Discovery of Potent Antimalarial Type II Kinase Inhibitors with Selectivity over Human Kinases.

While progress has been made in the effort to eradicate malaria, the disease remains a significant threat to global health. Acquired resistance to frontline treatments is emerging in Africa, urging a need for the development of novel antimalarial agents. Repurposing human kinase inhibitors provides a potential expedited route given the availability of a diverse array of kinase-targeting drugs that are approved or in clinical trials. Phenotypic screening of a library of type II human kinase inhibitors identified compound 1 as a lead antimalarial, which was initially developed to target human ephrin type A receptor 2 (EphA2). Here, we report a structure-activity relationship study and lead optimization of compound 1, which led to compound 33, with improved antimalarial activity and selectivity.

Visible light-activated prodrug system with a novel heavy-atom-free photosensitizer.

The use of light to activate prodrugs offers a promising method for the precise control of drug release, reducing drug-related side effects, and enhancing therapeutic effectiveness. We have created a novel prodrug system that utilizes a unique, heavy-atom-free photosensitizer to produce singlet oxygen, which then triggers the conversion of the prodrug into its active form. This system has been successfully demonstrated through the creation of "photo-unclick" prodrugs of paclitaxel (PTX), combretastatin A-4 (CA-4), and 10-hydroxy-7-ethylcamptothecin (SN-38). These prodrugs show decreased toxicity in the absence of light, but exhibit increased toxicity when exposed to red light.

Human Polo-like Kinase Inhibitors as Antiplasmodials.

Protein kinases have proven to be a very productive class of therapeutic targets, and over 90 inhibitors are currently in clinical use primarily for the treatment of cancer. Repurposing these inhibitors as antimalarials could provide an accelerated path to drug development. In this study, we identified BI-2536, a known potent human polo-like kinase 1 inhibitor, with low nanomolar antiplasmodial activity. Screening of additional PLK1 inhibitors revealed further antiplasmodial candidates despite the lack of an obvious orthologue of PLKs in Plasmodium. A subset of these inhibitors was profiled for their in vitro killing profile, and commonalities between the killing rate and inhibition of nuclear replication were noted. A kinase panel screen identified PfNEK3 as a shared target of these PLK1 inhibitors; however, phosphoproteome analysis confirmed distinct signaling pathways were disrupted by two structurally distinct inhibitors, suggesting PfNEK3 may not be the sole target. Genomic analysis of BI-2536-resistant parasites revealed mutations in genes associated with the starvation-induced stress response, suggesting BI-2536 may also inhibit an aminoacyl-tRNA synthetase.

Photostable Small-Molecule NIR-II Fluorescent Scaffolds that Cross the Blood-Brain Barrier for Noninvasive Brain Imaging.

The second near-infrared (NIR-II, 1000-1700 nm) fluorescent probes have significant advantages over visible or NIR-I (600-900 nm) imaging for both depth of penetration and level of resolution. Since the blood-brain barrier (BBB) prevents most molecules from entering the central nervous system, NIR-II dyes with large molecular frameworks have limited applications for brain imaging. In this work, we developed a series of boron difluoride (BF2) formazanate NIR-II dyes, which had tunable photophysical properties, ultrahigh photostability, excellent biological stability, and strong brightness. Modulation of the aniline moiety of BF2 formazanate dyes significantly enhances their abilities to cross the BBB for noninvasive brain imaging. Furthermore, the intact mouse brain imaging and dynamic dye diffusion across the BBB were monitored using these BF2 formazanate dyes in the NIR-II region. In murine glioblastoma models, these dyes can differentiate tumors from normal brain tissues. We anticipate that this new type of small molecule will find potential applications in creating probes and drugs relevant to theranostic for brain pathologies.

Xanthone-based solvatochromic fluorophores for quantifying micropolarity of protein aggregates.

Proper three-dimensional structures are essential for maintaining the functionality of proteins and for avoiding pathological consequences of improper folding. Misfolding and aggregation of proteins have been both associated with neurodegenerative disease. Therefore, a variety of fluorogenic tools that respond to both polarity and viscosity have been developed to detect protein aggregation. However, the rational design of highly sensitive fluorophores that respond solely to polarity has remained elusive. In this work, we demonstrate that electron-withdrawing heteroatoms with (d-p)-π* conjugation can stabilize lowest unoccupied molecular orbital (LUMO) energy levels and promote bathochromic shifts. Guided by computational analyses, we have devised a novel series of xanthone-based solvatochromic fluorophores that have rarely been systematically studied. The resulting probes exhibit superior sensitivity to polarity but are insensitive to viscosity. As proof of concept, we have synthesized protein targeting probes for live-cell confocal imaging intended to quantify the polarity of misfolded and aggregated proteins. Interestingly, our results reveal several layers of protein aggregates in a way that we had not anticipated. First, microenvironments with reduced polarity were validated in the misfolding and aggregation of folded globular proteins. Second, granular aggregates of AgHalo displayed a less polar environment than aggregates formed by folded globular protein represented by Htt-polyQ. Third, our studies reveal that granular protein aggregates formed in response to different types of stressors exhibit significant polarity differences. These results show that the solvatochromic fluorophores solely responsive to polarity represent a new class of indicators that can be widely used for detecting protein aggregation in live cells, thus paving the way for elucidating cellular mechanisms of protein aggregation as well as therapeutic approaches to managing intracellular aggregates.

