Selected Grantee Publications
Impaired Skeletal Development by Disruption of Presenilin-1 in Pigs and Generation of Novel Pig Models for Alzheimer's Disease
Uh et al., Journal of Alzheimer's Disease. 2024.
https://pubmed.ncbi.nlm.nih.gov/39177593/
This study explored the effects of presenilin 1 (PSEN1) disruption on vertebral malformations in male and female PSEN1 mutant pigs. Researchers observed significant skeletal impairments and early deaths in pigs with a PSEN1 null mutation, mirroring phenotypes seen in mouse models of Alzheimer’s disease (AD). This porcine model provides valuable insights into pathological hallmarks of PSEN1 mutations in AD, offering a robust platform of therapeutic exploration. The findings establish pigs as an essential translational model for AD, enabling advanced studies on pathophysiology and treatment development for human skeletal and neurological conditions. Supported by ORIP (U42OD011140), NHLBI, NIA, NIAID.
Gene Editing of Pigs to Control Influenza A Virus Infections
Kwon et al., Emerging Microbes & Infections. 2024.
https://pubmed.ncbi.nlm.nih.gov/39083026/
A reduction in the efficacy of vaccines and antiviral drugs for combating infectious diseases in agricultural animals has been observed. Generating genetically modified livestock species to minimize susceptibility to infectious diseases is of interest as an alternative approach. The researchers developed a homozygous transmembrane serine protease 2 (TMPRSS2) knockout (KO) porcine model to investigate resistance to two influenza A virus (IAV) subtypes, H1N1 and H3N2. TMPRSS2 KO pigs demonstrated diminished nasal cavity viral shedding, lower viral burden, and reduced microscopic lung pathology compared with wild-type (WT) pigs. In vitro culturing of primary bronchial epithelial cells (PBECs) demonstrated delayed viral replication in TMPRSS2 KO pigs compared with WT pigs. This study demonstrates the potential use of genetically modified pigs to mitigate IAV infections in pigs and limit transmission to humans. Supported by ORIP (U42OD011140), NHLBI, NIAID, and NIGMS.
The Splicing Factor hnRNPL Demonstrates Conserved Myocardial Regulation Across Species and Is Altered in Heart Failure
Draper et al., FEBS Letters. 2024.
https://pubmed.ncbi.nlm.nih.gov/39300280/
The 5-year mortality rate of heart failure (HF) is approximately 50%. Gene splicing, induced by splice factors, is a post-transcriptional modification of mRNA that may regulate pathological remodeling in HF. Researchers investigated the role of the splice factor heterogenous nuclear ribonucleoprotein-L (hnRNPL) in cardiomyopathy. hnRNPL protein expression is significantly increased in a male C57BL/6 transaortic constriction–induced HF mouse model and in clinical samples derived from canine or human HF patients. Cardiac-restricted knockdown of the hnRNPL homolog in Drosophila revealed systolic dysfunction and reduced life span. This study demonstrates a conserved cross-species role of hnRNPL in regulating heart function. Supported by ORIP (K01OD028205) and NHLBI.
Systematic Multi-trait AAV Capsid Engineering for Efficient Gene Delivery
Eid et al., Nature Communications. 2024.
https://doi.org/10.1038/s41467-024-50555-y
Engineering novel functions into proteins while retaining desired traits is a key challenge for developers of viral vectors, antibodies, and inhibitors of medical and industrial value. In this study, investigators developed Fit4Function, a generalizable machine learning (ML) approach for systematically engineering multi-trait adeno-associated virus (AAV) capsids. Fit4Function was used to generate reproducible screening data from a capsid library that samples the entire manufacturable sequence space. The Fit4Function data were used to train accurate sequence-to-function models, which were combined to develop a library of capsid candidates. Compared to AAV9, top candidates from the Fit4Function capsid library exhibited comparable production yields; more efficient murine liver transduction; up to 1,000-fold greater human hepatocyte transduction; and increased enrichment in a screen for liver transduction in macaques. The Fit4Function strategy enables prediction of peptide-modified AAV capsid traits across species and is a critical step toward assembling an ML atlas that predicts AAV capsid performance across dozens of traits. Supported by ORIP (P51OD011107, U42OD027094), NIDDK, NIMH, and NINDS.
