Skeletal Transformations of Terpenoid Forskolin Employing an Oxidative Rearrangement Strategy.

The skeletal transformations of diterpenoid forskolin were achieved by employing an oxidative rearrangement strategy. A library of 36 forskolin analogues with structural diversity was effectively generated. Computational analysis shows that 12 CTD compounds with unique scaffolds and ring systems were produced during the course of this work.

Identification of Piperazinyl-Difluoro-indene Derivatives Containing Pyridyl Groups as Potent FGFR Inhibitors against FGFR Mutant Tumor: Design, Synthesis, and Biological Evaluation.

The fibroblast growth factor receptor (FGFR) signaling pathway plays important roles in cellular processes such as proliferation, differentiation, and migration. In this study, we highlighted the potential of FGFR inhibitors bearing the (S)-3,3-difluoro-1-(4-methylpiperazin-1-yl)-2,3-dihydro-1H-indene scaffold containing a crucial 3-pyridyl group for the treatment of FGFR mutant cancers. The representative compound (S)-23, which was identified through comprehensive evaluation, exhibited potent antiproliferative activity with GI50 in the range of 6.4-10.4 nM against FGFR1 fusion protein-carrying, FGFR2-amplified, and FGFR2 mutant cancer cell lines and good antiproliferative activity against FGFR3 translocation and mutant FGFR4 cancer cell lines, as well as potency assessment against FGFR1-4 kinases. Moreover, compound (S)-23 exhibited favorable pharmacokinetic properties, low potential for drug-drug interactions, and very potent antitumor activity in MFE-296 xenograft mouse models with a TGI of 99.1% at the dose of 10 mg/kg. These findings demonstrate that compound (S)-23 is a potential therapeutic agent for FGFR mutant tumors.

Automatic Stub Avoidance for a Powered Prosthetic Leg over Stairs and Obstacles.

Passive prosthetic legs require undesirable compensations from amputee users to avoid stubbing obstacles and stairsteps. Powered prostheses can reduce those compensations by restoring normative joint biomechanics, but the absence of user proprioception and volitional control combined with the absence of environmental awareness by the prosthesis increases the risk of collisions. This paper presents a novel stub avoidance controller that automatically adjusts prosthetic knee/ankle kinematics based on suprasensory measurements of environmental distance from a small, lightweight, low-power, low-cost ultrasonic sensor mounted above the prosthetic ankle. In a case study with two transfemoral amputee participants, this control method reduced the stub rate during stair ascent by 89.95% and demonstrated an 87.5% avoidance rate for crossing different obstacles on level ground. No thigh kinematic compensation was required to achieve these results. These findings demonstrate a practical perception solution for powered prostheses to avoid collisions with stairs and obstacles while restoring normative biomechanics during daily activities.

Controlling Powered Prosthesis Kinematics over Continuous Transitions Between Walk and Stair Ascent.

One of the primary benefits of emerging powered prosthetic legs is their ability to facilitate step-over-step stair ascent by providing positive mechanical work. Existing control methods typically have distinct steady-state activity modes for walking and stair ascent, where activity transitions involve discretely switching between controllers and often must be initiated with a particular leg. However, these discrete transitions do not necessarily replicate able-bodied joint biomechanics, which have been shown to continuously adjust over a transition stride. This paper presents a phase-based kinematic controller for a powered knee-ankle prosthesis that enables continuous, biomimetic transitions between walking and stair ascent. The controller tracks joint angles from a data-driven kinematic model that continuously interpolates between the steady-state kinematic models, and it allows both the prosthetic and intact leg to lead the transitions. Results from experiments with two transfemoral amputee participants indicate that knee and ankle kinematics smoothly transition between walking and stair ascent, with comparable or lower root mean square errors compared to variations from able-bodied data.

Improving Amputee Endurance over Activities of Daily Living with a Robotic Knee-Ankle Prosthesis: A Case Study.

Robotic knee-ankle prostheses have often fallen short relative to passive microprocessor prostheses in time-based clinical outcome tests. User ambulation endurance is an alternative clinical outcome metric that may better highlight the benefits of robotic prostheses. However, previous studies were unable to show endurance benefits due to inaccurate high-level classification, discretized mid-level control, and insufficiently difficult ambulation tasks. In this case study, we present a phase-based mid-level prosthesis controller which yields biomimetic joint kinematics and kinetics that adjust to suit a continuum of tasks. We enrolled an individual with an above-knee amputation and challenged him to perform repeated, rapid laps of a circuit comprising activities of daily living with both his passive prosthesis and a robotic prosthesis. The participant demonstrated improved endurance with the robotic prosthesis and our mid-level controller compared to his passive prosthesis, completing over twice as many total laps before fatigue and muscle discomfort required him to stop. We also show that time-based outcome metrics fail to capture this endurance improvement, suggesting that alternative metrics related to endurance and fatigue may better highlight the clinical benefits of robotic prostheses.

