Study on the therapeutic effect of Kirschner wire tension band combined with anchor cross-stitch technique in the treatment of comminuted patellar inferopolar fractures.

PURPOSES: Fractures of the inferior patellar pole, unlike other patellar fractures, present challenges for traditional surgical fixation methods. This article introduces the clinical technique and outcomes of using Kirschner wire tension band combined with anchor screw cross-stitch fixation for comminuted inferior patellar pole fractures. METHODS: This retrospective case series study included 14 patients with comminuted inferior patellar pole fractures treated at our institution from September 1, 2020, to April 30, 2022. All patients underwent surgery using the Kirschner wire tension band with anchor screw cross-stitch technique. Follow-up assessments involved postoperative X-rays to evaluate fracture healing, as well as clinical parameters such as healing time, Visual Analog Scale (VAS) scores, range of motion (ROM), and Bostman scores. RESULTS: All patients were followed for an average of over 12 months, with no cases of internal fixation failure. Knee joint stability and function were excellent. X-rays revealed an average healing time of approximately 10.79 ± 1.53 weeks, hospitalization lasted 5.64 ± 1.15 days, surgery took approximately 37.86 ± 5.32 minutes, and intraoperative blood loss was 33.29 ± 8.15 ml. One patient experienced irritation from the internal fixation material. At the final follow-up, the Bostman score averaged 28.29 ± 0.83, knee joint flexion reached 131.07° ± 4.88°, all patients achieved full knee extension, and the VAS score was 0.36 ± 0.63. CONCLUSION: Kirschner wire tension band with anchor screw cross-stitch fixation for comminuted inferior patellar pole fractures delivered satisfactory clinical outcomes. This surgical method, characterized by its simplicity and reliability, is a valuable addition to clinical practice.

Copper-catalyzed atroposelective synthesis of C-O axially chiral compounds enabled by chiral 1,8-naphthyridine based ligands.

Axially chiral molecular scaffolds are widely present in therapeutic agents, natural products, catalysts, and advanced materials. The construction of such molecules has garnered significant attention from academia and industry. The catalytic asymmetric synthesis of axially chiral biaryls, along with other non-biaryl axially chiral molecules, has been extensively explored in the past decade. However, the atroposelective synthesis of C-O axial chirality remains largely underdeveloped. Herein, we document a copper-catalyzed atroposelective construction of C-O axially chiral compounds using novel 1,8-naphthyridine-based chiral ligands. Mechanistic investigations have provided good evidence in support of a mechanism involving synergistic interplay between a desymmetrization reaction and kinetic resolution process. The method described in this study holds great significance for the atroposelective synthesis of C-O axially chiral compounds, with promising applications in organic chemistry. The utilization of 1,8-naphthyridine-based ligands in copper catalysis is anticipated to find broad applications in asymmetric copper(i)-catalyzed azide-alkyne cycloadditions (CuAACs) and beyond.

Catalytic Asymmetric Construction of α,α-Diaryl Aldehydes via Oxo-Hydroarylation of Terminal Alkynes.

Chiral aldehydes containing a tertiary stereogenic center are versatile building blocks in organic chemistry. In particular, such structural motifs bearing an α,α-diaryl moiety are very challenging scaffolds and their efficient asymmetric synthesis is not reported. In this work, a phosphoric acid-catalyzed enantioselective synthesis of α,α-diaryl aldehydes from simple terminal alkynes is presented. This approach yields a wide range of highly enolizable α,α-diaryl aldehydes in good yields with excellent enantioselectivities. Facile transformations of the products, as well as an efficient synthesis of bioactive molecules, including an effective anti-smallpox agent and an FDA-approved antidepressant drug (+)-sertraline, are demonstrated.

Tertiary folds of the SL5 RNA from the 5' proximal region of SARS-CoV-2 and related coronaviruses.

