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In this prospective cohort study, we investigated cerebral glucose metabolism reductions on [18F]-fluorodeoxyglucose (FDG)-PET in progranulin (GRN) mutation carriers prior to frontotemporal dementia (FTD) onset.
Nine mutation carriers (age 51.5 ± 13.5 years) and 11 noncarriers (age 52.7 ± 9.5 years) from 5 families with FTD due to GRN mutations underwent brain scanning with FDG-PET and MRI and clinical evaluation. Normalized FDG uptake values were calculated with reference to the pons. PET images were analyzed with regions of interest (ROI) and statistical parametric mapping (SPM) approaches.
Compared with noncarriers, GRN mutation carriers had a lowered anterior-to-posterior (AP) ratio of FDG uptake (0.86 ± 0.09 vs 0.92 ± 0.05) and less left-right asymmetry, consistent with an overall pattern of right anterior cerebral hypometabolism. This pattern was observed regardless of whether they were deemed clinically symptomatic no dementia or asymptomatic. Individual ROIs with lowered FDG uptake included right anterior cingulate, insula, and gyrus rectus. SPM analysis supported and extended these findings, demonstrating abnormalities in the right and left medial frontal regions, right insular cortex, right precentral and middle frontal gyri, and right cerebellum. Right AP ratio was correlated with cognitive and clinical scores (modified Mini-Mental State Examination r = 0.74; Functional Rating Scale r = −0.73) but not age and years to estimated onset in mutation carriers.
The frontotemporal lobar degenerative process associated with GRN mutations appears to begin many years prior to the average age at FTD onset (late 50s–early 60s). Right medial and ventral frontal cortex and insula may be affected in this process but the specific regional patterns associated with specific clinical variants remain to be elucidated.
The multifaceted clinical syndrome of frontotemporal dementia (FTD) arises from degeneration of the frontal and temporal lobes (frontotemporal lobar degeneration). Mutations in the gene encoding progranulin (GRN), discovered in 2006,1,2 are found in 5%–20% of those with familial FTD (FTD-GRN).2,3 Despite the common haploinsufficiency mechanism1 and transactive response DNA-binding protein Mr 43 kD (TDP-43) neuropathology,4 there is phenotypic variation in FTD-GRN, with behavioral variant FTD (bvFTD), progressive nonfluent aphasia (PNFA), and corticobasal syndrome.5 Mean age at onset is 59–65 years, but can range from 35 to 87 years.6 In GRN mutation carriers with FTD, an asymmetric pattern of brain structural abnormalities is found, with severe gray matter loss involving frontal, anterior temporal, but also posterior temporal and inferior parietal regions.7–9 There is only very limited evidence on brain abnormalities in GRN mutation carriers before FTD onset, with reports of mild structural and functional abnormalities up to 2 decades prior to expected disease onset.10–12
In this study, we hypothesized that GRN mutation–associated neurodegenerative processes begin well before the onset of full-blown FTD and can be demonstrated as glucose metabolism reductions on [18F]-fluorodeoxyglucose (FDG)-PET. We prospectively acquired FDG-PET scans on GRN mutation carriers still free of FTD and noncarriers from the same families, and compared their glucose metabolism by global ratios, regions of interest (ROI), and statistical parametric mapping (SPM).
The 20 participants, recruited between 2006 and 2011 from 5 families with FTD due to a GRN mutation (table 1), included all subjects aged ≥19 years and at risk of disease by virtue of having a parent or a sibling with FTD seen through the University of British Columbia (UBC) Clinic for Alzheimer Disease and Related Disorders, Vancouver, Canada, since the 1980s. Consenting subjects received a comprehensive clinical examination at the time imaging was performed to determine that they were free of dementia. For 4 subjects, the examination was performed up to 6 months prior to or following imaging. GRN mutation status was determined after enrollment and was not known to the investigators performing clinical assessments or image analysis. The number of women and men differed significantly between mutation carriers (n = 9/7 women) and noncarriers (n = 11/4 women). Since prior PET studies of normal subjects have demonstrated sex-related differences in global and regional glucose metabolism,13,14 subject sex was included as a covariate in our analyses.
The study was approved by the UBC Ethics Board. Written informed consent was obtained from each participant.
