Kathryn Chase

Bio: Kathryn Chase is an academic researcher from University of Massachusetts Medical School. The author has contributed to research in topics: Huntingtin & Huntingtin Protein. The author has an hindex of 20, co-authored 28 publications receiving 5978 citations. Previous affiliations of Kathryn Chase include University of Massachusetts Amherst & Harvard University.

Aggregation of Huntingtin in Neuronal Intranuclear Inclusions and Dystrophic Neurites in Brain

Abstract: The cause of neurodegeneration in Huntington's disease (HD) is unknown. Patients with HD have an expanded NH2-terminal polyglutamine region in huntingtin. An NH2-terminal fragment of mutant huntingtin was localized to neuronal intranuclear inclusions (NIIs) and dystrophic neurites (DNs) in the HD cortex and striatum, which are affected in HD, and polyglutamine length influenced the extent of huntingtin accumulation in these structures. Ubiquitin was also found in NIIs and DNs, which suggests that abnormal huntingtin is targeted for proteolysis but is resistant to removal. The aggregation of mutant huntingtin may be part of the pathogenic mechanism in HD.

Therapeutic silencing of mutant huntingtin with siRNA attenuates striatal and cortical neuropathology and behavioral deficits

Abstract: Huntington's disease (HD) is an autosomal dominant disease caused by a CAG repeat expansion in the Htt gene (1). Mutant Htt causes neuronal death, dementia, and movement dysfunction; there is no effective treatment. In an inducible transgenic mouse model of HD, turning off transgene expression reversed neuropathology and motor deficits (2). Lowering mutant Htt gene expression in brain may treat HD. In mice, viral vector delivery of short hairpin RNAs (shRNAs) against mutant Htt gene exon 1 or genes that cause other neurodegenerative disorders reduced neuropathology and motor deficits (3–10). Brain delivery of adeno-associated virus (AAV)-shRNA against mutant Htt improved signs of disease in HD transgenic models (7, 11). In the inaugural study on RNAi targeting Htt in vivo, shRNA against Htt in AAV2, delivered to the N171–82Q transgenic model of HD, improved ambulation at 4 months and rotarod performance at 10 and 18 weeks after injection (7). Five and one-half months after shRNA administration, quantitative RT-PCR revealed a 50% reduction in striatal Htt mRNA. Statistical changes in quantification of Htt protein reduction and inclusions were not reported. AAV5 delivery of shRNA against Htt in the R6/1 murine model of HD showed a 25% decrease in Htt protein and an 80% reduction in Htt mRNA 10 weeks after shRNA injection (10). The shRNA delayed onset of clasping by 2 weeks (20–22 weeks), and treated mice had fewer clasps. No difference in rotarod performance was detected. Inclusion size and number decreased in the striatum, but not in the cortex, compared with the corresponding contralateral brain regions. The authors provided an important caveat that one of the shRNAs had off-target effects; the cause of the off-target effects was not established. shRNA in AAV2 or AAV5 was used to target EGFP to knock down EGFP-Htt in another transgenic model of HD (11). shRNA reversed pathology after the onset of pathologic changes; however, behaviors were not studied. Administration of large amounts of siRNA against Htt in a Lipofectamine 2000 suspension into the lateral ventricle of newborn R6/2 transgenic mice (exon 1 of Htt) reduced whole-brain levels of mutant Htt in two mice and Htt mRNA up to 7 days posttreatment, delayed the onset of clasping, rotarod, and open-field phenotypes, and improved survival (12). Statistical quantification of neuropathology was not reported. Thus, prior studies examining RNAi against Htt provided the groundwork for therapeutic gene silencing in HD. Most of the studies used viral delivery of shRNA, and the study using siRNA required liposome delivery to newborns, with the potential liposome neuronal toxicity. Caveats attend the use of shRNAs, which can be toxic when integrated into the host genome (13, 14), in part because shRNA production is unregulated. Long siRNAs (>29 nt) and shRNAs are prone to activate off-target gene expression (15). For patient safety, shRNA will need to be able to be switched off, currently a hurdle in viral delivery systems. An alternative strategy for HD therapy is the use of small-interfering RNAs (siRNAs), ≈21-nt RNA duplexes. siRNA has been administered into cerebroventricles, vasculature, intrathecal space, and parenchyma (16–20). siRNAs were found effective and safe when introduced into mice and non-human primates (19, 21, 22). Several limitations impede progress in using siRNAs as a treatment for HD: entry and effectiveness in adult neurons without the use of potentially toxic transfection reagents; a clear demonstration that gene silencing reduces protein expression; and an improvement in behavioral deficits and neuropathology, especially neuron survival. Because bioactive molecules conjugated to cholesterol have improved cellular uptake in vitro (23), LDL receptors have been detected in brain (24), and cholesterol conjugation enhances siRNA uptake in cells outside of the central nervous system (16), we speculated that cholesterol-conjugated (cc) siRNA might enter neurons in vivo. An in vivo, rapid-onset model of HD would be optimal to test gene silencing in brain. The current rapid mouse model of HD shows mutant Htt-induced pathology after 2 months. Transgenic mice expressing exon 1 of mutant Htt [R6/2 (25)] develop nuclear inclusions throughout the brain at 2 months of age and exhibit a rapidly progressing, severe phenotype. Other transgenic or knock-in mice expressing mutant Htt exhibit late-onset, mild phenotypes, often after 6 months of age (26, 27), and lack prominent neuronal loss. Neither model is ideal to test transient effects of a single injection of siRNA against Htt introduced directly to the striatum. We therefore developed an acute, in vivo, HD mouse model tailored to addressing the efficacy of siRNA. Here, we used AAV to deliver a 1,395-nt cDNA fragment of human mutant Htt into mouse striatum to evaluate the effectiveness of an siRNA targeting human Htt.