Local infiltration of analgesia and tranexamic acid is safe and efficacious in reducing blood loss and comparable to intra-articular tranexamic acid in total knee replacements.

INTRODUCTION: The use of peri-articular (PA) tranexamic acid (TXA) and its efficacy in comparison with intra-articular (IA) tranexamic acid has not been well explored in literature. This retrospective cohort study aims to compare the effects of IA and PA TXA with analgesic components in reducing blood loss and improving immediate post-operative pain relief and functional outcomes in unilateral primary total knee replacement (TKA) patients. METHODS: 63 patients who underwent a unilateral primary total knee replacement procedure were divided into 2 groups: 42 patients in the IA TXA delivery group, 21 patients in the PA TXA group. 1g of TXA was utilized for all patients. All patients had pericapsular infiltration consisting of 0.5ml of Adrenaline, 0.4ml of Morphine, 1g of Vancomycin, 1ml of Ketorolac and 15ml of Ropivacaine. Outcomes for blood loss, and surrogate markers for immediate functional recovery were measured. RESULTS: 54.0% of the patients were female, 46.0% male. The mean drop in post-operative Hb levels in the PA and IA group was 2.0g/dL and 1.6 g/dL respectively, and statistically insignificant (p=0.10). The mean HCT drop in the PA and IA group was 6.1% and 5.3% respectively and statistically insignificant (p=0.58). The POD 1 and discharge day flexion angles, POD 1 and POD 2 VAS scores, gait distance on discharge, and length of hospitalization stay were largely similar in both groups. CONCLUSION: Our study shows that both IA and PA TXA with analgesic components are equally efficient in reducing blood loss and improving immediate postoperative pain relief and functional outcomes.

Harnessing the power of antibodies to fight bone metastasis.

Antibody-based therapies have proved to be of great value in cancer treatment. Despite the clinical success of these biopharmaceuticals, reaching targets in the bone microenvironment has proved to be difficult due to the relatively low vascularization of bone tissue and the presence of physical barriers. Here, we have used an innovative bone-targeting (BonTarg) technology to generate a first-in-class bone-targeting antibody. Our strategy involves the use of pClick antibody conjugation technology to chemically couple the bone-targeting moiety bisphosphonate to therapeutic antibodies. Bisphosphonate modification of these antibodies results in the delivery of higher conjugate concentrations to the bone metastatic niche, relative to other tissues. In xenograft mice models, this strategy provides enhanced inhibition of bone metastases and multiorgan secondary metastases that arise from bone lesions. Specific delivery of therapeutic antibodies to the bone, therefore, represents a promising strategy for the treatment of bone metastatic cancers and other bone diseases.

Single-Atom Switching as a General Approach to Designing Colorimetric and Fluorogenic Probes for Mercury Ions.

By performing a single-atom replacement within common fluorophores, we have developed a facile and general strategy to prepare a broad-spectrum class of colorimetric and fluorogenic probes for the selective detection of mercury ions in aqueous environments. Thionation of carbonyl groups from existing fluorophore cores results in a great reduction of fluorescence quantum yield and loss of fluorescence emission. The resulting thiocaged probes are efficiently desulfurized to their oxo derivatives in the presence of mercury ions, leading to pronounced changes in chromogenic and fluorogenic signals. Because these probes exhibit high selectivity, excellent sensitivity, good membrane-permeability, and rapid responses towards mercury ions, they are suitable for visualization of mercury in both aqueous and intracellular environments.

Electronic Relaxation Pathways in Heavy-Atom-Free Photosensitizers Absorbing Near-Infrared Radiation and Exhibiting High Yields of Singlet Oxygen Generation.

