Mary K. Floeter - US grants

Over the past several years, our research has focused on physiological changes in cortical and spinal motor circuits in patients with primary lateral sclerosis (PLS). In collaborative studies with Dr. Ou Bai, we identified oscillatory EEG signals that preceded movement in PLS patients in whom the slow DC potentials generated by cortical neurons were diminished. In FY08, we report that single-trial changes in these oscillatory signals with imagined movement can provide a control signal for an EEG-based brain-computer interface in a small number of PLS patients. This work is being extended to amyotrophic lateral sclerosis (ALS) patients in the coming year. A second line of study is aimed at assessing the extent of (non-motor) frontal cortical dysfunction in PLS patients, and to compare them to patients with ALS. This 3-year protocol includes neuropsychological tests, psychiatric assessment, and quantitative MRI imaging, and is in its second year. Lastly, recognizing that PLS is likely to be complex disorder, we are collecting samples from PLS patients for the NINDS DNA repository for motor neuron diseases at Coriell, so that this rare population will be accessible to the general scientific community for further studies.

Clinical, laboratory and therapeutic studies were conducted to determine etiology (autoimmunity, neurotoxicity, genetics) of various neuromuscular diseases and design, or apply, effective therapies. Studies in FY07 involved patients with: a) inflammatory myopathies with emphasis on inclusion body myositis (IBM); b) inherited vacuolar myopathies with emphasis on hereditary IBM due to GNE mutations and desmin-related myopathies; c) demyelinating polyneuropathies; and d) the stiff-person syndrome(SPS). [unreadable] [unreadable] In inflammatory myopathies, the specificity of the T cell Receptors and the in situ clonal expansion of the endomysial T cells are examined longitudinally. The studies have shown that in IBM the T cells are driven by specific antigens. To search for putative antigen(s), T cell clones have been established from the endomysial T cell infiltrates; candidate immunodominant peptides that drive the T cell responses and serve as autoantigens are currently explored using combinatorial peptide libraries. It has been found that in IBM chemokines and costimulatory molecules such as ICOS, ICOS-L and PDI are upregulated and the muscle fiber may function as Antigen Presenting cell. Because cytokines share common antigenic determinants with the Alzheimer-like beta-APP amyloid deposits, an interrelationship between these molecules was explored. A linear relationship was found between cytokines, chemokines and amyloid-related degenerating molecules not only in vivo in the patient's muscles but also in vitro in human myotubes. At the clinical level, a longitudinal study examining the natural history of IBM has been completed. To suppress the myocytotoxic effect of T cells, a therapeutic and investigational clinical trial has begun using CAMPATH, a humanized monoclonal antibody that induces a sustained depletion of mature T cells allowing for toleragenic T cell responses. The study has been completed and is being analysed.[unreadable] [unreadable] In demyelinating neuropathies associated with IgM autoantibodies to MAG and glycolipids,a new controlled therapeutic study was performed using Rituximab, a humanized monoclonal antibody against B cell clones. The study is now completed and the data are being analysed. Preliminary analysis demonstrates that Rituximab is effective in the disease in some subgroups and exerts its action by enhancing immunoregulatory T cells.[unreadable] [unreadable] In an effort to find responsible autoantigens in patients with Stiff Person Syndrome (SPS), T cell clones were established from the CSF and being tested against combinatorial peptide libraries. Using proteomics in the patients' serum, the antigenic peptide GABARAP (GABA-Receptor Associated Protein), was found to be reduced. Further, anti-GABARAP antibodies were detected in up to 65% of SPS patients. These antibodies may destabilize the GABAA receptors and play a role in inducing the dysfunction of GABAergic pathways and the reduced level of GABA in the patients' brains as demonstrated with MRS spectroscopy. A new double-blind clinical trial using the B cell-depleting monoclonal antibody Rituximab has been completed and all 23 patients have been enrolled. The data is undergoing analysis. The origin of phobias, a common feature in SPS patients, was being explored using a series of neurocognitive measurements. It was found that the phobias are secondary to disability and not due to the primary disease.[unreadable] [unreadable] In patients with hereditary IBM due to mutations in the GNE gene we observed a defect in glycosylation of muscle proteins and reduction of alpha-dystroglycan. A pilot clinical trial using intravenous immunoglobulin in an effort to increase muscle glycosylation was completed in 4 patients. The results are promising and are currently analyzed. A phenotype/genotype correlation has been completed in patients with hereditary myopathies due to pathogenic mutations in the desmin gene. It has been concluded that desmin myopathy is a distinct disease affecting intermediate filaments and the type of mutations may dictate clinical severity or presence of cardiomyopathy.

