Over the past two decades, the number of clinical trials to modify MSA has increased. Unfortunately, most attempts have failed, possibly due to incomplete understanding of pathophysiology, inadequacies in preclinical animal models, and lack of early and accurate diagnosis of the disease.
Aberrant proteostasis
The impairment of protein processing and degradation has been implicated in the pathogenesis of α-synuclein aggregation based on some preclinical evidence [
148‐
150]. Therefore, enhancing the degradation of α-synuclein has also been adopted as one important treatment strategy for MSA.
Accumulating evidence suggests that the autophagy–lysosomal pathway is altered in MSA. By inhibiting the activity of mammalian target of rapamycin (mTOR), rapamycin (also known as Sirolimus) influenced a variety of essential cellular processes, such as protein synthesis and autophagy [
151,
152]. Rapamycin increased autophagy, decreased α-synuclein aggregation and provided partial neuroprotection in the SNpc of PLP-α-syn transgenic mice (PLP: proteolipid protein), a specific MSA mouse model [
153,
154]. However, a phase 2 trial on oral sirolimus for MSA was failed recently (NCT03589976).
Another degradation enhancer is rifampicin, an antibiotic which has been investigated to inhibit the α-synuclein fibrils and to disaggregate already-formed fibril [
155]. In addition, rifampicin decreased α-synuclein and neurodegeneration in MBP-α-syn transgenic mouse model of MSA [
156]. A large phase 3 trial in patients with early MSA was terminated after an interim analysis of the primary endpoint revealed that futility criteria were met (NCT01287221).
Besides, lithium exerted neuroprotection against rotenone-induced injuries partially through the autophagy pathway in an in vitro PD cell model [
157] and protected dopaminergic neurons likely via autophagy enhancement in an MPTP-administration mouse model of PD [
158], indicating that it may be beneficial in MSA. A phase 2 trial to evaluate efficacy, safety, and tolerability of lithium treatment in patients with MSA was terminated due to severe adverse effect from interim analysis, indicating that lithium treatment was not well-tolerated [
159] (NCT00997672).
Despite the failure of all clinical trials focusing on α-synuclein degradation enhancement to slow down the progression of the disease, several promising preclinical studies are still underway. On the other hand, it has been demonstrated that α-synuclein can be degraded via the autophagy and the ubiquitin–proteasome system (UPS) pathways [
160]. Targeting the proteasome degradation system may be a viable alternative, as suggested by the failure of clinical trials aimed at enhancing the autophagy-lysosomal pathway. Recently, by introducing the proteolysis targeting chimeric (PROTAC) concept and technology, a peptide fusion containing α-synuclein binding domain and a short strong proteasome-targeting motif was able to bind to α-synuclein and direct it to the proteasome for degradation [
161]. Additionally, one of Arvinas Company’s pipelines demonstrated that α-synuclein PROTAC protein degraders could degrade oligomeric forms of α-synuclein in their preclinical studies.
Inhibition of neuroinflammation
There was growing evidence that brain inflammation played a crucial role in the pathogenesis of MSA. In MSA, aggregated α-synuclein induced microglial activation and astrogliosis, stimulated the secretion of proinflammatory cytokines in microglia, and ultimately exacerbated the disease pathology [
58,
162,
163]. Therefore, suppression of microglial activation or inflammation as a whole has been viewed as a potential and promising approach in MSA.
Minocycline was a semi-synthetic, second-generation tetracycline analog which can effectively cross the blood–brain barrier and inhibit the microglial activation [
164,
165]. Notably, minocycline has been shown to provide neuroprotection in experimental models of multiple neurodegenerative diseases, including PD, Alzheimer’s Disease (AD) and HD [
166‐
168]. Contradictory preclinical evidence suggested that minocycline did not prevent lesion-induced neuronal damage in a rat model of striatonigral degeneration, a core pathology that may correlate with MSA, despite its microglial suppression [
169]. In addition, another study found that minocycline inhibited microglial activation, but exacerbated dopaminergic neuronal damage in a mouse model of PD administered with MPTP [
170]. A phase 3 for the evaluation of the efficacy and safety of Minocycline for treatment of MSA was completed (NCT00146809) but failed to demonstrate a clinical effect of minocycline on symptom severity as measured by clinical motor function. However, preliminary PET data suggested that minocycline may inhibit microglial activation [
171].
