**3. Other tracers**

#### **3.1. Tracers assessing the extrapyramidal system**

The exact role of the extrapyramidal system in the pathophysiology of ALS is still under debate. Although several large clinical studies reported the increased presence of extrapyramidal symptoms and signs (increased tonus, postural instability and backward falls) in ALS patients [54, 55], others even reported an increased incidence of Parkinson's disease in the ALS population [56]. The presence of extrapyramidal symptoms in ALS is correlated with the presence of a hexanucleotide repeat expansion in *C9orf72* in many patients [57, 58]. PET studies have tried to make a significant contribution in elucidating a possible link between ALS and extrapyramidal symptoms.

[18F]-fluorodopa was one of the first radioligands used to assess the integrity of the nigrostriatal tract with PET by quantifying dopadecarboxylase activity presynaptically, while nowadays dopamine transporter imaging due to its better access using [123I]-FP-CIT SPECT is the most widely used method to assess the extrapyramidal system. More novel and specific tracers such as [18F]-PE2I for PET dopamine transporters are already in use at several centres worldwide [59].

Literature data regarding extrapyramidal radionuclide imaging in ALS patients is still sporadic and seems to be discordant. While two earlier studies reported a dopaminergic deficit in sporadic and familial ALS patients without clinical extrapyramidal disease [60, 61], this was contradicted in a more recent small study investigating ALS patients with concomitant clinical parkinsonism ('ALS-parkinsonism') [62]. Another study did report dopaminergic deficits in two ALS-parkinsonism patients assessed by [18F]-FP-CIT PET [63]. These discrepancies are believed to be due to the heterogeneity of ALS-parkinsonism, and larger datasets are needed.

While in some cases the extrapyramidal signs are thought to be caused by true degeneration of the extrapyramidal system, in other cases, they are believed to be the result of cortical lesions. The latter concept is confirmed by a more recent study investigating UMN-ALS patients using [ 123I]-FP-CIT SPECT [64]. While a dopaminergic deficit was indeed evident in the majority of patients which even correlated with disease duration, there was no correlation with functional extrapyramidal scores (like UPDRS). This means that, although the neuropathological process in ALS extends towards the extrapyramidal system, some of the extrapyramidal signs noted in ALS patients are probably due to spasticity, which is a typical UMN feature.

#### **3.2. Tracers for neuroinflammation**

Neurons rely on several supportive cells, commonly named glial cells, to survive and exert their normal function. These glial cells, including astrocytes, microglia and oligodendrocytes, provide nutritional and trophic support to neurons, especially motor neurons. ALS is charac‐ terized by a neuroinflammatory reaction consisting of an activation of astrocytes and micro‐ glia. Several studies using ALS animal models have taught us that dysfunction of these cell types significantly contributes to motor neuron death, independent of intrinsic motor neuron dysfunction, leading to the view of ALS as a non-cell autonomous disease [65].

Two targets have been used to visualize this 'neuroinflammation' in ALS patients using radionuclide imaging. [11C]-DED, a deprenyl derivative, selectively binds the MAO-B enzyme which is primarily though not exclusively located in astrocytes. Because of the high back‐ ground activity of this ligand and newer derivatives, they are limited to research applications. The second, more frequently used target is the activated microglial cell, the macrophages of the central nervous system [66]. One of the most studied targets for activated microglia is the translocator protein (TSPO), a mitochondrial protein that is highly overexpressed in activated microglia [67]. Several radioligands such as [11C]-(R)-PK11195, [18F]-DPA-174 and [11C]-PBR28 can be used to quantify TSPO binding.

The major finding in TSPO PET studies in ALS is an increased binding in the motor cortex, highlighting this region as the primary focus of neuropathology [68–70]. The degree of neuroinflammation in the motor cortex is positively correlated with clinical UMN scores and probably negatively correlated with ALS-FRS. Other frequently involved regions are the prefrontal cortex, thalamus, pons and CST [68–71]. The latter two probably reflect the secon‐ dary neuroinflammation due to degeneration of the CST. Interestingly, some studies have shown that the inflammation can be detected on the individual patient level.

Thus, PET imaging assessing neuroinflammation has a high potential to become a specific UMN marker which can be used at the single patient level. Moreover, it could be an interesting method to monitor the effect of treatment selectively targeting the neuroinflammatory process.

