Glial cell line-derived neurotrophic factor (GDNF) family ligands (GFLs) include three other structurally related proteins in addition to GDNF: neurturin (NRTN), artemin (ARTN, also known as enovin and neublastin) and persephin (PSPN) and, also, a distant member, growth differentiation factor-15 (GDF15) [1
]. They all play a role in the development and maintenance of the nervous system, and GDNF is also important for kidney development and spermatogenesis [1
]. GDF15 is involved in appetite control [5
The founding member of GFLs, GDNF, was discovered as a potent survival factor for dopamine neurons [9
], and, therefore, it was tested in several clinical trials in patients with Parkinson’s disease (PD), as PD is characterized by the profound degeneration of dopamine neurons in the brain, resulting in motor symptoms in the disease [10
]. The efficacy of another GFL supporting dopamine neurons, NRTN, delivered via gene therapy-based approach was also evaluated in PD patients [16
]. However, the results of these clinical trials remain controversial; while, in small-scale open label ones, the positive effects of such treatments were seen, large-scale trials failed to reach their primary efficacy end point. The reasons explaining the inefficiency of GFLs in PD are reviewed in detail elsewhere [19
] and are mainly related to the inability of these proteins to cross through the blood–brain barrier and spread into tissues. These issues make it necessary to deliver GFL proteins directly into the brain to the target region of the midbrain dopamine neurons by means of complicated and expensive stereotaxic surgery, which limits the selection of patients into clinical trials to the ones with late-stage PD. However, late-stage PD patients have very little remaining target neurons to be supported and restored by GFLs [21
]. GFLs are able to support the survival of remaining neurons, regenerate their axons, restore the damaged ones (Figure 1
A) and improve the functional activity of viable neurons. However, GFLs are unable to revive dead cells or produce new neurons. Therefore, the treatment with GFLs should start as soon as possible, preferably immediately upon diagnosis, which is impossible—or, at least, immensely difficult—in PD patients at the moment due to ethical restrictions on brain surgery in early stage PD patients.
The characteristic histopathological feature of PD is the presence in the brain of protein aggregates called Lewy bodies, the main component of which is alpha-synuclein [24
]. Recent data indicate that the activation of GFL-dependent signaling can prevent the progression of alpha-synuclein pathology in the brain [27
ARTN, which has a well-established function in sensory neurons, was tested in patients with neuropathic pain (NP) [28
] and showed promising results in patients with painful lumbosacral radiculopathy nonresponsive to at least two standard treatments [30
]. However, in this trial, the dose response to ARTN was biphasic, and the lowest dose provided the highest pain relief, while the second most efficient dose was the highest one. This kind of dose-response relationship, along with the reported side effects and potential of the ARTN to attach to extracellular matrix components, producing high point concentrations and restricting tissue spreading, complicate the clinical use of the ARTN protein.
Due to the capability to support motoneurons, GDNF has also been tested for the ability to slow down the progression of amyotrophic lateral sclerosis (ALS), an incurable disease with very poor prognosis and an average survival below five years after the diagnosis [31
]. In patients suffering from ALS, motoneurons in the brain and spinal cord degenerate and die. This leads to progressive muscle atrophy and, finally, to respiratory failure, which is the most common cause of the death in ALS [32
]. In some studies in animal models of ALS, GFLs improved the disease manifestations, although the effect could be dependent on the delivery site, concern only some symptoms and be variable in the degree of improvement [34
], complicating the interpretation of the data and clinical translation. GDNF delivered to patients using stem cells was found to be safe and well-tolerated in a recent small-scale clinical trial, but the efficacy data are yet to be published [37
]. Taking into account the poor diffusion of GFL proteins in the tissue [38
] and widespreadness of ALS pathology, selecting the delivery paradigm for GFLs in ALS is especially difficult.
Retinitis pigmentosa is a hereditary eye disease characterized by the degeneration of retinal cells, resulting in a loss of the peripheral vision at first and blindness when the disease progresses [39
]. The data on the role of GDNF in retinal cell survival are controversial, but in some studies in animal models, the positive effects of this protein were seen [40
]. In retina, GFL receptors are expressed in Müller cells, the principal glial cells in this tissue, which have supportive and neuronal activity modulating functions [42
]. GFLs delivered to an eye exert neuroprotective effects in photoreceptors indirectly via the activation of Müller cells that secrete other trophic factors necessary for the survival of retinal cells, such as fibroblast growth factor-2 [43
GDNF also has a potential for the treatment of substance (drug) abuse and dependence via its effects in the dopamine system [44
]. GFLs may support basal forebrain neurons [46
] and promote dendrite arborization and synapse formation by hippocampal neurons [47
], which degenerate in Alzheimer’s disease (AD), although proof-of-concept for this indication needs further validation. The lentiviral vector-encoded GDNF delivered into the hippocampus before the onset of symptoms preserved the memory and learning ability in a mouse model of AD [48
]. However, further studies are needed to understand if the positive effects of GFLs can also be seen in mice with established AD and elucidate the mechanism of action of GFLs in this model. Human data for the conditions listed above are not yet available. GFLs exert trophic functions in the neurons belonging to autonomic nervous system (sympathetic, parasympathetic and enteric), which are important for multiple body functions, including digestion, breathing control, sweating, penile erection, etc. [4
], which provides interesting opportunities for further therapeutic developments. The use of GFLs proteins in associated conditions is complicated because of their poor diffusion, but alternatives with improved properties can be considered.
