that disrupt the function of Helios strong candidates for causing widespread immune-related and developmental abnormalities in humans. METHODS: We performed detailed phenotypic, genomic and functional investigations on two unrelated individuals with a phenotype of immune dysregulation combined with syndromic features including craniofacial differences, sensorineural hearing loss and congenital abnormalities. RESULTS: Genome sequencing revealed de novo heterozygous variants that alter the critical DNA-binding zinc fingers (ZFs) of Helios. Proband 1 had a tandem duplication of ZFs 2 and 3 in the DNA-binding domain of Helios (p.Gly136_Ser191dup) and Proband 2 had a missense variant impacting one of the key residues for specific base recognition and DNA interaction in ZF2 of Helios (p.Gly153Arg). Functional studies confirmed that both these variant proteins are expressed and that they interfere with the ability of the wild-type Helios protein to perform its canonical function-repressing IL2 transcription activity-in a dominant negative manner. CONCLUSION: This study is the first to describe dominant negative IKZF2 variants. These variants cause a novel genetic syndrome characterised by immunodysregulation, craniofacial anomalies, hearing impairment, athelia and developmental delay.

NOTCH1 loss of the TAD and PEST domain: An antimorph?

The Notch proteins play key roles in cell fate determination during development. Germline pathogenic variants in NOTCH1 predispose to a spectrum of cardiovascular malformations including Adams-Oliver syndrome and a wide variety of isolated complex and simple congenital heart defects. The intracellular C-terminus of the single-pass transmembrane receptor encoded by NOTCH1 contains a transcriptional activating domain (TAD) required for target gene activation and a PEST domain (a sequence rich in proline, glutamic acid, serine, and threonine), regulating protein stability and turnover. We present a patient with a novel variant encoding a truncated NOTCH1 protein without the TAD and PEST domain (NM_017617.4: c.[6626_6629del];[=], p.(Tyr2209CysfsTer38)) and extensive cardiovascular abnormalities consistent with a NOTCH1-mediated mechanism. This variant fails to promote transcription of target genes as assessed by luciferase reporter assay. Given the roles of the TAD and PEST domains in NOTCH1 function and regulation, we hypothesize that loss of both the TAD and the PEST domain results in a stable, loss-of-function protein that acts as an antimorph through competition with wild-type NOTCH1.

Comprehensive human amniotic fluid metagenomics supports the sterile womb hypothesis.

As metagenomic approaches for detecting infectious agents have improved, each tissue that was once thought to be sterile has been found to harbor a variety of microorganisms. Controversy still exists over the status of amniotic fluid, which is part of an immunologically privileged zone that is required to prevent maternal immune system rejection of the fetus. Due to this privilege, the exclusion of microbes has been proposed to be mandatory, leading to the sterile womb hypothesis. Since nucleic acid yields from amniotic fluid are very low, contaminating nucleic acid found in water, reagents and the laboratory environment frequently confound attempts to address this hypothesis. Here we present metagenomic criteria for microorganism detection and a metagenomic method able to be performed with small volumes of starting material, while controlling for exogenous contamination, to circumvent these and other pitfalls. We use this method to show that human mid-gestational amniotic fluid has no detectable virome or microbiome, supporting the sterile womb hypothesis.

Design, synthesis, and biological evaluation of diosgenin-indole derivatives as dual-functional agents for the treatment of Alzheimer's disease.

The complex pathogenesis of Alzheimer's disease (AD) has become a major obstacle in its treatment. An effective approach is to develop multifunctional agents that simultaneously target multiple pathological processes. Here, a series of diosgenin-indole compounds were designed, synthesized and evaluated for their neuroprotective effects against H2O2 (hydrogen peroxide), 6-OHDA (6-hydroxydopamine) and Aß (beta amyloid) damages. Preliminary structure-activities relationship revealed that the introduction of indole fragment and electron-donating group at C-5 on ring indole could be beneficial for neuroprotective activities. Results indicated that compound 5b was the most promising candidate against cellular damage induced by H2O2 (52.9 ± 1.9%), 6-OHDA (38.4 ± 2.4%) and Aß1-42 (54.4 ± 2.7%). Molecular docking study suggested the affinity for 5b bound to Aß1-42 was -40.59 kcal/mol, which revealed the strong binding affinity of 5b to Aß1-42. The predicted values of brain/blood partition coefficient (-0.733) and polar surface area (85.118 Å2) indicated the favorable abilities of BBB permeation and absorption of 5b. In addition, 5b significantly decreased ROS (reactive oxygen species) production induced by H2O2. In the following in vivo experiment, 5b obviously attenuated memory and learning impairments of Aß-injected mice. In summary, compound 5b could be considered as a promising dual-functional neuroprotective agent against AD.

