New Potent Sulfonamide-Based Inhibitors of S. aureus Biotin Protein Ligase.

The key regulatory metabolic enzyme, biotin protein ligase (BPL), is an attractive target for the development of novel antibiotics against multi-drug-resistant bacteria, such as Staphylococcus aureus. Here we report the synthesis and assay of a new series of inhibitors (6-9) against S. aureus BPL (SaBPL), where a component sulfonamide linker was used to mimic the acyl-phosphate group of the natural intermediate biotinyl-5'-AMP (1). A pivotal correlation between the acidity of the central NH of the sulfonamide linker of 6-9 and in vitro inhibitory activity against SaBPL was observed. Specifically, sulfonylcarbamate 8, with its highly acidic sulfonyl central NH, as evaluated by 1H NMR spectroscopy, showed exceptional potency (K i = 10.3 ± 3.8 nM). Furthermore, three inhibitors demonstrated minimum inhibitory concentrations of 16-32 µg/mL against clinical methicillin-resistant S. aureus (MRSA) strains.

Elucidating Synergies of Single-Atom Catalysts in a Model Thin Film Photoelectrocatalyst to Maximize Hydrogen Evolution Reaction.

Realization of the full potential of single-atom photoelectrocatalysts in sustainable energy generation requires careful consideration of the design of the host material. Here, a comprehensive methodology for the rational design of photoelectrocatalysts using anodic titanium dioxide (TiO2) nanofilm as a model platform is presented. The properties of these nanofilms are precisely engineered to elucidate synergies across structural, chemical, optoelectronic, and electrochemical properties to maximize the efficiency of the hydrogen evolution reaction (HER). These findings clearly demonstrate that thicker TiO2 nanofilms in anatase phase with pits on the surface can accommodate single-atom platinum catalysts in an optimal configuration to increase HER performance. It is also evident that the electrolyte temperature can further enhance HER output through thermochemical effect. A judicious design incorporating all these factors into one system gives rise to a ten-fold HER enhancement. However, the reusability of the host photoelectrocatalyst is limited by the leaching of the Pt atom, worsening HER. Density-functional theory calculations have provided insights into the mechanism underlying the experimental observations in terms of moderate hydrogen adsorption and enhanced gas generation. This improved understanding of the critical factors determining HER performance in a model photoelectrocatalyst paves the way for future advances in scalable and translatable photoelectrocatalyst technologies.

18 kDa Translocator protein (TSPO) is upregulated in rat brain after peripheral nerve injury and downregulated by diroximel fumarate.

Neuroimmune signaling is a key process underlying neuropathic pain. Clinical studies have demonstrated that 18 kDa translocator protein (TSPO), a putative marker of neuroinflammation, is upregulated in discrete brain regions of patients with chronic pain. However, no preclinical studies have investigated TSPO dynamics in the brain in the context of neuropathic pain and in response to analgesic treatments. We used positron emission tomography-computed tomography (PET-CT) and [18F]-PBR06 radioligand to measure TSPO levels in the brain across time after chronic constriction injury (CCI) of the sciatic nerve in both male and female rats. Up to 10 weeks post-CCI, TSPO expression was increased in discrete brain regions, including medial prefrontal cortex, somatosensory cortex, insular cortex, anterior cingulate cortex, motor cortex, ventral tegmental area, amygdala, midbrain, pons, medulla, and nucleus accumbens. TSPO was broadly upregulated across these regions at 4 weeks post CCI in males, and 10 weeks in females, though there were regional differences between the sexes. Using immunohistochemistry, we confirmed TSPO expression in these regions. We further demonstrated that TSPO was upregulated principally in microglia in the nucleus accumbens core, and astrocytes and endothelial cells in the nucleus accumbens shell. Finally, we tested whether TSPO upregulation was sensitive to diroximel fumarate, a drug that induces endogenous antioxidants via nuclear factor E2-related factor 2 (Nrf2). Diroximel fumarate alleviated neuropathic pain and reduced TSPO upregulation. Our findings indicate that TSPO is upregulated over the course of neuropathic pain development and is resolved by an antinociceptive intervention in animals with peripheral nerve injury.

Systemic and targeted activation of Nrf2 reverses doxorubicin-induced cognitive impairments and sensorimotor deficits in mice.