The Office of the Clinical Director (OCD) provides centralized oversight and support for conducting clinical research in the NINDS IRP, and provides neurological expertise and consultations for all NIH institutes that carry out clinical research in the Clinical Center. The OCD assures the quality of clinical care provided in NINDS clinical research protocols by coordinating the credentialing process and competency reviews of NINDS licensed health care providers. Quality assurance activities include chart reviews, monthly random audits of clinical research protocols for adherence with consent processes and medical documentation, and a monthly clinical care conference to review complications, occurrences, and adverse events. An annual orientation is held for new clinical fellows each July. The staff of the OCD are active participants in the Clinical Center's Quality Assurance Committee, communicating changes in hospital policies and practices to the NINDS clinical staff. The Neurology Consultation Service provides in-house physician consultations on a 24 hr/day, 7 day/week basis. The EEG Section and EMG section provide scheduled diagnostic testing each weekday, with emergency EEG coverage on a 24/7 basis. The EEG section also provides intraoperative monitoring services, and runs an active diagnostic sleep laboratory. Additional specialty consultative services, such as neuropsychological testing, pediatric neurological consultation, and muscle biopsies and consultations are made available on an as-needed basis. All clinical protocols of NINDS investigators are reviewed by a biostatistician and a scientific review committee to ensure that the research carried out meets high scientific standards. The OCD organizes the protocol review process, with monthly meetings of the convened committee, and ad hoc reviews arranged to provide expertise in specialty areas. The NINDS serves as the lead institute for the administration of the Combined Neuroscience Institutional Research Board, which reviews human subject protocols for six institutes that carry out neuroscience research at NIH. CNS IRB staff provide training and advice to investigators to navigate the protocol approval process. OCD contributes budget and personnel to support the work of the CNS IRB. The OCD carries out additional auditing and reviews on an as-needed basis through internal and external committees and contracts. In FY10, OCD supported the formation and training of a monitoring committee to provide external monitoring of clinical protocols with investigational new drugs(INDs) or devices (IDEs). The OCD also supported Good Clinical Practice Training for Principal Investigators (PIs) with protocols with INDs or IDEs, and arranged a lecture series on Database Management Systems for PIs. The OCD also provides support for common needs that cross multiple clinical research programs of NINDS. OCD personnel who provide scientific support include biostatisticians who advise on the design of clinical protocols and data analysis, and an imaging scientist to facilitate work of investigators using magnetic resonance imaging. Clinical care support may be provided on a limited or short-term basis to investigators to extend their program resources. The OCD also provides budgetary support for outside medical services, such as genetic testing for research purposes, and for patient services such as lodging and travel to participate in clinical protocols. The OCD organizes a weekly NINDS Grand Rounds from September to June each year with internal and outside speakers, and arranges continuing medical education credit for attendees. In addition, it coordinates residency and medical student elective rotations, to give trainees from other institutions opportunities to participate in clinical research. The OCD also contributes to the education of allied health professionals, with support of the annual review course for neuroscience nursing, and partnership with the Clinical Center for a periodic neuroscience nursing internship program. Recruitment activities are held at annual neurology meetings, and mailings and journal ads to recruit clinical fellows are carried out. The OCD also supports the development of new clinical investigators in the IRP with the Henry McFarland Clinical Transition to Independence Award, an award that bridges clinical fellowship and new investigator roles. In FY10, the first award was given.

Although primary lateral sclerosis (PLS) is generally considered to be a motor neuron disorder, its relationship to amyotrophic lateral sclerosis (ALS) and other motor neuron disorders is uncertain. PLS differs from ALS in its duration, with a median survival of more than a decade, in contrast to the median survival of 3-5 years in ALS. The long survival is closely linked to the restriction of disease to the corticospinal, or upper motor neurons, of the brain. We carried out a longitudinal study in PLS patients that showed that once each body region exhibited symptoms, measures of clinical severity of that region declined more rapidly in the first years and subsequently stabilized without continued progression. This finding suggests that loss of corticospinal input may progress in an intermittent fashion. It also indicates that progression is not linear, arguing against using the slope of clinical decline as an outcome in future clinical trials and highlighting the need for biological markers of disease activity. We have completed a multi-year cross-sectional study to evaluate the hypothesis that ALS and PLS are variants of a common neurodegenerative process that differ phenotypically because of regional differences in pathology. We first looked for overlap and differences in clinical dysfunction corresponding to non-motor cortical regions in PLS and ALS patients with similar extents of upper motor neuron dysfunction. Neuropsychological tests and psychiatric assessments showed that cognitive impairment is less common in PLS, and that PLS patients have a modestly greater extent of depression compared to ALS patients. To correlate clinical measures to regional anatomical measures, diffusion tensor MRI imaging and measures of cortical thickness have been obtained in ALS and PLS patients and healthy age-matched controls. We previously showed that quantitative measures of the corticospinal tract can be made from DTI images with good intra- and inter-rater reliability and test-retest reliability in controls. We are currently completing analysis of these data and are obtaining longitudinal imaging studies on a subset of the PLS and ALS patients in the cross-sectional study.

