Findings from the Swedish Study on Familial Alzheimer’s Disease Including the APP Swedish Double Mutation

INTRODUCTION

Familial Alzheimer’s disease (FAD) is a rare form of Alzheimer’s disease (AD), caused by autosomal dominant mutations in one of three known genes, the amyloid precursor protein (APP) gene [1–3], the presenilin 1 (PSEN1) gene [4], and the presenilin 2 (PSEN2) gene [5]. FAD mutations are usually close to 100% penetrant, leading to AD with an early and predictable age at onset of first cognitive symptoms [6].

The Swedish FAD study was initiated at the Karolinska Institutet in 1993 and has now been ongoing for 24 years. The participants in the study belong to four Swedish families, each carrying a different mutation leading to FAD: the PSEN1 H163Y mutation, the PSEN1 I143T mutation, the Swedish APP mutation (APPswe, KM670/671NL), and the arctic APP mutation (APParc, E693G). A total of 69 individuals from these families have participated in the FAD study through the years, some repeatedly, amounting in 169 separate examination occasions. The clinical signs and symptoms in the participating families have been described in previous publications [7–9]. The age at onset of cognitive symptoms in these families is 54±4 years for APPswe (based on 19 affected cases), 56±4 years for APParc (based on 12 affected cases), 51±7 years for PSEN1 H163Y (based on 11 affected cases), and 36±2 years for PSEN1 I143T (based on 5 affected cases).

The aim of the FAD study is to elucidate the pathological progress of AD through prospective collection of clinical and biomarker data from mutation carriers, with non-carriers from the same families serving as controls. The emphasis of the FAD study is on the preclinical stage of AD. Cognitively asymptomatic carriers of FAD mutations offer a unique opportunity to gather information on the preclinical stage of the disease, as they will develop the disease in the future with certainty and with a predictable age at symptom onset. Possible disease-modifying treatments for AD are now believed to be the most effective if initiated early in the course of the disease, preferably in the preclinical stage. Knowledge on the earliest detectable biomarker changes in this symptom free phase of AD is key when studying, and hopefully even applying, disease modification in the future.

All study procedures are in agreement with the Helsinki declaration and approved by the Regional Ethical Review Board in Stockholm, Sweden.

BIOMARKERS IN CEREBROSPINAL FLUID

The biomarkers amyloid-β (Aβ)₄₂, total tau-protein (t-tau), and phosphorylated tau-protein (p-tau) are routinely measured in the cerebrospinal fluid (CSF) of patients being evaluated for possible AD [10]. These markers offer support for diagnosing/excluding AD, with Aβ₄₂ typically decreasing and t-tau and p-tau increasing in AD. When measuring these three biomarkers in the CSF of 22 symptom-free participants from the FAD study (10 mutation carriers and 12 non-carriers), we observed a decrease in Aβ₄₂ 15–20 years before the expected onset of symptoms, while an increase in t-tau and p-tau was observed closer to the onset [11]. These findings are corroborated in other studies on the preclinical stage of FAD, that show a similar decrease in Aβ₄₂ in the CSF years before the onset of clinical symptoms [12–14]. CSF Aβ₄₂ is therefore a very early marker of AD pathology and useful both for early detection of the disease and potentially also for monitoring treatment response.

MAGNETIC RESONANCE IMAGING

Volumetric magnetic resonance imaging (MRI) is another well-established source of biomarkers in AD. The medial temporal atrophy score is widely used in the clinical setting to assess atrophy of the hippocampus [15–17]. A study by Bateman et al. detected a bilateral decrease in hippocampal volumes in carriers of FAD mutations 15 years before the expected onset of symptoms [12]. In another study on a different FAD cohort, Fox et al. reported similar results in 7 mutation carriers, albeit closer to the onset of cognitive symptoms [18]. When comparing 13 asymptomatic mutation carriers to 20 non-carriers from the Swedish FAD study, there was no significant difference in hippocampal volumes between the two groups. In this case, the mutation carriers had 9 years on average left to the onset of clinical symptoms. In the same study, however, there was a significant decrease in the volume of the left precuneus, left superior temporal gyrus, and left fusiform gyrus in the mutation carriers compared to the non-carriers [11].