Coronavirus genomes sequester their start codons within stem-loop 5 (SL5), a structured, 5' genomic RNA element. In most alpha- and betacoronaviruses, the secondary structure of SL5 is predicted to contain a four-way junction of helical stems, some of which are capped with UUYYGU hexaloops. Here, using cryogenic electron microscopy (cryo-EM) and computational modeling with biochemically determined secondary structures, we present three-dimensional structures of SL5 from six coronaviruses. The SL5 domain of betacoronavirus severe-acute-respiratory-syndrome-related coronavirus 2 (SARS-CoV-2), resolved at 4.7 Å resolution, exhibits a T-shaped structure, with its UUYYGU hexaloops at opposing ends of a coaxial stack, the T's "arms." Further analysis of SL5 domains from SARS-CoV-1 and MERS (7.1 and 6.4 to 6.9 Å resolution, respectively) indicate that the junction geometry and inter-hexaloop distances are conserved features across these human-infecting betacoronaviruses. The MERS SL5 domain displays an additional tertiary interaction, which is also observed in the non-human-infecting betacoronavirus BtCoV-HKU5 (5.9 to 8.0 Å resolution). SL5s from human-infecting alphacoronaviruses, HCoV-229E and HCoV-NL63 (6.5 and 8.4 to 9.0 Å resolution, respectively), exhibit the same coaxial stacks, including the UUYYGU-capped arms, but with a phylogenetically distinct crossing angle, an X-shape. As such, all SL5 domains studied herein fold into stable tertiary structures with cross-genus similarities and notable differences, with implications for potential protein-binding modes and therapeutic targets.

DNA adductomics aided rapid screening of genotoxic impurities using nucleosides and 3D bioprinted human liver organoids.

Current genotoxicity assessment methods are mainly employed to verify the genotoxic safety of drugs, but do not allow for rapid screening of specific genotoxic impurities (GTIs). In this study, a new approach for the recognition of GTIs has been proposed. It is to expose the complex samples to an in vitro nucleoside incubation model, and then draw complete DNA adduct profiles to infer the structures of potential genotoxic impurities (PGIs). Subsequently, the genotoxicity is confirmed in human by 3D bioprinted human liver organoids. To verify the feasibility of the approach, lansoprazole chloride compound (Lanchlor), a PGI during the synthesis of lansoprazole, was selected as the model drug. After confirming genotoxicity by Comet assay, it was exposed to different models to map and compare the DNA adduct profiles by LC-MS/MS. The results showed Lanchlor could generate diverse DNA adducts, revealing firstly its genotoxicity at molecular mechanism of action. Furthermore, the largest variety and content of DNA adducts were observed in the nucleoside incubation model, while the human liver organoids exhibited similar results with rats. The results showed that the combination of DNA adductomics and 3D bioprinted organoids were useful for the rapid screening of GTIs.

Tertiary folds of the SL5 RNA from the 5' proximal region of SARS-CoV-2 and related coronaviruses.

Coronavirus genomes sequester their start codons within stem-loop 5 (SL5), a structured, 5' genomic RNA element. In most alpha- and betacoronaviruses, the secondary structure of SL5 is predicted to contain a four-way junction of helical stems, some of which are capped with UUYYGU hexaloops. Here, using cryogenic electron microscopy (cryo-EM) and computational modeling with biochemically-determined secondary structures, we present three-dimensional structures of SL5 from six coronaviruses. The SL5 domain of betacoronavirus SARS-CoV-2, resolved at 4.7 Å resolution, exhibits a T-shaped structure, with its UUYYGU hexaloops at opposing ends of a coaxial stack, the T's "arms." Further analysis of SL5 domains from SARS-CoV-1 and MERS (7.1 and 6.4-6.9 Å resolution, respectively) indicate that the junction geometry and inter-hexaloop distances are conserved features across the studied human-infecting betacoronaviruses. The MERS SL5 domain displays an additional tertiary interaction, which is also observed in the non-human-infecting betacoronavirus BtCoV-HKU5 (5.9-8.0 Å resolution). SL5s from human-infecting alphacoronaviruses, HCoV-229E and HCoV-NL63 (6.5 and 8.4-9.0 Å resolution, respectively), exhibit the same coaxial stacks, including the UUYYGU-capped arms, but with a phylogenetically distinct crossing angle, an X-shape. As such, all SL5 domains studied herein fold into stable tertiary structures with cross-genus similarities, with implications for potential protein-binding modes and therapeutic targets.