Design, synthesis and biological evaluation of 3-substituted-2-thioxothiazolidin-4-one (rhodanine) derivatives as antitubercular agents against Mycobacterium tuberculosis protein tyrosine phosphatase B.

Mycobacterium tuberculosis infections still pose a serious threat to human health. Combination therapies are effective medical solutions to the problem. Mycobacterium tuberculosis is an intracellular pathogen that mainly depends on a virulence factor (Mycobacterium tuberculosis protein tyrosine phosphatase B, MptpB) for its survival in the host. Therefore, MptpB inhibitors are potential components of tuberculosis combination treatments. Herein, a new series of MptpB inhibitors bearing a rhodanine group were developed using a structure-based strategy based on the virtual screening hit. The new MptpB inhibitors displayed potent MptpB inhibitory activities and great improvements in cell membrane permeability. The optimal compounds reduced the bacterial burden in a dose-dependent manner in a macrophage infection model, especially, a combination of compound 20 and rifampicin led to a bacterial burden reduction of more than 95%, greater than the reductions achieved with compound 20 or rifampicin alone. This research provides new insights into the rational design of new MptpB inhibitors and verifies that the MptpB inhibitor has a promising potential as a component of tuberculosis treatment.

Synthesis of intramolecular cross-coupling analogues of forskolin.

A ring distortion strategy was applied to the synthesis of a series of intramolecular cross-coupled analogues of forskolin 1. Treatment with palladium acetate, forskolin underwent an intramolecular cross-coupling reaction to generate a novel cycloalkene ether 2 in 85% yield. Under the same conditions, a series of forskolin ester analogues 4a-4d were prepared from 1-OH ester derivatives of forskolin 3a-3d in 85-93% yields. Treating cycloalkene ether 2 with aryl iodides in the presence of a palladium catalyst afforded Z-isomers arylation products 5a-5e in a stereoselective manner in 70-85% yields.

Discovery of biphenyls bearing thiobarbiturate fragment by structure-based strategy as Mycobacterium tuberculosis protein tyrosine phosphatase B inhibitors.

Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) is an important virulence factor that blocks the host immune response and facilitates M. tuberculosis growth in host cells. MptpB inhibitors are potential components of tuberculosis combination treatment. Herein, we present the development of new biphenyls MptpB inhibitors with greatly improved MptpB inhibition based on our reported thiobarbiturate lead 6 by rational design with the structure-based strategy. The eight biphenyls bearing thiobarbiturate fragment target compounds showed more potent MptpB inhibition (IC50: 1.18-14.13 µM) than the lead compound 6. Further molecular docking studies showed that compounds 13, 26, 27 and 28 had multiple interactions with active sites. Among them, compound 13 exhibited dose-dependent increased antituberculosis activity in mouse macrophages. The results displayed that the strategy of modification utilizing biphenyl scaffold was efficient. Our study identifies biphenyls bearing thiobarbiturate fragment as new MptpB inhibitors and verifies the therapeutic potential of antimycobacterial agent targeting MptpB.

Cigarette smoke extract combined with LPS reduces ABCA3 expression in chronic pulmonary inflammation may be related to PPARγ/ P38 MAPK signaling pathway.

ABCA3 (ATP-binding cassette class A3) is a transmembrane transporter that plays a positive role in chronic pulmonary inflammation by regulating lipid metabolism. However, it is not completely clear whether ABCA3 and its signaling factors are involved in chronic pulmonary inflammation induced by the combination of CSE (cigarette smoke extract) and LPS (lipopolysaccharide). In this study, we used the method of combining CSE and LPS which was widely used to study lung inflammation-related diseases and has been proven effective in our group's studies to create in vivo and in vitro pulmonary inflammation models. The result showed that, after CSE in combination with LPS treatment, ABCA3 expression was downregulated in rat lung in vivo and in a human alveolar cell line in vitro. ABCA3 expression was upregulated, and related inflammatory factors were downregulated in the state of overexpression of PPARγ or inhibition of the p38 MAPK pathway, while PPARγ deletion or MAPK14 overexpression showed the opposite results. The level of PPARγ remained unchanged, and the expression of ABCA3 was upregulated in the state of the p38 MAPK pathway was inhibited under overexpression of PPARγ. These results indicate that CSE combined with LPS can result in downregulation of ABCA3 under conditions of inflammation, and that the p38 MAPK signaling pathway mediated by PPARγ can regulate the expression changes of ABCA3, thus providing new targets for treating chronic pulmonary inflammation.

Modeling the Transitional Kinematics Between Variable-Incline Walking and Stair Climbing.