Another study investigated the intravenous administration of immunoglobulins (IVIG). IVIG is a type of antibody mixture derived from human plasma that is believed to inhibit auto-reactive T-cells and then the production of cytokines. Even though the underlying mechanism remains poorly understood, this is utilized in a variety of immune-mediated neurological diseases. Activation of microglia and production of toxic cytokines suggested a role for neuroinflammation [
172] and the potential therapeutic benefit of IVIG in MSA. A phase 2, open-label pilot clinical study for the efficacy of IVIG had enrolled 9 MSA patients, and the UMSARS scores declined in the majority of this group of patients [
173] (NCT00750867). Despite this, to further confirm the efficacy of IVIG, larger confirmatory trials are still needed.
In addition, myeloperoxidase (MPO), a key enzyme involved in the production of reactive oxygen species by phagocytic cells, contributed to the oxidative stress implicated in the pathogenesis of the neurodegenerative disorders, such as AD, PD, multiple sclerosis, and MSA [
174‐
177]. In the MSA mouse model, it was demonstrated that MPO inhibition reduced motor impairment and rescued vulnerable neurons in the striatum, SNpc, cerebellar cortex, pontine nuclei, and inferior olives, which was accompanied by a reduction in microglial activation and intracellular aggregates of α-synuclein [
177]. In contrast, another research reported that MPO inhibition had no effect on motor impairments and neuronal loss in a mouse model of advanced MSA despite a significant decrease in microglial activation [
178]. Verdiperstat (also known as AZD3241, and BHV-3241) is a potent, selective, and irreversible inhibitor of MPO that suppresses microglial activation that was initially studied by AstraZeneca [
179]. A phase 1 study evaluating the safety of AZD3241 in healthy participants revealed this compound has a good safety profile (NCT01457807). A phase 2 PET study in PD demonstrated that AZD3241 was safe and well tolerated and reported amelioration of microglial activation, supporting proof of the mechanism of AZD3241 and extending further study of AZD3241 in PD or MSA [
180] (NCT01527695). A phase 2 12-week trial of AZD3241 clinical trial in patient with MSA to assess the effect on microglia activation as measured by PET using 11-
CPBR28 tracer, a tracer that binds to the transporter protein (TSPO) in activated glia as primary outcome was completed and no significant changes from baseline, or between groups, were detected (NCT02388295). In 2018, Biohaven licensed AZD3241 from AstraZeneca and further developed by a phase 3 clinical trial of BHV-3241 (referred to AZD3241) in patients with MSA, given the supportive phase 2 exploratory efficacy outcomes. However, this phase 3 study to evaluate the efficacy and safety of BHV-3241 in subjects with MSA was recently completed and failed to meet its primary and key secondary endpoints (NCT03952806). Recently, Biohaven began a small study evaluating the newer TSPO PET ligand 18
FPBR06, before and after Verdiperstat treatment [
181]. This study enrolled 19 MSA, studied the effect of Verdiperstat on microglial activation in well-characterized MSA patients and was completed in January 2022 (NCT04616456).
Neuroprotective therapies
It is well-known that glutamate-induced neurotoxicity plays a crucial role in the neuronal damage and death underlying a broad spectrum of central nervous system disorders. Several glutamate receptor antagonists have been investigated for their neuroprotective effect in CNS disorders. Riluzole, an anti-glutamatergic agent approved by the FDA as a disease-modifying therapy for amyotrophic lateral sclerosis (ALS), is believed to have neuroprotective properties [
182]. Observations of a reduction in behavioral deficits and striatal degeneration in the double lesion rat model of MSA-P administered by Riluzole supported the neuroprotective effect [
183]. Despite promising preclinical results, a placebo-controlled cross-over trial in 10 probable MSA patients showed no significant anti-parkinsonian effects after administrated Riluzole [
184]. A subsequent, large-scale phase 3 trial of Riluzole in MSA (NNIPPS) failed to meet the primary endpoint which is the difference of 36-month survival rate between the placebo group and the treatment group [
185] (NCT00211224). Other neuronal excitability modulators are currently under investigation, such as Tllsh-2910, a specific NMDA receptor modulator. NMDA receptors in the cerebellum have unique properties that distinguish their function and modulation from those in other brain regions [
186]. A phase 3 clinical trial of Tllsh-2910 in patients with MSA is ongoing and its primary endpoint is the improvement rating of ataxia (NCT03901638).