Theoretical framework and experimental solution for the air-water interface adsorption problem in cryoEM.

As cryogenic electron microscopy (cryoEM) gains traction in the structural biology community as a method of choice for determining atomic structures of biological complexes, it has been increasingly recognized that many complexes that behave well under conventional negative-stain electron microscopy tend to have preferential orientation, aggregate or simply mysteriously "disappear" on cryoEM grids, but the reasons for such misbehavior are not well understood, limiting systematic approaches to solving the problem. Here, we have developed a theoretical formulation that explains these observations. Our formulation predicts that all particles migrate to the air-water interface (AWI) to lower the total potential surface energy - rationalizing the use of surfactant, which is a direct solution to reducing the surface tension of the aqueous solution. By conducting cryogenic electron tomography (cryoET) with the widely-tested sample, GroEL, we demonstrate that, in a standard buffer solution, nearly all particles migrate to the AWI. Gradual reduction of the surface tension by introducing surfactants decreased the percentage of particles exposed to the surface. By conducting single-particle cryoEM, we confirm that applicable surfactants do not damage the biological complex, thus suggesting that they might offer a practical, simple, and general solution to the problem for high-resolution cryoEM. Application of this solution to a real-world AWI adsorption problem with a more challenging membrane protein, namely, the ClC-1 channel, has led to its first near-atomic structure using cryoEM.

Molecular Editing of Pyrroles via a Skeletal Recasting Strategy.

Heterocyclic scaffolds are commonly found in numerous biologically active molecules, therapeutic agents, and agrochemicals. To probe chemical space around heterocycles, many powerful molecular editing strategies have been devised. Versatile C-H functionalization strategies allow for peripheral modifications of heterocyclic motifs, often being specific and taking place at multiple sites. The past few years have seen the quick emergence of exciting "single-atom skeletal editing" strategies, through one-atom deletion or addition, enabling ring contraction/expansion and structural diversification, as well as scaffold hopping. The construction of heterocycles via deconstruction of simple heterocycles is unknown. Herein, we disclose a new molecular editing method which we name the skeletal recasting strategy. Specifically, by tapping on the 1,3-dipolar property of azoalkenes, we recast simple pyrroles to fully substituted pyrroles, through a simple phosphoric acid-promoted one-pot reaction consisting of dearomative deconstruction and rearomative reconstruction steps. The reaction allows for easy access to synthetically challenging tetra-substituted pyrroles which are otherwise difficult to synthesize. Furthermore, we construct N-N axial chirality on our pyrrole products, as well as accomplish a facile synthesis of the anticancer drug, Sutent. The potential application of this method to other heterocycles has also been demonstrated.

Enantioselective De Novo Synthesis of α,α-Diaryl Ketones from Alkynes.

Chiral α,α-diaryl ketones are structural motifs commonly present in bioactive molecules, and they are also valuable building blocks in synthetic organic chemistry. However, catalytic asymmetric synthesis of α,α-diaryl ketones bearing a tertiary stereogenic center remains largely unsolved. Herein, we report a catalytic de novo enantioselective synthesis of α,α-diaryl ketones from simple alkynes via chiral phosphoric acid (CPA) catalysis. A broad range of enolizable α,α-diaryl ketones are prepared in good yields and with excellent enantioselectivities. The described protocol also serves as an efficient deuteration method for the preparation of enantiomerically enriched deuterated α,α-diaryl ketones. Using the methodology reported, bioactive molecules, including one of the best-selling anti-breast cancer drugs, tamoxifen, are readily synthesized.

Saponin Nanoparticle Adjuvants Incorporating Toll-Like Receptor Agonists Improve Vaccine Immunomodulation.

