D-peptide-magnetic nanoparticles fragment tau fibrils and rescue behavioral deficits in a mouse model of Alzheimer's disease.

Amyloid fibrils of tau are increasingly accepted as a cause of neuronal death and brain atrophy in Alzheimer's disease (AD). Diminishing tau aggregation is a promising strategy in the search for efficacious AD therapeutics. Previously, our laboratory designed a six-residue, nonnatural amino acid inhibitor D-TLKIVW peptide (6-DP), which can prevent tau aggregation in vitro. However, it cannot block cell-to-cell transmission of tau aggregation. Here, we find D-TLKIVWC (7-DP), a d-cysteine extension of 6-DP, not only prevents tau aggregation but also fragments tau fibrils extracted from AD brains to neutralize their seeding ability and protect neuronal cells from tau-induced toxicity. To facilitate the transport of 7-DP across the blood-brain barrier, we conjugated it to magnetic nanoparticles (MNPs). The MNPs-DP complex retains the inhibition and fragmentation properties of 7-DP alone. Ten weeks of MNPs-DP treatment appear to reverse neurological deficits in the PS19 mouse model of AD. This work offers a direction for development of therapies to target tau fibrils.

Oolonghomobisflavans from Camellia sinensis disaggregate tau fibrils across Alzheimer's disease models.

Alzheimer's disease (AD) is a common debilitating neurodegenerative disease with limited treatment options. Amyloid-ß (Aß) and tau fibrils are well-established hallmarks of AD, which can induce oxidative stress, neuronal cell death, and are linked to disease pathology. Here, we describe the effects of Oolonghomobisflavan A (OFA) and Oolonghomobisflavan B (OFB) on tau fibril disaggregation and prionogenic seeding. Transcriptomic analysis of OF-treated animals reveals the induction of a proteostasis-enhancing and health-promoting signature. OFA treatment reduced the burden of Tau protein aggregation in a C. elegans model expressing pathogenic human tau ("hTau-expressing") and promoted Tau disaggregation and inhibited seeding in assays using ex vivo brain-derived paired helical filament tau protein fibrils from Alzheimer's disease brain donors. Correspondingly, treatment with OF improved multiple fitness and aging-related health parameters in the hTau-expressing C. elegans model, including reproductive output, muscle function, and importantly, reversed the shortened lifespan stemming from pathogenic Tau expression. Collectively, this study provides new evidence supporting the neuroprotective effects of OFs and reveal a new therapeutic strategy for targeting AD and other neurodegenerative diseases characterized by tauopathy.

Structure-based design of nanobodies that inhibit seeding of Alzheimer's patient-extracted tau fibrils.

Despite much effort, antibody therapies for Alzheimer's disease (AD) have shown limited efficacy. Challenges to the rational design of effective antibodies include the difficulty of achieving specific affinity to critical targets, poor expression, and antibody aggregation caused by buried charges and unstructured loops. To overcome these challenges, we grafted previously determined sequences of fibril-capping amyloid inhibitors onto a camel heavy chain antibody scaffold. These sequences were designed to cap fibrils of tau, known to form the neurofibrillary tangles of AD, thereby preventing fibril elongation. The nanobodies grafted with capping inhibitors blocked tau aggregation in biosensor cells seeded with postmortem brain extracts from AD and progressive supranuclear palsy (PSP) patients. The tau capping nanobody inhibitors also blocked seeding by recombinant tau oligomers. Another challenge to the design of effective antibodies is their poor blood-brain barrier (BBB) penetration. In this study, we also designed a bispecific nanobody composed of a nanobody that targets a receptor on the BBB and a tau capping nanobody inhibitor, conjoined by a flexible linker. We provide evidence that the bispecific nanobody improved BBB penetration over the tau capping inhibitor alone after intravenous administration in mice. Our results suggest that the design of synthetic antibodies that target sequences that drive protein aggregation may be a promising approach to inhibit the prion-like seeding of tau and other proteins involved in AD and related proteinopathies.

A heterologous expression platform in Aspergillus nidulans for the elucidation of cryptic secondary metabolism biosynthetic gene clusters: discovery of the Aspergillus fumigatus sartorypyrone biosynthetic pathway.

