TAU x α-synuclein

A protein called alpha-synuclein that accumulates and ravages the brains of some people with Parkinson’s disease is the target of several experimental drugs. Until recently, the only way to detect it was by opening someone’s brain when they died. That’s starting to change. Amprion, a 30-person biotech based in San Diego, has developed a way to detect small amounts of alpha-synuclein from spinal fluid. A study published last year in The Lancet Neurology suggests the method could help confirm suspected diagnoses and even predict Parkinson’s before common symptoms arise. And now Amprion is scaling up to commercialize the test. CEO and co-founder Russell Lebovitz told Endpoints News in an exclusive interview that Amprion has raised $6 million in the first portion of its Series B, which will help it hire 15 more employees and expand its salesforce and operations throughout the US. The round was led by Formation Venture Engineering, with participation from Eli Lilly. Lilly’s interest is notable because the pharma company helped pioneer the use of diagnostics to detect amyloid in the brains of people with Alzheimer’s disease. The drugmaker is also developing drugs for Parkinson’s, and the company’s scientific leaders have mentioned alpha-synuclein as an area of interest for future neurodegenerative research at the company. “They have recognized, as have a lot of the drug companies, that this is going to be important in the future,” Lebovitz said. The technology underlying Amprion’s test was forged in the early 2000s in the aftermath of the mad cow disease crisis. Claudio Soto, a biochemist and professor of neurology at the UT Health Houston McGovern Medical School in Houston, developed a way to detect prions, the misfolded proteins that cause bovine spongiform encephalopathy in cattle and Creutzfeldt-Jakob disease in humans. The infectious proteins cause natural versions of the proteins to misfold, leaving tainted and toxic prions in their wake as they spread through the brain. That natural amplification formed the basis of Soto’s test. He added highly purified and properly folded proteins to spinal fluid or blood from a patient suspected of having the disease. If prions were present in the sample, they would quickly convert the normal proteins into diseased ones. He compared the method to PCR, the technique used to make many copies of DNA for easier sequencing. Soto formed a startup with Lebovitz in 2007 that aimed to refine the technique, now dubbed a seed amplification assay. The mad cow crisis abated, but the duo believed the technique would be valuable to help detect other misfolded proteins, such as amyloid and tau in Alzheimer’s and alpha-synuclein in Parkinson’s. The first ten years were slow going. The test originally took nearly two weeks to run, but they got it down to 20 hours. And a fluorescent dye that only lights up in the presence of misfolded proteins made the results easy to interpret. The tests are currently run out of its CLIA-certified lab in San Diego. The company is developing a simplified kit of the test that other labs can run, and hopes to submit it to the FDA in about 18 months. By 2019, Amprion had developed an alpha-synuclein test for Parkinson’s that it believed would be scalable. The company raised a $13.7 million Series A over the next three years and began offering the test to doctors at major medical centers and research institutions in 2023, Lebovitz said. The study published last year revealed that the test had a sensitivity of about 88%, meaning that about 12 in 100 people with Parkinson’s had a false negative result. Among the 1,100 healthy volunteers, the test had a 97% specificity, meaning about 3 in 100 people without the disease had a false positive. The test performed more poorly for certain subsets of Parkinson’s disease, including those with mutations in the LRRK2 gene linked to the disease. Drugmakers and regulators alike are increasingly turning to biomarkers to decide which patients to include in a clinical trial and how to assess whether the drug is working. In August, the FDA issued a letter of support encouraging “further study and use of” seed amplification assays for alpha-synuclein. Amprion has worked on similar amplification assays for amyloid and tau, but these tests are still experimental. The company offers clinical tests for amyloid, tau and another brain biomarker called neurofilament using machines sold by Fujirebio. With the new funding, Leboviz said “we can hit every major medical center doing cognitive and motor disorders in the US.” He’s hoping to raise another $9 million in the Series B round this year to help the company open a lab in Europe. But Amprion’s efforts to scale up come at a challenging time. About two weeks ago, an investigation published in Science cast doubt on several research papers supporting the role of alpha-synuclein in Parkinson’s disease. More than 130 papers published by National Institutes of Health neuroscientist Eliezer Masliah had “suspect images,” including doctored and duplicated results, according to the report. Some of the questionable studies were cited as evidence underpinning the alpha-synuclein-targeting antibody prasinezumab, which is being developed by Prothena Biosciences and Roche. Lebovitz said that other scientists have consistently demonstrated alpha-synuclein’s role in Parkinson’s and other neurodegenerative diseases irrespective of Masliah’s studies. “None of Amprion’s data relies on any work ever performed or published by Eliezer Masliah,” Lebovitz said. “We have never met with Dr. Masliah, and he has never provided any advice or other counsel.”