While cancer survivorship has increased due to advances in treatments, chemotherapy often carries long-lived neurotoxic side effects which reduce quality of life. Commonly affected domains include memory, executive function, attention, processing speed and sensorimotor function, colloquially known as chemotherapy-induced cognitive impairment (CICI) or "chemobrain". Oxidative stress and neuroimmune signaling in the brain have been mechanistically linked to the deleterious effects of chemotherapy on cognition and sensorimotor function. With this in mind, we tested if activation of the master regulator of antioxidant response nuclear factor E2-related factor 2 (Nrf2) alleviates cognitive and sensorimotor impairments induced by doxorubicin. The FDA-approved systemic Nrf2 activator, diroximel fumarate (DRF) was used, along with our recently developed prodrug 1c which has the advantage of specifically releasing monomethyl fumarate at sites of oxidative stress. DRF and 1c both reversed doxorubicin-induced deficits in executive function, spatial and working memory, as well as decrements in fine motor coordination and grip strength, across both male and female mice. Both treatments reversed doxorubicin-induced loss of synaptic proteins and microglia phenotypic transition in the hippocampus. Doxorubicin-induced myelin damage in the corpus callosum was reversed by both Nrf2 activators. These results demonstrate the therapeutic potential of Nrf2 activators to reverse doxorubicin-induced cognitive impairments, motor incoordination, and associated structural and phenotypic changes in the brain. The localized release of monomethyl fumarate by 1c has the potential to diminish unwanted effects of fumarates while retaining efficacy.

A New Gramicidin S Analogue with Potent Antibacterial Activity and Negligible Hemolytic Toxicity.

Antibiotic resistance is an urgent threat to global health, with the decreasing efficacy of conventional drugs underscoring the urgency for innovative therapeutic strategies. Antimicrobial peptides present as promising alternatives to conventional antibiotics. Gramicidin S is one such naturally occurring antimicrobial peptide that is effective against Staphylococcus aureus, with a minimum inhibitory concentration (MIC) of 4 µg/mL (3.6 µM). Despite this potent activity, its significant hemolytic toxicity restricts its clinical use to topical applications. Herein, we present rational modifications to the key ß-strand and ß-turn regions of gramicidin S to concurrently mitigate hemolytic effects, while maintaining potency. Critically, peptide 9 displayed negligible hemolytic toxicity, while possessing significant antibacterial potency against a panel of methicillin-sensitive and methicillin-resistant S. aureus clinical isolates (MIC of 8 µg/mL, 7.2 µM). Given the substantial antibacterial activity and near absence of cytotoxicity, 9 presents as a potential candidate for systemic administration in the treatment of S. aureus bacteremia/sepsis.

Detection of Volatile Organic Compounds through Spectroscopic Signatures in Nanoporous Fabry-Pérot Optical Microcavities.

Increasingly complex modern gas-monitoring scenarios necessitate advanced sensing capabilities to detect and identify a diverse range of gases under varying conditions. There is a rising demand for individual sensors with multiple responses capable of recognizing gases, identifying components in mixtures, and providing stable responses. Inspired by gas sensors employing multivariable response principles, we develop a nanoporous anodic alumina high-order microcavity (NAA-HOµCV) gas sensor with multiple optical outputs for discriminative gas detection. The NAA-HOµCV architecture, formed by a Fabry-Pérot microcavity with distributed Bragg reflector (DBR) mirrors and an extended-length microcavity layer supporting multiple resonant modes, serves as an effective solid-state fingerprint platform for distinguishing volatile organic compound (VOC) gases. Our research reveals that the coupling strength of light into resonant modes and their evolution depend on the thickness of the DBR mirrors and the dimension of the microcavity layer, which allows us to optimize the discriminative sensing capability of the NAA-HOµCV sensor through structural engineering of the microcavity and photonic crystal mirrors. Gas-sensing experiments conducted on the NAA-HOµCV sensor demonstrate real-time discrimination between physiosorbed VOC gases (isopropanol, ethanol, or acetone) in reversible gas sensing. It also achieves superior ppb-level sensing in irreversible gas sensing of model silane molecules. Our study presents promising avenues for designing compact, cost-effective, and highly efficient gas sensors with tailored properties for discriminative gas detection.

Engineering of Solid-State Nanoporous Laser through Dendrimer-Encapsulated Fluorophores.