RNA Landscapes of Brain and Brain-Derived Extracellular Vesicles in Simian Immunodeficiency Virus Infection and Central Nervous System Pathology
Huang et al., The Journal of Infectious Diseases. 2024.
https://pubmed.ncbi.nlm.nih.gov/38079216/
Brain tissue–derived extracellular vesicles (bdEVs) act locally in the central nervous system (CNS) and may indicate molecular mechanisms in HIV CNS pathology. Using brain homogenate (BH) and bdEVs from male pigtailed macaques, researchers identified dysregulated RNAs in acute and chronic infection. Most dysregulated messenger RNAs (mRNAs) in bdEVs reflected dysregulation in source BH, and these mRNAs are disproportionately involved in inflammation and immune responses. Additionally, several circular RNAs were differentially abundant in source tissue and might be responsible for specific differences in small RNA levels in bdEVs during simian immunodeficiency virus (SIV) infection. This RNA profiling shows potential regulatory networks in SIV infection and SIV-related CNS pathology. Supported by ORIP (U42OD013117), NCI, NIAID, NIDA, NIMH, and NINDS.
Evolution of the Clinical-Stage Hyperactive TcBuster Transposase as a Platform for Robust Non-Viral Production of Adoptive Cellular Therapies
Skeate et al., Molecular Therapy. 2024.
https://pubmed.ncbi.nlm.nih.gov/38627969/
In this study, the authors report the development of a novel hyperactive TcBuster (TcB-M) transposase engineered through structure-guided and in vitro evolution approaches that achieve high-efficiency integration of large, multicistronic CAR-expression cassettes in primary human cells. This proof-of-principle TcB-M engineering of CAR-NK and CAR-T cells shows low integrated vector copy number, a safe insertion site profile, robust in vitro function, and improved survival in a Burkitt lymphoma xenograft model in vivo. Their work suggests that TcB-M is a versatile, safe, efficient, and open-source option for the rapid manufacture and preclinical testing of primary human immune cell therapies through delivery of multicistronic large cargo via transposition. Supported by ORIP (F30OD030021), NCI, NHLBI, and NIAID.
AAV5 Delivery of CRISPR/Cas9 Mediates Genome Editing in the Lungs of Young Rhesus Monkeys
Liang et al., Human Gene Therapy. 2024.
https://pubmed.ncbi.nlm.nih.gov/38767512/
Genome editing in somatic cells and tissues has the potential to provide long-term expression of therapeutic proteins to treat a variety of genetic lung disorders. However, delivering genome-editing machinery to disease-relevant cell types in the lungs of primates has remained a challenge. Investigators of this article are participating in the NIH Somatic Cell Genome Editing Consortium. Herein, they demonstrate that intratracheal administration of a dual adeno-associated virus type 5 vector encoding CRISPR/Cas9 can mediate genome editing in rhesus (male and female) airways. Up to 8% editing was observed in lung lobes, including a housekeeping gene, GAPDH, and a disease-related gene, angiotensin-converting enzyme 2. Using single-nucleus RNA-sequencing, investigators systematically characterized cell types transduced by the vector. Supported by ORIP (P51OD01110, U42OD027094, S10OD028713), NCATS, NCI, and NHLBI.
Engineered IgM and IgG Cleaving Enzymes for Mitigating Antibody Neutralization and Complement Activation in AAV Gene Transfer
Smith et al., Molecular Therapy. 2024.
https://www.sciencedirect.com/science/article/pii/S1525001624003058?via%3Dihub=
Recombinant adeno-associated viral (AAV) vectors have emerged as the leading platform for therapeutic gene transfer, but systemic dosing of AAV vectors poses potential risk of adverse side effects, including complement activation triggered by anti-capsid immunity. In this study, investigators discovered an IgM cleaving enzyme (IceM) that degrades human IgM, a key trigger in the anti-AAV immune cascade. They engineered a fusion enzyme (IceMG) with dual proteolytic activity against human IgM and IgG. Antisera from animals treated with IceMG show decreased ability to neutralize AAV and activate complement. These studies have implications for improving the safety of AAV gene therapies and offer broader applications, including for organ transplantation and autoimmune diseases. Supported by ORIP (P51OD011107, U42OD027094), NHLBI, and NIAID.