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Full-Text Articles in Life Sciences

Crispr-Sonic: Targeted Somatic Oncogene Knock-In Enables Rapid In Vivo Cancer Modeling, Haiwei Mou, Deniz M. Ozata, Jordan L. Smith, Ankur Sheel, Suet-Yan Kwan, Soren Hough, Alper Kucukural, Zachary Kennedy, Yueying Cao, Wen Xue Apr 2019

Crispr-Sonic: Targeted Somatic Oncogene Knock-In Enables Rapid In Vivo Cancer Modeling, Haiwei Mou, Deniz M. Ozata, Jordan L. Smith, Ankur Sheel, Suet-Yan Kwan, Soren Hough, Alper Kucukural, Zachary Kennedy, Yueying Cao, Wen Xue

RNA Therapeutics Institute Publications

CRISPR/Cas9 has revolutionized cancer mouse models. Although loss-of-function genetics by CRISPR/Cas9 is well-established, generating gain-of-function alleles in somatic cancer models is still challenging because of the low efficiency of gene knock-in. Here we developed CRISPR-based Somatic Oncogene kNock-In for Cancer Modeling (CRISPR-SONIC), a method for rapid in vivo cancer modeling using homology-independent repair to integrate oncogenes at a targeted genomic locus. Using a dual guide RNA strategy, we integrated a plasmid donor in the 3'-UTR of mouse beta-actin, allowing co-expression of reporter genes or oncogenes from the beta-actin promoter. We showed that knock-in of oncogenic Ras and ...


Diverse Lipid Conjugates For Functional Extra-Hepatic Sirna Delivery In Vivo, Annabelle Biscans, Andrew H. Coles, Reka A. Haraszti, Dimas Echeverria, Matthew R. Hassler, Maire F. Osborn, Anastasia Khvorova Feb 2019

Diverse Lipid Conjugates For Functional Extra-Hepatic Sirna Delivery In Vivo, Annabelle Biscans, Andrew H. Coles, Reka A. Haraszti, Dimas Echeverria, Matthew R. Hassler, Maire F. Osborn, Anastasia Khvorova

RNA Therapeutics Institute Publications

Small interfering RNA (siRNA)-based therapies are proving to be efficient for treating liver-associated disorders. However, extra-hepatic delivery remains challenging, limiting therapeutic siRNA utility. We synthesized a panel of fifteen lipid-conjugated siRNAs and systematically evaluated the impact of conjugate on siRNA tissue distribution and efficacy. Generally, conjugate hydrophobicity defines the degree of clearance and the liver-to-kidney distribution profile. In addition to primary clearance tissues, several conjugates achieve significant siRNA accumulation in muscle, lung, heart, adrenal glands and fat. Oligonucleotide distribution to extra-hepatic tissues with some conjugates was significantly higher than with cholesterol, a well studied conjugate, suggesting that altering conjugate ...


Hydrophobicity Drives The Systemic Distribution Of Lipid-Conjugated Sirnas Via Lipid Transport Pathways, Maire F. Osborn, Andrew H. Coles, Annabelle Biscans, Reka A. Haraszti, Loic Roux, Sarah M. Davis, Socheata Ly, Dimas Echeverria, Matthew R. Hassler, Bruno M. D. C. Godinho, Mehran Nikan, Anastasia Khvorova Feb 2019

Hydrophobicity Drives The Systemic Distribution Of Lipid-Conjugated Sirnas Via Lipid Transport Pathways, Maire F. Osborn, Andrew H. Coles, Annabelle Biscans, Reka A. Haraszti, Loic Roux, Sarah M. Davis, Socheata Ly, Dimas Echeverria, Matthew R. Hassler, Bruno M. D. C. Godinho, Mehran Nikan, Anastasia Khvorova

RNA Therapeutics Institute Publications

Efficient delivery of therapeutic RNA beyond the liver is the fundamental obstacle preventing its clinical utility. Lipid conjugation increases plasma half-life and enhances tissue accumulation and cellular uptake of small interfering RNAs (siRNAs). However, the mechanism relating lipid hydrophobicity, structure, and siRNA pharmacokinetics is unclear. Here, using a diverse panel of biologically occurring lipids, we show that lipid conjugation directly modulates siRNA hydrophobicity. When administered in vivo, highly hydrophobic lipid-siRNAs preferentially and spontaneously associate with circulating low-density lipoprotein (LDL), while less lipophilic lipid-siRNAs bind to high-density lipoprotein (HDL). Lipid-siRNAs are targeted to lipoprotein receptor-enriched tissues, eliciting significant mRNA silencing in ...


Rac1 Activity Is Modulated By Huntingtin And Dysregulated In Models Of Huntington's Disease, Adelaide Tousley, Anastasia Khvorova, Neil Aronin, Kimberly B. Kegel-Gleason Feb 2019

Rac1 Activity Is Modulated By Huntingtin And Dysregulated In Models Of Huntington's Disease, Adelaide Tousley, Anastasia Khvorova, Neil Aronin, Kimberly B. Kegel-Gleason

RNA Therapeutics Institute Publications

BACKGROUND: Previous studies suggest that Huntingtin, the protein mutated in Huntington's disease (HD), is required for actin based changes in cell morphology, and undergoes stimulus induced targeting to plasma membranes where it interacts with phospholipids involved in cell signaling. The small GTPase Rac1 is a downstream target of growth factor stimulation and PI 3-kinase activity and is critical for actin dependent membrane remodeling.

OBJECTIVE: To determine if Rac1 activity is impaired in HD or regulated by normal Huntingtin.

METHODS: Analyses were performed in differentiated control and HD human stem cells and HD Q140/Q140 knock-in mice. Biochemical methods included ...