Now showing items 21-40 of 26600

    • Within-host evolutionary dynamics and tissue compartmentalization during acute SARS-CoV-2 infection

      Farjo, Mireille; Koelle, Katia; Martin, Michael A; Gibson, Laura L; Walden, Kimberly K O; Rendon, Gloria; Fields, Christopher J; Alnaji, Fadi G; Gallagher, Nicholas; Luo, Chun Huai; et al. (2024-01-04)
      We detail the within-host evolutionary dynamics of SARS-CoV-2 during acute infection in 31 individuals using daily longitudinal sampling. We characterized patterns of mutational accumulation for unvaccinated and vaccinated individuals, and observed that temporal variant dynamics in both groups were largely stochastic. Comparison of paired nasal and saliva samples also revealed significant genetic compartmentalization between tissue environments in multiple individuals. Our results demonstrate how selection, genetic drift, and spatial compartmentalization all play important roles in shaping the within-host evolution of SARS-CoV-2 populations during acute infection.
    • Experiences of cisgender youth with a transgender and/or nonbinary sibling

      Godwin, Eli G; Moore, Lb M; Sansfaçon, Annie Pullen; Nishman, Melissa MacNish; Rosal, Milagros C; Katz-Wise, Sabra L (2024-01-03)
      While recent research has begun to address the effects of family support on transgender and/or nonbinary youth (TNY), almost no studies have directly examined how cisgender siblings in families with TNY navigate their sibling's gender disclosure and affirmation within both their families and their larger communities. We conducted an exploratory secondary analysis of in-person, semi-structured interviews with 15 adolescent and young adult siblings (age 13-24 years) of TNY from the northeastern United States from the baseline wave of the community-based, longitudinal, mixed methods Trans Teen and Family Narratives Project. Interview transcripts were analyzed using immersion/crystallization and template organizing approaches. Analyses yielded three main themes: gender-related beliefs and knowledge, peri- and post-disclosure family dynamics, and assessing responses to their sibling. Subthemes included anticipation of their sibling's TN identity, expectations post-disclosure, participants' level of involvement in gender-related family processes, perceptions of changes in family relationships, concern for their sibling (including a high degree of attunement to gender-affirming name and pronoun usage), and concern for themselves. Findings from this study suggest the need to engage directly with siblings of TNY to further elucidate their intrapersonal, intra-familial, and extra-familial experiences related to having a TN sibling and determine their unique support needs. Implications for families, clinicians, and communities are discussed.
    • UMCCTS Newsletter, January 2024

      UMass Center for Clinical and Translational Science (2024-01-02)
      This is the January 2024 issue of the UMass Center for Clinical and Translational Science Newsletter containing news and events of interest.
    • Vitamin B produced by gut bacteria modulates cholinergic signalling

      Kang, Woo Kyu; Florman, Jeremy T; Araya, Antonia; Fox, Bennett W; Thackeray, Andrea; Schroeder, Frank C; Walhout, Albertha J M; Alkema, Mark J (2024-01-02)
      A growing body of evidence indicates that gut microbiota influence brain function and behaviour. However, the molecular basis of how gut bacteria modulate host nervous system function is largely unknown. Here we show that vitamin B12-producing bacteria that colonize the intestine can modulate excitatory cholinergic signalling and behaviour in the host Caenorhabditis elegans. Here we demonstrate that vitamin B12 reduces cholinergic signalling in the nervous system through rewiring of the methionine (Met)/S-adenosylmethionine cycle in the intestine. We identify a conserved metabolic crosstalk between the methionine/S-adenosylmethionine cycle and the choline-oxidation pathway. In addition, we show that metabolic rewiring of these pathways by vitamin B12 reduces cholinergic signalling by limiting the availability of free choline required by neurons to synthesize acetylcholine. Our study reveals a gut-brain communication pathway by which enteric bacteria modulate host behaviour and may affect neurological health.
    • Unintentional Ketamine Overdose Via Telehealth