Important to mention is that, in many diseases caused by neuronal degeneration or damage, neuroinflammation plays a prominent role [53
]. A characteristic feature of neuroinflammation is an activation of microglial cells in the nervous system. Recent data indicate that microglial cells express GFL receptors, and GFLs can reduce microglia activation by inhibiting p38 MAPK signaling [56
]. Therefore, targeting GFL receptors can also diminish the deteriorative effects of microglial cells in neurodegenerative disorders and chronic pain. Important to remember, however, is that microglial cells can play not only proinflammatory but, also, an anti-inflammatory, protective role in the nervous system, depending on their phenotype [57
]. Further studies are needed to characterize the influence of GFLs on specific subtypes of inflammatory cells and the consequences of such effects on the tissue and organism levels.
GDF15 is aside from the other GFLs in its function. While it is important for the survival of motor, sensory and dopamine neurons [59
], it plays a role in the inflammatory and cardiovascular systems, exerts nephroprotective effects and it has attracted major attention in the recent three years as a factor controlling appetite [5
GDF15 overexpression or infusions of recombinant growth factor resulted in a decrease in food intake, improved glucose tolerance and, also, stimulated weight loss in mice fed by both standard and high-fat diets [6
]. The prevalence of overweight, obesity, metabolic diseases and associated comorbidities is rising all over the world with a speed that made it be referred to as global pandemic. According to the data provided in a systematic analysis for the Global Burden of Disease Study 2013, 36.9% of adult men and 38% of women in the world have a body mass index ≥ 25 kg/m2
]. The proportion of overweight children is also rising and exceeds 10% in developed and 20% in developing countries [65
]. Obesity treatments with proven efficacy include invasive gastric bypass surgery and the lipase inhibitor orlistat, which works only in a subset of patients [66
]. Therefore, new noninvasive drugs to treat obesity are in high demand, and GDF15 appears as an attractive therapeutic option [67
]. Interestingly, NRTN supports pancreatic β cells and, thus, may be useful in the management of diabetes [68
]. This review focuses mainly on the role of GFLs in neurodegeneration; extensive data on GDF15 functions in obesity and NRTN in diabetes are provided elsewhere [5
To summarize, GFLs have a clear therapeutic potential for disease modification in several diseases and conditions caused by neurodegeneration (Figure 1
B). However, their clinical use is complicated because of poor pharmacokinetic characteristics, high price, variability in biological activity between batches, instability and immunogenic potential [10
]. Besides, at least some neurodegenerative disorders are characterized by widespread pathologies. For example, PD patients, in addition to the degeneration of midbrain dopamine neurons, have other neuronal populations affected, e.g., enteric neurons, noradrenergic neurons, etc. In Alzheimer’s disease, neurons in several brain regions die. Sensory and motor neurons, degenerating in NP and ALS, are also located in different parts in the body. GFLs have a high binding affinity to the extracellular matrix, and therefore, their diffusion in the organism is limited [38
]. Alternatives such as more diffusible neurotrophic factors, peptides and peptidomimetics, as well as small molecules targeting GFL receptors, may provide valuable tools to utilize GFLs’ potential to combat neurodegeneration and support the survival of different neuronal populations in different parts of the body. The current review will describe peptides and small molecules mimicking the biological effects of GFLs in cultured cells and animal models of relevant neurodegenerative diseases and discuss their advantages and limitations for the therapy of neurodegeneration.
Despite great promise in the treatment of neurodegenerative disorders and neuropathic pain, the clinical development of GFLs have, so far, achieved only limited success. This is due to the pharmacokinetics of these proteins, which are unable to cross tissue barriers and spread very poorly into tissues. In classic neurodegenerative disorders such as Parkinson’s disease, this means that GFL proteins have to be surgically delivered into the brain, and even in this case, they cover only a small portion of the affected tissue, as they diffuse poorly. In NP and eye diseases, these proteins can be delivered systemically or locally, respectively, but the target tissue exposure to GFL proteins remains problematic. Therapeutic efficacy issues of GFLs can be solved by the development of alternatives with better pharmacological characteristics. Several exciting molecules with improved clinical translation potentials, such as chemical compounds and peptides, targeting components of GFL receptor complexes have been described over the last two decades. At the moment, it is unknown which strategy would be the best: positive allosteric modulation of GFLs signaling or targeting RET, NCAM or GFRa coreceptors. Available molecules should be further optimized to improve their potency and other properties. Moreover, their safety has to be evaluated in extensive preclinical experiments. However, available data show that GFL receptors are druggable targets, and the modulation of their biological activity may be beneficial for the survival and well-being of retinal cells and sensory, dopamine and possibly other neuronal populations. Other major possible indications for the applications of such compounds are ALS and obesity, where in vivo proof of concept is yet to be established. Trophic effects of GFLs in the autonomic nervous system also justify testing molecules targeting GFL receptors in conditions associated with intestinal tract motility, excessive sweating and salivation, erectile dysfunction, etc. Despite promising preliminary data, many questions regarding the clinical translation of compounds targeting GFL receptors remain to be answered. In particular, the administration regimen for such compounds has to be carefully considered. Taking into account potential side effects as a result of the continuous long-term activation of RET and recent clinical trials data with intermittent delivery, we think that the best treatment paradigm will be systemic administration with regular intervals (e.g., once-monthly or weekly). However, further research is needed to best utilize the potential of GFLs and their receptors for the treatment of neurodegeneration and to establish a treatment paradigm for the developed compounds.