GT-02287 Demonstrates Disease Modifying Capacity in Both GBA1 and Idiopathic Parkinson’s Disease Models GT-02287 Improves Mitochondrial Function and Provides Neuroprotective Effect in GBA1-Parkinson’s Disease Models GT-02287 Prevents Tau Accumulation in a Cellular Model BETHESDA, Md., Oct. 07, 2024 (GLOBE NEWSWIRE) -- Gain Therapeutics, Inc. (Nasdaq: GANX) (“Gain”, or the “Company”), a clinical-stage biotechnology company leading the discovery and development of the next generation of allosteric small molecule therapies, today announced the presentation of new evidence supporting the disease-modifying activity of GT-02287 in preclinical animal models of both GBA-1 and idiopathic Parkinson’s disease during a late-breaker poster session yesterday at the Society for Neuroscience (SfN) 2024 conference that is being held October 5-9 in Chicago, IL. In animal models of GBA-1 and idiopathic Parkinson’s disease, rescue of motor and cognitive function associated with administration of GT-02287 was observed to be persistent upon wash out of GT-02287 without significant effect in performance for more than a week following discontinuation when compared to the animals in which treatment was continued. Another presentation for GT-02287 yesterday described its association with improved mitochondrial function and neuroprotection in GBA1-Parkinson’s disease models. Finally, a second late-breaker was presented describing the reduction in Tau accumulation associated with GT-02287 in both GBA-1 mutation and wild type cell lines, suggesting GT-02287 may have broader potential as a treatment for tauopathies such as Alzheimer’s disease. “The robust preclinical development dossier for GT-02287 presented further strengthen our belief that GT-02287 has the potential to be a disease-modifying therapy for people suffering from Parkinson’s disease. The data demonstrate rescue of motor deficit and prevention of development of deficits in cognition which persisted even once the compound was removed. Moreover, GT-02287 demonstrated a positive effect on mitochondrial health as well as on lysosomal health which further demonstrates that the molecule acts throughout the disease cascade resulting from dysfunction of the lysosomal enzyme glucocerebrosidase (GCase), which GT-02287 was designed to target. Additionally, GT-02287 prevented Tau accumulation in human cellular models which demonstrates its potential to be a treatment for Alzheimer’s disease and other tauopathies,” commented Joanne Taylor, Ph.D., Senior Vice President of Research at Gain “The preservation of motor and cognitive improvement in animal models of Parkinson’s disease associated with GT-02287 after its discontinuation coupled with the neuroprotective effects after delayed administration support the potential of GT-02287 to slow or stop the progression of Parkinson’s disease,” said Gene Mack, Interim CEO and CFO of Gain Therapeutics. “The potential of GCase as a therapeutic target beyond Parkinson’s Disease is an encouraging finding that we can incorporate into our future development plans for GT-02287.” The late-breaking poster titled, “GT-02287, a clinical stage GCase regulator, demonstrates disease modifying capacity in both GBA1 and idiopathic Parkinson’s disease models,” was presented on-site October 6 at 3pm CT. In animal models of both GBA1 and idiopathic Parkinson’s disease, GT-02287 was administered after a clinical phenotype was established and then withdrawn from half the animals for more than one week prior to conclusion of the study. Neuromuscular function, motor coordination, and activities of daily living/cognition were measured in wire hang, beam walk, and nest building tests, respectively, throughout the study. GT-02287 was shown to rescue deficits in neuromuscular function and motor coordination in both models as well as to prevent the development of deficits in cognition and activities of daily living. Notably, withdrawal of GT-02287 for more than one week did not significantly affect performance in any of the tests, suggesting a disease modifying effect. The poster titled, “GT-02287, a clinical stage GCase regulator, improves mitochondrial function and provides a neuroprotective effect in GBA1-Parkinson's disease models,” was presented on-site October 6 at 10am CT. In an in vitro model - in which cultured rat mesencephalic