      Johnson, Brett E; Borges, Eric S; Gaspari, Romolo J; Galletta, Gayle M; Lai, Jeffrey T (2024-01-01)
      The use of ketamine in psychiatry has expanded to at-home ketamine-assisted therapy (KAT) via telemedicine. We report a case of massive unintentional ketamine overdose during at-home KAT resulting in hypoxemic respiratory failure, successfully treated with atropine.
    • Raising Cultural Awareness and Addressing Barriers to Breast Imaging Care for Black Women

      Jackson, Tatianie; Wahab, Rifat A; Bankston, Karen; Mehta, Tejas S (2023-12-23)
      Health care disparities, which are differences in the attainment of full health potential among population groups, have been documented across medical conditions, clinical settings, and diagnostic and treatment modalities. Deeply rooted health care disparities due to many factors have affected how Black women (BW) view medical care including screening mammography. This article explores health care disparities around breast cancer in BW and how patient distrust, provider biases, race, and social determinants of health continue to have negative effects on breast cancer outcomes in BW, despite medical advances in breast cancer detection and management. In addition, this article addresses the importance of culturally competent care for BW around breast cancer awareness, screening, and treatment, and offers strategies to address disparities and rebuild trust.
    • Developing Mood-Based Computer-Tailored Health Communication for Smoking Cessation: Feasibility Randomized Controlled Trial

      Lee, Donghee N; Sadasivam, Rajani S; Stevens, Elise M (2023-12-22)
      Background: Computer-tailored health communication (CTHC), a widely used strategy to increase the effectiveness of smoking cessation interventions, is focused on selecting the best messages for an individual. More recently, CTHC interventions have been tested using contextual information such as participants' current stress or location to adapt message selection. However, mood has not yet been used in CTCH interventions and may increase their effectiveness. Objective: This study aims to examine the association of mood and smoking cessation message effectiveness among adults who currently smoke cigarettes. Methods: In January 2022, we recruited a web-based convenience sample of adults who smoke cigarettes (N=615; mean age 41.13 y). Participants were randomized to 1 of 3 mood conditions (positive, negative, or neutral) and viewed pictures selected from the International Affective Picture System to induce an emotional state within the assigned condition. Participants then viewed smoking cessation messages with topics covering five themes: (1) financial costs or rewards, (2) health, (3) quality of life, (4) challenges of quitting, and (5) motivation or reasons to quit. Following each message, participants completed questions on 3 constructs: message receptivity, perceived relevance, and their motivation to quit. The process was repeated 30 times. We used 1-way ANOVA to estimate the association of the mood condition on these constructs, controlling for demographics, cigarettes per day, and motivation to quit measured during the pretest. We also estimated the association between mood and outcomes for each of the 5 smoking message theme categories. Results: There was an overall statistically significant effect of the mood condition on the motivation to quit outcome (P=.02) but not on the message receptivity (P=.16) and perceived relevance (P=.86) outcomes. Participants in the positive mood condition reported significantly greater motivation to quit compared with those in the negative mood condition (P=.005). Participants in the positive mood condition reported higher motivation to quit after viewing smoking cessation messages in the financial (P=.03), health (P=.01), quality of life (P=.04), and challenges of quitting (P=.03) theme categories. We also compared each mood condition and found that participants in the positive mood condition reported significantly greater motivation to quit after seeing messages in the financial (P=.01), health (P=.003), quality of life (P=.01), and challenges of quitting (P=.01) theme categories than those in the negative mood condition. Conclusions: Our findings suggest that considering mood may be important for future CTHC interventions. Because those in the positive mood state at the time of message exposure were more likely to have greater quitting motivations, smoking cessation CTHC interventions may consider strategies to help improve participants' mood when delivering these messages. For those in neutral and negative mood states, focusing on certain message themes (health and motivation to quit) may be more effective than other message themes.
    • Clinician Attitudes Toward Suicide Prevention Practices and Their Implementation: Findings From the System of Safety Study

