RESUMEN
OBJECTIVE: To investigate neurodegenerative and inflammatory biomarkers in people with amyotrophic lateral sclerosis (PALS), evaluate their predictive value for ALS progression rates, and assess their utility as pharmacodynamic biomarkers for monitoring treatment effects. METHODS: De-identified, longitudinal plasma, and cerebrospinal fluid (CSF) samples from PALS (n = 108; 85 with samples from ≥2 visits) and controls without neurological disease (n = 41) were obtained from the Northeast ALS Consortium (NEALS) Biofluid Repository. Seventeen of 108 PALS had familial ALS, of whom 10 had C9orf72 mutations. Additional healthy control CSF samples (n = 35) were obtained from multiple sources. We stratified PALS into fast- and slow-progression subgroups using the ALS Functional Rating Scale-Revised change rate. We compared cytokines/chemokines and neurofilament (NF) levels between PALS and controls, among progression subgroups, and in those with C9orf72 mutations. RESULTS: We found significant elevations of cytokines, including MCP-1, IL-18, and neurofilaments (NFs), indicators of neurodegeneration, in PALS versus controls. Among PALS, these cytokines and NFs were significantly higher in fast-progression and C9orf72 mutation subgroups versus slow progressors. Analyte levels were generally stable over time, a key feature for monitoring treatment effects. We demonstrated that CSF/plasma neurofilament light chain (NFL) levels may predict disease progression, and stratification by NFL levels can enrich for more homogeneous patient groups. INTERPRETATION: Longitudinal stability of cytokines and NFs in PALS support their use for monitoring responses to immunomodulatory and neuroprotective treatments. NFs also have prognostic value for fast-progression patients and may be used to select similar patient subsets in clinical trials.
Asunto(s)
Esclerosis Amiotrófica Lateral/diagnóstico , Esclerosis Amiotrófica Lateral/metabolismo , Citocinas/metabolismo , Progresión de la Enfermedad , Proteínas de Neurofilamentos/metabolismo , Adulto , Anciano , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/fisiopatología , Bancos de Muestras Biológicas , Biomarcadores/sangre , Biomarcadores/líquido cefalorraquídeo , Proteína C9orf72/genética , Citocinas/sangre , Citocinas/líquido cefalorraquídeo , Femenino , Humanos , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Proteínas de Neurofilamentos/sangre , Proteínas de Neurofilamentos/líquido cefalorraquídeo , PronósticoRESUMEN
OBJECTIVE: We measured the levels of mutant huntingtin (mtHtt) and total huntingtin (tHtt) in blood leukocytes from Prospective Huntington At-Risk Observational Study (PHAROS) subjects at 50% risk of carrying the Huntington disease mutation using a homogeneous time-resolved fluorescence (HTRF) assay to assess its potential as a biomarker. METHODS: Peripheral blood mononuclear cells from consenting PHAROS subjects were analyzed by HTRF using antibodies that simultaneously measured mtHtt and tHtt. mtHtt levels were normalized to tHtt, double-stranded DNA, or protein and analyzed according to cytosine-adenine-guanine repeat length (CAGn), demographics, predicted time to clinical onset or known time since clinical onset, and available clinical measures. RESULTS: From 363 assayed samples, 342 met quality control standards. Levels of mtHtt and mt/tHtt were higher in 114 subjects with expanded CAG repeats (CAG ≥ 37) compared with 228 subjects with nonexpanded CAG repeats (CAG <37) (p < 0.0001). Analysis of relationships to predicted time to onset or to phenoconversion suggested that the HTRF signal could mark changes during the Huntington disease prodrome or after clinical onset. CONCLUSIONS: The HTRF assay can effectively measure mtHtt in multicenter sample sets and may be useful in trials of therapies targeting huntingtin.
Asunto(s)
Transferencia Resonante de Energía de Fluorescencia/métodos , Enfermedad de Huntington/sangre , Enfermedad de Huntington/patología , Leucocitos Mononucleares/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Observación , Adulto , Ensayos Clínicos como Asunto , Método Doble Ciego , Femenino , Lóbulo Frontal/metabolismo , Lóbulo Frontal/patología , Humanos , Proteína Huntingtina , Enfermedad de Huntington/genética , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Mutación/genética , Proteínas del Tejido Nervioso/genética , Cambios Post Mortem , Estudios Retrospectivos , Expansión de Repetición de Trinucleótido/genéticaRESUMEN
A means for measuring levels of soluble huntingtin proteins in clinical samples is essential for assessing the biological effects of potential mutant huntingtin (mtHtt) modifying treatments being developed for Huntington's disease (HD). We have optimized a previously described cell-based Homogeneous Time Resolved Fluorescence method that can measure soluble mtHtt and its ratio to the total Htt (tHtt) in blood buffy coats [1]. The results of the optimization and assay qualification indicate the assay to be specific for mtHtt in HD compared to Control subjects, highly sensitive, and technically and biologically reproducible. We therefore generated a Good Laboratory Practice Standard Operating Procedure which we validated, using 30 HD and 8 control buffy coat samples in which significant differences in mtHtt levels were found. We intend to deploy the assay to evaluate sample sets from observational and therapeutic studies enrolling HD subjects to further validate soluble mtHtt measurement by HTRF as a biomarker for HD and to explore its potential uses.
