1. SAFETY EFFICIENCY REPROGRAMMING Integrating viral vectors*
Excisable viral vectors*
piggyBac
transposon
Non-integrating
vectors
Reprogramming
proteins
REPROGRAMMING FACTORS
MOUSE: Oct4 (Nr5a2), Klf4 (Esrrb, Klf2 or Klf5), Sox2 (Sox1), c-Myc (N-Myc or L-Myc)
HUMAN: Oct4, Klf4, Sox2 , c-Myc, Nanog, Lin28, hTERT, SV40LT
STARTING CELL POPULATION
MOUSE: embryonic and adult fibroblasts, neural stem cells, hepatocytes, gastric epithelial cells, B- cells
HUMAN: keratinocytes, skin fibroblasts, embryonic fibroblasts, CD34+ peripheral blood cells
SAFETY
EFFICIENCY
Easy to use; reproducible; allows secondary iPSC generation
Increased risk of insertional mutagenesis; possibility of transgene reactivation; incomplete silencing (lentiviruses)
Efficient Cre-mediated removal of transgenes
Exogenous DNA (primarily viral LTR) remains integrated following excision
Precise transposon excision leaves no ‘footprints’ of exogenous DNA
Time-consuming; excision may be inefficient
Low frequency of integration
Repeated transfection/infection required; possibility of random insertion of vector fragments
Eliminates risks associated with genome modification; non-DNA approach
Very slow process
Small molecules
can substitute for a reprogramming factor and/or enhance reprogramming
efficiency
Cell types with high endogenous levels of
a reprogramming factor require a minimal cocktail of factors
REPROGRAMMING
Differentiated
somatic cell
iPSCs
SMALL
MOLECULES#
VPA (histone deacetylase inhibitor); BIX (histone methyltransferase inhibitor); vitamin C; 5’-azaC (DNA methyltransferase inhibitor); BIO (GSK3 inhibitor); ALK5 and RepSox (inhibitors of TGF-β signaling); BayK8644 (L-type Calcium channel agonist); kenpaullone (kinase inhibitor)
advantages
disadvantages