      Larkin, Celine; Kiefe, Catarina I; Morena, Alexandra L; Rahmoun, Mhd B; Lazar, Peter; Sefair, Ana Vallejo; Boudreaux, Edwin D (2023-12-21)
      Objective: The authors aimed to assess clinicians' attitudes toward suicide-related practices and their implementation, across roles and settings, before implementation of the Zero Suicide model in a health care system. Methods: Clinicians (N=5,559) were invited to complete a survey assessing demographic characteristics; confidence and self-reported suicide-related practice; leadership buy-in; and attitudes toward suicide prevention, safety planning, and continuous quality improvement (CQI). Results: Of 1,224 respondents, most felt confident conducting suicide screening but less confident performing other suicide-related care. Provider role and care setting were significantly associated with confidence (p<0.001, Kruskal-Wallis H test) and practice (p<0.001, Kruskal-Wallis H test) of providing suicide prevention care, with behavioral health providers and providers in the emergency department (ED) reporting the highest confidence. Attitudes toward safety planning were more positive among women (p<0.001, t test) and behavioral health providers (p<0.001, F test) than among their counterparts or peers. Positive attitudes toward CQI were significantly associated with male sex (p=0.01), non-White race (p=0.03), younger age (p=0.02), fewer years working in health care (p<0.001), administrative role (p<0.001), working in the ED (p<0.001), outpatient settings (p<0.02), and medical provider role (p<0.001). Conclusions: Behavioral health providers and those in the ED reported feeling prepared to deliver suicide-related care, with nurses feeling less confident and less supported. Initiatives to improve suicide-related care should account for clinical role and care setting during planning. CQI could help engage a broader range of clinicians in suicide-related care improvements.
    • More Short Writings #2

      Reynolds, Maggie (2023-12-21)
      Introduction: Maggie Reynolds, UMass Chan '26, is the last of the students in my Structural Inequity, Advocacy, and Justice (SIAJ) Pathway to share their writing from our recent humanities session. She has written several pieces about the inequities she sees around her as she volunteers and interacts with people in Worcester. She describes the context of each piece below.
    • ATXN2 is a target of N-terminal proteolysis

      Chitre, Monika; Emery, Patrick (2023-12-21)
      Spinocerebellar ataxia 2 (SCA2) is a neurodegenerative disorder caused by the expansion of the poly-glutamine (polyQ) tract of Ataxin-2 (ATXN2). Other polyQ-containing proteins such as ATXN7 and huntingtin are associated with the development of neurodegenerative diseases when their N-terminal polyQ domains are expanded. Furthermore, they undergo proteolytic processing events that produce N-terminal fragments that include the polyQ stretch, which are implicated in pathogenesis. Interestingly, N-terminal ATXN2 fragments were reported in a brain extract from a SCA2 patient, but it is currently unknown whether an expanded polyQ domain contributes to ATXN2 proteolytic susceptibility. Here, we used transient expression in HEK293 cells to determine whether ATXN2 is a target for specific N-terminal proteolysis. We found that ATXN2 proteins with either normal or expanded polyQ stretches undergo proteolytic cleavage releasing an N-terminal polyQ-containing fragment. We identified a short amino acid sequence downstream of the polyQ domain that is necessary for N-terminal cleavage of full-length ATXN2 and sufficient to induce proteolysis of a heterologous protein. However, this sequence is not required for cleavage of a short ATXN2 isoform produced from an alternative start codon located just upstream of the CAG repeats encoding the polyQ domain. Our study extends our understanding of ATXN2 posttranslational regulation by revealing that this protein can be the target of specific proteolytic cleavage events releasing polyQ-containing products that are modulated by the N-terminal domain of ATXN2. N-terminal ATXN2 proteolysis of expanded polyQ domains might contribute to SCA2 pathology, as observed in other neurodegenerative disorders caused by polyQ domain expansion.
    • Exploring patients' experiences with telehealth in obstetrics care during the COVID-19 pandemic: A qualitative study

