[¹⁸F]Florbetapir PET/MR imaging to assess demyelination in multiple sclerosis

Multiple Sclerosis (MS) usually presents with episodes of transient neurological deficits, called ‘relapses’ [1]. Relapses mirror the occurrence of focal areas of acute demyelination within the brain, optic nerves and spinal cord [2]. Demyelination can cause axonal loss, which over time causes permanent disability [3]. Demyelination can be qualitatively detected as areas of high signal on T2-weighted magnetic resonance imaging (MRI) sequences or as low signal in T1-weighted spin-echo images on MRI scans [4]. Advanced MRI techniques might provide a better insight into multiple sclerosis pathology. In example, diffusion tensor imaging enables us to detect microstructural tissue damages in brain regions [5, 6], whereas arterial spin labelling provides a quantitative measure of the cerebral blood flow. However, conventional MRI, diffusion tensor imaging and arterial spin labelling only represent indirect measures of tissue modifications and lack specificity towards myelin integrity. Therefore, although previous studies have reported the association between brain microstructural changes and physical disability in MS, the effect of demyelination on physical disability might not be disentangled for other pathological factors such as axonal loss or gliosis for the low specificity of MRI techniques.

Over the past years, positron emission tomography (PET) imaging has become a promising modality to monitor the biological processes underlying MS pathology [7]. Radioligands developed to measure amyloid-β pathology in grey matter for Alzheimer’s disease, such as [¹¹C]PIB, [¹⁸F]florbetaben and [¹⁸F]florbetapir [8‐10] also showed affinity for myelin protein. This is due to the fact that these stilbene and benzothiazole derivatives have a very flat structure, interacting with the secondary structure of myelin basic protein similarly to the way they interact with amyloid [11, 12]. In fact, in healthy white matter, myelin basic protein has a particular two-dimensional secondary structure as it acts as a hinge between lipid bilayers. In MS, this structure is damaged which, consequently, leads to a decreased binding of amyloid-β PET tracers [13]. Therefore, stilbene and benzothiazole derivatives, firstly used to explore amyloid accumulation throughout grey matter in neurodegenerative disease, might be a better marker of demyelination compared to MRI in MS [14].

Bodini and colleagues, used [¹¹C]PIB PET to image myelin using dynamic acquisitions that has a short half-life (about 20 min) that limits its clinical viability, whereas Fluorine-18 labelled PET tracers have longer half-life (about 120 min), which allows for a clinical use in patients MS. However, by combining PET and MRI Bodini and colleagues demonstrated that myelin changes, detected with amyloid PET imaging, within lesions was associated with physical disability [15]. Therefore, suggesting a relationship between myelin disruption and outcome.

In this study we adopted a hybrid PET-MR approach using [¹⁸F]florbetapir PET to quantify myelin content in MS patients. We investigated the relationship between [¹⁸F]florbetapir uptake and MRI measures, e.g. diffusion tensor imaging and arterial spin labelling together with standard measures of cortical grey matter volume. Finally, we compared SUV_70–90 analyses with the DVR kinetic modelling to estimate the clinical applicability of PET imaging through simplified semi-quantitative PET measures.

In this study, we demonstrated that [¹⁸F]florbetapir PET is a valid tool to detect the pattern of demyelination in MS, with a comparable accuracy between SUV_70–90 PET measure and the computational DVR analysis methods. Combining PET with advanced MRI techniques, we showed that myelin damage was correlated with microstructural white matter changes and lower perfusion rate, providing additional information.

We showed that [¹⁸F]florbetapir binding from dynamic DVR and static SUV measures have comparable accuracy in differentiating myelin damaged tissue from NAWM. [¹⁸F]florbetapir quantification with SUV has a number of advantages including short scan-time, with acquisition within a predefined time window and lower costs compared to the dynamic continuous acquisition. Bodini and colleagues demonstrated that longitudinal [¹¹C]PIB PET imaging scans quantified through the DVR, might depict not only demyelination but also remyelination over time [15]. Hence SUV measures could be helpful when assessing treatment efficacy in clinical trials and in the management of patients with MS. However, the longitudinal reproducibility of Logan DVR and SUV for [¹⁸F]florbetapir PET might deserve further studies to be confirmed.

