The need for regenerative or neuroprotective therapies – Approaches and

As mentioned above, PD can be diagnosed with the onset of clinical motor symptoms. A problem is that the first motor symptoms occur late in disease progression when approximately 60 % of the nigrostriatal projections are already degenerated (Jellinger 2012). By now, there is no reliable biomarker known that might predict disease onset. Thus, it is of major importance to develop treatment strategies that prevent further degeneration of remaining dopaminergic SNpc neurons or induce regeneration of the nigrostriatal system.

One major problem in the injured or diseased CNS is the prevention of neuronal regeneration by extracellular cues (e.g. glial responses) and a limited intrinsic regeneration capacity in adult neurons (Fitch & Silver 2008). Nevertheless, there is evidence that also adult neurons have the ability to sprout and thus, regeneration of the adult CNS is in principle possible. In toxin-induced animal models for PD spontaneous re-growth of dopaminergic nigrostriatal projections could be observed in different species.

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17 In rats that received 6-OHDA injections in the SNpc vigorous axonal sprouting could be observed 4 months post-lesion (Finkelstein et al. 2000). In MPTP-intoxicated monkeys and mice, the extent of spontaneous regeneration was linked to the dimension of degeneration (Elsworth et al. 1999; Mitsumoto et al. 1998). These examples demonstrate that the adult dopaminergic nigrostriatal system in principle has the ability to regenerate after a lesion, but by now it is not known whether compensatory sprouting occurs in PD patients or how it could be facilitated or induced.

Recently, several approaches to induce regeneration of the nigrostriatal system or avoid further degeneration of SNpc neurons were proposed. Among them was extensive research on grafting of fetal dopaminergic neurons or embryonic/induced pluripotent stem cells (ES; IPS). Implantation of fetal dopaminergic neurons into toxin-induced animal models for PD led to improved motor symptoms and increased striatal innervation (Bankiewicz et al. 1990; Redmond et al. 1986). After these promising preclinical studies grafts of fetal dopaminergic neurons were also implanted into the brains of patients with PD. The grafts can survive in the brains of PD patients but the degree of symptomatic improvement is highly variable and strongly dependent on different factors. Additionally, severe side effects like dyskinesia can occur (Freed et al. 2001; Piccini et al. 2005). With the emergence of advanced stem cell technology transplantation of undifferentiated ES or IPS became a novel possibility in PD research. Stem cells are the optimal source for cell replacement therapy because they are self-renewing and multipotent. It has been shown that mouse ES transplanted to 6-OHDA lesioned rats can spontaneously differentiate into dopaminergic neurons (Bjorklund et al. 2002). Transplantation of neuronal precursor cells derived from monkey ES attenuated MPTP-induced neurological symptoms in monkeys (Takagi et al. 2005). As undifferentiated ES always bear the risk of tumor formation after transplantation there is the aim to differentiate dopaminergic neurons out of IPS. These cells can be sorted and might improve the safety of cell replacement therapies (Sundberg et al. 2013).

Another important approach to facilitate regeneration and provide neuroprotection of the nigrostriatal system is the application or expression of neurotrophic factors. Neurotrophic factors are important for the development and maintenance of neurons. Several neurotrophic factors have been investigated for their neuroprotective potential for dopaminergic neurons. The neurotrophins that have a

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18 selective effect on dopaminergic neurons are glial cell-line derived neurotrophic factor (GDNF), neurturin, growth / differentiation factor 5 (GDF5), mesencephalic astrocyte-derived neurotrophic factor (MANF) and cerebral dopaminergic neurotrophic factor (CDNF) (Sullivan & Toulouse 2011). The most promising factor is GDNF, belonging to the GDNF family of ligands, which has been extensively studied in vivo and in vitro and was already tested in clinical trials. GDNF has been shown to promote the differentiation and survival of dopaminergic neurons in culture. Additionally, treatment with GDNF could prevent those neurons from degeneration upon 1-methyl-4-phenylpyridinium ion (MPP+) treatment in vitro (Hou et al. 1996; Krieglstein et al. 1995; Lin et al. 1993). Similar neuroprotection and functional effects could also been observed in different animal models of PD, consequently it acts also on the adult dopaminergic neurons (Cheng et al.

1998; Kearns & Gash 1995; Kordower et al. 2000). The administration site and mode (transfusion of the recombinant protein or expression by viral vectors) are crucial for the effectiveness of GDNF delivery (Sullivan & Toulouse 2011). After first successful trials of intrastriatal GDNF delivery in human PD patients (Patel et al. 2005; Slevin et al. 2005), another randomized placebo-controlled trial reported no relief of motor symptoms (Lang et al. 2006). In addition, safety issues were raised when antibodies against GDNF were found in approximately 10 % of patients that received striatal GDNF infusion, which could lead to adverse effects (Tatarewicz et al. 2007). General consensus is that GDNF is a valuable treatment option, but needs further preclinical studies to overcome safety issues and develop sustainable effects.

Another possibility to achieve protection or regeneration of dopaminergic neurons is the pharmacological modulation of intrinsic signaling and disease relevant pathways.

Potential candidates to prevent neurodegeneration are inhibition of apoptotic and necrotic cell death pathways as well as application of ROS scavengers and energy mimetics and inhibitors of alpha-synuclein toxicity (Dawson & Dawson 2002).

Furthermore modulation of kinase activity (e.g. Akt, JNK, LRRK2, MAPK, ROCK) is a promising treatment option for PD because there is evidence that inhibition or activation of certain kinases is preventing neurodegeneration in preclinical studies (Burke 2007;

Deng et al. 2011; Saal et al. 2015; Tönges et al. 2012).

Despite all efforts, the development of an applicable tool to prevent dopaminergic neuron cell death and facilitate regeneration of the nigrostriatal system in PD was still not

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19 successful and new ideas for treatment strategies are urgently needed. It is known that endogenous microRNAs (miRNAs) exert different functions in the CNS. As they usually target not only one but multiple proteins and pathways, they might be powerful tools for understanding PD pathology and development of new treatment strategies.

1.3 miRNAs – Biogenesis, function and role in neurodegenerative diseases