Our current research focuses on patients with motor neuron disorders, particularly disorders affecting corticospinal (upper) motor neurons such as primary lateral sclerosis (PLS). It has been proposed that PLS, amyotrophic lateral sclerosis (ALS), and frontotemporal dementia (FTD) are related disorders that represent different phenotypes on the same spectrum of neurodegenerative disorders. Pathological studies by other investigators found the same proteins within neuronal inclusions in brains of ALS and a subset of FTD patients, and in genetic studies, a mutation in the gene C9ORF72 was recently found to account for a significant portion of familial ALS and FTD. It has been hypothesized that PLS is a variant of these disorders in which the degenerative process remains largely limited to the motor cortex, failing to progress to frontotemporal cortex and the spinal cord. If this hypothesis is correct, understanding what limits the extent of disease in PLS patients may provide clues to mechanisms for halting progression of neurodegeneration. The first aim of this project is to understand the relationship between PLS and other disorders on the motor neuron disease spectrum. To determine whether clinical phenotypes are a function of the brain regions undergoing degeneration, work over the last few years has utilized quantitative magnetic resonance imaging (MRI). During FY13, we completed and published an analysis of white matter tracts associated with cognitive performance in ALS and PLS patients, using a semi-automated atlas-based analysis to compare diffusion tensor imaging (DTI) measures in eleven white matter association tracts. Changes in diffusivity measures in the superior longitudinal fasciculus, cingulum, and portions of the corpus callosum were found to be associated with performance on particular executive function and memory tests. Performance was not related to focal changes in grey matter volume, highlighting the importance of the integrity of axonal tracts for proper temporal encoding of information in cognitive networks. In collaboration with the NINDS Cognitive Neuroscience section, we are now extending that work to include comparisons with FTD patients studied with the same methods, with a goal of identifying imaging measures that discriminate between patients with different diagnoses and clinical presentations. We have also begun examining whether other non-motor manifestations, such as pseudobulbar affect, can be explained by alterations in particular white matter networks. To examine the question whether disease begins focally in PLS and spreads thorough axonal networks or by propagation to adjacent areas of the brain and spinal cord, we began a chart review of the cohort of PLS patients followed in our clinic to document how symptoms spread. A second aim of the project is to identify reliable, non-invasive markers for detection and measuring progression of brain involvement in motor neuron disorders. During FY13, we published a longitudinal MRI study in ALS and PLS patients that found that changes in DTI metrics and measures of cortical thickness and atrophy progress over different time scales. We hypothesized that MRI findings represent a sequence of structural changes that occur with axonal breakdown and clearance, followed by measureable thinning and atrophy of cortical grey matter. Differences between ALS and PLS patient groups may represent early and late stages of the same process, with longer time for structural remodeling to evolve in PLS patients. Changes in white matter DTI measures occurred relatively early. We are currently looking to see whether functional MRI resting state networks show alterations in motor neuron disease, and will be assessing whether functional changes can be detected earlier than changes in diffusion measures. To date, most imaging studies in motor neuron disease describe differences between groups of patients and controls. It is a challenge to develop imaging measures that are indicators of the state of upper motor neuron integrity in individual patients. As a first step, we are currently obtaining multi-modal imaging in repeated sessions in a cohort of healthy controls in order to determine the intersession variability of candidate imaging markers. These data will be used to select the most informative techniques for prospectively assessing patients with motor neuron disorders in the future. In addition to the two primary aims of this project, we participated in two collaborative studies in FY13. We continued as a site in a collaborative study to examine the role of oxidative stress in progression of motor neuron diseases organized by Columbia University. The study was fully enrolled at our site and all patients completed their annual visits. Two patients completed the entire study, and our site is on track for completion in two years. Additionally, we began a new collaboration with the NINDS Section on Infections of the Nervous System to look for evidence of endogenous retroviruses in motor neuron disease patients. Lastly, in collaboration with the NIA Neuromuscular Diseases Section, we have laid the groundwork for a natural history and biomarker study of symptomatic and presymptomatic carriers of the C9ORF72 gene mutation. The human subject research protocol has now been approved by the IRB and we anticipate enrollment of the first patient before the end of FY13.

