Next Generation Drugs Flashcards
(20 cards)
What are microRNAs (miRNAs)
Special class of non-coding RNAs that promote mRNA degradation and/or repress translation to fine-tune gene expression
What are some miRNA characteristics?
-When mature: ~22 nucleotides long
-Complementary in sequence to particular regions of mRNAs
-Found broadly in plants and animals
Process of forming miRNA
-Begin as long pri-microRNA (pri-miRNA)
-Drosha (endonuclease) processes pri-miRNA into pre-miRNA, which then exits the nucleus
-Dicer (endonuclease) processes pre-miRNA into miRNA and short RNA complement
-RNA helicase removes short RNA complement
-Mature mRNA is incorporated into RNA induced silencing complex (RISC)
Basics of how miRNA promotes mRNA degradation and/or repress translation
-Mature miRNA is incorporated into RISC
-Target RNA is bound to RISC
-Complementary determines whether translational repression of mRNA cleavage occurs
-If near-perfect complementarity, mRNA is cleaved
-If partial complementarity, translation is repressed
What are siRNAs
-Small interfering RNAs
-Synthetic
-Cleave segments of mRNA to silence mRNA
How are siRNAs formed and how do they work
-Double-stranded RNA is cleaved into 21-25 nt long fragments
—–Each has a 2 nt overhang at 3’ end
—–5’ phosphate
—–Mediated by Dicer homodimer
-siRNA is transferred to RISC and a helicase that separates the two strands
—–Guide RNA stays bound
—–Other strand is the passenger RNA, gets cleaved and discarded
-Guide RNA recruits the RISC complex to an mRNA with complementary sequence
-RISC component ‘Argonaute’ cleaves the mRNA opposite the bound guide RNA
What is RNAi?
Stands for RNA interference, includes siRNA and miRNA
What is the argonaute and what does it do?
-Argonaute proteins are a component of the RISC
-Facilitate cleavage of target mRNA
-Argonautes are RNAses
-Use miRNA and siRNA as guide RNA
What are ASOs
-Antisense oligonucleotides
-Single-stranded, synthetic RNA sequence which selectively binds via complementary base-pairing to mRNA
-Alters splicing to either include a skipped exon or exclude and econ
What does derivatization (modifications, etc) of RNA do in ASOs?
Increase nuclease resistance
What is a major challenge of ASO therapy?
Their short half-life, dsRNA and ssRNA are susceptible to breakdown by RNAses
What are some derivatizations of RNA in ASOs?
-Replacing the nonbridging oxygen at each phosphate with:
—–Dimethylamine (methylphosphonate)
—–Sulfur (phosphorothioate)
-Replace ribose and bridging oxygen with linked nitrogen
What are thiomorpholino oligonucleotides (TMOs) and what makes them useful?
-Derivatized versions of RNA
-Less susceptible to RNAse degradation
-Well-suited for use as antisense oligonucleotide drugs
How are TMOs applied to RNA?
-TMOs are not loaded on RISC like siRNA or miRNA
-Altered chemistry prevents loading
-Instead, TMOs bind mRNA directly
What are some mechanisms that TMOs can work with?
What can TMOs do?
-Inhibition of translation
-Inhibition of splicing
What disease does nusinersen work on and what is the cause of the disease?
-Spinal muscular atrophy (SMA)
-Typically, the cause is a mutation of SMN1 gene
-Humans have a near identical copy of SMN1
What does nusinersen do?
-Replaces SMN1 production by changing SMN2 splicing
-Antisense oligonucleotide promotes inclusion of exon 7 in SMN2
What are ProTaCs?
-Proteolysis Targeting Chimeras
-Targets proteins that are a product of a gene
-Ligand of the protein of interest and ligand of E3 ubiquitin ligase are covalently linked
-Ligand of protein of interest binds the protein, and E3 ligand recruits E3 for ubiquitination
-Some E3 ligases do not care exactly what they ubiquinate, as long as they are close
Thalidomide-based ProTaCs
-Thalidomide: degrades IKZF and SALL4
-Found that is makes off-target degradation of SALL4, which looks like IKZF1/3