Dendrimers─nanosized macromolecules that can function as hosts for encapsulation of guest molecules─provide new avenues to engineer gain media for lasing systems. In this context, this study investigates the interplay between the geometric features of a model porous scattering medium, nanoporous anodic alumina (NAA), and the chemical features of a model fluorophore-dendrimer encapsulation system to maximize random lasing. The inner surface of the NAA platforms is functionalized with fluorophore molecules encapsulated within dendrimers via an electrostatic interaction. The resulting solid-state composite structures emit well-resolved, intense random lasing when subjected to optical pumping. By engineering fluorophore-dendrimer and geometric features of scattering medium, we can precisely tune the characteristics of random lasing emissions. It is found that lasing structures with low porosity and thickness functionalized with fluorophore molecules encapsulated in second-generation dendrimers provide the best platforms for lasing generation, resulting in a strongly polarized laser at ∼594 nm that has a high quality-gain product of ∼1588 au, a polarization quality of ∼0.86, and a lasing threshold of ∼0.05 mJ pulse-1. Comparative analysis indicates that dendrimers achieve 2.5 times better random lasing than conventional surfactants due to improved encapsulation and minimization of photobleaching. Our results reveal the importance of the fluorophore encapsulation method and design of scattering media in the engineering of random lasing platforms for applications in optical and optoelectrical systems.

Tailoring Tamm Plasmon Resonances in Dielectric Nanoporous Photonic Crystals.

The fields of plasmonics and photonic crystals (PCs) have been combined to generate model light-confining Tamm plasmon (TMM) cavities. This approach effectively overcomes the intrinsic limit of diffraction faced by dielectric cavities and mitigates losses associated with the inherent properties of plasmonic materials. In this study, nanoporous anodic alumina PCs, produced by two-step sinusoidal pulse anodization, are used as a model dielectric platform to establish the methodology for tailoring light confinement through TMM resonances. These model dielectric mirrors feature highly organized nanopores and narrow bandwidth photonic stopbands (PSBs) across different positions of the spectrum. Different types of metallic films (gold, silver, and aluminum) were coated on the top of these model dielectric mirrors. By structuring the features of the plasmonic and photonic components of these hybrid structures, the characteristics of TMM resonances were studied to elucidate effective approaches to optimize the light-confining capability of this hybrid TMM model system. Our findings indicate that the coupling of photonic and plasmonic modes is maximized when the PSB of the model dielectric mirror is broad and located within the midvisible region. It was also found that thicker metal films enhance the quality of the confined light. Gas sensing experiments were performed on optimized TMM systems, and their sensitivity was assessed in real time to demonstrate their applicability. Ag films provide superior performance in achieving the highest sensitivity (S = 0.038 ± 0.001 nm ppm-1) based on specific binding interactions between thiol-containing molecules and metal films.

Towards a High-Affinity Peptidomimetic Targeting Proliferating Cell Nuclear Antigen from Aspergillus fumigatus.

Invasive fungal infections (IFIs) are prevalent in immunocompromised patients. Due to alarming levels of increasing resistance in clinical settings, new drugs targeting the major fungal pathogen Aspergillus fumigatus are required. Attractive drug targets are those involved in essential processes like DNA replication, such as proliferating cell nuclear antigens (PCNAs). PCNA has been previously studied in cancer research and presents a viable target for antifungals. Human PCNA interacts with the p21 protein, outcompeting binding proteins to halt DNA replication. The affinity of p21 for hPCNA has been shown to outcompete other associating proteins, presenting an attractive scaffold for peptidomimetic design. p21 has no A. fumigatus homolog to our knowledge, yet our group has previously demonstrated that human p21 can interact with A. fumigatus PCNA (afumPCNA). This suggests that a p21-based inhibitor could be designed to outcompete the native binding partners of afumPCNA to inhibit fungal growth. Here, we present an investigation of extensive structure-activity relationships between designed p21-based peptides and afumPCNA and the first crystal structure of a p21 peptide bound to afumPCNA, demonstrating that the A. fumigatus replication model uses a PIP-box sequence as the method for binding to afumPCNA. These results inform the new optimized secondary structure design of a potential peptidomimetic inhibitor of afumPCNA.

