Fig. 6: Xenograft transplant with HSPCs exhibiting high-frequency CCR5 editing confers resistance to CCR5-tropic HIV infection.

NSG mice were transplanted with either 1 × 10⁶ mock edited (GFP gRNA) or CCR5 edited (TB48 + TB50 dual gRNA) HSPCs following sublethal irradiation of 200 cGy. 24 weeks after transplant, mice demonstrating sufficient human T cell reconstitution (>10 CD4⁺ T cells/uL of blood) were challenged intraperitoneally with 20,000 TCID₅₀ of CCR5-tropic HIV_JRCSF. Viral loads and CD4⁺ T cell counts were assessed weekly by qRT-PCR and flow cytometry, respectively. Following two negative viral loads, uninfected mice were re-challenged with 100,000 TCID₅₀ HIV_JRCSF. A Viral load in individual mock edited (n = 11, green) and CCR5 edited (n = 6, blue) mice via qRT-PCR from weekly blood draws following HIV_JRCSF challenge. ^‡denotes animal euthanized prior to experimental endpoint due to health concerns. B Average frequency of human CD4⁺ T cells within total T cells in the blood of mock edited (green, n = 11) and CCR5 edited (blue, n = 5) mice following HIV_JRCSF challenge. *denotes p < 0.05 and statistics are generated from a one-tailed Student’s t-test. Error bars represent ± SEM. C Log₁₀-transformed number of HIV DNA copies per 1 × 10⁶ human cells as measured in genomic DNA extracted from pooled lymph nodes by a multiplexed droplet-digital PCR assay. The limit of detection (LOD) for the assay is set as the threshold to detect a single HIV DNA copy in the total number of human cells assayed per mouse. Lines represent the mean. D A single CCR5 edited humanized mouse that was resistant to both HIV_JRCSF challenges was subsequently challenged with 20,000 TCID₅₀ of HIV_NL4-3 via the intraperitoneal route and log₁₀-transformed HIV RNA copies per mL are plotted.