Universal allogeneic CAR T cells engineered with Sleeping Beauty transposons and CRISPR-CAS9 for cancer immunotherapy

Applications of CRISPR technology in cellular immunotherapy

“…To address this issue, various groups are currently adopting CRISPR technology to generate universal CAR‐T cells using allogeneic T cells. Typically, to prevent graft‐versus‐host disease (GvHD) and host rejection of the graft, TRAC and/or TRBC , which encode the constant chain of TCRa or TCRb, respectively, as well as B2M , which is essential for HLA‐I expression, and class II major histocompatibility complex trans‐activator (CIITA) for HLA‐II expression are disrupted 107,108,117,118 . It was also shown that deletion of CD52 on CAR‐T cells may support anti‐CD52‐mediated depletion of autologous T cells in patients 119 …”

“…Typically, to prevent graft-versus-host disease (GvHD) and host rejection of the graft, TRAC and/or TRBC, which encode the constant chain of TCRa or TCRb, respectively, as well as B2M, which is essential for HLA-I expression, and class II major histocompatibility complex transactivator (CIITA) for HLA-II expression are disrupted. 107,108,117,118 It was also shown that deletion of CD52 on CAR-T cells may support anti-CD52-mediated depletion of autologous T cells in patients. 119 However, simply disrupting TCR and/or HLA-I on allogeneic CAR-T cells might not yield optimal outcomes.…”

Molecular and therapeutic effect of CRISPR in treating cancer

“…For reduction of off-the-shelf allogeneic CAR T cell therapy with sleeping beauty (SB) transposons and CRISPR Cas 9 with minicircle (mc) DNA plasmid for transfection increase efficacy. By inactivation of TCR by CRISPR Cas9-ribonucleoparticles (RNPs) for CD19 targeted CAR reduce allo-reactivity due to the reduction of endogenous TCR expression and SB-CD19-28z.CAR T/TCR KO T cells for a increase memory phenotype thus improve the chances of using CAR T cells for therapy with reduce GVHD [ 60 ].…”

Uncertainty quantification for gene delivery methods: A roadmap for pDNA manufacturing from phase I clinical trials to commercialization

“…[54] The sleeping beauty (SB) transposon system is the most used one, with clinical trials reporting encouraging results for the production of CD-19-specific CAR T-cells. [18][19][20][21][22]55] The most commonly used method for the delivery of transposon-derived plasmid vectors is electroporation. [33,54] The main benefits of non-virally modified CAR T-cells are the shorter upstream process owing to the simplicity of the transfection process and the overall decrease in cost compared to the viral-based counterpart.…”

“…[13,14] At the same time, their production and scale-up under current Good Manufacturing Practices (cGMP) are characterized by complex, costly, and time-intensive manufacturing processes, leading to logistical challenges that impact their cost and availability. [10,[15][16][17] To overcome these challenges, attention has shifted to non-viral delivery methods as an attractive alternative to viral vectors, [18][19][20][21][22][23][24][25][26][27] offering several advantages such as simpler manufacturing processes, increased yields, and reduced costs, [12,17,28] while also addressing safety and immunogenicity concerns that are associated with viral vectors. [29,30] Non-viral methods include the delivery of naked pDNA as well as physical and chemical pDNA delivery methods.…”