Aspergillus fumigatus is a serious human pathogen causing life-threatening Aspergillosis in immunocompromised patients. Secondary metabolites (SMs) play an important role in pathogenesis, but the products of many SM biosynthetic gene clusters (BGCs) remain unknown. In this study, we have developed a heterologous expression platform in Aspergillus nidulans, using a newly created genetic dereplication strain, to express a previously unknown BGC from A. fumigatus and determine its products. The BGC produces sartorypyrones, and we have named it the spy BGC. Analysis of targeted gene deletions by HRESIMS, NMR, and microcrystal electron diffraction (MicroED) enabled us to identify 12 products from the spy BGC. Seven of the compounds have not been isolated previously. We also individually expressed the polyketide synthase (PKS) gene spyA and demonstrated that it produces the polyketide triacetic acid lactone (TAL), a potentially important biorenewable platform chemical. Our data have allowed us to propose a biosynthetic pathway for sartorypyrones and related natural products. This work highlights the potential of using the A. nidulans heterologous expression platform to uncover cryptic BGCs from A. fumigatus and other species, despite the complexity of their secondary metabolomes.

Selective inhibition of hsp90 paralogs: Structure and binding studies uncover the role of helix 1 in Grp94-selective ligand binding.

Grp94 is the endoplasmic reticulum paralog of the hsp90 family of chaperones, which have been targeted for therapeutic intervention via their highly conserved ATP binding sites. The design of paralog-selective inhibitors relies on understanding the structural elements that mediate each paralog's response to inhibitor binding. Here, we determined the structures of Grp94 and Hsp90 in complex with the Grp94-selective inhibitor PU-H36, and of Grp94 with the non-selective inhibitor PU-H71. In Grp94, the 8-aryl moiety of PU-H36 is inserted into Site 2, a conditionally available side pocket, but in Hsp90 it occupies Site 1, a non-selective side pocket that is accessible in all hsp90 paralogs. The structure of Grp94 in complex with the non-selective PU-H71 shows only Site 1 binding. Large conformational shifts involving helices 1, 4 and 5 of the N-terminal domain of Grp94 are associated with the engagement of the Site 2 pocket for ligand binding. To understand the origins of Site 2 pocket engagement, we tested the binding of Grp94-selective ligands to chimeric Grp94/Hsp90 constructs. These studies show that helix 1 of the Grp94 N-terminal domain is the discriminating element that allows for remodeling of the ATP binding pocket and exposure of the Site 2 selective pocket.

A heterogeneously integrated lithium niobate-on-silicon nitride photonic platform.

The availability of thin-film lithium niobate on insulator (LNOI) and advances in processing have led to the emergence of fully integrated LiNbO3 electro-optic devices. Yet to date, LiNbO3 photonic integrated circuits have mostly been fabricated using non-standard etching techniques and partially etched waveguides, that lack the reproducibility achieved in silicon photonics. Widespread application of thin-film LiNbO3 requires a reliable solution with precise lithographic control. Here we demonstrate a heterogeneously integrated LiNbO3 photonic platform employing wafer-scale bonding of thin-film LiNbO3 to silicon nitride (Si3N4) photonic integrated circuits. The platform maintains the low propagation loss (<0.1 dB/cm) and efficient fiber-to-chip coupling (<2.5 dB per facet) of the Si3N4 waveguides and provides a link between passive Si3N4 circuits and electro-optic components with adiabatic mode converters experiencing insertion losses below 0.1 dB. Using this approach we demonstrate several key applications, thus providing a scalable, foundry-ready solution to complex LiNbO3 integrated photonic circuits.

Low complexity domains of the nucleocapsid protein of SARS-CoV-2 form amyloid fibrils.

The self-assembly of the Nucleocapsid protein (NCAP) of SARS-CoV-2 is crucial for its function. Computational analysis of the amino acid sequence of NCAP reveals low-complexity domains (LCDs) akin to LCDs in other proteins known to self-assemble as phase separation droplets and amyloid fibrils. Previous reports have described NCAP's propensity to phase-separate. Here we show that the central LCD of NCAP is capable of both, phase separation and amyloid formation. Within this central LCD we identified three adhesive segments and determined the atomic structure of the fibrils formed by each. Those structures guided the design of G12, a peptide that interferes with the self-assembly of NCAP and demonstrates antiviral activity in SARS-CoV-2 infected cells. Our work, therefore, demonstrates the amyloid form of the central LCD of NCAP and suggests that amyloidogenic segments of NCAP could be targeted for drug development.

