RNA-Based Therapeutics and Genome Editing Quiz

Overview of RNA-based Therapeutics and Genome Editing

• Ribozymes are RNA sequences that function as catalysts to cleave specific mRNA structures, and can be directed to a specific mRNA by introducing short flanking oligos.
• Aptamers are single-stranded DNA or RNA-based sequences that fold up to adopt a unique 3D structure, allowing them to bind a specific target molecule with high specificity and binding affinity.
• Macugen is an FDA-approved aptamer that binds vascular endothelial growth factor and is indicated for the treatment of neovascular age-related macular degradation.
• CRISPR-Cas9 is a bacterial immune system that confers immunity in three steps: acquisition, expression, and interference.
• CRISPR-Cas9 can be exploited as a genome-editing tool by using a single-guide RNA molecule to target and cleave specific DNA sequences.
• Editing events occur when the cell tries to repair the damage via either non-homologous end-joining or homology-directed repair.
• CTX001 is a promising CRISPR/Cas9-based genome-editing therapy currently undergoing late-stage clinical trials for two monogenic diseases: Transfusion-dependent β-thalassemia and Sickle Cell Disease.
• Both diseases result from mutations to the β-globin gene, and CTX001 functions to elevate patient’s HbF, thereby alleviating transfusion requirements for TDT patients and painful and debilitating sickle crises for SCD patients.
• CTX001 is co-run by Vertex Pharmaceuticals and CRISPR Therapeutics, founded in 2013 by Emmanuelle Charpentier.
• Ribozymes have been targeted against hepatitis C, SARS-CoV-2, and Influenza in preclinical trials.
• Aptamers may prove useful for affinity-based purification, diagnostics, and therapeutics, similar to antibodies.
• CRISPR/Cas9-based genome-editing therapies have potential for treating a wide range of genetic diseases.

Types of Stem Cells and Their Characteristics

• There are three classes of stem cells: totipotent, pluripotent, and multipotent.
• Stem cells are classified based on their source: embryonic, adult, and induced pluripotent (iPS) cells.
• Embryonic stem (ES) cells are derived from pre-implant-stage human embryos, usually from in vitro fertilization procedures.
• ES cells can multiply while remaining undifferentiated, but have limitations such as difficulties maintaining lines and potential for tumor formation.
• Somatic cell nuclear transfer (SCNT), also known as therapeutic cloning, is a process to create new cell lines, but it has ethical and technical issues.
• Adult stem cells are undifferentiated cells found in tissues or organs, can differentiate to yield major cell types, and are used to maintain and repair tissue.
• Adult stem cells have advantages over ES cells, such as overcoming ethical difficulties and allowing for autologous transplantation.
• Adult stem cells have been used to successfully treat over 60 human diseases.
• Induced pluripotent stem (iPS) cells are artificially derived from non-pluripotent cells by inducing the expression of certain genes.
• iPS cells are typically derived through transfection of stem cell-associated genes into non-pluripotent cells using viral vectors.
• iPS cells have similarities to ES cells, but their generation has technical challenges such as inefficient process and genomic instability.
• The process of generating iPS cells involves isolating and culturing host cells, introducing ES-specific genes into the cells, and harvesting and culturing the cells to generate iPS cells.