Generation of HIV-1 resistant T cells from induced pluripotent stem cell using CRISPR/Cas9 system

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The Step 1 of the study, "to establish anti-HIV-1 CRISPR-Cas9", has been completed successfully with its results published in the journal Viruses in 2020, as well as selected as the cover story of the journal's #11 issue. Here, we showed that our CRISPR construct blocked infectious HIV-1 NL4-3 replication in T cell culture for up to 45 days. In Step 2, "to transduce iPSC with CRISPR-Cas9", even though we were successful in introducing the transgene, we now aim to increase the transduction efficiency to ensure constitutive CRISPR-Cas9 and gRNA expression even after iPSC differentiation, Here, we made a new lentivector with Cocal virus (COCV) pseudotyped-envelope to replace VSV-G, which has been reported to increase transduction efficiency in hematopoietic stem cells. For Step 3, "to differentiate iPSC into T cells", we will adopt the "feeder-free" protocol which was recently published. However, the current lab of the grantee is not an iPSC lab, so setbacks are likely and differentiation into HSC and T cells may be more difficult and take longer time than first expected.