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are adult-onset neurodegenerative disorders in which up to one-third of patients have overlapping clinical and pathological features. A repeat expansion mutation in the C9orf72 gene is the commonest cause of familial ALS and familial FTD in the US and also accounts for a 5-8% of ALS cases thought to be sporadic. In 2013, we began a prospective longitudinal study of persons who carry the C9orf72 mutation that includes asymptomatic carriers, patients with ALS, and patients with FTD. The first goal is to understand the natural history of C9orf72-related disease: how quickly weakness and cognitive dysfunction progress, whether clinical presentation influences survival, and whether subtle motor or cognitive abnormalities are detectable prior to the onset of definite symptoms. The second aim of the study is to explore candidate biomarkers of disease progression. Biomarkers that signal improvement or decline in disease activity before clinically evident deterioration are need for clinical trials. Imaging, physiology, and biofluids are being collected at 3 visits over 18 months. Phone surveys are conducted for 36 months. Biospecimens obtained are being shared with collaborators within and outside NIH, to facilitate translational research. Enrollment was completed in FY2018, with 50 C9orf72 mutation carriers. This included 31 patients with ALS/ALS-FTD and 15 asymptomatic carriers. To evaluate clinical progression, measures of motor, cognitive and behavioral function were obtained at baseline, and at follow-up visits at 6 and 18 months. Clinical, imaging, and physiological findings from the first half of the enrolled participants have been published. Imaging shows more extensive volume loss of extramotor areas in C9orf72 carriers with cognitive dysfunction. Ventricular enlargement and spread of white matter diffusion alterations could be detected over a 6-month period. Biospecimens were shared with collaborators at academic institutions and pharmaceutical companies, who found stable CSF levels of the repeat-associated dipeptide in longitudinal specimens, supporting its use as a pharmacodynamic marker. Collaborators found that levels of CSF phosphorylated neurofilament heavy chain were predictive of survival. To examine whether oligogenic inheritance may be a factor affecting the diversity of clinical phenotypes, exome sequencing was carried out in FY18, and analysis is ongoing. Additional activities include preparation of de-identified clinical datasets for data sharing from those participants who have completed the protocol.

The traditional classification of neurodegenerative disorders into discrete diseases has been challenged by scientific discoveries showing overlapping clinical symptoms, genes, and pathology. A clinical continuum is particularly notable for neurodegenerative disorders associated with the intracellular aggregation of misfolded TDP-43 or microtubule-associated tau proteins. Patients can have a mixture of cognitive, language, and motor symptoms. Examples of these disorders include amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), corticobasal syndrome (CBS), progressive supranuclear palsy (PSP), primary progressive aphasia (PPA), and HD-negative Huntington. Some causative genes have been found for each disorder, although most patients have sporadic disease. Interestingly, the same genetic mutation can produce different clinical phenotypes. The link between the gene mutations and pathology in these disorders may lie in the molecular and cellular processes affected by mutant gene products. As the field of neurology moves more towards the molecular characterization of neurodegenerative disorders, there is a need to understand clinical phenomenology in the context of pathology and genetic or epigenetic defects that are the drivers of disease. Given the broad spectrum of neurological symptoms in this family of disorders, the goal of this clinic is to bring together an outstanding and energetic group of physicians and researchers with a wide range of clinical expertise and scientific approaches. The objective is to gain knowledge that will set the stage for precision medicine in which therapy will be tailored to the underlying cause of each patient's disease. Considerable work was carried out during FY17 to prepare for the clinic, including hiring a nurse practitioner, obtaining IRB approval to carry out clinical screening and collection of biospecimens on patients with neurodegenerative disorders, staff training, and establishing case report forms, recruitment materials, and an electronic data system. In FY18, enrollment of patients began in a monthly screening clinic. Clinical staff also provided support for two ongoing protocols. The clinic holds a monthly conference for the neurologists, neuropsychologists, and health personnel involved in the clinic to discuss individual patients and arrive at a consensus.