The Potential Use of the CRISPR-Cas System for HIV-1 Gene Therapy

Table 1

Strategies and results of CRISPR-Cas9-based gene therapy for HIV-1.

Stage	Components	Target to gRNA	Method of analysis	Results	Authors
Infection (CCR5)	Cas9+3 gRNA (CR1, CR2, and CR3)	CCR5 gene	CCR5 expression	Negative cells for CCR5 expression were 10.8% (CR1), 67.7% (CR2), and 36.7% (CR3)	[35]
Infection (CCR5)	Cas9+8 gRNA (sgR5-3 and sgR5-10)	CCR5 gene	CCR5 expression	Presence of 74.1% and 63.8% of indels in the CCR5 gene	[37]
Infection (CCR5)—in vivo	NPG rat	—	Viral RNA quantification	Reduction of viral RNA in peripheral blood from animals that received HSPCs with edited CCR5	[38]
Infection (CCR5Δ32)	Cas9+gRNA	CCR5 gene	Mutations into CCR5 gene	Presence of Δ32 deletion in the CCR5 coreceptor and reduction of viral replication	[44]
Infection (CCR5Δ32)	Cas9+2 gRNA	CCR5 gene	Mutations into CCR5 gene	Presence of Δ32 deletion in the CCR5 coreceptor between 20% and 60% of the cells	[41]
Infection (CCR5Δ32)—in vivo	Allogeneic stem cells transplantation with CCR5Δ32 mutation	—	CXCR4 and CCR5 gene expression	Interruption of antiretrovirals and reduction to undetectable levels of viral RNA	[45]
Infection (CCR5Δ32)—in vivo	Allogeneic stem cell transplantation with CCR5Δ32 mutation	—	CXCR4 and CCR5 gene expression	Reduction to undetectable levels of viral charge	[46]
Infection (CXCR4)	Cas9+10 gRNA	Conserved sites of CXCR4 gene	CXCR4 expression	Reduction of CXCR4 expression by 23.5% and 45%	[48]
Infection (CXCR4)	Cas9+gRNA	CXCR4 gene	CXCR4 expression	Reduction of CXCR4 expression by 60%	[50]
Infection (CXCR4)	Cas9+2 gRNA	CXCR4 gene	CXCR4 expression	Reduction of CXCR4 expression by 18.4% and 12.0%	[49]
Infection (CXCR4 and CCR5)	Cas9+2 gRNA	CXCR4 and CCR5 gene	CXCR4 and CCR5 gene expression	Presence of indels in 40.57% of the CXCR4 gene and 32.95% of the CCR5 gene	[52]
Infection (stable immunity)	Cas9+gRNA (stable in the cellular genome)	LTR (U3 and R) and rev	GFP reporter gene expression	Threefold HIV-1 expression reduction and resistance to viral infection	[54]
Viral infection	Cas9 (stable in the cellular genome)+2 gRNA	LTR	Viral particles	Rupture of viral genetic material and reduction of luciferase activity	[55]
Viral infection	Cas9+2 gRNA (stable in the cellular genome)	LTR	GFP reporter gene expression	Prevention of new HIV-1 infection by immunizing cells	[56]
Viral replication	Cas9-NLS+gRNA	LTR, gag, pol, tat, and rev	YFP reporter gene and Gag expression	Reduction of 57-89% in YFP expression and decline in Gag expression	[53]
Viral replication	Cas9+3 gRNA	LTR, gag e pol	Luciferase reporter gene expression	Reduction of 23% and 96% in luciferase expression with different combinations of gRNAs	[57]
Viral replication	Cas9+gRNA	LTR	HIV-1 copy number quantification; Gag and p24 expression	Reduction in HIV-1 copy number between 56% and 92% and decline in Gag and p24 expression in all patients	[55]
Viral replication	Cas9+gRNA	LTR and viral genes	p24 expression	Reduction in p24 expression	[58]
Viral replication	Cas9+gRNA	LTR and viral genes	Viral replication	Reduction of viral replication, which depends on the combinations of gRNAs used	[59]
Viral replication	Cas9+2 gRNA (stable in the cellular genome)	Gag, pol, env, and rev	Viral replication	Reduction of viral particles and number of infection cells; Reduction of reverse transcriptase activity	[60]
Viral integration	Cas9+gRNA	GFP coding region and LTR	GFP reporter gene expression	Reduction in GFP expression	[62]
Viral integration	Cas9-NLS+gRNA	LTR (R and U5)	Initial, late, and integrated HIV-1 DNA	Reduction of three- to fivefold of integrated viral DNA, twofold of late DNA, and no significant change in early DNA	[53]
Latency	Cas9+2 gRNA	LTR (TAR and NF-κB)	GFP reporter gene expression and MIF	Reduction in GFP and MIF expression for up to 20.0% potentiated after some transductions	[62]
Latency	Cas9+gRNA	GFP coding region and LTR	GFP reporter gene expression	Reduction of GFP expression, regardless of the amount of integrated virus, potentiated after some transductions	[54]
Latency	Cas9+10 gRNA	LTR, pol, tat, and rev	Activation of viral expression by GFP and p24	Reduction of 3- to 10-fold of GFP expression and 20-fold of p24 expression	[63]
Latency	Cas9+2 gRNA	LTR	GFP reporter gene expression	Reduction of GFP-positive cells to 4.5%	[62]
Latency	Cas9+4 gRNA	LTR (U3 region)	GFP reporter gene expression	Reduction of GFP and elimination of the proviral fragment, preventing reactivation and viral replication	[56]
Latency	Cas9+2 gRNA	—	GFP reporter gene expression and viral mRNA	Reduction of GFP and viral mRNA expression	[38]
Latency	Cas9+gRNA	gag, env, pol, vif, rev, and LTR	GFP reporter gene expression	Reduction in GFP expression between 48% and 92%, potentiated with multiple transductions	[62]
Latency	Cas9+43 gRNA	gag, env, pol, vif, rev, and LTR	P24 expression	Reduction of p24 expression, potentiated with multiple transductions	[62]
Latency (in vivo)	Cas9+2 gRNA; Tg26 transgenic mice	LTR and gag	DNA extracted from the liver lymphocytes, heart, spleen, lung, kidney, brain and blood	Presence of a 1323 bp and 368 bp fragments, with an additional fragment of 183 bp	[65]
Latency (in vivo)	Cas9+2 gRNA; Rats	LTR and gag	DNA extracted from liver, lymphocytes, heart, spleen, lung, kidney, brain, and blood	Elimination of viral fragment and reduction of viral mRNA	[65]
Latency (in vivo)	Cas9+gRNA; NCr nude mouse; BLT humanized mice	LTR and gag	Rupture of genomic DNA in several organs and tissues	Reduction of viral expression; Fragmentary deletion of provirus in several organs and tissues in the LTR and gag regions	[68]
Latency (in vivo)	Cas9+2 gRNA; NRG rats	LTR	Viral DNA	Reduction in more 90% of viral DNA and elimination of viral fragment present between target sites	[70]