      Alkawaldeh, Mohammad; Alkhawaldeh, Asma; Yeboah, Tracy (2023-12-20)
      Aim: The aim of this study was to evaluate patients' experiences with telehealth provision of obstetrics and gynecology care during the COVID-19 pandemic qualitatively. Design and setting: In this study, a qualitative research design, namely descriptive phenomenology, was employed. Participants were recruited from the OB department at UMass Memorial Medical center in Worcester, MA, between 6/2020 and 7/2020. Methods: Between June 2020 and July 2020, in-depth interviews were conducted with 18 women receiving care at the Obstetrics and Maternal and Fetal Medicine clinics. Data were analyzed using qualitative thematic analysis, as outlined by Braun and Clarke. Results: Telehealth is a feasible and safe health-care tool that is available during these unprecedented times. This study provided qualitative evidence based on patients' perspectives and experiences. Participants' meanings in relation to their experiences of using telehealth services emerged from the data in four themes: the overall experience of using modern telehealth platforms, telehealth and its perceived benefits, telehealth and its perceived challenges, and telehealth and its potential future use. Conclusion: While this study highlights areas in telehealth implementation that require improvement, the overall positive experiences and consistent perceived benefits of most participants suggests that telehealth can be an important tool in healthcare delivery for appropriate patients and situations moving forward in a post-pandemic world. Impact: During the global pandemic, telehealth has been recognized to have the potential to play a critical role in healthcare delivery. Establishing qualitative evidence-based practices in the emerging field of telehealth for OB services is pivotal to mitigate potential safety, feasibility, and cost issues that could be associated with the rapid adoption of telehealth. Yet, this qualitative study However highlighted several challenges that are necessary to be addressed in order for telehealth to meet maximum effectiveness and functionality in the future.
    • Genetic ablation of Sarm1 attenuates expression and mislocalization of phosphorylated TDP-43 after mouse repetitive traumatic brain injury

      Dogan, Elif O; Bouley, James; Zhong, Jianjun; Harkins, Ashley L; Keeler, Allison M; Bosco, Daryl A; Brown, Robert H; Henninger, Nils (2023-12-20)
      Traumatic brain injury (TBI), particularly when moderate-to-severe and repetitive, is a strong environmental risk factor for several progressive neurodegenerative disorders. Mislocalization and deposition of transactive response DNA binding protein 43 (TDP-43) has been reported in both TBI and TBI-associated neurodegenerative diseases. It has been hypothesized that axonal pathology, an early event after TBI, may promote TDP-43 dysregulation and serve as a trigger for neurodegenerative processes. We sought to determine whether blocking the prodegenerative Sarm1 (sterile alpha and TIR motif containing 1) axon death pathway attenuates TDP-43 pathology after TBI. We subjected 111 male Sarm1 wild type, hemizygous, and knockout mice to moderate-to-severe repetitive TBI (rTBI) using a previously established injury paradigm. We conducted serial neurological assessments followed by histological analyses (NeuN, MBP, Iba-1, GFAP, pTDP-43, and AT8) at 1 month after rTBI. Genetic ablation of the Sarm1 gene attenuated the expression and mislocalization of phosphorylated TDP-43 (pTDP-43) and accumulation of pTau. In addition, Sarm1 knockout mice had significantly improved cortical neuronal and axonal integrity, functional deficits, and improved overall survival after rTBI. In contrast, removal of one Sarm1 allele delayed, but did not prevent, neurological deficits and neuroaxonal loss. Nevertheless, Sarm1 haploinsufficient mice showed significantly less microgliosis, pTDP-43 pathology, and pTau accumulation when compared to wild type mice. These data indicate that the Sarm1-mediated prodegenerative pathway contributes to pathogenesis in rTBI including the pathological accumulation of pTDP-43. This suggests that anti-Sarm1 therapeutics are a viable approach for preserving neurological function after moderate-to-severe rTBI.
    • U.S. nursing home leadership experiences with COVID-19 and its impact on residents and staff: A qualitative analysis

