Table 3.

Gene-editing strategies.

Nuclease	Mechanism	Effect	DM1 Model	Advantages	Limitations	Ref
ZNF	Induction of DNA double strand breaks at CAG/CTG repeats	Repeat contractions	Yeast cells carrying CTG repeats	Permanent reduction in CTG repeats; good cleavage efficiency	Repeat rearrangements	[171]
ZNF	Induction of DNA double strand breaks at CAG/CTG repeats	Repeat contractions and duplications	Mammalian cells carrying CTG repeats	Permanent reduction in CTG repeats; good cleavage efficiency	Repeat duplications	[172]
TALEN	Induction of DNA double strand breaks at CAG/CTG repeats	Repeat contractions	Yeast cells carrying CTG repeats	Permanent reduction in CTG repeats; good cleavage efficiency, no mutations	Application limited to yeast cells	[173,174]
TALEN	Insertion of a polyA signal upstream CTG repeats	Production of shorter DMPK transcripts (no CUG)	DM1-patient-derived iPSCs	Elimination of toxic CUG repeats from DMPK transcript	Production of truncated DMPK protein; retention of CTG at DMPK locus	[43,175]
SpCas9 D10A	Induction of DNA single strand breaks at CAG/CTG repeats	Repeat contractions	Human cells carrying CAG/CTG repeats	Permanent reduction in CTG repeats	Cell-type dependent efficiency	[176]
dCas9	Block DNA transcription at CTG repeats	DMPK transcription inhibition	DM1-patient-derived cells; HSALR mice	Suppression of CUG-repeat-transcript production	Decreased DMPK protein production; retention of CTG at DMPK locus; need of repeated treatment	[124]
RCas9	Cleaving single-strand RNA at CUG repeats	CUG-repeated transcript degradation	DM1-patient-derived cells; HSALR mice	Elimination of CUG-repeat transcript	Decreased DMPK protein production; retention of CTG at DMPK locus; need of repeated treatment	[125,177]
SpCas9 D10A	Insertion of a polyA signal upstream CTG repeats	Production of shorter DMPK transcripts (no CUG)	DM1-patient-derived iPSCs	Elimination of toxic CUG repeats from DMPK transcript	Production of truncated DMPK protein; retention of CTG at DMPK locus	[178]
SpCas9 or SaCas9	Induction of two DNA double strand breaks at CTG-repeats flanking regions	Deletion of CTG expanded region	DM1-patient-derived iPSCs	Permanent elimination of toxic CTG repeats; no off-targets	Low efficiency using SpCas9; higher efficiency but frequent inversions using SaCas9	[178]
SpCas9	Induction of two DNA double strand breaks at CTG-repeats flanking regions	Deletion of CTG expanded region	DM1-mouse-derived myoblasts; DM1-patient derived myoblasts	Permanent elimination of CTG repeats; no off-targets	On-target indels, inversions, large deletions	[179]
SpCas9	Induction of two DNA double strand breaks at CTG-repeats flanking regions	Deletion of CTG expanded region	DM1-patient-derived MYOD1-converted fibroblasts	Permanent elimination of CTG repeats; no off-targets	On-target indels, inversions	[40]
SpCas9	Induction of two DNA double strand breaks at CTG-repeats flanking regions	Deletion of CTG expanded region	DM1-patient-derived primary myoblasts; DM1-patient-derived iPSCs	Permanent elimination of CTG repeats; good editing efficiency in iPSCs; no off-targets	On-target indels, partial deletions	[180]
SaCas9	Induction of two DNA double strand breaks at CTG-repeats flanking regions	Deletion of CTG expanded region	DM1-patient-derived myoblasts; DMSXL mice	Permanent elimination of CTG repeats; good editing efficiency in DM1 cells; no off-targets	On-target indels; low editing efficiency in mice skeletal muscle	[123]
eSpCas9	Induction of two DNA double strand breaks at CTG-repeats flanking regions	Inducible deletion of CTG expanded region	DM1-patient-derived MYOD1-converted fibroblasts; DMSXL mice	Permanent elimination of CTG repeats; well-regulated editing induction; no off-targets	On-target indels, inversions, large deletions	[122]

Abbreviations. dCas9 = deactivated Cas9; eSpCas9 = enhanced Streptococcus pyogenes Cas9; RCas9 = RNA-targeting Cas9; SaCas9 = Staphylococcus aureus Cas9; SpCas9 = Streptococcus pyogenes Cas9; SpCas9 D10A = SpCas9 nickase.