Over the past few decades, the development of potent and safe immune-activating adjuvant technologies has become the heart of intensive research in the constant fight against highly mutative and immune evasive viruses such as influenza, SARS-CoV-2, and HIV. Herein, we developed a highly modular saponin-based nanoparticle platform incorporating toll-like receptor agonists (TLRas) including TLR1/2a, TLR4a, TLR7/8a adjuvants and their mixtures. These various TLRa-SNP adjuvant constructs induce unique acute cytokine and immune-signaling profiles, leading to specific Th-responses that could be of interest depending on the target disease for prevention. In a murine vaccine study, the adjuvants greatly improved the potency, durability, breadth, and neutralization of both COVID-19 and HIV vaccine candidates, suggesting the potential broad application of these adjuvant constructs to a range of different antigens. Overall, this work demonstrates a modular TLRa-SNP adjuvant platform which could improve the design of vaccines for and dramatically impact modern vaccine development.

Diastereo- and atroposelective synthesis of N-arylpyrroles enabled by light-induced phosphoric acid catalysis.

The C-N axially chiral N-arylpyrrole motifs are privileged scaffolds in numerous biologically active molecules and natural products, as well as in chiral ligands/catalysts. Asymmetric synthesis of N-arylpyrroles, however, is still challenging, and the simultaneous creation of contiguous C-N axial and central chirality remains unknown. Herein, a diastereo- and atroposelective synthesis of N-arylpyrroles enabled by light-induced phosphoric acid catalysis has been developed. The key transformation is a one-pot, three-component oxo-diarylation reaction, which simultaneously creates a C-N axial chirality and a central quaternary stereogenic center. A broad range of unactivated alkynes were readily employed as a reaction partner in this transformation, and the N-arylpyrrole products are obtained in good yields, with excellent enantioselectivities and very good diastereoselectivities. Notably, the N-arylpyrrole skeletons represent interesting structural motifs that could be used as chiral ligands and catalysts in asymmetric catalysis.

Enhancing the Catalytic Activity of Glycolate Oxidase from Chlamydomonas reinhardtii through Semi-Rational Design.

Glycolate oxidase is a peroxisomal flavoprotein catalyzing the oxidation of glycolate to glyoxylate and plays crucial metabolic roles in green algae, plants, and animals. It could serve as a biocatalyst for enzymatic production of glyoxylate, a fine chemical with a wide variety of applications in perfumery, flavor, and the pharmaceutical and agrochemical industries. However, the low catalytic activity of native glycolate oxidase and low levels of active enzyme in heterologous expression limit its practical use in industrial biocatalysis. Herein, the glycolate oxidase from Chlamydomonas reinhardtii (CreGO) was selected through phylogenetic tree analysis, and its low level of soluble expression in E. coli BL21(DE3) was improved through the use of the glutathione thioltransferase (GST), the choice of the vector pET22b and the optimization of induction conditions. The semi-rational design of the fusion enzyme GST-Gly-Ser-Gly-CreGO led to the superior variant GST-Gly-Ser-Gly-CreGO-Y27S/V111G/V212R with the kcat/Km value of 29.2 s-1·mM-1, which was six times higher than that of the wild type. In contrast to GST-Gly-Ser-Gly-CreGO, 5 mg/mL of crude enzyme GST-Gly-Ser-Gly-CreGO-Y27S/V111G/V212R together with 25 µg/mL of catalase catalyzed the oxidation of 300 mM of methyl glycolate for 8 h, increasing the yield from 50.4 to 93.5%.

Multi-Enzymatic Cascade for Efficient Deracemization of dl-Pantolactone into d-Pantolactone.

d-pantolactone is an intermediate in the synthesis of d-pantothenic acid, which is known as vitamin B5. The commercial synthesis of d-pantolactone is carried out through the selective resolution of dl-pantolactone catalyzed by lactone hydrolase. In contrast to a kinetic resolution approach, the deracemization of dl-pantolactone is a simpler, greener, and more sustainable way to obtain d-pantolactone with high optical purity. Herein, an efficient three-enzyme cascade was developed for the deracemization of dl-pantolactone, using l-pantolactone dehydrogenase from Amycolatopsis methanolica (AmeLPLDH), conjugated polyketone reductase from Zygosaccharomyces parabailii (ZpaCPR), and glucose dehydrogenase from Bacillus subtilis (BsGDH). The AmeLPLDH was used to catalyze the dehydrogenated l-pantolactone into ketopantolactone; the ZpaCPR was used to further catalyze the ketopantolactone into d-pantolactone; and glucose dehydrogenase together with glucose fulfilled the function of coenzyme regeneration. All three enzymes were co-expressed in E. coli strain BL21(DE3), which served as the whole-cell biocatalyst. Under optimized conditions, 36 h deracemization of 1.25 M dl-pantolactone d-pantolactone led to an e.e.p value of 98.6%, corresponding to productivity of 107.7 g/(l·d).