Heavy-atom-free photosensitizers (HAF-PSs) based on thionation of carbonyl groups of readily accessible organic compounds are rapidly emerging as a versatile class of molecules. However, their photochemical properties and electronic relaxation mechanisms are currently unknown. Investigating the excited-state dynamics is essential to understand their benefits and limitations and to develop photosensitizers with improved photochemical properties. Herein, the photochemical and electronic-structure properties of two of the most promising HAF-PSs developed to date are revealed. It is shown that excitation of thio-4-(dimethylamino)naphthalamide and thionated Nile Red with near-infrared radiation leads to the efficient population of the triplet manifold through multiple relaxation pathways in hundreds of femtoseconds. The strong singlet-triplet couplings in this family of photosensitizers should enable a broad range of applications, including in photodynamic therapy, photocatalysis, photovoltaics, organic LEDs, and photon up-conversion.

Oxime as a general photocage for the design of visible light photo-activatable fluorophores.

Photoactivatable fluorophores have been widely used for tracking molecular and cellular dynamics with subdiffraction resolution. In this work, we have prepared a series of photoactivatable probes using the oxime moiety as a new class of photolabile caging group in which the photoactivation process is mediated by a highly efficient photodeoximation reaction. Incorporation of the oxime caging group into fluorophores results in loss of fluorescence. Upon light irradiation in the presence of air, the oxime-caged fluorophores are oxidized to their carbonyl derivatives, restoring strong fluorophore fluorescence. To demonstrate the utility of these oxime-caged fluorophores, we have created probes that target different organelles for live-cell confocal imaging. We also carried out photoactivated localization microscopy (PALM) imaging under physiological conditions using low-power light activation in the absence of cytotoxic additives. Our studies show that oximes represent a new class of visible-light photocages that can be widely used for cellular imaging, sensing, and photo-controlled molecular release.

Creation of Bacterial cells with 5-Hydroxytryptophan as a 21st Amino Acid Building Block.

While most organisms utilize 20 canonical amino acid building blocks for protein synthesis, adding additional candidates to the amino acid repertoire can greatly facilitate the investigation and manipulation of protein structures and functions. In this study, we report the generation of completely autonomous organisms with a 21st ncAA, 5-hydroxytryptophan (5HTP). Like 20 canonical amino acids, 5-hydroxytryptophan can be biosynthesized in vivo from simple carbon sources and is subsequently incorporated into proteins in response to the amber stop codon. Using this unnatural organism, we have prepared a single-chain immunoglobulin variable fragment conjugated with a fluorophore and demonstrated the utility of these autonomous cells to monitor oxidative stress. Creation of this and other cells containing the 21st amino acid will provide an opportunity to generate proteins and organisms with novel activities, as well as to determine the evolutionary consequences of using additional amino acid buildings.

Single-atom replacement as a general approach towards visible-light/near-infrared heavy-atom-free photosensitizers for photodynamic therapy.

Photodynamic therapy has become an emerging strategy for the treatment of cancer. This technology relies on the development of photosensitizers (PSs) that convert molecular oxygen to cytotoxic reactive oxygen species upon exposure to light. In this study, we have developed a facile and general strategy for obtaining visible light/near-infrared-absorbing PSs by performing a simple sulfur-for-oxygen replacement within existing fluorophores. Thionation of carbonyl groups within existing fluorophore cores leads to an improvement of the singlet oxygen quantum yield and molar absorption coefficient at longer wavelengths (deep to 600-800 nm). Additionally, these thio-based PSs lack dark cytotoxicity but exhibit significant phototoxicity against monolayer cancer cells and 3D multicellular tumor spheroids with IC50 in the micromolar range. To achieve tumor-specific delivery, we have conjugated these thio-based PSs to an antibody and demonstrated their tumor-specific therapeutic activity.

Higher doses of topical tranexamic acid safely improves immediate functional outcomes and reduces transfusion requirement in total knee arthroplasty.

BACKGROUND: Topical administration of tranexamic acid (TXA) in patients undergoing total knee arthroplasty (TKA) is increasingly popular as it avoids the risks related with systemic absorption of the medication. Previous studies have established the efficacy of TXA in TKA, however here are limited direct comparison studies available and the dosing regimens vary. Hence, there is no consensus on an optimal dose. Our objective is to compare blood loss, transfusion requirement and immediate post-operative function between high (2 g) and low (1 g) dose tranexamic acid in patients undergoing TKA. METHODS: This is a retrospective cohort study of 104 patients undergoing total knee arthroplasty in a single institution under a single surgeon. In total, 61 and 43 patients receiving 1 g and 2 g of topical TXA respectively. Blood loss as estimated from the difference in haemoglobin (Hb) and haematocrit (HCT) levels post-surgery and number of blood transfusions required were compared between groups. Immediate post-operative function and complications were also measured. RESULTS: Patient characteristics were mostly similar between groups. The transfusion requirements were higher in 1 g group compared to the 2 g group (0.11 vs 0.00, p = 0.034). The mean post op day 1 (POD1) range of motion higher in the 1 g group vs 2 g group (72.1 vs 63.7, p = 0.035). The 2 g group had a lower POD1 pain score compared to the 1 g group (4.02 vs 5.43, p < 0.01). There was no statistically significant difference in complications that were related to the administration of TXA between the two groups. CONCLUSION: Higher dose of topical TXA is safe, helps improve immediate post-operative functional outcomes and reduces transfusion requirements.

