Development of a novel RNA interference method to reduce Cryptosporidium infection
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Principal Investigator: Alejandro Castellanos-Gonzalez Organization: UNIVERSITY OF TEXAS MED BR GALVESTON Fiscal Year: 2021 Award: $197,500 Funding agency: National Institute of Allergy and Infectious Diseases Project Summary/Abstract The World Health Organization reports diarrhea kills around 760,000 children under five every year. Cryptosporidium is a leading cause of diarrhea morbidity and mortality. Therefore novel treatments against this pathogen are urgently needed. The limitations of tools used to genetically manipulate gene expression in this parasite have been identified as a major hurdle for drug and vaccine development. To circumvent this gap, we developed a rapid method to silence genes in this parasite using pre-assembled complexes (minimal RISC complexes, ready to silence) of Cryptosporidium single-stranded RNA and the human enzyme Argonaute 2 (ssRNA/Ago). We used this method to determine the role of selected genes during Cryptosporidium infection. We identified four genes (Actin, Nucleoside diphosphate kinase, Rhomboid protein 1 and transcription factor Ap2) that block proliferation and egress of Cryptosporidium parasites. Therefore, we hypothesized that ssRNA/Ago based therapy may be optimized in order to treat Cryptosporidium infection. In this project our goal is to develop a method to block parasite proliferation on infected intestinal cells. Our specific aims are: 1) Developing a novel method to reduce Cryptosporidium infection by silencing essential genes and to test the silencing with ssRNA/Ago complexes under physiological conditions and 2) Demonstrate the feasibility to deliver ssRNA-Ago complexes in the intestine. The characterization of these mechanisms will be essential to explore the feasibility to use RNA interference technology as novel alternative for the treatment of Crypotosporidiosis Terms: <0-11 years old><2 year old><2 years of age><2-(Acetolyloxy)-N-(5-nitro-2-thiazolyl)benzamide><AP2><AP2 Protein><AP2TF Protein><Actins><Activator Protein 2><Address><Affect><Age><Cell Body><Cells><Cessation of life><Child><Child Youth><Children (0-21)><Complex><Cryptosporidiosis><Cryptosporidium><Cryptosporidium infection><Data><Death><Deoxynucleoside Diphosphate Kinases><Development><Diarrhea><Dose><Drugs><Encapsulated><Enteral><Enteric><Enzyme Gene><Enzymes><Essential Genes><FDA approved><Face><Freeze Drying><Freeze Dryings><Frequencies><Future><Gene Expression><Gene Inactivation><Gene Silencing><Genes><Goals><Hand><Human><Immune Cell Activation><Immune response><Immune system><Immunocompromised><Immunocompromised Host><Immunocompromised Patient><Immunological response><Immunosuppressed Host><Individual><Intestinal><Intestines><Liquid substance><Lyophilization><Malnutrition><Medication><Messenger RNA><Methods><Mice><Mice Mammals><Microscopy><Modern Man><Molecular><Morbidity><Morbidity - disease rate><Mucous body substance><Mucus><Murine><Mus><Non-Polyadenylated RNA><Nucleic Acids><Nucleoside Diphosphokinases><Nucleoside-Diphosphate Kinase><Nutritional Deficiency><Oral><Parasites><Pharmaceutic Preparations><Pharmaceutical Preparations><Physiologic><Physiological><Position><Positioning Attribute><Post-Transcriptional Gene Silencing><Posttranscriptional Gene Silencing><Pre-Clinical Model><Preclinical Models><Proteins><Quelling><RNA><RNA Gene Products><RNA Interference><RNA Interference Therapy><RNA Silencing><RNA interference therapeutics><RNA interference-based therapy><RNAi><RNAi therapeutics><RNAi therapy><RNAi-based therapeutics><RNAi-based therapy><Reporting><Research><Ribonucleic Acid><Role><Sequence-Specific Posttranscriptional Gene Silencing><Short interfering RNA><Small Interfering RNA><System><TFAP2 Protein><Tablets><Technology><Testing><Toddler><Transfection><Undernutrition><Validation><World Health Organization><activator protein AP-2><age 2 years><aged 2 years><aged two years><ages><allergic/immunologic body system><allergic/immunologic organ system><alternative treatment><base><bowel><develop a vaccine><development of a vaccine><developmental><diarrheal disease><diarrheal illness><dietary deficiency><drug development><drug/agent><effective therapy><effective treatment><enhancer-binding protein AP-2><enteral pathogen><enteric pathogen><enteropathogen><experience><experiment><experimental research><experimental study><faces><facial><fluid><gene manipulation><genetic manipulation><genetically manipulate><genetically perturb><host response><immune activation><immune system response><immunoresponse><immunosuppressed patient><in vivo><infected with Cryptosporidium><intestinal pathogen><intestine pathogen><liquid><mRNA><malnourished><mortality><mouse model><mucous><murine model><nano particle><nano particle delivery><nano-sized particle><nanoparticle><nanoparticle delivered><nanoparticle delivery><nanosized particle><nitazoxanide><novel><nutrition deficiency><nutrition deficiency disorder><nutritional deficiency disorder><pathogen><public health relevance><rapid method><rapid technique><rhomboid><siRNA><siRNA delivery><siRNA therapy><siRNA-based therapeutic><social role><therapeutic siRNA><tool><transcription factor AP-2><transcription factor AP2><transcriptional silencing><two year old><two years of age><vaccine development><vaccine formulation><youngster>