Interfering with aggregated α-synuclein in advanced melanoma leads to a major upregulation of MHC class II proteins.

Melanoma is the most serious and deadly form of skin cancer and with progression to advanced melanoma, the intrinsically disordered protein α-synuclein is upregulated to high levels. While toxic to dopaminergic neurons in Parkinson's disease, α-synuclein is highly beneficial for primary and metastatic melanoma cells. To gain detailed insights into this exact opposite role of α-synuclein in advanced melanoma, we performed proteomic studies of high-level α-synuclein-expressing human melanoma cell lines that were treated with the diphenyl-pyrazole small-molecule compound anle138b, which binds to and interferes with the oligomeric structure of α-synuclein. We also performed proteomic and transcriptomic studies of human melanoma xenografts that were treated systemically with the anle138b compound. The results reveal that interfering with oligomerized α-synuclein in the melanoma cells in these tumor xenografts led to a substantial upregulation and expression of major histocompatibility complex proteins, which are pertinent to enhancing anti-melanoma immune responses.

First Satellite Tagging of the Northern Fur Seals (Callorhinus ursinus) on the Tyuleniy Island, the Sea of Okhotsk.

In October 2018, three Northern fur seals (two adult females and one juvenile male) were deployed with satellite tags on the Tyuleniy Island in the Sea of Okhotsk. The operational time of the tags ranged from 33 to 203 days. The adult females started their winter migration in the first half of November; the initial stage of their winter migration occurred in the Japan/East Sea, which they entered through the La Perouse Strait. The juvenile male left the rookery in mid-October, crossed the Sea of Okhotsk in a north-western direction and returned to the south. The male had reached the coastal areas of Hokkaido Island, Japan by the end of November. From the Sea of Okhotsk, the male entered the Pacific Ocean through the Yekaterina Strait and subsequently entered the Japan/East Sea via the Tsugaru Strait. The winter foraging of the male occurred within the north-eastern part of the Japan/East Sea just off the Tsugaru Strait. After 3 months, the male returned to the Pacific and remained off the Sanriku Coast (Honshu Island).

Anle138b interaction in α-synuclein aggregates by dynamic nuclear polarization NMR.

Small molecules that bind to oligomeric protein species such as membrane proteins and fibrils are of clinical interest for development of therapeutics and diagnostics. Definition of the binding site at atomic resolution via NMR is often challenging due to low binding stoichiometry of the small molecule. For fibrils and aggregation intermediates grown in the presence of lipids, we report atomic-resolution contacts to the small molecule at sub nm distance via solid-state NMR using dynamic nuclear polarization (DNP) and orthogonally labelled samples of the protein and the small molecule. We apply this approach to α-synuclein (αS) aggregates in complex with the small molecule anle138b, which is a clinical drug candidate for disease modifying therapy. The small central pyrazole moiety of anle138b is detected in close proximity to the protein backbone and differences in the contacts between fibrils and early intermediates are observed. For intermediate species, the 100 K condition for DNP helps to preserve the aggregation state, while for both fibrils and oligomers, the DNP enhancement is essential to obtain sufficient sensitivity.

Mercury content in the fur of sea otters (Enhydra lutris) from the Commander Islands.

The sea otter (Enhydra lutris) is a keystone species in the ecosystem which is currently in depression in Russia. The objectives of this study were to: (1) establish if the sea otters from the Commander Islands have hazardous levels of mercury (Hg) in their fur; (2) assess Hg pollution in sea otters during a period of high abundance and population depression; (3) identify the age and sex differences in sea otters by Hg content. The sea otters were classified from no to low risk for Hg health effects. Differences in Hg content during periods of low and high population size were not statistically significant. Hg concentrations in adult sea otters were significantly higher than in the young, and higher in males than in females. This study presents the first data on Hg content in sea otters' fur and the first estimate of Hg contamination for the Commander Islands population.

The clinical drug candidate anle138b binds in a cavity of lipidic α-synuclein fibrils.