We showed that [¹⁸F]florbetapir binding was decreased in focal white matter lesions of the MS patients, defined on conventional MRI, compared to the tracer uptake in the white matter of the healthy controls. T1 lesions showed the lowest value, confirming that they are characterized by extensive myelin destruction [29]. Post-mortem studies have already reported that T1 hypointensity reflects histopathological modifications, such as axonal loss, extracellular oedema and demyelination [30]. The extent of each of these pathological processes could not be evaluated through conventional MRI. To overcome this obstacle, several advanced MRI techniques have been applied in MS. Among them, the diffusion tensor imaging is the most promising technique to assess microstructural changes while measuring the water motility within tissues. However, the diffusion tensor imaging measures the global structural modifications within damaged tissues including demyelination, axonal damage, microglia activation [31] and gliosis without being able to differentiate them [32, 33]. [¹⁸F]florbetapir PET imaging can measure the extent of demyelination without including biases from other pathological changes, such as gliosis or axonal loss, and we demonstrated that DVR provides additional information on tissue damage compared to DTI measures. Further studies including other advanced MRI techniques such as magnetization transfer imaging might also evaluate the specificity of PET imaging over MRI in detecting demyelination.

[¹⁸F]florbetapir binding was similar for MS patients and healthy controls in NAWM. The absence of difference in myelin content of grey and white matter that we found for the two groups appears not to be in line with previous studies that have shown tissue changes within both grey and white matter outside the lesions detected by conventional MRI through diffusion tensor imaging, magnetisation transfer imaging and spectroscopy [32, 34‐36]. However, this discrepancy further underlines that such MRI techniques are able to detect global pathological changes, including axonal degeneration, gliosis and inflammation throughout the brain, but are not specific measures of myelin integrity. Noteworthy, it is also important to underline that demyelination arises to different extent from neuroinflammation. Therefore, although microglia activation was reported throughout the normal appearing white matter [37], demyelination might not occur simultaneously. However, we reported that the white matter tissue surrounding the edge of demyelinating lesions, detected through conventional MRI, is also affected by demyelination at different degrees. A post-mortem study by De Groot and colleagues reported reduced myelin density in areas in close proximity to the demyelinating damaged areas in brain specimens from MS patients [30]. Furthermore, Bodini and colleagues reported a similar pattern of demyelination through a [¹¹C]PIB PET imaging study [15]. In this latter study, authors evaluated up to 2 mm rim surrounding T2 lesions. Here, we reported that myelin disruption is a process going further beyond the borders detected through conventional MRI images, up to almost 1 cm. This aspect is important when assessing the extent of demyelination throughout the CNS. Slowly evolving lesions are becoming a hallmark for disease progression in MS [38]. Noticeably, our finding suggested that [¹⁸F]florbetapir PET imaging might detect with a single scan a larger areas of damaged tissue compared to conventional MRI and, thus, might be considered a better predictor of disease progression. Moreover, the increased size of a lesion may also depend on an ineffective and incomplete process of remyelination and does not necessarily implies the reactivation of the lesion. Interestingly, longitudinal [¹⁸F]florbetapir PET scans might shed further light in the biological bases of the remyelinating process around the lesions.

We would expect a difference in cortical grey matter [¹⁸F]florbetapir binding between MS patients and healthy controls, because cortical demyelinating lesions are becoming a hallmark in MS pathology [36]. According to this hypothesis, we found that the myelin content within the cerebellar cortex for MS patients was lower than the one in the cerebellar cortex of healthy controls. This finding also highlight that the cerebellum grey matter cortex must not be chosen as reference region for amyloid-β PET imaging in MS as opposed to other neurodegenerative disorders, such as Alzheimer’s disease [39, 40]. This caveat also confirms that the supervised cluster analysis is crucial when selection of the reference region is needed. Moreover, as previously stated, also SUV could be helpful when assessing treatment efficacy; however, its longitudinal reproducibility should to be explored. The extent of demyelination within both T2 and T1 lesions was not associated with cortical or white matter volume. Although several studies have pointed out that the lesion load measured through conventional MRI correlates with the grey matter volume in MS [41, 42], our results suggested that demyelination specifically measured with [¹⁸F]florbetapir PET imaging within MS lesions per-se is not the cause of brain atrophy. Supposedly, the balance between remyelination, preserved axonal integrity and the wallerian degeneration of axons traversing lesions might impact the total brain volume and the cortical thickness [43]. Moreover, axonal injury in MS is not restricted to white matter lesions but can occur also in the NAWM, independently from focal cerebral demyelination [35]. Therefore, longitudinal amyloid-β PET imaging studies, eventually coupled with [¹¹C]flumazenil PET, specifically measuring neurodegeneration [44], are needed to further elucidate the impact of demyelination throughout the CNS on brain and cortical axonal loss. In addition, a combined [¹¹C]flumazenil and amyloid-β PET imaging study might also shed further light on the interplay between demyelination, eventually remyelination and axonal degeneration. A recent paper, using a complex approach involving optical coherence tomography, conventional and advanced MRI, reported that demyelination over a specific white matter tract (i. e. optic radiation) precedes neurodegeneration [45]. Using a PET approach using two probes specifically targeting the two main pathological processes occurring in MS patients, might provide the unique opportunity to evaluate the dynamics of such processes with a high degree of specificity and to evaluate the impact of the interplay between these two phenomena on clinical disability.

