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Principal Investigator: XUE-RU WU
Organization: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
Fiscal Year: 2024
Award: $207,000
Funding agency: National Institute of Diabetes and Digestive and Kidney Diseases
This proposal is submitted in response to PAR-23-119: “Catalytic Tool and Technology Development in
Kidney, Urologic, and Hematologic Diseases (R21 Clinical Trial Not Allowed)”. The main goal of our application
is to test the feasibility and define the strategies, reagents, parameters and methodology to perform genome
editing in the urothelium of live mice in order to enhance and accelerate the research of urothelial biology and
diseases. Urothelium of the bladder is a frequent site of chemical and radiation damages, inflammation,
infection and uncontrolled proliferation. Unlike many other epithelial tissues that are difficult to access,
urothelium is readily accessible through transurethral catheterization, a standard procedure for diagnosis,
monitoring and treatment of urothelium-related disorders. Despite its easy accessibility, urothelial research has
been greatly hampered by the lack of efficient, cost-effective and broadly applicable technological approaches.
In the past, the Principal Investigator's laboratory and those of other investigators have relied heavily on
genetically engineered mouse models to investigate gene functions and the cellular and molecular
mechanisms that govern urothelial growth, differentiation and disease pathogenesis. While highly useful and
informative, this technological platform is inherently time-consuming, labor-intensive and costly, and
consequently no longer adequate to fulfill the increasing research needs of the urothelial field. To tackle this
deficiency, we have decided to test whether it is feasible to perform genome editing in vivo in urothelium of live
mice by combining the easy accessibility of the bladder urothelium with the CRISPR/Cas9 technological
platform. We will approach this progressively by (1) delivering gRNAs via transurethral catheterization into
mice that constitutively express Cas9 to knock out, truncate or mutate a select set of genes of interest in
urothelium; (2) restricting gene knockouts specifically to basal urothelial cells versus suprabasal urothelial cells
using conditional Cas9 expression systems; and (3) performing gene knockouts in wild-type mice using all-in-
one adenoviruses bearing Cas9 and gRNAs or using all-in-one plasmids coupled with ultrasound-guided
percutaneous needle-electrode electroporation of the bladder. The success of these experiments should open
doors to brand new possibilities to target genes of interest in urothelium and model a wide spectrum of
urothelium-related human diseases.
Terms: <21+ years old><Acceleration><Adenoviridae><Adenoviruses><Adult><Adult Human><Area><Asparagine><Basal Cell><Basal Transcription Factor><Basal transcription factor genes><Biology><Biomedical Engineering><Bladder><Bladder Transitional Cell Epithelium><Bladder Urinary System><Bladder Urothelium><Blood Diseases><Body Tissues><CD105><CD105 Gene><CRISPR><CRISPR/Cas system><Cell Compartmentation><Cell Compartmentations><Cellular Matrix><Chemicals><Clinical Trials><Clustered Regularly Interspaced Short Palindromic Repeats><Consumption><Coupled><Cytokeratin><Cytoplasm><Cytoplasmic Domain><Cytoplasmic Tail><Cytoskeletal System><Cytoskeleton><D-Mannose><Development><Diagnosis><Differentiation and Growth><Disease><Disorder><E coli><E. coli><END Gene><ENG gene><Effectiveness><Elasticity><Electrodes><Electroporation><Endoglin Gene><Engineered Gene><Epithelium><Escherichia coli><Exposure to><Funding Opportunities><GEM model><GEMM model><Gene Alteration><Gene Mutation><General Transcription Factor Gene><General Transcription Factors><Generalized Growth><Generations><Genes><Genetic><Genetically Engineered Mouse><Gln><Glutamine><Goals><Growth><Guide RNA><HHT1><HHT1 Gene><HUMPPARG><Hematologic Diseases><Hematological Disease><Hematological Disorder><Infection><Inflammation><Injury><Investigators><Kidney Diseases><Kidney Pelvis><Knock-out><Knockout><L-Asparagine><L-Glutamine><Laboratories><LoxP-flanked allele><Mannopyranose><Mannopyranoside><Mannose><Membrane><Metabolic Glycosylation><Methodology><Mice><Mice Mammals><Modeling><Molecular><Monitor><Mucosa><Mucosal Tissue><Mucous Membrane><Murine><Mus><Mutate><NIDDK><NR1C3><National Institute of Diabetes and Digestive and Kidney Diseases><Needles><Nephropathy><ORW Gene><ORW1><ORW1 Gene><Occluding Junctions><PPARG><PPARG gene><PPARG1><PPARG2><Pathogenesis><Permeability><Plasmids><Point Mutation><Principal Investigator><Procedures><Progenitor Cells><Proliferating><Q Levoglutamide><Q. Levoglutamide><Radiation induced damage><Reagent><Receptor Protein><Renal Disease><Renal pelvis><Research><Research Personnel><Researchers><Role><Site><Specificity><Surface><System><Technology><Testing><Tight Junctions><Time><Tissue Growth><Tissues><Transcription Factor Proto-Oncogene><Transcription factor genes><UP2><UP3><UPII><UPIIIA><UPK2><UPK2 gene><UPK3><UPK3 gene><UPK3A><Ureter><Urethra><Urethral Catheterization><Urinary Tract Diseases><Urinary tract><Urination><Urine><Urologic Diseases><Urologic Disorder><Urological Diseases><Urological Disorders><Uroplakin 2><Uroplakin 3><Uroplakin 3A><Urothelial Cell><Urothelium><Wild Type Mouse><Zonula Occludens><adulthood><bio-engineered><bio-engineers><bioengineering><biological engineering><blood disorder><cost><cost effective><developmental><electroporative delivery><experiment><experimental research><experimental study><experiments><feasibility testing><floxed><floxed allele><gRNA><gene electrotransfer><gene function><genetically engineered mouse model><genetically engineered murine model><genome editing><genomic editing><glycosylation><human disease><in vivo><injuries><interest><intracellular skeleton><intravesical><kidney disorder><knockout gene><membrane structure><micturition><new technology><novel><novel technologies><ontogeny><overexpress><overexpression><protein biomarkers><protein markers><radiation damage><receptor><renal disorder><response><social role><stem cells><success><tech development><technology development><technology platform><technology system><tool><tool development><transcription factor><ultrasound><urethral><urinary bladder><urinary tract disorder><uroplakin II><uroplakin III><wildtype mouse>