Twin drug design, synthesis and evaluation of diosgenin derivatives as multitargeted agents for the treatment of vascular dementia.

A novel series of multitargeted molecules were designed and synthesized by combining the pharmacological role of cholinesterase inhibitor and antioxidant of steroid as potential ligands for the treatment of Vascular Dementia (VD). The oxygen-glucose deprivation (OGD) model was used to evaluate these molecules, among which the most potent compound ML5 showed the highest activity. Firstly, ML5 showed appropriate inhibition of cholinesterases (ChEs) at orally 15 mg/kg in vivo. The further test revealed that ML5 promoted the nuclear translocation of Nrf2. Furthermore, ML5 has significant neuroprotective effect in vivo model of bilateral common carotid artery occlusion (BCCAO), significantly increasing the expression of Nrf2 protein in the cerebral cortex. In the molecular docking research, we predicted the ML5 combined with hAChE and Keap1. Finally, compound ML5 displayed normal oral absorption and it was nontoxic at 500 mg/kg, po, dose. We can draw the conclusion that ML5 could be considered as a new potential compound for VD treatment.

Design, synthesis and evaluation of diosgenin carbamate derivatives as multitarget anti-Alzheimer's disease agents.

In order to produce an effective and multi-targeted clinical drug that could prevent progressive neurodegeneration, a series of diosgenin carbamate derivatives were designed, synthesized and tested for their anti-inflammatory, antioxidant and anti-Aß activities. The results demonstrated that compound M15 was the most promising derivative against inflammatory (NO inhibition 22.7 ± 2.2%,10 µM) and cellular damage induced by H2O2 (SH-SY5Y cell protection = 75.3 ± 3.4%, 10 µM) or Aß (astrocytes protection = 70.2 ± 6.5%, 10 µM). Molecular docking studies revealed the strong binding affinity of M15 to the active site of nNOS, Aß42 and pro-inflammatory proteins. Western blot demonstrated that M15 decreased IL-1ß, IL-6 and TNF-α level, which may contribute to its anti-inflammatory effects. In addition, M15 maintained mitochondrial function as well as cell viability through reducing H2O2-induced ROS production. The results indicated that oral administration of M15 attenuated memory deficits and played a neuroprotective effect on subcutaneous (s.c.) D-gal aging mice. In summary, M15 could be considered as a potential multifunctional neuroprotective agent due to the effects of anti-inflammatory, antioxidant and anti-Aß activities.

Synthesis and biological evaluation of 3-carbamate smilagenin derivatives as potential neuroprotective agents.

Studies indicated that smilagenin, isolated from Anemarrhena asphodeloides Bunge, could improve cognitive impairment and exhibit neuroprotective activity. On the basis of the structure of smilagenin, a series of derivatives were synthesized and evaluated for their neuroprotective effects of H2O2-induced, oxygen glucose deprivation-induced neurotoxicity in SH-SY5Y cells and LPS-induced NO production in RAW264.7 cells. Structure activity relationship of derivatives revealed that benzyl-substituted piperazine formate derivatives showed the potent neuroprotective activity such as A12. These findings may provide new insights for the development of neuroprotective agents against Alzheimer's disease.

Design, synthesis and biological evaluation of 3-piperazinecarboxylate sarsasapogenin derivatives as potential multifunctional anti-Alzheimer agents.

A series of multifunctional 3-piperazinecarboxylate sarsasapogenin derivatives were designed and synthesized against Alzheimer's disease (AD). The protection against H2O2-triggered oxidative stress in PC12 cells, and inhibition on LPS-induced NO production in RAW264.7 cell lines in vitro by these derivatives were firstly evaluated. Most of the compounds showed better antioxidant and antiinflammatory activities compared with sarsasapogenin, especially AA34 and AA36. Structure-activity relationships revealed that benzyl group, electron-donating group and intramolecular hydrogen bond might be beneficial to enhancing their neuroprotective activities. Moreover, Aß42 was the optimum predicted target based on the high 3D molecular similarity between compound AA36 and caprospinol. In the following experiments, AA36 significantly protected PC12 cells from Aß-induced damage and improved learning and memory impairments in Aß-injected mice. Thus AA36 is regarded as a potent anti-AD agent and N-substituted piperazinecarboxylate can be served as a promising structural unit for anti-AD drug design.

[Differential diagnosis of hepatocellular carcinoma and hepatic hemangiomas based on radiomic features of gadoxetate disodium-enhanced magnetic resonance imaging].