Although emerging powered prostheses can enable people with lower-limb amputation to walk and climb stairs over different task conditions (e.g., speeds and inclines), the control architecture typically uses a finite-state machine to switch between activity-specific controllers. Because these controllers focus on steady-state locomotion, powered prostheses abruptly switch between controllers during gait transitions rather than continuously adjusting leg biomechanics in synchrony with the users. This paper introduces a new framework for powered prosthesis control by modeling the lower-limb joint kinematics over a continuum of variable-incline walking and stair climbing, including steady-state and transitional gaits. Steady-state models for walking and stair climbing represent joint kinematics as continuous functions of gait phase, forward speed, and incline. Transition models interpolate kinematics as convex combinations of the two steady-state models, with an additional term to account for kinematics that fall outside their convex hull. The coefficients of this convex combination denote the similarity of the transitional kinematics to each steady-state mode, providing insight into how able-bodied individuals continuously transition between ambulation modes. Cross-validation demonstrates that the model predictions of untrained kinematics have errors within the range of physiological variability for all joints. Simulation results demonstrate the model's robustness to incline estimation and mode classification errors.

Recent ring distortion reactions for diversifying complex natural products.

Covering: 2013-2022.Chemical diversification of natural products is an efficient way to generate natural product-like compounds for modern drug discovery programs. Utilizing ring-distortion reactions for diversifying natural products would directly alter the core ring systems of small molecules and lead to the production of structurally complex and diverse compounds for high-throughput screening. We review the ring distortion reactions recently used in complexity-to-diversity (CtD) and pseudo natural products (pseudo-NPs) strategies for diversifying complex natural products. The core ring structures of natural products are altered via ring expansion, ring cleavage, ring edge-fusion, ring spiro-fusion, ring rearrangement, and ring contraction. These reactions can rapidly provide natural product-like collections with properties suitable for a wide variety of biological and medicinal applications. The challenges and limitations of current ring distortion reactions are critically assessed, and avenues for future improvements of this rapidly expanding field are discussed. We also provide a toolbox for chemists for the application of ring distortion reactions to access natural product-like molecules.

Structural insights into the binding of nanobody Rh57 to active RhoA-GTP.

RhoA protein is a small GTPase that acts as a molecular switch. When bound to guanosine triphosphate (GTP), RhoA can activate several key signal pathways. Recently, nanobody Rh57 specific binding with GTP bound active RhoA was discovered and developed as a BRET biosensor without cytotoxicity. To further clarify the nanobody Rh57's mechanism of action, we co-expressed, purified, and crystallized the RhoA-Rh57 nanobody complex and solved the structure by X-ray diffraction with a resolution of 2.76 Å. The structure showed that the interaction is mainly through hydrogen bonds, salt bridges, aromatic-aromatic interactions, and hydrophobic interactions. The involved regions include CDR3 and non-hypervariable loop of Rh57, and the SWI switch loops of RhoA, respectively. The different SWI conformation of inactivated RhoA-GDP prevented the Rh57's binding. The possible explanation of Rh57 as a non-cytotoxic BRET intracellular tracer is that Rh57's binding did not overlap with downstream PRK1 and thus did not interfere with the downstream signaling pathway. Our research provides an in-depth understanding of how nanobodies recognize activated RhoA-GTP while not binding inactivated RhoA-GDP. This structural information may also provide critical information for further optimization of relevant nanobodies.

Structural insights into the binding of nanobodies LaM2 and LaM4 to the red fluorescent protein mCherry.

Red fluorescent proteins (RFPs) are powerful tools used in molecular biology research. Although RFP can be easily monitored in vivo, manipulation of RFP by suitable nanobodies binding to different epitopes of RFP is still desired. Thus, it is crucial to obtain structural information on how the different nanobodies interact with RFP. Here, we determined the crystal structures of the LaM2-mCherry and LaM4-mCherry complexes at 1.4 and 1.9 Å resolution. Our results showed that LaM2 binds to the side of the mCherry ß-barrel, while LaM4 binds to the bottom of the ß-barrel. The distinct binding sites of LaM2 and LaM4 were further verified by isothermal titration calorimetry, fluorescence-based size exclusion chromatography, and dynamic light scattering assays. Mutation of the residues at the LaM2 or LaM4 binding interface to mCherry significantly decreased the binding affinity of the nanobody to mCherry. Our results also showed that LaM2 and LaM4 can bind to mCherry simultaneously, which is crucial for recruiting multiple operation elements to the RFP. The binding of LaM2 or LaM4 did not significantly change the chromophore environment of mCherry, which is important for fluorescence quantification assays, while several GFP nanobodies significantly altered the fluorescence. Our results provide atomic resolution interaction information on the binding of nanobodies LaM2 and LaM4 with mCherry, which is important for developing detection and manipulation methods for RFP-based biotechnology.

Real-Time Activity Recognition With Instantaneous Characteristic Features of Thigh Kinematics.