Fas-associated factor 1 (FAF1) is an apoptosis-related Fas-binding protein. It has been reported that FAF1 levels were significantly elevated in PD and were responsible for neuronal cell death [
187,
188]. In addition, FAF1 induced α-synuclein accumulation in dopaminergic neurons [
189]. KM-819 is a novel FAF1 inhibitor and could act as a neuroprotective agent. The effect of KM-819 in dopaminergic neurons of MPTP mouse model of PD was investigated. The study manifested the neurorestorative effect of KM-819 in striatal dopamine neurons of MPTP model via restoring autophagic α-synuclein degradation [
190,
191], implying that KM-819 may have therapeutic potential for synucleinopathies. A phase 1 first-in-human trial to investigate the safety, tolerability, pharmacokinetics, and pharmacodynamics of KM-819 in healthy subjects has been completed and revealed favorable safety, tolerability, and pharmacokinetics results [
192] (NCT03022799). Recently, a phase 2 trial to further evaluate the safety and efficacy of KM-819 as a disease-modifying therapy to slow down the progression of PD was planned [
193].
Lipid and fatty acid homeostases are crucial for neuronal functions in the brain, the second-most lipid-rich organ. A lipidomic analysis of α-synuclein neurotoxicity revealed that stearoyl-CoA desaturase (SCD) is essential for α-syn-induced neurotoxicity and that inhibiting SCD may be a novel therapeutic strategy [
194‐
197]. A phase 1 study of SCD inhibitor YTX-7739 in healthy subjects has been completed and the result showed it was safe and well-tolerated. A following phase 1b safety and biomarker study has been conducted on PD patients, and the result reported no serious safety events, and a reduction in fatty acid desaturation in blood and CSF was observed. However, the FDA has placed a partial clinical hold on multi-dose clinical trials of YTX-7739 for some reason [
198] (Trial NL9172).
A genetic relationship between
COQ2 mutations (
V393A variant), which resulted in decreased production of Coenzyme Q10 (CoQ10) as an electron carrier in the mitochondrial respiratory chain and MSA-C type was exclusively established in Japanese population [
25]. In addition, multiple mitochondrial dysregulations were observed in iPSC-derived dopaminergic neurons from MSA patients [
199,
200]. In a report of a 3-year follow-up of high-dose ubiquinol supplementation in a case of familial MSA with
COQ2 mutations, the clinical rating scale scores remained stable, but mitochondrial oxidative metabolism improved [
201]. A phase 2 multicenter study to evaluate efficacy and safety of high-dose ubiquinol (drug name: MSA-01) supplementation in MSA patients was completed (UMIN000031771). The results showed that high-dose ubiquinol was well-tolerated and led to a significantly smaller decline of UMSARS part 2 score compared with placebo in MSA patients, indicating orally administered ubiquinol have clinical benefits in patients with MSA [
202].
During brain development, it is well-known that growth hormone promotes proliferation of neural precursors, neurogenesis, and gliogenesis, indicating its potential neuroprotective or neurotrophic effects [
203]. A randomized, double-blinded, placebo-controlled pilot study in patients with MSA has been conducted and showed no treatment differences for any efficacy measures, but some trend improvements in terms of UMSARS total score and cardiovascular reflex autonomic testing. Maybe a large-scale trial and higher doses will be required for further studies [
204].
Rasagiline is a monoamine oxidase type B (MAO-B) inhibitor that is approved for the symptomatic treatment of PD [
205] and may have disease-modifying effect for PD, but because of the previous clinical studies have revealed that the treatment outcomes differ when different drug doses had been administered, further phase 3 trials are required [
206] (NCT00256204). A promising preclinical study in the MSA transgenic mouse model with GCI pathology revealed improvement in motor deficits and significant reduction of neuronal loss [
207]. However, a phase 2 trial to evaluate the efficacy, safety, and tolerability of Rasagiline in MSA-P patients failed, measured by UMSARS [
208] (NCT00977665). In addition, another MAO-B inhibitor Safinamide was investigated to measure the treatment-emergent adverse events (TEAE) and serious adverse events (SAE) in MSA-P patients in the phase 2 stage (NCT03753763).
Boosting the levels of neurotrophic factors, such as brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) in the brain, is an integral part of neuroprotective strategies. In animal research on the MSA transgenic mouse model, selective serotonin reuptake inhibitor Fluoxetine has been demonstrated to increase GDNF and BDNF [
209,
210]. However, in a phase 2 trial in MSA patients failed to show improvement [
211] (NCT01146548). Another trial on GDNF is focusing on gene therapy and currently recruiting to assess the incidence of TEAE and SAE within 3 years in a phase 1 trial of AAV2-GDNF in MSA (NCT04680065).
