RNA Interference in Medicine

Question 1

RNAi

Answer 1

Down-regulation of gene expression by small RNA molecules that either inhibit translation or cause degradation of mRNAs of genes

Question 2

miRNAs

Answer 2

Transcribed from genes

Targets mRNAs of specific gene or set of genes

Question 3

miRNA production

Answer 3

Pri-miR has hairpin turns…cut by DROSHA…exported by exportin 5…pre-miR has hairpin cut by dicer…helicase unzips…incorporated into RISC

Question 4

RISC mechanism

Answer 4

Incorporates a single strand of miRNA into its sequence…if completelely complementary targets for destruction…if almost completely complementary, represses translation

Question 5

shRNAs

Answer 5

Short hairpin RNAs that can be introduced into the cell…bypasses the DROSHA and just cleaved by dicer

Question 6

siRNAs come from where and miRNAs come from where

Answer 6

siRNAs from shRNAs and dsRNAs

miRNAs from pre-miRNA transcripts

Question 7

Mediate RNAi by

Answer 7

Introducing processed miRNAs and siRNAs into cells…bypasses DROSHA and dicer

Question 8

miRNAs can come from

Answer 8

Individual genes or from introns…after removed from introns, cut by DROSHA and dicer and used as miRNA

Question 9

miRNAs can be altered in

Answer 9

Cancers

Question 10

miRNAs down regulated in cancers and how to treat

Answer 10

Ones that function to decrease expression of proliferation proteins like RAS…can artifically over-express miRNA to fix this

Question 11

miRNAs up regulated in cancers and how to treat

Answer 11

Ones that decrease tumor suppressor proteins…inhibt these miRNAs

Question 12

Antagomirs

Answer 12

anti-miRNAs

Can block activity of an miRNA of a dz

Perfectly complementary to the miRNA that you want to block

Bind to single strand form

Question 13

siRNAs can be administered how in order to get to one area?

Answer 13

Conjugated to a certain molecule

Question 14

single RNAi treatment is good for

Answer 14

2 weekas

Question 15

siRNAs are stabilized by modifying what and why

Answer 15

Methylating or fluorinating that 2’OH in order to make it unrecognizable to ribonucleases

Question 16

AAV delivered therapies

Answer 16

shRNAs inside and DNA is episomal so avoid integrating into the genome

Will not be passed from cell to cell

Good at going to many cells in the body

Long lasting in a give ncell

Question 17

RNAi to block plasma cholesterol

Answer 17

Use antagomir against miR-122

miR-122 reduces expression of DNA binding proteins that inhibits transcription of cholesterol biosyntehsis genes, resulting in higher cholesterol biosyntehsis

Question 18

RNAi is particularly good for

Answer 18

Dominant-negative dz (many neurodegen dzs)

Question 19

How to tell if dz is dom-negative

Answer 19

If you put normal protein back in and disease symptoms still present

Question 20

Direct delivery of genes

Answer 20

Therapeutic gene packaged into virus and injected into the patient

Question 21

Cell-based delivery of genes

Answer 21

Therapeutic gene inserted into virus…grown in culture with some type of stem cell…reintroduced into patient…allows amplification

Question 22

Benefits of AAVs

Answer 22

No disease association and provoke limited innate immunity

Question 23

Retroviruses

Answer 23

Have genomes of single-stranded RNA

Question 24

Reverse transcriptase is

Answer 24

RNA-dependent DNA polymerase

Most replication polymerases are DNA-dependent DNA polymerase

Question 25

How can you restrict expression of inserted genes?

Answer 25

Use a tissue specific promotor

Question 26

Gene therapy would NOT work in

Answer 26

Dominant negative disease

Question 27

Gene therapy works great with

Answer 27

Loss of function disease

Question 28

Gene therapy of CF

Answer 28

inhale AAV with gene sequence…infects lung cells and now grow normally

Question 29

Gene therapy for blood clotting

Answer 29

Liver chosen as target because where most serum proteins are made (Factor 8)

Question 30

Gene therapy in diabetes

Answer 30

Transfers gene that make cells differentiate into missing pancreas B-cells…remove cells, induce, transplant

Question 31

Gene therapy for ADA-SCID

Answer 31

Remove T cells and infect T cells with retrovirus…Use ADA cDNA, will correct gene…amplify and transplant

Toxic level of adenosine and deoxyadenosine decreases

Question 32

Gene therapy for choroideremia

Answer 32

CHM gene mutation…gene into a virus and injected beneath the retina

Question 33

Retinitis pigmentoasa

Answer 33

Inherited degen dz in which photoreceptor cells die

Question 34

Retinitis pigmentosum gene therapy

Answer 34

Use PEDF (pigment epithelium-derived factor) which is a growth factor in order to induce growht

Question 35

Oncolytic virus mechnaisms

Answer 35

Oncolytic virus replicates in cancer cells…cancer cells rupture and then virus can infect other cells thus amplifying the effect…cannot replicate in normal cells