Tetrazine as a general phototrigger to turn on fluorophores.

Light-activated fluorescence affords a powerful tool for monitoring subcellular structures and dynamics with enhanced temporal and spatial control of the fluorescence signal. Here, we demonstrate a general and straightforward strategy for using a tetrazine phototrigger to design photoactivatable fluorophores that emit across the visible spectrum. Tetrazine is known to efficiently quench the fluorescence of various fluorophores via a mechanism referred to as through-bond energy transfer. Upon light irradiation, restricted tetrazine moieties undergo a photolysis reaction that generates two nitriles and molecular nitrogen, thus restoring the fluorescence of fluorophores. Significantly, we find that this strategy can be successfully translated and generalized to a wide range of fluorophore scaffolds. Based on these results, we have used this mechanism to design photoactivatable fluorophores targeting cellular organelles and proteins. Compared to widely used phototriggers (e.g., o-nitrobenzyl and nitrophenethyl groups), this study affords a new photoactivation mechanism, in which the quencher is photodecomposed to restore the fluorescence upon light irradiation. Because of the exclusive use of tetrazine as a photoquencher in the design of fluorogenic probes, we anticipate that our current study will significantly facilitate the development of novel photoactivatable fluorophores.

Amidyl Radical Directed Remote Allylation of Unactivated sp3 C-H Bonds by Organic Photoredox Catalysis.

The development of visible-light-mediated allylation of unactivated sp3 C-H bonds is reported. The remote allylation was directed by the amidyl radical, which was generated by photocatalytic fragmentation of a pre-functionalized amide precursor. Both aromatic and aliphatic amide derivatives could successfully deliver the remote C-H allylation products in good yields. A variety of electron deficient allyl sulfone systems could be used as δ-carbon radical acceptor.

Studies toward the Synthesis of Iriomoteolide-2a: Construction of the C(6)-C(28) Fragment.

The synthesis of an appropriately functionalized advanced C(6-28) fragment (3) of the marine macrolide iriomoteolide-2a (1) has been achieved in a highly efficient manner. The C(6)-C(18) fragment of 1 is prepared via a radical cyclization of a vinyl ether intermediate and palladium-promoted hydrostannylation/iodination. Paterson aldol reaction and Peterson olefination are used to construct the C(19)-C(28) fragment. The union of the C(6)-C(18) and C(19)-C(28) fragments is accomplished via a Suzuki-Miyaura coupling reaction.

Regio- and Stereospecific Construction of 3a-(1H-Indol-3-yl)pyrrolidinoindolines and Application to the Formal Syntheses of Gliocladins B and C.

A one-pot regio- and stereospecific strategy for the construction of 3a-(3-indolyl)-hexahydropyrrolo[2,3-b]indoles based on the condensation of an indole and an in situ generated cyclopropylazetoindoline has been developed. This unified strategy works with a variety of substituted indoles to produce 3a-(3-indolyl)hexahydropyrrolo[2,3-b]indole products in high yields. The utility of this transformation was highlighted in the formal total syntheses of gliocladins B and C.

Silver-catalysed direct amination of unactivated C-H bonds of functionalized molecules.

Carbon-nitrogen bond formation from inert C-H bonds is an ideal organic transformation and a highly desirable method for the synthesis of N-containing molecules due to its high efficiency and atom economy. In this report, we develop a general reaction to achieve an unprecedented selective intramolecular amination of unactivated C-H bond in the absence of complex directing groups. Functionalized heterocyclic products are built up from readily available linear amines through simple and reliable silver catalysis, representing a new silver-based C-H functionalization. This method displays preference for primary sp(3) C-H bonds and exhibits distinct chemo- and regioselectivity compared to existing methods of direct amination (Hofmann-Löffler-Freytag reaction and nitrene insertion). The study highlights the manipulation of unfunctionalized groups in organic molecules to furnish complex structural units in the natural and bioactive molecules.

Cutaneous hypoesthesia following plate fixation in clavicle fractures.

