Clinical pharmacology II Flashcards
HIV lecture
What are the pitfalls of HIV treatment? (3)
- Lifelong treatment
- Many side-effects
- Not curative
What are the main characteristics of HIV? (2)
- Lentivirus - ssRNA
- Reverse transcriptase
Which receptors can HIV use for entry in the cell?
CCR5 or CXCR4
Describe the viral life cycle of HIV briefly (5)
- Entry
- RNA -> DNA
- DNA integration in the nucleus
- Transcription + translation
- Budding
Which cells can be infected by HIV? (3)
- CD4+ T cells
- Macrophages
- DCs
HIV: After how many days are many progeny virions produced?
~2 days
Antiretroviral therapy does not eliminate the latent reservoir. How many cells remain latently infected?
1 in 100.000 cells
What is meant with the effect of the classical/sterilizing HIV cure?
Replication competent HIV provirus is completely eliminated from the blood
- Individual who received a SCT from a donor who has natural immunity to HIV
- Individual who has cleared the virus in the absence of ART
Which gene therapy strategies can be explored in combating HIV? (2)
- RNAi
- CRISPR-Cas
To what end is RNAi used to combat HIV?
Combination of production of HIV-resistant anti-HIV immune effector cells
To what end is CRISPR-Cas used to combat HIV?
Specific inactivation of integrated HIV DNA from infected cells
Current gene therapy approaches for HIV focus on?
Ex vivo transduction of HPSC or T cells followed by rein fusion of the modified cells -> highly inefficient
What is the advantage of using RNAi to combat HIV?
You don’t have to introduce an external protein -> we have all the tools in our body
What are miRNAs?
Small molecules (20-25nt) which regulate expression of genes
- siRNA is complementary to target mRNA
- siRNA is loaded
- siRNA is cleaved
What are examples of exogenous RNAi inducers? (2)
- Virus
- Vector
Where can you use RNAi in de viral life cycle? (2)
- Directly after viral RNA enters the cell (not efficient)
- Translation after exiting the nucleus
miRNA biosynthesis (12:30 min)
Partial complementarity vs perfect complementarity
Describe the basic CRISPR-Cas mechanism (2)
- Cas (nuclease) binds to short CRISPR RNA
- crDNA targets complementary viral RNA/DNA sequences
What are the three nucleases?
- Cas9
- Cas12
- Cas13d
What are the characteristics of Cas9? (3)
- Targets dsDNA
- Blunt ends
- PAM requirement
What are the characteristics of Cas12? (3)
- Targets dsDNA
- Stagger ends
- PAM requirement
What are the characteristics of Cas13d? (2)
- Targets ts ssRNA
- No PAM requirement
How can the CRISPR-Cas cut be repaired? (2)
- NHEJ
- Homology-directed repair (HDR)
Using either RNAi or CRISPR-Cas, you target either…? (2)
- Sequences essential for HIV-1
- Conserved HIV-1 sequences
True or false: nearly all sgRNAs to different HIV protein exhibit potent suppressive capacity
True