We observed a reduced cerebral arterial blood perfusion in white matter regions with a higher extent of demyelination. MS lesions and normal appearing white matter are characterized by cerebral hemodynamic impairment even in absence of blood-brain-barrier damage [46], resulting in reduced arterial blood supply. Previous pathological studies have already reported hemodynamic changes in MS including vessels’ thrombosis and tissue hypoxia due to fibrin deposition in demyelinating plaques. Specifically, the high venular density and the reduced arterial flow in the damaged white matter tissue suggest that hemodynamic changes could contribute to tissue damage. Yet, the precise impact of vascular factors on MS pathology has not been fully elucidated. Our data about cerebral blood flow are partly in line with previous findings, especially when compared with studies using the same methodology [47, 48], which will further support the impact of vascular changes on MS pathology. Longitudinal PET/MRI studies could further investigate whether arterial blood flow reduction is the primary cause or the result of the inflammatory-driven tissue necrosis. It is worthy to mention that both DVR and SUV depend on an intact blood-brain barrier and changes in blood-brain barrier integrity occurring in gadolinium enhancing lesions could potentially introduce a bias. However, in the present study only one single gadolinium enhancing lesion was present in one patient and, in addition, DVR is a dynamic measure which take into account the time activity curve of tracer distribution. Furthermore, changes in CBF do not appear to have a significant effect on DVR values estimated using nonlinear modelling or graphical analysis [49]. Therefore, [¹⁸F]florbetapir DVR measures are unlikely to be significantly influenced by cerebral arterial blood perfusion.

We did not find any correlations between the extent of white matter lesions demyelination and the disease severity, assessed through the MSSS, disease duration and annualized relapse rate and physical disability assessed through the EDSS. However, we found a correlation between physical disability assessed through the Timed 25-ft walk and the extent of myelin loss within T2 lesions. Bodini and colleagues showed that both the EDSS and the MSSS were not related to the tracer DVR in T2 lesions at a cross-sectional evaluation [15]. Physical disability was rather related to the extent of lesions demyelination over a follow-up period of four months [15]. We might speculate that physical disability is actually driven by the balance between demyelination and remyelination within MS lesion with a consequent axonal loss. Moreover, EDSS is a measure of physical disability with a low degree of specificity because it is mainly related to walking abilities, especially when it turns to the highest point. Timed 25-ft walk is actually more precise than EDSS and might explain the discrepancy in our finding.

In conclusion, our study reported that [¹⁸F]florbetapir PET imaging might be able to assess in vivo myelin pathology with a higher extent of specificity compared to MRI. The static SUV_70–90 PET imaging evaluation provides similar information to the dynamic PET. Dynamic PET imaging requires long dynamic scans and, hence, high costs. Conversely, the SUV might be calculated through a static acquisition lasting for twenty minutes. This makes the technique highly appealing to implement in clinical settings. Clinicians could use the PET imaging as a tool to measure the subjective remyelinating potential, with the ultimate goal of personalizing treatment. They could also rely on its quantitative measures when assessing the effect of a drug targeting remyelination, e.g. opicinumab [50]. Finally, although further longitudinal evaluations are needed to test the reproducibility of the technique, measures derived from PET imaging (mean DVR or mean SUV) are able to measure the accumulation of tissue changes beyond the defined borders of the lesion.