FDG-PET scanning was performed at UBC on a high-resolution research tomograph (Siemens/CTI, Knoxville, TN) in 3D mode, with an isotropic resolution of 2.3 mm3. Overnight fasting subjects' plasma glucose was measured and confirmed to be <180 mg/dL prior to starting the scanning procedure. Subjects were scanned supine in a quiet room, instructed to remain awake with eyes open or closed. The imaging plane was parallel to the orbito-meatal line. An individually fitted thermoplastic mask was utilized to minimize head motion. A 10-minute transmission scan using a 137Cs source was performed first to acquire data required for radiation attenuation corrected, and followed by an injection of 185 MBq of [18F]-2-fluoro-2-d-glucose IV over 60 seconds (total effective dose 5 mSv). The emission scan was commenced 30 minutes after tracer injection. Data were acquired in list mode, framed into 6 5-minute frames prior to reconstruction with the ordinary Poisson expectation maximization algorithm, and corrected for detector normalization, dead time, scatter, and attenuation. Frame to frame realignment was performed to account for subject head motion during the scan and a single time-integrated image was created by averaging the realigned dynamic PET images acquired 40 to 60 minutes after tracer injection. Blood sampling throughout the scan was not performed, and raw average regional FDG uptake values (Bq/mL/min) were utilized in the image analysis.
Volumetric head MRI scans were obtained within 24 hours of the PET scans on a 1.5-T GE Signa scanner (GE Medical Systems, Milwaukee, WI) also located at UBC. Two subjects had MRI and PET scanning ~12 months apart due to system or subject conditions. A T1-weighted 3D volumetric fast spoiled gradient echo inversion recovery prepared sequence was acquired in the coronal or sagittal plane (echo time = 4.8 ms; repetition time = 10.3 ms; inversion time = 450 ms; field of view 25 cm; matrix size 256 × 256; 170 contiguous 1.0-mm-thick slices). All scans were assessed with an MRI visual scale to obtain ratings of frontal and anterior lobe atrophy.15
Given the challenges of a small sample, individual variability in FDG uptake, and the likelihood of mild abnormalities in the mutation carriers, we analyzed PET images with both ROI and SPM approaches to maximize our ability to detect true regionalized carrier/noncarrier differences. We utilized ROI analysis to screen for differences in predefined brain regions. We then applied voxel-wise whole-brain SPM to independently validate the ROI findings. ROI analyses were performed utilizing MEDx. A custom ROI template was first created in coronal view on the MNI305 template16 normalized to the T1-MRI template in SPM8 (Wellcome Department of Cognitive Neurology, Institute of Neurology, University College London, http://www.fil.ion.ucl.ac.uk/spm), with reference to the resliced and normalized MRICron Brodmann template. Twelve ROIs were developed bilaterally: 1) anterior cerebral cortex included anterior cingulate cortex (Brodmann area [BA] 32), orbitofrontal cortex (BA-11), gyrus rectus (BA-25), dorsolateral prefrontal cortex (BA-46), anterior insular cortex (BA-48), and temporal pole (BA-38); 2) posterior cerebral cortex included middle temporal gyrus (BA-21), superior temporal gyrus (BA-22), angular gyrus (BA-39), and superior parietal lobule (BA-7); 3) subcortical included head of caudate12 and dorsomedial thalamus.17 For cortical ROIs, 3D 8-mm circles were placed on the area of interest on 3 consecutive slices of the Brodmann, then transferred to the MNI305 template. For subcortical ROIs and the reference ROI (the pons), the circles were placed directly on the MNI305 template. The pons was defined on 5 consecutive slices with a 12-mm circle.
Using SPM8 methods, individual MRI scans were coregistered to their PET scans, and both scans spatially normalized to the T1 MRI template. The custom ROI template was then overlaid onto the individual MRI scans. Small adjustments were made to correct for registration errors and to ensure maximal inclusion of tissue in the ROI. Finally, the individually adjusted ROI template was overlaid on the MRI-coregistered and normalized static PET image. Raw regional average FDG uptake values were extracted and averaged. Intrarater reliability on a subset of 10 scans was high (intraclass correlation coefficient [ICC] range 0.70–0.98), with lower ICCs only for superior parietal lobule (0.43) and temporal pole (0.50). Inter-rater reliability on the same scan subset was also high (ICC range 0.70–0.91), with lower ICCs for superior parietal lobule (0.52) and gyrus rectus (0.61).