dopaminergic neurons were treated with conduritol beta epoxide (CBE) to cause partial knockdown of GCase activity comparable to that seen in PD patients carrying heterozygous GBA1 mutations (GBA1 is the gene encoding GCase)- treatment with GT-02287 reduced the level of mitochondrial reactive oxygen species (ROS) as well as ameliorating lysosomal pathology, reducing α-synuclein aggregation, and providing a neuroprotective effect. In an in vivo model in which mice were subjected to toxic insult by CBE and injection of α-synuclein preformed fibrils (PFFs) into the striatum – delayed administration of GT-02287 reduced levels of mitochondrial protein Miro1, an important maker for mitophagy; aggregated α-synuclein and plasma neurofilament light chain (NfL), a marker of neurodegeneration; as well as completely restoring motor function to control levels. The late-breaking poster titled, “GT-02287, a GCase modulator and Gain Therapeutics’ PD drug candidate prevents Tau accumulation in a cellular model,” was presented on-site October 6 at 1pm CT. Lysosomal Tau accumulation was quantified in human fibroblasts, carrying either the L444P mutant or wild-type (WT) GBA engineered to express labelled Tau protein, at basal conditions as well in the presence of Alzheimer’s brain-derived Tau seeds. Basal and seed-induced Tau accumulation was significantly higher in Tau-GBA L444P cells, which express defective GCase, compared to Tau WT GBA cells. Treatment with GT-02287 reduced Tau accumulation in both cells carrying the L444P GBA mutation and those carrying wild-type GBA, demonstrating GT-02287’s potential as a disease-modifying treatment for Alzheimer’s disease and other tauopathies. This work was carried out in collaboration with scientists in the group of Professor Paolo Paganetti at the Laboratories for Translational Research of the Ente Ospedaliero Cantonale (LRT-EOC) in Bellinzona, Switzerland. This study was conducted under the support of an Innosuisse-Swiss Innovation Agency grant with the LRT-EOC and the Università della Svizzera italiana-affiliated Institute for Research in Biomedicine in Bellinzona, Switzerland. PDFs of the posters presented at Neuroscience 2024 are available on the Science and Technology section of the Company’s website at https://gaintherapeutics.com/science-and-technology/posters. About GT-02287Gain Therapeutics’ lead drug candidate, GT-02287, is in clinical development for the treatment of Parkinson’s disease with or without a GBA1 mutation. The orally administered, brain-penetrant small molecule is an allosteric protein modulator that restores the function of the lysosomal protein enzyme glucocerebrosidase (GCase) which becomes misfolded and impaired due to mutations in the GBA1 gene, the most common genetic abnormality associated with PD, or other age-related stress factors. In preclinical models of PD, GT-02287 restored GCase enzymatic function, reduced aggregated α-synuclein, neuroinflammation and neuronal death, increased dopamine levels and improved motor function and cognitive performance. Additionally, GT-02287 significantly reduced plasma neurofilament light chain (NfL) levels, an emerging biomarker for neurodegeneration. Compelling preclinical data in mouse models of GBA1-PD, including that presented at FENS Forum 2024 in June describing improvement in cognitive performance in addition to motor performance after administration of GT-02287, suggests that GT-02287 may have the potential to slow the progression of Parkinson’s disease. Gain’s lead program in Parkinson’s disease has been awarded funding support from The Michael J. Fox Foundation for Parkinson’s Research (MJFF) and The Silverstein Foundation for Parkinson’s with GBA, as well as from the Eurostars-2 joint program with co-funding from the European Union Horizon 2020 research and Innosuisse – Swiss Innovation Agency. About Gain Therapeutics, Inc.Gain Therapeutics, Inc. is a clinical-stage biotechnology company leading the discovery and development of next generation allosteric therapies. Gain’s lead drug candidate, GT-02287 is currently being evaluated for the treatment of Parkinson’s disease with or without a GBA1 mutation. Results from a Phase 1 study of GT-02287 in healthy volunteers demonstrated favorable safety and tolerability, plasma exposure in the projected therapeutic range, CNS exposure, and