Other modalities of MRI are also of interest in mapping the pathology of AD. By using diffusion tensor imaging, we observed white matter changes in the form of increased mean diffusivity in the left inferior longitudinal fasciculus, left cingulum and bilaterally in the superior longitudinal fasciculus in seven asymptomatic mutation carriers (compared to 20 non-carriers). When 3 symptomatic mutation carriers were included in the analysis, the affected areas became wider, suggesting early and progressive loss of myelination. In the same study, whole brain grey matter volume was analyzed and did not differ between the two groups [19].

Finally, 10 mutation carriers (3 of whom were symptomatic) and 13 non-carriers underwent resting-state functional MRI to assess functional connectivity in the default mode network (DMN). The DMN is a neuronal network that is active during rest and deactivates during active cognitive tasks. A decrease in functional connectivity has previously been observed in patients with mild cognitive impairment and dementia due to sporadic AD [20–22]. When all of the 10 mutation carriers were included in the analysis there was a decrease in functional connectivity in the right inferior parietal lobule, the right precuneus and the left posterior cingulate cortex. This decrease in functional connectivity did not reach significance when the symptomatic mutation carriers were excluded [23]. These findings suggest that amyloid and tau pathology interfere with neuronal and synaptic functions in the DMN, though this does not seem to be an early event in the disease cascade. However, lack of power to detect significant changes due to the small sample size may confound these results.

POSITRON EMISSION TOMOGRAPHY

In 2011, revised diagnostic criteria for AD were proposed by the National Institute on Aging – Alzheimer’s Association workgroups [24]. This is the first time that biomarkers are included in the diagnostic criteria for AD, but presently their use is generally only recommended for research purposes. The diagnostic criteria include biomarkers derived from the CSF as well as from imaging with positron emission tomography (PET) using ¹⁸F-fluorodeoxyglucose (FDG) as well as PET ligands binding directly to amyloid [24].

In an FDG-PET study from 2009, 6 asymptomatic carriers of the PSEN1 H163Y mutation were included, who were on average 20 years from expected symptom onset at baseline. The control group consisted of 23 non-carriers. Statistical parametric mapping revealed a trend of decreased thalamic glucose metabolism at baseline, which reached significance in the right thalamus at follow-up, 2 years later [25]. These findings suggest that metabolic changes are a very early event in AD, and this is in agreement with findings from other similar FDG-PET studies on carriers of FAD mutations [26, 27].

Inflammation has been implicated as a causative factor in AD and this has been supported by the discovery of reactive astrocytes surrounding amyloid plaques in brain tissue on autopsy [28–30]. ¹¹C-deuterium-L-deprenyl (DED) is a PET ligand that binds to monoamine oxidase B (MAO-B) on the outer mitochondrial membrane in astrocytes and its binding indicates reactive astrocytosis [31, 32]. When looking at DED binding in asymptomatic (n = 6) and symptomatic (n = 3) carriers of FAD mutations, as well as in patients with sporadic AD (n = 7) and mild cognitive impairment (n = 11), the highest level of DED binding was observed in the asymptomatic FAD mutation carriers. Conversely, DED binding was low in mutation carriers with cognitive symptoms. In the same study, an increase in the retention of ¹¹C-Pittsburg compound-B (PIB), an amyloid ligand, occurred early in the preclinical stage of FAD and predominantly in the anterior and posterior cinguli and the basal ganglia. These areas of increased PIB binding differed from the areas of increased DED binding [33]. This suggests that astrocytosis might be a response to non-fibrillar Aβ or even early plaque deposition and not to fibrillar Aβ as visualized by PIB binding. Furthermore, that astrocytosis occurs upstream of clinical symptoms and the formation of Aβ fibrils.

The findings from the multi tracer PET study described above were later replicated and further characterized with regards to temporality in a longitudinal study using the same tracers [34]. By using linear mixed-effects models, fibrillary Aβ plaque deposition was first observed in the striatum of asymptomatic FAD mutation carriers 17 years before the expected symptom onset. At about the same time, astrocytosis was significantly increased and then steadily declined. Diverging from the astrocytosis pattern, Aβ plaque deposition increased with disease progression. Glucose metabolism steadily declined from 10 years after initial Aβ plaque deposition. The prominent initially high and then declining astrocytosis in FAD mutation carriers, contrasting with the increasing Aβ plaque load during disease progression, suggests that astrocyte activation is most prominent in the early stages of AD pathology.