We estimated regional FDG uptake values using 2 approximate methods.18 A normalized FDG uptake value was calculated for each ROI in reference to the pons: raw average ROI/raw average reference ROI. A standard FDG uptake value (SUV) was also calculated for each ROI including the pons: (raw average ROI/[injected dose/weight, kg]). We utilized averaged normalized regional FDG uptake values to compute global ratios that quantified the relationship of anterior-to-posterior (AP = averaged anterior ROI uptake divided by averaged posterior ROI uptake, bilaterally) and left-to-right (LR = averaged left-sided ROI uptake divided by the averaged right-sided ROI uptake).19
FDG-PET SPM was conducted using SPM8 implemented in Matlab 2010b (MathWorks, Inc., Natick, MA). The spatially normalized PET images were normalized to the pons value and smoothed with a Gaussian kernel with a full-width at half maximum of 5 mm, a parameter chosen to maintain a reasonable signal-to-noise ratio, correct for subtle individual anatomical variations, and detect small-scale features. A full factorial statistical model was used to assess differences between GRN mutation carriers and noncarriers. To account for individual differences, we also included analysis of covariance (ANCOVA)-by-subject factors.
DNA was extracted from peripheral blood and GRN sequencing performed at the Mayo Clinic, Jacksonville, Florida, using standard protocols.1,20 Subjects' mutation status was determined as carrier or noncarrier.
The clinical assessment included cognitive screening, motor assessments, and functional and behavioral assessments (table 1). A neurologist and neuropsychologist evaluated subjects for FTD21 and dementia22 diagnostic criteria, and determined that they did not meet these. Based on clinical impression, rating scale, and test results, they classified subjects as asymptomatic if no abnormalities were found or as clinically symptomatic no dementia (CSND) if clinical or cognitive symptoms suggestive of FTD were identified but had no impact on function. CSND is a descriptive term that has not been validated prospectively in FTD.
Demographic and clinical group differences between GRN mutation carriers and noncarriers were examined with t test and χ2 test. FDG uptake values were compared using ANCOVA to adjust for the confounding effect of sex. In primary analyses, we compared carrier/noncarrier groups on the global AP and LR ratios. In secondary analyses, we examined normalized FDG uptake values and SUVs for individual ROIs. Significance was set at p < 0.05. We utilized our ROI comparisons as a screening method for regional differences and did not correct for multiple tests. We then utilized SPM to independently validate the ROI findings. We used receiver operating characteristic curves to further evaluate FDG uptake group differences. We performed a count of ROIs where the z scores for normalized FDG uptake values (ROIsubject − ROImean noncarriers/SDnoncarriers) were ≤ −1. We examined the relationship between FDG uptake, demographic, and clinical measures with Spearman rank correlations. All statistics were performed using SPSS package (v 17.0, Chicago, IL).
GRN mutation carriers and noncarriers differed on sex but not age, education, cognitive, motor, functional, and neuropsychiatric measures. In both groups, frontal and anterior temporal lobe was deemed normal or borderline in appearance (no atrophy) on MRI, with the exception of one carrier. Five carriers (56%, mean age = 48) and 3 noncarriers (27%, mean age = 54) were classified as CSND. Overall, members from 5 GRN families participated, with unequal family representation in the mutation carrier and noncarrier groups. Years to estimated onset (subject age – mean age at FTD onset in the family) were −7 (7 years younger than mean onset in family) for mutation carriers (range −22 to +16). This is a very rough estimate given the variability of age at onset in each family (table 1).
Table 2 shows FDG uptake results. Mutation carriers had total (bilateral), left, and right lower AP ratios than noncarriers. They also had a higher LR ratio, with near equal left- and right-sided FDG uptake compared to noncarriers, who had greater right lateralization. Averaged right anterior uptake was significantly lower for mutation carriers (1.43 vs 1.56, F = 4.561, p = 0.048). The pattern of carrier/noncarrier differences on the AP and LR ratios was present for both sexes (figure 1). Differences were of comparable magnitude for CSND or asymptomatic subjects (data not shown). To address the potential confound of carrier/noncarrier family membership differences, a subanalysis of members from family R8 only, with 6 subjects in each group, was performed on global ratios, with group differences the same as for the entire sample.