target engagement and modulation of GCase enzyme. Gain’s unique approach enables the discovery of novel, allosteric small molecule modulators that can restore or disrupt protein function. Deploying its highly advanced Magellan™ platform, Gain is accelerating drug discovery and unlocking novel disease-modifying treatments for untreatable or difficult-to-treat disorders including neurodegenerative diseases, rare genetic disorders and oncology. Forward-Looking StatementsThis release contains “forward-looking statements” made pursuant to the safe harbor provisions of the Private Securities Litigation Reform Act of 1995. These statements are typically preceded by words such as “believes,” “expects,” “anticipates,” “intends,” “will,” “may,” “should,” or similar expressions. These forward-looking statements reflect management’s current knowledge, assumptions, judgment and expectations regarding future performance or events. Although management believes that the expectations reflected in such statements are reasonable, they give no assurance that such expectations will prove to be correct or that those goals will be achieved, and you should be aware that actual results could differ materially from those contained in the forward-looking statements. Forward-looking statements are subject to a number of risks and uncertainties, including, but not limited to, risks associated with market conditions and the satisfaction of customary closing conditions related to the offering and uncertainties related to the offerings and the use of proceeds from the offerings. For a further description of the risks and uncertainties that could cause actual results to differ from those expressed in these forward-looking statements, as well as risks relating to the Company’s business in general, please refer to the Company’s prospectus supplement to be filed with the SEC, and the documents incorporated by reference therein, including the Company’s Form 10-K for the year ended December 31, 2023 and Form 10-Q for the quarter ended June 30, 2024. All forward-looking statements are expressly qualified in their entirety by this cautionary notice. You are cautioned not to place undue reliance on any forward-looking statements, which speak only as of the date of this release. We have no obligation, and expressly disclaim any obligation, to update, revise or correct any of the forward-looking statements, whether as a result of new information, future events or otherwise. Investor Contacts:Apaar Jammu and Chuck Padalaajammu@gaintherapeutics.comchuck@lifesciadvisors.com Media Contacts:Russo PartnersNic Johnson and Elio Ambrosionic.johnson@russopartnersllc.comelio.ambrosio@russopartnersllc.com(760) 846-9256

*仅供医学专业人士阅读参考神经退行性疾病的重要标志之一，就是病理性蛋白质聚集体积累。α-突触核蛋白（α-syn）和tau蛋白聚集体均表现出“类朊病毒”特性，在细胞间扩散并诱导临近神经元的蛋白质错误折叠，从而在相连脑区传播神经元损伤。小胶质细胞是大脑先天免疫系统的一部分，在清除病理性蛋白质聚集体方面发挥着关键作用。人们认为，神经元细胞死亡后将病理性蛋白质聚集体释放到细胞外，之后才会被小胶质细胞识别并清除。但是，来自卢森堡大学的研究团队发现，小胶质细胞可以通过隧道纳米管（TNT）与远距离的神经元建立连接，并通过TNT进行细胞器和蛋白质的快速交换。病理条件下，小胶质细胞通过TNT将神经元中的毒性蛋白质聚集体转移，并为神经元输送健康线粒体，从而有效减轻神经元的病理负担，恢复神经元功能。研究发表在《神经元》杂志上。α-syn或tau聚集体可谓是让神经元吃尽了苦头。RNA测序显示，共有951个基因表达水平在毒性蛋白质聚集体存在时发生变化，尤其是线粒体功能相关基因，氧化应激水平增加，神经元功能受损甚至死亡。在正常情况下，皮质神经元和小胶质细胞共培养，可以在细胞之间形成TNT，不同于小胶质细胞的其他伪足，TNT是自由悬浮状态，并且可以促进连接细胞之间的细胞成分交换。在病理条件下，毒性蛋白质聚集体积累使悬浮TNT数量增加，这些TNT中含有α-syn和tau免疫反应颗粒，表示正在进行毒性蛋白质的物质交换。α-syn聚集体通过TNT从神经元转移到小胶质细胞与单独培养的含有α-syn或tau的神经元相比，与小胶质细胞共培养可以显著降低神经元中的毒性蛋白质水平。那么，毒性蛋白质被转移走之后的神经元，恢复健康了吗？α-syn或tau积累与神经元网络活动下降或神经损伤有关。研究人员评估了神经元的Ca2+振荡，发现当小胶质细胞与神经元发生物理接触之后，原本α-syn诱导降低的Ca2+振荡频率重新升高。小胶质细胞通过输送线粒体支持α-syn负担的神经元不仅如此，TNT还可以将小胶质细胞中的健康线粒体运输至神经元中，弥补α-syn诱导的神经元线粒体破碎，使神经元中的ROS水平降低，有效减少神经元死亡。接下来，研究人员尝试在小鼠和人体中验证TNT的存在。将含有α-syn的皮质神经元移植到小鼠皮质中，虽然可以看到被标记的α-syn从神经元转移至小胶质细胞，但是这条转移道路的特征和研究人员观察到的TNT存在一些出入。无奈之下，研究人员在皮层网络机器智能（MICrONS）联盟数据集筛选了小鼠大脑皮层的连续切片电子显微镜图像，终于发现了TNT样开放式管状连接。在健康捐赠者和痴呆症患者的死后脑组织中，也观察到神经元和小胶质细胞之间存在大量TNT样接触。根据以上实验结果，研究人员已经肯定，小胶质细胞的TNT是一种新型神经保护机制。那么在最需要神经保护的地方，也就是神经退行性疾病中，TNT是否正常发挥作用了呢？在已知的神经退行性疾病相关基因变异中，携带LRRK2突变的小胶质细胞线粒体输送增加，但是对α-syn聚集体的转移减少；携带TREM2突变的小胶质细胞清除tau聚集体的效率下降，线粒体输送未受影响。总体而言，这些遗传变异损害了小胶质细胞对神经元的保护作用。作者表示，“我们知道小胶质细胞在清除蛋白质聚集物方面发挥作用，但是最近才知道它们是通过纳米管远距离作业”，“这项研究不仅加深了我们对TNT进行细胞间通讯的理解，还挑战了小胶质细胞是神经炎症诱因的传统观点，强调了一种新的神经保护机制”。参考文献：Scheiblich H, Eikens F, Wischhof L, et al. Microglia rescue neurons from aggregate-induced neuronal dysfunction and death through tunneling nanotubes[J]. Neuron, 2024, 112(18): 3106-3125. e8.本文作者｜王雪宁