      Dubé, Catherine E; Nielsen, Natalia; McPhillips, Emily; Lee Hargraves, J; Cosenza, Carol; Jesdale, Bill; Lapane, Kate L (2023-12-19)
      Objectives: To explore experiences of U.S. (United States) nursing home leadership during the COVID-19 pandemic in their efforts to address resident loneliness and social isolation and to elicit stories about personal and professional impacts on themselves and staff. Design: Qualitative inquiry via three optional open-ended questions appended to a national self-administered survey of American nursing home leaders was employed. Textual data was analyzed using an iterative reflexive thematic approach. Setting and participants: A stratified sample frame defined by facility size (beds: 30-99, 100+) and quality ratings (1, 2-4, 5) was employed. Web survey links and paper surveys were sent to 1,676 nursing home directors of nursing between February and May 2022. Results: Open text responses were collected from 271 nursing homes. Broad themes included: 1) Addressing needs of residents & families; 2) Challenges; and 3) Personal experiences of nursing home leadership/staff. Respondents described trauma to residents, staff, and leadership. Resident loneliness was addressed using existing and newer technologies and innovative indoor and outdoor activities. Residents experienced fear, illness, loss, and sometimes death. Isolation from family and lack of touch were particularly difficult. Regulations were seen as punitive while ignoring emotional needs of residents. Staffing challenges and pressures to do more with less created additional stress. Leadership and staff made significant sacrifices resulting in physical, social, and emotional consequences. Beneficial outcomes included staff bonding, professional growth, and permanent implementation of new interventions. Conclusions and implications: New and creative interventions were successfully implemented to address social isolation and loneliness. Improved Wi-Fi and other nursing home infrastructure upgrades are needed to maintain them. Reimagining often conflicting overlapping federal, state, and local regulations, grounding them in good clinical judgement, and incentivizing performance improvement should be considered. Trauma experienced by staff needs to be addressed to deal with current and future workforce needs.
    • A programmable dual-targeting di-valent siRNA scaffold supports potent multi-gene modulation in the central nervous system [preprint]

      Belgrad, Jillian; Tang, Qi; Hildebrand, Samuel; Summers, Ashley; Sapp, Ellen; Echeverria, Dimas; O'Reilly, Dan; Luu, Eric; Bramato, Brianna; Allen, Sarah; et al. (2023-12-19)
      Di-valent short interfering RNA (siRNA) is a promising therapeutic modality that enables sequence-specific modulation of a single target gene in the central nervous system (CNS). To treat complex neurodegenerative disorders, where pathogenesis is driven by multiple genes or pathways, di-valent siRNA must be able to silence multiple target genes simultaneously. Here we present a framework for designing unimolecular "dual-targeting" di-valent siRNAs capable of co-silencing two genes in the CNS. We reconfigured di-valent siRNA - in which two identical, linked siRNAs are made concurrently - to create linear di-valent siRNA - where two siRNAs are made sequentially attached by a covalent linker. This linear configuration, synthesized using commercially available reagents, enables incorporation of two different siRNAs to silence two different targets. We demonstrate that this dual-targeting di-valent siRNA is fully functional in the CNS of mice, supporting at least two months of maximal target silencing. Dual-targeting di-valent siRNA is highly programmable, enabling simultaneous modulation of two different disease-relevant gene pairs (e.g., Huntington's disease: MSH3 and HTT; Alzheimer's disease: APOE and JAK1) with similar potency to a mixture of single-targeting di-valent siRNAs against each gene. This work potentiates CNS modulation of virtually any pair of disease-related targets using a simple unimolecular siRNA.
    • An exploration of influenza A virus entry factors using CRISPR-based gene editing

      Kyawe, Pyae Phyo (2023-12-18)
      Influenza A virus (IAV) is a respiratory pathogen with a segmented negative-sense RNA genome that is capable of causing epidemics and pandemics. During the 2021-2022 influenza season, approximately 9 million people in the U.S. were infected with influenza, resulting in an estimated 5,000 deaths. The error-prone nature of the IAV polymerase results in antigenic drift and antigenic shift which contribute to low vaccine efficacy and escape from antivirals. Furthermore, the host factors required for the complete IAV infectious cycle have not been fully identified. The aim of this dissertation is to examine the host factors that may contribute to IAV infectivity in human lung cells. My goal is to understand how changes in the expression levels of host factors can impact influenza infection by CRISPR-mediated knockout or overexpression of target genes. Utilizing CRISPR screens, several candidates, whose up- or down-regulation resulted in reduced IAV infection in the human A549 cell line were identified. I confirmed that the knockout of CMAS or overexpression of B4GALNT2 inhibited IAV infection. In addition, I tested whether overexpression of two candidates from the CRISPR activation screen – DEFB127 and ADAR1 – would inhibit IAV and non-IAV viruses. Surprisingly, overexpression of the two candidates had minimal impact on IAV in A549 cells, but overexpression of ADAR1 had a pro-viral effect on other viruses. Taken together, these data provide insight into host factors modulating IAV infection and how CRISPR-mediated gene modulation can be utilized to further understand the IAV life cycle and for development of therapeutic agents for flu.
    • Experimental and Computational Methods for Identifying Death-Regulatory Genes from Chemo-Genetic Profiles