Ultrafast tunable lasers using lithium niobate integrated photonics.

Early works1 and recent advances in thin-film lithium niobate (LiNbO3) on insulator have enabled low-loss photonic integrated circuits2,3, modulators with improved half-wave voltage4,5, electro-optic frequency combs6 and on-chip electro-optic devices, with applications ranging from microwave photonics to microwave-to-optical quantum interfaces7. Although recent advances have demonstrated tunable integrated lasers based on LiNbO3 (refs. 8,9), the full potential of this platform to demonstrate frequency-agile, narrow-linewidth integrated lasers has not been achieved. Here we report such a laser with a fast tuning rate based on a hybrid silicon nitride (Si3N4)-LiNbO3 photonic platform and demonstrate its use for coherent laser ranging. Our platform is based on heterogeneous integration of ultralow-loss Si3N4 photonic integrated circuits with thin-film LiNbO3 through direct bonding at the wafer level, in contrast to previously demonstrated chiplet-level integration10, featuring low propagation loss of 8.5 decibels per metre, enabling narrow-linewidth lasing (intrinsic linewidth of 3 kilohertz) by self-injection locking to a laser diode. The hybrid mode of the resonator allows electro-optic laser frequency tuning at a speed of 12 × 1015 hertz per second with high linearity and low hysteresis while retaining the narrow linewidth. Using a hybrid integrated laser, we perform a proof-of-concept coherent optical ranging (FMCW LiDAR) experiment. Endowing Si3N4 photonic integrated circuits with LiNbO3 creates a platform that combines the individual advantages of thin-film LiNbO3 with those of Si3N4, which show precise lithographic control, mature manufacturing and ultralow loss11,12.

Small molecules disaggregate alpha-synuclein and prevent seeding from patient brain-derived fibrils.

The amyloid aggregation of alpha-synuclein within the brain is associated with the pathogenesis of Parkinson's disease (PD) and other related synucleinopathies, including multiple system atrophy (MSA). Alpha-synuclein aggregates are a major therapeutic target for treatment of these diseases. We identify two small molecules capable of disassembling preformed alpha-synuclein fibrils. The compounds, termed CNS-11 and CNS-11g, disaggregate recombinant alpha-synuclein fibrils in vitro, prevent the intracellular seeded aggregation of alpha-synuclein fibrils, and mitigate alpha-synuclein fibril cytotoxicity in neuronal cells. Furthermore, we demonstrate that both compounds disassemble fibrils extracted from MSA patient brains and prevent their intracellular seeding. They also reduce in vivo alpha-synuclein aggregates in C. elegans. Both compounds also penetrate brain tissue in mice. A molecular dynamics-based computational model suggests the compounds may exert their disaggregating effects on the N terminus of the fibril core. These compounds appear to be promising therapeutic leads for targeting alpha-synuclein for the treatment of synucleinopathies.

Structure-based discovery of small molecules that disaggregate Alzheimer's disease tissue derived tau fibrils in vitro.

Alzheimer's disease (AD) is the consequence of neuronal death and brain atrophy associated with the aggregation of protein tau into fibrils. Thus disaggregation of tau fibrils could be a therapeutic approach to AD. The small molecule EGCG, abundant in green tea, has long been known to disaggregate tau and other amyloid fibrils, but EGCG has poor drug-like properties, failing to fully penetrate the brain. Here we have cryogenically trapped an intermediate of brain-extracted tau fibrils on the kinetic pathway to EGCG-induced disaggregation and have determined its cryoEM structure. The structure reveals that EGCG molecules stack in polar clefts between the paired helical protofilaments that pathologically define AD. Treating the EGCG binding position as a pharmacophore, we computationally screened thousands of drug-like compounds for compatibility for the pharmacophore, discovering several that experimentally disaggregate brain-derived tau fibrils in vitro. This work suggests the potential of structure-based, small-molecule drug discovery for amyloid diseases.