Current supervised learning or deep learning-based activity recognition classifiers can achieve high accuracy in recognizing locomotion activities. Most available techniques use a high-dimensional space of features, e.g., combinations of EMG, kinematics and kinetics, and transformations over those signals. The associated classification rules are therefore complex; the machine tries to understand the human, but the human does not understand the machine. This paper presents an activity recognition system that uses signals from a thigh-mounted IMU and a force sensitive resistor to classify transitions between sitting, walking, stair ascending, and stair descending. The system uses the thigh's orientation and velocity with foot contact information at specific moments within a given activity as the features to classify transitions to other activities. We call these Instantaneous Characteristic Features (ICFs). Because these ICFs are biomechanically intuitive, they are easy for the user to understand and thus control the activity transitions of wearable robots. We assessed our classification algorithm offline using an existing dataset with 10 able-bodied subjects and online with another 10 able-bodied subjects wearing a real-time system. The offline study analyzed the effect of subject-dependency and ramp inclinations. The real-time classification accuracy was evaluated before and after training the subjects on the ICFs. The real-time system achieved overall pre-subject-training and post-subject-training error rates of 0.59% ± 0.24% and 0.56% ± 0.20%, respectively. We also evaluated the feasibility of our ICFs for amputee ambulation by analyzing a public dataset with the open-source bionic leg. The simplicity of these classification rules demonstrates a new paradigm for activity recognition where the human can understand the machine and vice-versa.

Structural insights into two distinct nanobodies recognizing the same epitope of green fluorescent protein.

Green fluorescent protein (GFP) and its derivatives are widely used in biomedical research, and the manipulation of GFP-tagged proteins by GFP-specific binders is highly desired. However, structural information on how these binders bind with GFP is still lacking. In this study, we determined the crystal structure of the nanobody Nb2 complexed with superfolder GFP (sfGFP) at a resolution of 2.2 Å. Interestingly, although the complementarity-determining regions (CDRs) of Nb2 and LaG16 sequences were only 29.7% identical, they both bound to the same epitope of GFP and existed in the same orientation. Structural analysis indicated that they achieved similar binding characteristics through different mechanisms. We further verified the kinetics and thermodynamics of binding by biolayer interferometry (BLI) and isothermal titration calorimetry (ITC). Nb2 showed a slightly higher binding affinity for sfGFP than LaG16. The stability of GFP-specific nanobodies was verified by nano differential scanning fluorimetry (nanoDSF). Nb2 exhibited the highest melting temperature (Tm); thus, Nb2 is a promising GFP nanobody candidate for use in applications requiring harsh testing conditions. We also compared the binding sites of available GFP nanobodies and showed that some of them can simultaneously bind with GFP and assemble into multifunctional complexes to manipulate GFP-tagged target proteins. Our results provide atomic-scale binding information for Nb2-sfGFP, which is important for the further development of GFP-nanobody based fusion protein manipulation techniques.

Efficient and stereoselective one-pot synthesis of benzo[b]oxazolo[3,4-d][1,4]oxazin-1-ones.

An efficient and mild one-pot convergent synthesis protocol has been developed for benzo[b]oxazolo[3,4-d][1,4]oxazin-1-one derivatives through the Mitsunobu reaction and sequential cyclization. Various tricyclic fused benzoxazinyl-oxazolidinones (20 examples) were obtained in good to excellent yields and high enantioselectivities with facile operation. Furthermore, four stereoisomers were afforded respectively in high ee values (>97.8%) via using different chiral 2,3-epoxy-4-trityloxybutanol. This methodology has been applied to the synthesis of key intermediates of drug candidates.

Chlorambucil-conjugated platinum(IV) prodrugs to treat triple-negative breast cancer in vitro and in vivo.

Modification of platinum (II) into lipophilic platinum (IV) compounds by introducing biologically active molecules were widely employed to develop new platinum-based prodrugs in the past decade. In this paper, two chlorambucil platinum (IV) complexes, CLB-Pt and CLB-Pt-CLB, were synthesized and displayed very potent antiproliferative activity against all the tested cancer cell lines, such as A549, HeLa and MCF-7, especially to treat the well-known refractory triple-negative breast cancer. CLB-Pt-CLB significantly improved cell-killing effect in triple-negative subtype MDA-MB-231 cells, and showed much stronger cytotoxicity than either monotherapy or combination of cisplatin and chlorambucil. CLB-Pt-CLB prodrug entered cells in dramatically increased amount compared with cisplatin and enhanced DNA damage, inducing cancer cell apoptosis. It exhibited high anticancer activity and no observable toxicity in BALB/c nude mice bearing MDA-MB-231 tumors. The chlorambucil moiety not only greatly assisted the passive diffusion of CLB-Pt-CLB into cells, but also produced the synergism with cisplatin in targeting DNA.