Aggregation of amyloidogenic proteins is a characteristic of multiple neurodegenerative diseases. Atomic resolution of small molecule binding to such pathological protein aggregates is of interest for the development of therapeutics and diagnostics. Here we investigate the interaction between α-synuclein fibrils and anle138b, a clinical drug candidate for disease modifying therapy in neurodegeneration and a promising scaffold for positron emission tomography tracer design. We used nuclear magnetic resonance spectroscopy and the cryogenic electron microscopy structure of α-synuclein fibrils grown in the presence of lipids to locate anle138b within a cavity formed between two ß-strands. We explored and quantified multiple binding modes of the compound in detail using molecular dynamics simulations. Our results reveal stable polar interactions between anle138b and backbone moieties inside the tubular cavity of the fibrils. Such cavities are common in other fibril structures as well.

Safety, tolerability and pharmacokinetics of the oligomer modulator anle138b with exposure levels sufficient for therapeutic efficacy in a murine Parkinson model: A randomised, double-blind, placebo-controlled phase 1a trial.

BACKGROUND: Synucleinopathies such as Parkinson ́s disease (PD), Dementia with Lewy bodies (DLB) and Multiple System Atrophy (MSA) are characterized by deposition of misfolded and aggregated α-synuclein. Small aggregates (oligomers) of α-synuclein have been shown to be the most relevant neurotoxic species and are targeted by anle138b, an orally bioavailable small molecule compound which shows strong disease-modifying effects in animal models of synucleinopathies. METHODS: Anle138b was studied in a single-centre, double-blind, randomised, placebo-controlled single ascending dose (SAD) and multiple ascending dose (MAD) study in healthy subjects. Eligible participants were randomly assigned (1:1 for sentinel subjects and 1:5 for main group) to placebo or anle138b (dose range 50 mg to 300 mg per day), respectively. In addition, the effect of food on the pharmakokinetics of anle138b in healthy subjects was examined in doses of 150 mg per day. Participants were randomized to treatment sequence (fed→fasted) or (fasted→fed). Treatment was administered orally in hard gelatine capsules containing either 10 mg or 30 mg of anle138b or excipient only. The primary endpoints were safety and tolerability, the secondary endpoint was pharmakokinetics. Data from all randomized individuals were evaluated. CLINICALTRIALS: gov-identifier: NCT04208152. EudraCT-number: 2019-004218-33. FINDINGS: Between December 17th, 2019 and June 27th, 2020 196 healthy volunteers were screened and 68 participants were enrolled. Of these, all completed the study per protocol. There were no major protocol deviations. Adverse events in this healthy volunteer trial were mostly mild and all fully recovered or resolved prior to discharge. From baseline to completion of the trial no medically significant individual changes were observed in any system organ class. Already at multiple doses of 200 mg, exposure levels above the fully effective exposure in the MI2 mouse Parkinson model were observed. INTERPRETATION: The favourable safety and PK profile of anle138b in doses resulting in exposures above the fully effective plasma level in a mouse Parkinson model warrant further clinical trials in patients with synucleinopathies. FUNDING: This study was funded by MODAG GmbH and by the Michael J. Fox foundation for Parkinson's Research.

Correction: Migrations of young spotted seals (Phoca largha) from Peter the Great Bay, Sea of Japan/East Sea, and the pattern of their use of seasonal habitats.

[This corrects the article DOI: 10.1371/journal.pone.0244232.].

Steller Sea Lion (Eumetopias jubatus) at Tuleny Island, Russia in 2018: Abundance, Composition, and Entanglement.