Structure-guided design and synthesis of ATP-competitive N-acyl-substituted sulfamide d-alanine-d-alanine ligase inhibitors.

d-Alanine-d-alanine ligase (Ddl) catalyses the ATP-dependent formation of d-Ala-d-Ala, a critical component in bacterial cell wall biosynthesis and is a validated target for new antimicrobial agents. Here, we describe the structure-guided design, synthesis, and evaluation of ATP-competitive N-acyl-substituted sulfamides 27-36, 42, 46, 47 as inhibitors of Staphylococcus aureus Ddl (SaDdl). A crystal structure of SaDdl complexed with ATP and d-Ala-d-Ala (PDB: 7U9K) identified ATP-mimetic 8 as an initial scaffold for further inhibitor design. Evaluation of 8 in SaDdl enzyme inhibition assays revealed the ability to reduce enzyme activity to 72 ± 8 % (IC50 = 1.6 mM). The sulfamide linker of 8 was extended with 2-(4-methoxyphenyl)ethanol to give 29, to investigate further interactions with the d-Ala pocket of SaDdl, as predicted by molecular docking. This compound reduced enzyme activity to 89 ± 1 %, with replacement of the 4-methoxyphenyl group in 29 with alternative phenyl substituents (27, 28, 31-33, 35, 36) failing to significantly improve on this (80-89 % remaining enzyme activity). Exchanging these phenyl substituents with selected heterocycles (42, 46, 47) did improve activity, with the most active compound (42) reducing SaDdl activity to 70 ± 1 % (IC50 = 1.7 mM), which compares favourably to the FDA-approved inhibitor d-cycloserine (DCS) (IC50 = 0.1 mM). To the best of our knowledge, this is the first reported study of bisubstrate SaDdl inhibitors.

Understanding the Intrinsic Rectification Properties of Nanoporous Anodic Alumina by Selective Chemical Etching.

The distribution of oxygen and aluminum vacancies across the hemispherical barrier oxide layer (BOL) of nanoporous anodic alumina (NAA) relies intrinsically on the electric field-driven flow of electrolytic species and the incorporation of electrolyte impurities during the growth of anodic oxide through anodization. This phenomenon provides new opportunities to engineer BOL's inherited ionic current rectification (ICR) fingerprints. NAA's characteristic ICR signals are associated with the space charge density gradient across BOL and electric field-induced ion migration through hopping from vacancy to vacancy. In this study, we engineer the intrinsic space charge density gradient of the BOL of NAA under a range of anodizing potentials in hard and mild anodization regimes. Real-time characterization of the ICR fingerprints of NAA during selective etching of the BOL makes it possible to unravel the distribution pattern of vacancies through rectification signals as a function of etching direction and time. Our analysis demonstrates that the space charge density gradient varies across the BOL of NAA, where the magnitude and distribution of the space charge density gradient are revealed to be critically determined by anodizing the electrolyte, regime, and potential. This study provides a comprehensive understanding of the engineering of ion transport behavior across blind-hole NAA membranes by tuning the distribution of defects across BOL through anodization conditions. This method has the potential to be harnessed for developing nanofluidic devices with tailored ionic rectification properties for energy generation and storage and sensing applications.

Fortuitous In Vitro Compound Degradation Produces a Tractable Hit against Mycobacterium tuberculosis Dethiobiotin Synthetase: A Cautionary Tale of What Goes In Does Not Always Come Out.

We previously reported potent ligands and inhibitors of Mycobacterium tuberculosis dethiobiotin synthetase (MtDTBS), a promising target for antituberculosis drug development (Schumann et al., ACS Chem Biol. 2021, 16, 2339-2347); here, the unconventional origin of the fragment compound they were derived from is described for the first time. Compound 1 (9b-hydroxy-6b,7,8,9,9a,9b-hexahydrocyclopenta[3,4]cyclobuta[1,2-c]chromen-6(6aH)-one), identified by an in silico fragment screen, was subsequently shown by surface plasmon resonance to have dose-responsive binding (KD = 0.6 mM). Clear electron density was revealed in the DAPA substrate binding pocket when 1 was soaked into MtDTBS crystals, but the density was inconsistent with the structure of 1. Here, we show that the lactone of 1 hydrolyzes to a carboxylic acid (2) under basic conditions, including those of the crystallography soak, with a subsequent ring opening of the component cyclobutane ring forming a cyclopentylacetic acid (3). Crystals soaked directly with authentic 3 produced an electron density that matched that of crystals soaked with presumed 1, confirming the identity of the bound ligand. The synthetic utility of fortuitously formed 3 enabled the subsequent compound development of nanomolar inhibitors. Our findings represent an example of chemical modification within drug discovery assays and demonstrate the value of high-resolution structural data in the fragment hit validation process.

Exploring Photoswitchable Binding Interactions with Small-Molecule- and Peptide-Based Inhibitors of Trypsin.