BACKGROUND: We report the functional impact and natural history of cutaneous hypoesthesia after plate internal fixation for mid shaft clavicle fractures with a horizontal skin incision from year 2009 to 2011. MATERIALS AND METHODS: 38 patients had complete followup data with an average followup period of 23.2 months (range 8-43 months). The impact of supraclavicular nerve injury was studied by assessment of the incidence, functional impact and natural history of numbness with detailed review of the case records and phone questionnaires. RESULTS: The incidence of postoperative numbness was 55.3% (n = 21). Most patients reported the numbness to be at its worst within the first operative month. At the time of worst numbness, 28.6% (n = 6/21) of patients reported the numbness to be severe while 42.9% (n = 9/21) reported moderate numbness and 28.6% (n = 6/21) reported mild numbness. Fifteen of these patients described increased awareness of numbness during contact with straps or clothes. Two patients were significantly bothered by this numbness; 4 patients stated that it was a moderate bother while 7 patients considered it a mild bother. A total of 8 patients reported that they were not bothered at all by the numbness. An overwhelming majority of affected patients (90.5%, n = 19/21) reported an improvement in the severity of numbness felt over time. At the last followup, the incidence of numbness declined from 55.3% to 36.8% with 7 patients reporting complete resolution of numbness. The numbness however was found to persist in 66.7% of patients. Only 1 patient reported continued severe numbness. The awareness of numbness with straps and clothing was severe in 5 patients. None of the patients were significantly bothered by this numbness. CONCLUSIONS: Cutaneous sensory loss is a common occurrence following plate fixation of the clavicle and might have been under reported in the literature. The numbness improves in the vast majority, but commonly persists to some degree for up to 2 years and maybe permanent. However, only a small minority eventually considered the numbness a significant "bother" and to affect them while wearing clothing or when in contact with shoulder straps.

Surgically treated cervical myelopathy: a functional outcome comparison study between multilevel anterior cervical decompression fusion with instrumentation and posterior laminoplasty.

BACKGROUND CONTEXT: Multilevel cervical myelopathy can be treated with anterior cervical discectomy and fusion (ACDF) or corpectomy via the anterior approach and laminoplasty via the posterior approach. Till date, there is no proven superior approach. PURPOSE: To elucidate any potential advantage of one approach over the other with regard to clinical midterm outcomes in this study. STUDY DESIGN: A prospective, 2-year follow-up of patients with cervical myelopathy treated with multilevel anterior cervical decompression fusion and plating and posterior laminoplasty. PATIENT SAMPLE: In total, 116 patients were studied. Sixty-four patients underwent ACDF two levels and above or anterior cervical corpectomy and fusion one level and above. Fifty-two patients underwent posterior cervical surgery (laminoplasty C3-C6 and C3-C7). OUTCOME MEASURES: Self-report measures: Japan Orthopedic Association (JOA) score, JOA recovery rate, visual analog scale for neck pain (VASNP), neck disability index (NDI), and American Academy of Orthopaedic Surgeons (AAOS) neurogenic symptom score (AAOS-NSS). Physiologic measures: range of motion (ROM) flexion and extension of neck. Functional measures: short-form 36 (SF-36) score comprising physical functioning, physical role function, bodily pain, general health, vitality, social role function, emotional role function, and mental health scales. METHODS: Comparison of the JOA scores, JOA recovery rates, NDI scores, SF-36 scores, VASNP, and ROM preoperatively to 2 years. Chi-square and two-sided Student t tests were used to analyze the variables. RESULTS: Posterior surgery took an hour shorter (p<.05) and had better improvement in JOA scores at early follow-up of 6 months (p=.025). Anterior surgery group had better improvement of NDI scores at early follow-up of 6 months (p=.024) and was associated with less blood loss intraoperatively compared with posterior surgery. There was no statistical difference between the two groups for JOA scores, JOA recovery rates, SF-36 quality-of-life scores, NDI, AAOS-NSS, VAS neck pain, and ROM at 2 years. Complications were higher for anterior surgery group: two hematoma postoperation, one vocal cord paresis, and one new onset C6/C7 dermatome numbness versus one dura leak in posterior surgery group. CONCLUSIONS: Our study showed that patients with multilevel disease treated with laminoplasty do well and compare favorably with patients treated with an anterior approach. Notably, posterior surgery was associated with shorter operating time, better improvement in JOA scores at 6 months, and a tendency toward lesser complications. Posterior surgery was not associated with increased neck disability and neck pain at 2 years. Anterior surgery had better NDI improvement at early follow-up. There is a need for a larger study that is prospectively randomized with long-term follow-up before we can confidently advocate one approach over the other in the management of cervical myelopathy.