      Honeywell, Megan E (2023-12-18)
      A common approach to understanding how drugs induce their therapeutic effect is identifying the genetic determinants of drug sensitivity. This can be achieved following systematic loss- or gain-of-function genetic perturbations with CRISPR/Cas9. Because these “chemo-genetic profiles” are generally performed in a pooled format, inference of gene function is subject to several confounding influences, including variation in growth rates between clones or variation in the degree of coordination between growth and death. To overcome these issues, we developed an analysis method called MEDUSA (Method for Evaluating Death Using a Simulation-assisted Approach). MEDUSA uses time-resolved measurements and model driven constraints to reveal the combination of growth and death rates that generated the drug-treated clonal abundance. We find that MEDUSA is uniquely effective at identifying death regulatory genes, and we apply MEDUSA to determine how DNA damage-induced lethality varies in the presence and absence of p53. We find that loss of p53 switches the mechanism of DNA damage-induced death from apoptosis to a non-apoptotic form of death called MPT-driven necrosis. We find that activation of MPT by DNA damage requires high respiration, and that cell death can be exacerbated by modulating NAD+ in p53-deficient cells. These findings demonstrate the accuracy and utility of MEDUSA, both for determining the genetic dependencies of lethality and for revealing opportunities to promote the lethality of chemotherapies in a cancer specific manner.
    • Pediatric healthcare professionals' attitudes and beliefs about weight stigma: A descriptive study

      Turner, Samantha L (2023-12-15)
      Purpose: Children face weight-based stigma from their healthcare providers at a disconcerting rate, and efforts to mitigate this have been scant. This study aimed to quantify pediatric healthcare professionals' attitudes and beliefs about weight stigma and to determine stigma reduction interventions that are most supported by pediatric healthcare providers. Design and methods: Participants completed two validated instruments which measured implicit and explicit weight bias, respectively. They then completed a researcher-designed questionnaire to assess their attitudes and beliefs about weight stigma, and demographic questions. ANOVA models were used to examine associations between bias measures and participant characteristics, chi-square analyses were used to examine associations between questionnaire responses and participant characteristics, and Spearman's rank was used to determine correlations between weight bias and questionnaire responses. Results: Participants exhibited moderate-to-high levels of implicit and explicit weight bias (mean Implicit Association Test score = 0.59, mean Crandall Anti-Fat Attitudes Score = 38.95). Associations were noted between implicit bias and years in practice (p < 0.05), and implicit bias and occupation (p < 0.05). There was a significant correlation between explicit bias and multiple questionnaire items, suggesting that healthcare providers with greater weight bias are aware of those biases and are ready to take action to address them. Conclusion: Though pediatric healthcare exhibit weight-based biases, they are invested in taking steps to mitigate these biases and their impact on patients. Practice implications: The results of this study can inform the design of future interventions that aim to reduce healthcare-based weight bias, thus improving the quality of pediatric healthcare.
    • Evaluation of ChatGPT and Google Bard Using Prompt Engineering in Cancer Screening Algorithms