The Steller sea lion (SSL, Eumetopias jubatus) inhabits the North Pacific Ocean off both the North American and Asian coasts. The abundance of the species in Asia declined by more than half in the second part of the 20th century. Decline recurred in the second decade of the 21th century after a short period of restoration. In contrast with the total dynamics of SSL in Asia, the reproductive aggregation on Tuleny I. (Sea of Okhotsk) has been growing almost continuously since the beginning of its formation in the late 1980s. Long-term monitoring of SSL at Tuleny I. always covered only summer reproductive seasons. We surveyed Tuleny I. in October 2018, and counted 1058 non-pup sea lions and 396 pups. The majority of tagged animals encountered at the rookery were of local origin. About one-third of the summer non-pup sea lions' number could remain at the rookery until the middle of October, which coincides with the seasonal appearance of sea lions off the coast of Japan. The sex-age structure was characterized by total absence of adult males and reduced proportion of subadult males and juveniles. It contrasts with autumn behavior of SSLs in the rookery of the northern Sea of Okhotsk. We observed that 0.7% of the inspected sea lions were entangled in marine debris. The proportion of entangled animals is lower in comparison with that of the whole population of Tuleny I. due to reduction in the ratios of males and juveniles, which entangle in foreign objects more often than adult females.

Migrations of young spotted seals (Phoca largha) from Peter the Great Bay, Sea of Japan/East Sea, and the pattern of their use of seasonal habitats.

We studied the migrations of young spotted seals during their annual cycle. In May 2017, we attached satellite tags (SPOT-293A) to three individuals (two underyearlings and one yearling) captured at their breeding ground in Peter the Great Bay, western Sea of Japan/East Sea. The operational time of the installed tags ranged from 207 to 333 days; a total of 27195 locations were uploaded. All three seals migrated east and further north along the coast of the mainland. The average daily migration speed of the seals ranged between 70 and 135 km/day. The yearling moved faster than the underyearlings. During early August, they arrived at their summer habitats, which were located in the northern part of the Tatar Strait (Sea of Japan/East Sea) for the underyearling seals and in Aniva Bay (Sea of Okhotsk) for the yearling seal. While moving from the place of tagging to the summer feeding grounds, the seals covered a distance of 2300 to 3100 km. From August to October, each seal permanently stayed within the same isolated area. The reverse migration of all three seals began in November. When the seals traveled south, they used the same routes by which they had moved north in the spring, but they moved at a faster speed. By December, two seals returned to their natal islands, where both stayed until their transmitters stopped sending signals (in March 2018).

[11C]MODAG-001-towards a PET tracer targeting α-synuclein aggregates.

PURPOSE: Deposition of misfolded alpha-synuclein (αSYN) aggregates in the human brain is one of the major hallmarks of synucleinopathies. However, a target-specific tracer to detect pathological aggregates of αSYN remains lacking. Here, we report the development of a positron emission tomography (PET) tracer based on anle138b, a compound shown to have therapeutic activity in animal models of neurodegenerative diseases. METHODS: Specificity and selectivity of [3H]MODAG-001 were tested in in vitro binding assays using recombinant fibrils. After carbon-11 radiolabeling, the pharmacokinetic and metabolic profile was determined in mice. Specific binding was quantified in rats, inoculated with αSYN fibrils and using in vitro autoradiography in human brain sections of Lewy body dementia (LBD) cases provided by the Neurobiobank Munich (NBM). RESULTS: [3H]MODAG-001 revealed a very high affinity towards pure αSYN fibrils (Kd = 0.6 ± 0.1 nM) and only a moderate affinity to hTau46 fibrils (Kd = 19 ± 6.4 nM) as well as amyloid-ß1-42 fibrils (Kd = 20 ± 10 nM). [11C]MODAG-001 showed an excellent ability to penetrate the mouse brain. Metabolic degradation was present, but the stability of the parent compound improved after selective deuteration of the precursor. (d3)-[11C]MODAG-001 binding was confirmed in fibril-inoculated rat striata using in vivo PET imaging. In vitro autoradiography showed no detectable binding to aggregated αSYN in human brain sections of LBD cases, most likely, because of the low abundance of aggregated αSYN against background protein. CONCLUSION: MODAG-001 provides a promising lead structure for future compound development as it combines a high affinity and good selectivity in fibril-binding assays with suitable pharmacokinetics and biodistribution properties.