OBJECTIVE: To evaluate the feasibility of using radiomic features for differential diagnosis of hepatocellular carcinoma (HCC) and hepatic cavernous hemangioma (HHE). METHODS: Gadoxetate disodium-enhanced magnetic resonance imaging data were collected from a total of 135 HCC and HHE lesions. The radiomic texture features of each lesion were extracted on the hepatobiliary phase images, and the performance of each feature was assessed in differentiation and classification of HCC and HHE. In multivariate analysis, the performance of 3 feature selection algorithms (namely minimum redundancy-maximum relevance, mRmR; neighborhood component analysis, NCA; and sequence forward selection, SFS) was compared. The optimal feature subset was determined according to the optimal feature selection algorithm and used for testing the 3 classifier algorithms (namely the support vector machine, RBF-SVM; linear discriminant analysis, LDA; and logistic regression). All the tests were repeated 5 times with 10-fold cross validation experiments. RESULTS: More than 50% of the radiomic features exhibited strong distinguishing ability, among which gray level co-occurrence matrix feature S (3, -3) SumEntrp showed a good classification performance with an AUC of 0.72 (P<0.01), a sensitivity of 0.83 and a specificity of 0.57. For the multivariate analysis, 15 features were selected based on the SFS algorithm, which produced better results than the other two algorithms. Testing of these 15 selected features for their average cross-validation performance with RBF-SVM classifier yielded a test accuracy of 0.82∓0.09, an AUC of 0.86∓0.12, a sensitivity of 0.88∓0.11, and a specificity of 0.76∓0.18. CONCLUSION: The radiomic features based on gadoxetate disodium-enhanced magnetic resonance images allow efficient differential diagnosis of HCC and HHE, and can potentially provide important assistance in clinical diagnosis of the two diseases.

Alleviation of cancerous pain by external compress with Xiaozheng Zhitong Paste.

OBJECTIVE: To observe the clinical effectiveness of a topical application of Xiaozheng Zhitong: Paste (, XZP) in alleviating the cancerous pain of patients with middle/late stage cancer METHODS: By: adopting a random number table, 124 patients enrolled were randomized into the treatment group (64 patients) and the control group (60 patients). In addition to the basic therapy [including the three-ladder (3L) analgesia] used in both groups, topical application of XZP was given to patients in the treatment group for pain alleviation. The analgesic efficacy was recorded in terms of pain intensity, analgesia initiating time and sustaining time, and the optimal analgesic effect revealing time. Meanwhile, the quality of life (QOL) and adverse reactions that occurred in patients were recorded as well. RESULTS: The total effective rate in the treatment group was: 84.38% (54/64), and in the control group it was 88.33% (53/60), showing no significant difference between them (P>0.05), but the analgesia initiating time and the optimal analgesia effect revealing time in the treatment group were significantly shorter (both P<0.01). Moreover, XZP was better in improving patients' QOL, showing more significant improvements in the treatment group than those in the control group in aspects of mental condition, walking capacity, working capacity, social acceptability, sleep and joy of living (P<0.05 or P<0.01). Lower incidence of adverse reactions, such as nausea, vomiting, mouth dryness, dizziness, etc., especially constipation, was noted in the treatment group (P<0.05 or P<0.01). CONCLUSION: Applying an external compress: of XZP showed a synergistic action with 3L analgesia for shortening the initiating time and the optimal effect revealing time, and could evidently enhance patients' QOL with fewer adverse reactions.

Detection and characterisation of CTX-M and CMY-2 beta-lactamases among Escherichia coli isolates from farm animals in Guangdong Province of China.

The objective of this study was to characterise the beta-lactamase genes of cephalosporin-resistant Escherichia coli isolated from farm animals in Guangdong Province of China. Of 592 E. coli isolates recovered from farm animals from 2003-2005, 50 (8.4%) showed cephalosporin resistance. Polymerase chain reaction and sequencing analysis showed that 14 isolates (2.4%) from chickens, ducks, pigs and partridges were positive for the bla(CTX-M) gene (10 for bla(CTX-M-14) and 4 for bla(CTX-M-27)). CMY-2 was detected for the first time in mainland China in six E. coli isolates (1.0%) from chickens and goose. Except for one isolate, bla(CTX-M)- and bla(CMY-2)-containing isolates also harboured the bla(TEM-1b) gene. Conjugation experiments demonstrated that the bla(CTX-M) and bla(TEM) genes could be transferred to E. coli DH5alpha. Pulsed-field gel electrophoresis (PFGE) showed that the 14 CTX-M-producing isolates belonged to 12 different types. Two isolates (one from a chicken, the other from a pig) containing CTX-M-14 showed indistinguishable PFGE patterns, indicating clonal dissemination of this strain among animals from different farms. This study describes for the first time the emergence of CTX-M- and CMY-2-producing E. coli among farm animals in China, with the CTX-M-9 group being the predominant extended-spectrum beta-lactamase detected.