Broad and Durable Humoral Responses Following Single Hydrogel Immunization of SARS-CoV-2 Subunit Vaccine.

Most vaccines require several immunizations to induce robust immunity, and indeed, most SARS-CoV-2 vaccines require an initial two-shot regimen followed by several boosters to maintain efficacy. Such a complex series of immunizations unfortunately increases the cost and complexity of populations-scale vaccination and reduces overall compliance and vaccination rate. In a rapidly evolving pandemic affected by the spread of immune-escaping variants, there is an urgent need to develop vaccines capable of providing robust and durable immunity. In this work, a single immunization SARS-CoV-2 subunit vaccine is developed that can rapidly generate potent, broad, and durable humoral immunity. Injectable polymer-nanoparticle (PNP) hydrogels are leveraged as a depot technology for the sustained delivery of a nanoparticle antigen (RND-NP) displaying multiple copies of the SARS-CoV-2 receptor-binding domain (RBD) and potent adjuvants including CpG and 3M-052. Compared to a clinically relevant prime-boost regimen with soluble vaccines formulated with CpG/alum or 3M-052/alum adjuvants, PNP hydrogel vaccines more rapidly generated higher, broader, and more durable antibody responses. Additionally, these single-immunization hydrogel-based vaccines elicit potent and consistent neutralizing responses. Overall, it is shown that PNP hydrogels elicit improved anti-COVID immune responses with only a single administration, demonstrating their potential as critical technologies to enhance overall pandemic readiness.

Subcutaneous delivery of an antibody against SARS-CoV-2 from a supramolecular hydrogel depot.

Prolonged maintenance of therapeutically-relevant levels of broadly neutralizing antibodies (bnAbs) is necessary to enable passive immunization against infectious disease. Unfortunately, protection only lasts for as long as these bnAbs remain present at a sufficiently high concentration in the body. Poor pharmacokinetics and burdensome administration are two challenges that need to be addressed in order to make pre- and post-exposure prophylaxis with bnAbs feasible and effective. In this work, we develop a supramolecular hydrogel as an injectable, subcutaneous depot to encapsulate and deliver antibody drug cargo. This polymer-nanoparticle (PNP) hydrogel exhibits shear-thinning and self-healing properties that are required for an injectable drug delivery vehicle. In vitro drug release assays and diffusion measurements indicate that the PNP hydrogels prevent burst release and slow the release of encapsulated antibodies. Delivery of bnAbs against SARS-CoV-2 from PNP hydrogels is compared to standard routes of administration in a preclinical mouse model. We develop a multi-compartment model to understand the ability of these subcutaneous depot materials to modulate the pharmacokinetics of released antibodies; the model is extrapolated to explore the requirements needed for novel materials to successfully deliver relevant antibody therapeutics with different pharmacokinetic characteristics.

Theoretical framework and experimental solution for the air-water interface adsorption problem in cryoEM.