Huntingtin localization in brains of normal and Huntington's disease patients.

Abstract: The immunohistochemical localization of huntingtin was examined in the Huntington's disease (HD) brain with an antibody that recognizes the wild-type and mutant proteins. Neuronal staining was reduced in areas of the HD striatum depleted of medium-sized neurons; large striatal neurons, which are spared in HD, retained normal levels of huntingtin expression. Neuronal labeling was markedly reduced in both segments of the globus pallidus including in brains with minimal loss of pallidal neurons. In some HD cortical and striatal neurons with normal looking morphology, huntingtin was associated with punctate cytoplasmic granules that at the ultrastructural level resembled the multivesicular body, an organelle involved in retrograde transport and protein degradation. Some immunoreactive processes showed blebbing and segmentation similar to that induced experimentally by hypoxic-ischemic or excitotoxic injury. Huntingtin staining was more concentrated in the perinuclear cytoplasm and reduced or absent in processes of atrophic cortical neurons. Nuclear staining was also evident. Fibers in the subcortical white matter of HD patients had significantly increased huntingtin immunoreactivity compared with those of controls. Results suggest that there may be changes in the neuronal expression and transport of wild-type and/or mutant huntingtin at early and late stages of neuronal degeneration in affected areas of the HD brain.

Protein aggregation and neurodegenerative disease.

Abstract: Neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS) and prion diseases are increasingly being realized to have common cellular and molecular mechanisms including protein aggregation and inclusion body formation. The aggregates usually consist of fibers containing misfolded protein with a beta-sheet conformation, termed amyloid. There is partial but not perfect overlap among the cells in which abnormal proteins are deposited and the cells that degenerate. The most likely explanation is that inclusions and other visible protein aggregates represent an end stage of a molecular cascade of several steps, and that earlier steps in the cascade may be more directly tied to pathogenesis than the inclusions themselves. For several diseases, genetic variants assist in explaining the pathogenesis of the more common sporadic forms and developing mouse and other models. There is now increased understanding of the pathways involved in protein aggregation, and some recent clues have emerged as to the molecular mechanisms of cellular toxicity. These are leading to approaches toward rational therapeutics.

Aggregation of Huntingtin in Neuronal Intranuclear Inclusions and Dystrophic Neurites in Brain

Abstract: The cause of neurodegeneration in Huntington's disease (HD) is unknown. Patients with HD have an expanded NH2-terminal polyglutamine region in huntingtin. An NH2-terminal fragment of mutant huntingtin was localized to neuronal intranuclear inclusions (NIIs) and dystrophic neurites (DNs) in the HD cortex and striatum, which are affected in HD, and polyglutamine length influenced the extent of huntingtin accumulation in these structures. Ubiquitin was also found in NIIs and DNs, which suggests that abnormal huntingtin is targeted for proteolysis but is resistant to removal. The aggregation of mutant huntingtin may be part of the pathogenic mechanism in HD.

Knocking down barriers: advances in siRNA delivery

Abstract: In the 10 years that have passed since the Nobel prize-winning discovery of RNA interference (RNAi), billions of dollars have been invested in the therapeutic application of gene silencing in humans. Today, there are promising data from ongoing clinical trials for the treatment of age-related macular degeneration and respiratory syncytial virus. Despite these early successes, however, the widespread use of RNAi therapeutics for disease prevention and treatment requires the development of clinically suitable, safe and effective drug delivery vehicles. Here, we provide an update on the progress of RNAi therapeutics and highlight novel synthetic materials for the encapsulation and intracellular delivery of nucleic acids.