Normalized FDG uptake values for right anterior cingulate, anterior insula, and gyrus rectus were lower for mutation carriers. No group differences were observed for left anterior ROIs or any posterior or subcortical ROI. ROI findings were identical for male and female subjects. When SUVs were considered, no significant carrier/noncarrier differences emerged for the pons, thus confirming the validity of this region as the reference ROI (5044.9 and 4717.5, respectively, F = 0.077, p = 0.79). There was a broader pattern of lowered anterior FDG uptake than with normalized values in mutation carriers, involving additionally left anterior cingulate, orbitofrontal cortex, and temporal pole, and left and right caudate. SPM analyses in table 3 showed extensive FDG uptake reductions in mutation carriers in the frontal lobe, particularly on the right side. We first examined carrier/noncarrier differences significant at the voxel level but no region survived the family-wise correction for multiple comparisons of pcorr < 0.05. We then searched for cluster level differences. Here we used regions with a statistical threshold of puncorr < 0.0001 (T value > 4.88) with a minimum size of kappa = 100 voxels significant at the cluster level (pcorr < 0.05). Seven significant clusters are shown in table 3 and figure 2, A and B. When we searched for differences with mutation carriers > noncarriers, no significant findings emerged.
The right AP ratio had the highest area under the curve (AUC) = 0.78 (95% confidence interval [CI] 0.57–0.99). When a right AP ratio cutoff of 0.88/0.89 was utilized, sensitivity was 73%, specificity 78% in classifying carriers and noncarriers. Among single ROIs, the right anterior insula achieved an AUC = 0.76 (95% CI 0.54–0.98). Mutation carriers had nearly twice as many anterior ROIs with reduced FDG uptake (z ≤ −1) than noncarriers (3.7 vs 1.9, respectively, p = 0.05). There was considerable inter- and intraindividual variability in regional patterns of FDG uptake in carriers (figure e-1 on the Neurology® Web site at www.neurology.org), greater in anterior than posterior ROIs. In mutation carriers, right AP ratio was correlated with modified Mini-Mental State Examination and Functional Rating Scale scores (r = 0.74, p = 0.02; r = −0.73, p = 0.03) but not with diagnostic status, age, or years to estimated onset. When plotted against years to estimated onset (figure e-2), the ratio showed high variability among mutation carriers >10 years from estimated onset.
This study has revealed brain glucose metabolism abnormalities in GRN mutation carriers from 4 different families, on average 7 years younger than mean age at onset in their family, with about half deemed asymptomatic and half CSND. Globally, there was a lowered AP ratio resulting from reduced glucose metabolism in frontal and anterior temporal lobe regions. There was also less left-right asymmetry in our GRN mutation carriers, which we interpret as an indication of more marked metabolic abnormalities in the right hemisphere in light of reports of increased right-sided regional or global tracer uptake in normal subjects on FDG-PET and HMPAO SPECT.14,23 These global abnormalities were present in both asymptomatic and CSND carriers. Concurrent localization with ROI and SPM analyses highlighted right medial and ventral regions in the frontal lobe. Inspection of individual patterns showed frequent abnormalities in the caudate. Unbiased SPM further revealed a right posterior cerebellar region with lowered glucose metabolism. No abnormalities were noted in posterior cerebral regions. The right lowered AP ratio was a sensitive and specific marker of GRN mutation carrier status over a wide range of ages, and correlated with cognitive and clinical scales.
Our findings generally align with existing neuroimaging evidence on subjects with GRN-associated FTD, who typically display a pattern of severe, asymmetric atrophy involving frontal, temporal, and parietal lobes.7,9 None of our cases showed the parietal lobe involvement reported in established disease,8 in line with a longitudinal case study suggesting earlier abnormalities in frontotemporal than posterior regions. The pattern of right-sided frontal abnormalities we observed may be more associated with future bvFTD than other clinical variants. In fact, left-sided lateral temporal abnormalities on FDG-PET have been indicated as early manifestations of PNFA.11 It is possible that our use of FDG-PET, a highly sensitive marker of metabolic dysfunction,24,25 allowed us to detect very early signs of GRN-related neurodegeneration that have not been detected on volumetric MRI of predementia mutation carriers.10 Several authors have stressed the severity of the neurodegeneration associated with GRN mutations.7,19,26,27 This observation is consistent with either a particularly aggressive pathophysiologic process or one extended over many years. Our findings are in agreement with the latter interpretation.