Some reported that the Insulin/IGF-1 signaling pathway contributed to the regulation of neuronal excitability, nerve cell metabolism, and cell survival, thereby supporting their neurotrophic function in a number of neurodegenerative diseases [
212,
213]. Increased insulin and IGF-1 plasma concentrations in MSA patients and decreased IGF-1 brain levels in MSA transgenic mouse model supported the hypothesis that impaired insulin/IGF-1 signaling existing in MSA pathology [
214‐
216]. A phase 2 study evaluating the efficacy of intranasal Insulin in patients with MSA (
n = 1) and PD (
n = 15) demonstrated an improvement based on UPDRS scale and the only MSA patient in this study received insulin-treatment remained symptom stable without disease progression [
217] (NCT02064166).
Exendin-4, an FDA-approved antidiabetic glucagon-like pepdide-1 (GLP-1) analog, has been shown to protect the nigral dopaminergic neurons survival and reduce the α-synuclein load, but motor benefits in a MSA transgenic mouse model have not been observed [
216]. A phase 2 randomized, open label study to evaluate the safety and efficacy of Exenatide (synthetic exendin-4) in patients with MSA is ongoing (NCT04431713).
Most recently, another neuroprotective target sphingosine-1-phosphate receptor 5 (S1P
5) was focused and new clinical trial was initiated. S1P
5 is predominantly expressed in nervous system and playing a role in neurodegenerative disorders [
218]. A phase 2 study of S1P
5 agonists ONO-2808 in patients with MSA is ongoing to assess the safety, tolerability, pharmacokinetics, pharmacodynamics, and potential efficacy as well (NCT05923866).
So far, the majority of completed clinical trials reported negative outcomes. The only clinical trial with positive outcomes is a trial of mesenchymal stem cells (MSC) treatment. As MSC is multipotent, it has been investigated as a potential neuroprotective or neurotrophic therapy [
219‐
222]. Preclinical studies of MSC treatment in transgenic MSA mouse models supported that intravenously infused MSCs have a potent effect on immunomodulation and neuroprotection [
223,
224]. The first clinical trial was an open-label, single-center study evaluating the feasibility and safety of therapy with autologous MSCs through consecutively intra-arterial and three repeated intravenous injections. The treatment group demonstrated significant improvement on UMSARS than the control group in all visits throughout the 12-month study period without serious adverse effects [
225,
226]. However, the open-label design has been challenged. After that, a phase 2 trial of autologous MSCs in patients with MSA was completed and showed a smaller increase in total and part II UMSARS scores in MSA patients receiving autologous bone marrow derived MSCs (bmMSCs) via intra-arterial routes, indicating MSC therapy could delay the progression of neurological deficits in patients with MSA [
227] (NCT00911365). However, because it was common to observe small ischemic lesions using magnetic resonance imaging (MRI) following intra-arterial infusion, safety concerns were raised to MSC therapies. Therefore, one recent phase 1 MSC clinical trial added the observation of small ischemic lesion as a safety measurement. The study reported that no ischemic lesions on diffusion-weighted images in any of the study participants, suggesting that a single intra-arterial administration of autologous bmMSCs is a safe and promising neuroprotective strategy in patients with MSA-C (NCT03265444). In addition, another ongoing long-term observational study also monitors the incidence of adverse events and the efficacy of subjects who participated in the above phase 1 trial to evaluate the safety and tolerability of autologous bone marrow derived MSCs in patients with MSA for up to 60 months after administration (NCT04495582).
MSCs derived from autologous fat (fMSCs) were also being investigated as a potential treatment to interfere MSA progression. A phase 1/2 open-label study evaluating the safety, tolerability, and efficacy of intrathecal injection of autologous MSCs in MSA patients was completed and revealed intrathecal MSCs was safe, well-tolerated, but associated with painful implantation at high doses [
228] (NCT02315027). A phase 2 trial of intrathecally administered autologous fMSCs in patients with MSA is ongoing (NCT05167721).
In addition, a recent interventional phase 1/2a study is underway to assess the safety and efficacy of intrathecal administration of low-/high-dose allogenic human oral mucosa stem cells (hOMSCs) in MSA patients in early to moderate stage (NCT05698017).
Although the aforementioned clinical trials achieved their primary endpoints and represent a potentially significant advance in the treatment, the safety of intra-arterial injection remains controversial due to reports of micro-strokes and increased mortality.