As cryogenic electron microscopy (cryoEM) gains traction in the structural biology community as a method of choice for determining atomic structures of biological complexes, it has been increasingly recognized that many complexes that behave well under conventional negative-stain electron microscopy tend to have preferential orientation, aggregate or simply mysteriously "disappear" on cryoEM grids. However, the reasons for such misbehavior are not well understood, which limits systematic approaches to solving the problem. Here, we have developed a theoretical formulation that explains these observations. Our formulation predicts that all particles migrate to the air-water interface (AWI) to lower the total potential surface energy-rationalizing the use of surfactant, which is a direct solution to reduce the surface tension of the aqueous solution. By performing cryogenic electron tomography (cryoET) on the widely-tested sample, GroEL, we demonstrate that, in a standard buffer solution, nearly all particles migrate to the AWI. Gradually reducing the surface tension by introducing surfactants decreased the percentage of particles exposed to the surface. By conducting single-particle cryoEM, we confirm that suitable surfactants do not damage the biological complex, thus suggesting that they might provide a practical, simple, and general solution to the problem for high-resolution cryoEM. Applying this solution to a real-world AWI adsorption problem involving a more challenging membrane protein, namely, the ClC-1 channel, has resulted in its near-atomic structure determination using cryoEM.

Phosphorylcholine-conjugated gold-molecular clusters improve signal for Lymph Node NIR-II fluorescence imaging in preclinical cancer models.

Sentinel lymph node imaging and biopsy is important to clinical assessment of cancer metastasis, and novel non-radioactive lymphographic tracers have been actively pursued over the years. Here, we develop gold molecular clusters (Au25) functionalized by phosphorylcholine (PC) ligands for NIR-II (1000-3000 nm) fluorescence imaging of draining lymph nodes in 4T1 murine breast cancer and CT26 colon cancer tumor mouse models. The Au-phosphorylcholine (Au-PC) probes exhibit 'super-stealth' behavior with little interactions with serum proteins, cells and tissues in vivo, which differs from the indocyanine green (ICG) dye. Subcutaneous injection of Au-PC allows lymph node mapping by NIR-II fluorescence imaging at an optimal time of ~ 0.5 - 1 hour postinjection followed by rapid renal clearance. Preclinical NIR-II fluorescence LN imaging with Au-PC affords high signal to background ratios and high safety and biocompatibility, promising for future clinical translation.

Nickel-Hydride-Catalyzed Diastereo- and Enantioselective Hydroalkylation of Cyclopropenes.

Cyclopropanes are structural motifs that are widely present in natural products and bioactive molecules, and they are also tremendously useful building blocks in synthetic organic chemistry. Asymmetric synthesis of cyclopropane derivatives has been an intensively researched area over the years, but efficient asymmetric preparation of alkylcyclopropane scaffolds remains a challenging topic. Herein, we report a nickel-hydride-catalyzed enantioselective and diastereoselective hydroalkylation of cyclopropenes for facile synthesis of chiral alkylcyclopropane motifs. The reported method is efficient and versatile, taking place under mild reaction conditions, and having broad applicability and excellent functional group tolerance.

Structure of the trypanosome paraflagellar rod and insights into non-planar motility of eukaryotic cells.

Eukaryotic flagella (synonymous with cilia) rely on a microtubule-based axoneme, together with accessory filaments to carryout motility and signaling functions. While axoneme structures are well characterized, 3D ultrastructure of accessory filaments and their axoneme interface are mostly unknown, presenting a critical gap in understanding structural foundations of eukaryotic flagella. In the flagellum of the protozoan parasite Trypanosoma brucei (T. brucei), the axoneme is accompanied by a paraflagellar rod (PFR) that supports non-planar motility and signaling necessary for disease transmission and pathogenesis. Here, we employed cryogenic electron tomography (cryoET) with sub-tomographic averaging, to obtain structures of the PFR, PFR-axoneme connectors (PACs), and the axonemal central pair complex (CPC). The structures resolve how the 8 nm repeat of the axonemal tubulin dimer interfaces with the 54 nm repeat of the PFR, which consist of proximal, intermediate, and distal zones. In the distal zone, stacked "density scissors" connect with one another to form a "scissors stack network (SSN)" plane oriented 45° to the axoneme axis; and ~370 parallel SSN planes are connected by helix-rich wires into a paracrystalline array with ~90% empty space. Connections from these wires to the intermediate zone, then to overlapping layers of the proximal zone and to the PACs, and ultimately to the CPC, point to a contiguous pathway for signal transmission. Together, our findings provide insights into flagellum-driven, non-planar helical motility of T. brucei and have broad implications ranging from cell motility and tensegrity in biology, to engineering principles in bionics.