#### **3.3. Tracers reflecting neuronal loss and/or dysfunction**

**3. Other tracers**

32 Update on Amyotrophic Lateral Sclerosis

extrapyramidal symptoms.

**3.2. Tracers for neuroinflammation**

[59].

[

**3.1. Tracers assessing the extrapyramidal system**

The exact role of the extrapyramidal system in the pathophysiology of ALS is still under debate. Although several large clinical studies reported the increased presence of extrapyramidal symptoms and signs (increased tonus, postural instability and backward falls) in ALS patients [54, 55], others even reported an increased incidence of Parkinson's disease in the ALS population [56]. The presence of extrapyramidal symptoms in ALS is correlated with the presence of a hexanucleotide repeat expansion in *C9orf72* in many patients [57, 58]. PET studies have tried to make a significant contribution in elucidating a possible link between ALS and

[18F]-fluorodopa was one of the first radioligands used to assess the integrity of the nigrostriatal tract with PET by quantifying dopadecarboxylase activity presynaptically, while nowadays dopamine transporter imaging due to its better access using [123I]-FP-CIT SPECT is the most widely used method to assess the extrapyramidal system. More novel and specific tracers such as [18F]-PE2I for PET dopamine transporters are already in use at several centres worldwide

Literature data regarding extrapyramidal radionuclide imaging in ALS patients is still sporadic and seems to be discordant. While two earlier studies reported a dopaminergic deficit in sporadic and familial ALS patients without clinical extrapyramidal disease [60, 61], this was contradicted in a more recent small study investigating ALS patients with concomitant clinical parkinsonism ('ALS-parkinsonism') [62]. Another study did report dopaminergic deficits in two ALS-parkinsonism patients assessed by [18F]-FP-CIT PET [63]. These discrepancies are believed to be due to the heterogeneity of ALS-parkinsonism, and larger datasets are needed. While in some cases the extrapyramidal signs are thought to be caused by true degeneration of the extrapyramidal system, in other cases, they are believed to be the result of cortical lesions. The latter concept is confirmed by a more recent study investigating UMN-ALS patients using

123I]-FP-CIT SPECT [64]. While a dopaminergic deficit was indeed evident in the majority of patients which even correlated with disease duration, there was no correlation with functional extrapyramidal scores (like UPDRS). This means that, although the neuropathological process in ALS extends towards the extrapyramidal system, some of the extrapyramidal signs noted

Neurons rely on several supportive cells, commonly named glial cells, to survive and exert their normal function. These glial cells, including astrocytes, microglia and oligodendrocytes, provide nutritional and trophic support to neurons, especially motor neurons. ALS is charac‐ terized by a neuroinflammatory reaction consisting of an activation of astrocytes and micro‐ glia. Several studies using ALS animal models have taught us that dysfunction of these cell types significantly contributes to motor neuron death, independent of intrinsic motor neuron

in ALS patients are probably due to spasticity, which is a typical UMN feature.

dysfunction, leading to the view of ALS as a non-cell autonomous disease [65].

Although the neurodegenerative process in ALS is non-cell autonomous, markers of selective motor neuron death would be of significant value. FDG PET assesses glucose metabolism in general, hence encompassing various processes like neuronal dysfunction, atrophy, micro‐ gliosis and astrocytosis. Therefore, the capabilities of FDG PET to selectively assess motor neuron degeneration are limited.

Two tracers have been used so far to specifically assess neuronal loss and/or dysfunction. [11C] flumazenil, which selectively binds GABA-A receptors expressed by neurons, is the most widely used. It is, however, unclear whether this ligand primarily visualizes pyramidal neurons (like motor neurons) or interneurons. Although an early study reported an almost generalized decreased signal in the cortex [72], more recent studies revealed a more selected involvement of primary motor and motor association cortices, which was even correlated with a clinical UMN score [73, 74]. One study used the radioligand [11C]-WAY100635 targeting the 5-HT1A serotonin receptor which is expressed in pyramidal neurons [75]. They also reported a generalized cortical decrease, which was most pronounced in the motor cortex and fronto‐ temporal regions.

So, these neuronal radioligands seem to be able to specifically highlight the primary patho‐ physiological process of ALS, which is degeneration and dysfunction of the UMNs. Hence, it seems promising to further investigate the diagnostic and prognostic value of these potential UMN markers.