Based on the specific pattern of medial frontal hypometabolism in our GRN mutation carriers, we speculate that most will eventually develop bvFTD. Hypometabolism in medial frontal networks involving anterior medial frontal cortex, gyrus rectus, and insula has been depicted as the neural substrate for behavioral impairments including apathy, disinhibition, and eating abnormalities in subjects with FTD.28 In brain-behavioral studies of FTD, the right orbitofrontal cortex has emerged as a regulator of behavior in relation to a predominantly right-sided network involving the insula and the striatum. The disruption of this network is believed to account for abnormalities in social, emotional, and impulse regulation.29 Our finding of bilateral caudate involvement further strengthens this suggestion. Our finding of hypometabolism in the cerebellar semilunar lobule may not be comparable to the extensive bilateral cerebellar atrophy in FTD associated with C9ORF72 mutations.8 The most reasonable interpretation in early FTD-GRN is crossed cerebellar diaschisis.
The strengths of this study are the careful characterization of GRN mutation carriers and noncarriers from the same pool of families and the use of multiple methods for the analysis of PET images. We did not undertake partial volume (PV) correction of the PET images because of ongoing debates as to appropriate methods, and instead applied a visual rating scale to assess atrophy on MRI. While mutation carriers did not display ratings different from noncarriers, the potential of PV effects remains and limits the interpretation of our findings solely in terms of glucose metabolic abnormalities. Other limitations included our small sample size and the unequal representation of the GRN families in carrier/noncarrier groups. This may have introduced biases due to genetic discordances beyond the GRN gene mutation with effects on cerebral glucose metabolism.30 The finding of carrier/noncarrier differences in members from a single family, however, reduces the likelihood of these. Limitations also include the use of parametric methods (ANCOVA) we had to resort to because of a significant sex imbalance between our groups: the predominance of women in our mutation carrier and men in our noncarrier group represented a serious risk of Type II error because of the normal tendency for women to have higher glucose metabolic rates globally and in ventral and medial regions of the frontal lobe, of direct interest in the current study.13,14
This study documents GRN mutation–related brain metabolic abnormalities prior to onset of full-blown FTD, in the absence of visible frontal and anterior temporal atrophy on MRI. Future longitudinal studies are needed to document the localization and progression of abnormalities to full-blown FTD in larger samples. FDG-PET is available as a clinical test in differentiating FTD from Alzheimer disease. Our study supports its potential utility in detecting brain metabolic changes prior to dementia onset. Thus, FDG-PET may be useful in future therapeutic trials of progranulin modulation in GRN mutation carriers, where strict monitoring of neurodegeneration is required.31
C. Jacova: drafting the manuscript, study concept, statistical analysis, image analysis, interpretation of data. G.Y.R. Hsiung: revising the manuscript, study concept, image analysis, and interpretation of data. I. Tawankanjanachot: revising the manuscript, image analysis. K. Dinelle: revising the manuscript, image analysis. S. McCormick: revising the manuscript, image analysis. H. Lee: revising the manuscript, image analysis. P. Sengdy: revising the manuscript, data acquisition. P. Bouchard-Kerr: revising the manuscript, data acquisition. M. Baker: revising the manuscript, genetic analysis. R. Rademakers: revising the manuscript, study concept, genetic analysis. V. Sossi: revising the manuscript, study concept, image analysis. A.J. Stoessl: revising the manuscript, study concept, image analysis. H.H. Feldman: revising the manuscript, study design, concept and supervision, interpretation of data, obtaining funding. I.R.A. Mackenzie: revising the manuscript, study design, concept and supervision, interpretation of data, obtaining funding.
Supported by funding from the Canadian Institutes of Health Research operating grant #179009 and #74580, the Pacific Alzheimer's Research Foundation (center grant C06-01), and the National Institutes of Health R01 NS065782 to Rosa Rademakers in support of research on frontotemporal dementia. Claudia Jacova and Robin Hsiung are supported by funding through the Ralph Fisher Professorship in Alzheimer's Research (Alzheimer Society of British Columbia, Canada). Robin Hsiung is supported by a CIHR Clinical Genetics Investigatorship award.