NEUROPSYCHOLOGY

Signs of cognitive decline through repeated neuropsychological tests are yet another biomarker of interest for early detection of AD. In 2017, Almkvist et al. published the results of neuropsychological assessments of the participants in the Swedish FAD study [35]. The participating mutation carriers were in different stages of FAD, from 28 years before the expected onset of symptoms until 12 years past the expected onset, spanning four decades of the disease. The age at symptom onset is a recurring concept in studies on FAD and is derived from the average age at onset of the first subjective cognitive symptoms in affected individuals in each FAD family. This family specific age at onset is currently widely used in research to estimate the expected onset age of asymptomatic mutation carriers [36].

The study by Almkvist et al. included 35 mutation carriers and 44 non-carriers who underwent a comprehensive neuropsychological assessment. A decline in performance on the Rey Auditory Verbal Learning test, an episodic memory test, was observed in the mutation carrier group 10 years prior to the expected symptom onset. This change was closely followed by a decline in performance in tests assessing executive function (Digit Symbol) and visuospatial ability (Block Design). These results are of particular interest as they imply that an objective decline on neuropsychological tests, covering several areas of cognition, precedes the subjective symptoms experienced by the patient.

CONCLUDING REMARKS

A hypothetical model of biomarker changes in the preclinical stage of AD was published by Jack et al. in 2010 [37]. According to this model, the earliest changes are observed in biomarkers reflecting the accumulation of Aβ, both in the CSF and on amyloid PET. These changes are followed by changes in biomarkers reflecting tau pathology (in the CSF and on FDG-PET). At the end of the preclinical stage, structural changes on MRI can be observed and shortly thereafter a decline in memory, heralding the onset of mild cognitive impairment.

The earliest biomarker changes observed in the asymptomatic mutation carriers from the Swedish FAD study were a decrease in CSF Aβ as well as increased binding of PIB on PET, corresponding nicely to the model proposed above. Interestingly however, an increase in DED binding on PET and a decrease in thalamic glucose metabolism on FDG-PET, reflecting inflammation and neuronal death, were early events as well. These changes on PET were further characterized in a longitudinal study showing that DED and PIB binding diverged as the preclinical stage progressed, with a decrease in DED binding and an increase in the uptake of PIB as the age at symptom onset approached. Later in the preclinical stage, around 10 years from symptom onset, there was a decline in episodic memory and atrophy was detected in several areas in the left cerebral hemisphere on volumetric MRI. White matter changes on MRI were also observed in the preclinical stage. Finally, an increase in CSF tau and p-tau was observed close to the onset of symptoms. This places the CSF markers of tau pathology downstream of both volumetric MRI and cognitive decline assessed by neuropsychological tests.

The sample size in the FAD study is small, due to the rarity of this disease, which detracts somewhat from the robustness of the acquired data. Also, the data presented here is mostly cross-sectional which reduces the certainty of our conclusions on the temporality of events in the preclinical stage of AD. However, the results from this valuable group of patients add to the current base of knowledge on biomarker changes in this stage of the disease and warrant further investigation in larger cohorts of FAD mutation carriers as well as in sporadic AD. Our future goals are to use the longitudinal collected data and make comparisons between the biomarkers and thereby provide further insights into the chain of pathological changes in preclinical and clinical stages of AD. Furthermore, developing biomarkers in serum and plasma will be an important goal to replace or complement the more invasive and technically demanding CSF and PET based assessments.

Finally, as part of the Swedish FAD study, all subjects provide fibroblasts and current studies have shown promising results regarding the potential use of patient derived cells for both basic scientific studies of cellular mechanisms of neurodegeneration as well as a possible tool for treatment [38]. We have thus also initiated a cell modeling program where we hope to elucidate some of the possible mechanisms of reduced penetrance in PSEN1 H163Y mutation carriers as well as study the effects of autophagy dysfunction in neurodegeneration as observed on autopsy of AD and other tauopathies