The ability to photochemically activate a drug, both when and where needed, requires optimisation of the difference in biological activity between each isomeric state. As a step to this goal, we report small-molecule- and peptide-based inhibitors of the same protease-trypsin-to better understand how photoswitchable drugs interact with their biological target. The best peptidic inhibitor displayed a more than fivefold difference in inhibitory activity between isomeric states, whereas the best small-molecule inhibitor only showed a 3.4-fold difference. Docking and molecular modelling suggest this result is due to a large change in 3D structure in the key binding residues of the peptidic inhibitor upon isomerisation; this is not observed for the small-molecule inhibitor. Hence, we demonstrate that significant structural changes in critical binding motifs upon irradiation are essential for maximising the difference in biological activity between isomeric states. This is an important consideration in the design of future photoswitchable drugs for clinical applications.

Designing Fluorescent Nuclear Permeable Peptidomimetics to Target Proliferating Cell Nuclear Antigen.

Human proliferating cell nuclear antigen (PCNA) is a critical mediator of DNA replication and repair, acting as a docking platform for replication proteins. Disrupting these interactions with a peptidomimetic agent presents as a promising avenue to limit proliferation of cancerous cells. Here, a p21-derived peptide was employed as a starting scaffold to design a modular peptidomimetic that interacts with PCNA and is cellular and nuclear permeable. Ultimately, a peptidomimetic was produced which met these criteria, consisting of a fluorescein tag and SV40 nuclear localization signal conjugated to the N-terminus of a p21 macrocycle derivative. Attachment of the fluorescein tag was found to directly affect cellular uptake of the peptidomimetic, with fluorescein being requisite for nuclear permeability. This work provides an important step forward in the development of PCNA targeting peptidomimetics for use as anti-cancer agents or as cancer diagnostics.

Smart Wearable Patches Using Light-Controlled Activation and Delivery of Photoswitchable Antimicrobial Peptides.

A novel strategy to treat Staphylococcus aureus (S. aureus) skin infections is presented, where UV light is used to facilitate concomitant light-controlled activation and delivery of an antimicrobial therapeutic agent. Specifically, a new photoswitchable gramicidin S analogue was immobilized onto a polymeric wearable patch via a photocleavable linker that undergoes photolysis at the same wavelength of light required for activation of the peptide. Unlike toxic gramicidin S, the liberated active photoswitchable peptide exhibits antimicrobial activity against S. aureus while being ostensibly non-haemolytic to red blood cells. Moreover, irradiation with visible light switches off the antimicrobial properties of the peptide within seconds, presenting an ideal strategy to regulate antibiotic activity for localized bacterial infections with the potential to mitigate resistance.

PPARγ Corepression Involves Alternate Ligand Conformation and Inflation of H12 Ensembles.

Inverse agonists of peroxisome proliferator activated receptor γ (PPARγ) have emerged as safer alternatives to full agonists for their reduced side effects while still maintaining impressive insulin-sensitizing properties. To shed light on their molecular mechanism, we characterized the interaction of the PPARγ ligand binding domain with SR10221. X-ray crystallography revealed a novel binding mode of SR10221 in the presence of a transcriptionally repressing corepressor peptide, resulting in much greater destabilization of the activation helix, H12, than without corepressor peptide. Electron paramagnetic resonance provided in-solution complementary protein dynamic data, which revealed that for SR10221-bound PPARγ, H12 adopts a plethora of conformations in the presence of corepressor peptide. Together, this provides the first direct evidence for corepressor-driven ligand conformation for PPARγ and will allow the development of safer and more effective insulin sensitizers suitable for clinical use.

Uncovering the link between the SpnIII restriction modification system and LuxS in Streptococcus pneumoniae meningitis isolates.

Streptococcus pneumoniae is capable of randomly switching their genomic DNA methylation pattern between six distinct bacterial subpopulations (A-F) via recombination of a type 1 restriction-modification locus, spnIII. These pneumococcal subpopulations exhibit phenotypic changes which favor carriage or invasive disease. In particular, the spnIIIB allele has been associated with increased nasopharyngeal carriage and the downregulation of the luxS gene. The LuxS/AI-2 QS system represent a universal language for bacteria and has been linked to virulence and biofilm formation in S. pneumoniae. In this work, we have explored the link between spnIII alleles, the luxS gene and virulence in two clinical pneumococcal isolates from the blood and cerebrospinal fluid (CSF) of one pediatric meningitis patient. The blood and CSF strains showed different virulence profiles in mice. Analysis of the spnIII system of these strains recovered from the murine nasopharynx showed that the system switched to different alleles commensurate with the initial source of the isolate. Of note, the blood strain showed high expression of spnIIIB allele, previously linked with less LuxS protein production. Importantly, strains with deleted luxS displayed different phenotypic profiles compared to the wildtype, but similar to the strains recovered from the nasopharynx of infected mice. This study used clinically relevant S. pneumoniae strains to demonstrate that the regulatory network between luxS and the type 1 restriction-modification system play a key role in infections and may support different adaptation to specific host niches.