C. Jacova is a recipient of a British Columbia Ministry of Health grant and salary support from the Ralph Fisher Professorship in Alzheimer's Research through the Alzheimer Society of British Columbia, Canada. She is a founding member of the International Society for Frontotemporal Dementia. G. Hsiung is the recipient of a Clinical Genetics Investigatorship award from the CIHR. Dr. Hsiung has received research support as a clinical trials site investigator from Baxter, Bristol-Myers-Squibb, Elan, Janssen-AI, Pfizer, Hoffman-La Roche, and Genentech. I. Tawankanjanachot received salary support in 2008-2009 for this study through the Pacific Alzheimer Research Foundation #C06-01 grant. K. Dinelle and S. McCormick report no disclosures. M. Gonzalez is recipient of support from the Natural Sciences and Engineering Council of Canada and from the Parkinson Society of Canada. H. Lee reports no disclosures. P. Sengdy receives salary support from Canadian Institutes of Health Research operating grant #179009. P. Bouchard-Kerr receives salary support from Canadian Institutes of Health Research operating grant #179009. M. Baker reports no disclosures. R. Rademakers receives research support from the NIH (R01 NS065782, R01 AG02651, R01 NS080882, and P50 AG16574), the ALS Therapy Alliance, CurePSP, and the Consortium for Frontotemporal Degeneration Research. Dr. Rademakers further received honoraria for lectures or educational activities not funded by industry. She serves on the medical advisory board of the Association for Frontotemporal Degeneration and is a founder of the International Society for Frontotemporal Dementia. V. Sossi reports no disclosures. A. Stoessl's research is supported by the Canadian Institutes of Health Research, the Michael J. Fox Foundation, the Pacific Alzheimer Research Foundation, and the Canada Research Chairs program. He has served on advisory boards or committees (unpaid) for Canada Research Chairs, the Movement Disorders Society, the Parkinson Study Group, and Association for Parkinsonism & Related Disorders and on editorial boards for Annals of Neurology, Lancet Neurology, Parkinsonism & Related Disorders, and Translational Neurodegeneration. He has received speaker fees from Abbott, Medscape, and Teva and has served as a consultant or on advisory boards for Abbott, Biogen Idec, Bioscape Imaging, Medgenesis Kyowa/ProStrakan, Ono Pharma, and UCB. H. Feldman was a full-time paid employee of Bristol-Myers Squibb on leave from University of British Columbia from 2009 to 2011. He received stocks and stock options in this role from Bristol-Myers Squibb. In 2012, he returned to University of British Columbia, Division of Neurology, Faculty of Medicine. In the past 2 years, Dr. Feldman has served as paid consultant and received travel expenses for Scientific Advisory Boards with Lilly (2012) and Kyowa Hakko Kirin (2012). He received an honorarium for lecturing from Danon Nutricia (2012). He also received honoraria and related travel expenses for consulting and/or lecturing for non-industry activities including the Bluefield Project on Frontotemporal dementia (2012), the Alzheimer Association of Saudi Arabia (2012), the Alzheimer Society of Manitoba (2012), the Department of Neurology Columbia University, and the Institute for Clinical and Economic Review Massachusetts General Hospital's Institute for Technology Assessment. He received travel grant or travel support for lecturing at the 2011 World Congress of Neurology, the Alzheimer Society of Canada Biomedical Peer Review Panel, New York Academy of Science Alzheimer Disease Leadership Council, Latin American Cognitive Impairment Meeting, Alzheimer Association Research Roundtable, Yale University Department of Neurology, and Fidelity Biosciences. He has served as a member of the editorial board or associate editor of Dementia and Geriatric Cognitive Disorders and Journal of Neurological Sciences. He is a co-patent holder of US Serial Number PCT/2007/070008 Detecting and Treating Dementia and has received royalties for the book Atlas of Alzheimer's Disease (2007), published by Informa Health. He has received funding for this research from grants from the Canadian Institutes of Health Research #00725-000 (2008-2012), #179009 (2008-2013), #74580 (2005-2008), #73376 (2005-2006), and the Pacific Alzheimer Research Foundation #C06-01 (2006-2011). I. Mackenzie is a recipient of a Canadian Institutes of Health Research operating grant (#179009). Go to Neurology.org for full disclosures.