      Nguyen, Daniel; Swanson, Daniel; Newbury, Alex; Kim, Young H (2023-12-15)
      Large language models (LLMs) such as ChatGPT and Bard have emerged as powerful tools in medicine, showcasing strong results in tasks such as radiology report translations and research paper drafting. While their implementation in clinical practice holds promise, their response accuracy remains variable. This study aimed to evaluate the accuracy of ChatGPT and Bard in clinical decision-making based on the American College of Radiology Appropriateness Criteria for various cancers. Both LLMs were evaluated in terms of their responses to open-ended (OE) and select-all-that-apply (SATA) prompts. Furthermore, the study incorporated prompt engineering (PE) techniques to enhance the accuracy of LLM outputs. The results revealed similar performances between ChatGPT and Bard on OE prompts, with ChatGPT exhibiting marginally higher accuracy in SATA scenarios. The introduction of PE also marginally improved LLM outputs in OE prompts but did not enhance SATA responses. The results highlight the potential of LLMs in aiding clinical decision-making processes, especially when guided by optimally engineered prompts. Future studies in diverse clinical situations are imperative to better understand the impact of LLMs in radiology.
    • Advancing RNA-Targeting Therapeutics by Oligonucleotide Engineering: RNA Activation, Off-Target Effect, and Co-Targeting Nuclear and Cytoplasmic RNA

      Wang, Feng (2023-12-15)
      Traditional protein-targeting therapeutics by small molecules can target ~3000 proteins, representing <2% of the human genome. In comparison, ~80% of human genome is transcribed into RNAs, which are mostly targetable by oligonucleotides. Thus, RNA-targeting therapeutics enable a much bigger spectrum of drug targets. With nearly 20 FDA-approved oligonucleotide drugs and numerous ongoing clinical trials, RNA-targeting therapeutics by oligonucleotides have significantly addressed unmet medical needs. However, challenges persist in gene expression enhancement and comprehensive dual-RNA modulation. Here, I set out to address these two challenges. Firstly, I identified two ASOs designed to target intron 1 of FXN pre-mRNA, leading to a ~2-fold increase in FXN mRNA levels in multiple cell models. Despite rigorous controls such as two RNA quantification assays and normalization by multiple housekeeping genes, the FXN activation by these two ASOs wasn’t driven by direct binding to the FXN pre-mRNA or a mutual off-target transcript. Surprisingly, it’s found dependent on guanosine-rich motifs in the full PS backbone, suggesting non-base-paired off-target effects. Secondly, we developed siRNASO, an oligonucleotide scaffold integrating the functionalities of siRNA and ASO into a single molecule. siRNASOs demonstrated potent single RNA silencing and comprehensive dual-RNA modulation, including RNA silencing, splicing modulation, and RNA editing in vitro and in vivo. Notably, siRNASO exhibits excellent tolerability in the mouse CNS, suggesting its potential as a therapeutic platform for CNS disorders. Overall, the research in this thesis will serve as a valuable foundation for future research and therapeutic applications, significantly contributing to the advancement of RNA-targeting therapeutics.
    • A role for mutations in AK9 and other genes affecting ependymal cells in idiopathic normal pressure hydrocephalus

      Yang, Hong Wei; Lee, Semin; Berry, Bethany C; Yang, Dejun; Zheng, Shaokuan; Carroll, Rona S; Park, Peter J; Johnson, Mark D (2023-12-15)
      Idiopathic normal pressure hydrocephalus (iNPH) is an enigmatic neurological disorder that develops after age 60 and is characterized by gait difficulty, dementia, and incontinence. Recently, we reported that heterozygous CWH43 deletions may cause iNPH. Here, we identify mutations affecting nine additional genes (AK9, RXFP2, PRKD1, HAVCR1, OTOG, MYO7A, NOTCH1, SPG11, and MYH13) that are statistically enriched among iNPH patients. The encoded proteins are all highly expressed in choroid plexus and ependymal cells, and most have been associated with cilia. Damaging mutations in AK9, which encodes an adenylate kinase, were detected in 9.6% of iNPH patients. Mice homozygous for an iNPH-associated AK9 mutation displayed normal cilia structure and number, but decreased cilia motility and beat frequency, communicating hydrocephalus, and balance impairment. AK9+/- mice displayed normal brain development and behavior until early adulthood, but subsequently developed communicating hydrocephalus. Together, our findings suggest that heterozygous mutations that impair ventricular epithelial function may contribute to iNPH.