Structural Study of Potent Triazole-Based Inhibitors of Staphylococcus aureus Biotin Protein Ligase.

The rise of multidrug-resistant bacteria, such as Staphylococcus aureus, has highlighted global urgency for new classes of antibiotics. Biotin protein ligase (BPL), a critical metabolic regulatory enzyme, is an important target that shows significant promise in this context. Here we report the in silico docking, synthesis, and biological assay of a new series of N1-diphenylmethyl-1,2,3-triazole-based S. aureus BPL (SaBPL) inhibitors (8-19) designed to probe the adenine binding site and define whole-cell activity for this important class of inhibitor. Triazoles 13 and 14 with N1-propylamine and -butanamide substituents, respectively, were particularly potent with K i values of 10 ± 2 and 30 ± 6 nM, respectively, against SaBPL. A strong correlation was apparent between the K i values for 8-19 and the in silico docking, with hydrogen bonding to amino acid residues S128 and N212 of SaBPL likely contributing to potent inhibition.

A New 1,2,3-Triazole Scaffold with Improved Potency against Staphylococcus aureus Biotin Protein Ligase.

Staphylococcus aureus, a key ESKAPE bacteria, is responsible for most blood-based infections and, as a result, is a major economic healthcare burden requiring urgent attention. Here, we report in silico docking, synthesis, and assay of N1-diphenylmethyl triazole-based analogues (7-13) designed to interact with the entire binding site of S. aureus biotin protein ligase (SaBPL), an enzyme critical for the regulation of gluconeogenesis and fatty acid biosynthesis. The second aryl ring of these compounds enhances both SaBPL potency and whole cell activity against S. aureus relative to previously reported mono-benzyl triazoles. Analogues 12 and 13, with added substituents to better interact with the adenine binding site, are particularly potent, with Ki values of 6.01 ± 1.01 and 8.43 ± 0.73 nM, respectively. These analogues are the most active triazole-based inhibitors reported to date and, importantly, inhibit the growth of a clinical isolate strain of S. aureus ATCC 49775, with minimum inhibitory concentrations of 1 and 8 µg/mL, respectively.

Designer D-peptides targeting the N-terminal region of α-synuclein to prevent parkinsonian-associated fibrilization and cytotoxicity.

The deposition of α-synuclein (αS) aggregates in the gut and the brain is ever present in cases of Parkinson's disease. While the central non-amyloidogenic-component (NAC) region of αS plays a critical role in fibrilization, recent studies have identified a specific sequence from within the N-terminal region (NTR, residues 36-42) as a key modulator of αS fibrilization. Due to the lack of effective therapeutics which specifically target αS aggregates, we have developed a strategy to prevent the aggregation and subsequent toxicity attributed to αS fibrilization utilizing NTR targeting peptides. In this study, L- and D-isoforms of a hexa- (VAQKTV-Aib, 77-82 NAC) and heptapeptide (GVLYVGS-Aib, 36-42 NTR) containing a self-recognition component unique to αS, as well as a C-terminal disruption element, were synthesized to target primary sequence regions of αS that modulate fibrilization. The D-peptide that targets the NTR (NTR-TP-D) was shown by ThT fluorescence assays and TEM to be the most effective at preventing fibril formation and elongation, as well as increasing the abundance of soluble monomeric αS. In addition, NTR-TP-D alters the conformation of destabilised monomers into a less aggregation-prone state and reduces the hydrophobicity of αS fibrils via fibril remodelling. Furthermore, both NTR-TP isoforms alleviate the cytotoxic effects of αS aggregates in both Neuro-2a and Caco-2 cells. Together, this study highlights how targeting the NTR of αS using D-isoform peptide inhibitors may effectively combat the deleterious effects of αS fibrilization and paves the way for future drug design to utilise such an approach to treat Parkinson's disease.