The feasibility of microRNA gene silencing has been shown to provide a critical new method for cellular therapy. Creative Biolabs is recognized as the world leader in providing the most diverse portfolio of standard and custom microRNA gene silencing-based chimeric antigen receptor (CAR) T-Cell studies. Currently, we apply our expertise in the design and development services of microRNA-mediated CARs for clients all over the world.
In the past few decades, many attempts in gene engineering have opened up a new perspective for future CAR design and optimization. As a global CRO company dedicated to CAR T therapy research and development, Creative Biolabs has gone beyond the original design of CAR to improve safety and efficacy by combining our unique microRNA gene silencing technology with CAR expression. In general, our microRNA silencing domains are usually designed by simultaneously silencing the function of the MHC-TCR complex and inhibitory T cell receptors. The optimized gene silencing domain sequences are incorporated into CAR design to improve CAR T-cell persistence. Meanwhile, we can also achieve effective knockout of multiple targeted genes in CAR constructs by modifying the sequence of various miR-30 microRNA hairpins from a single transcript.
Fig.1 The Workflow of MicroRNA Gene Silencing-Based CAR T-Cell Development Platform.
Previous studies have indicated that the persistence of different CAR T cells after the infusion is associated with the mitochondrial adaptation of CAR T cells. Therefore, microRNA gene editing techniques in mitochondria, especially gene knockout, have shown great research potential. In Creative Biolabs, we strongly believe that manipulating microRNA regulation through certain targeted gene silencing is an ideal regulatory strategy for improving the effectiveness of CAR-T cell therapy. As a result, a large number of genes, either highly expressed in T cells or have an important role in T cell regulation, have been used as candidate microRNA targets. Furthermore, microRNAs often target multiple genes. Therefore, our microRNA gene silencing technology platform has been optimized to enable multiple genes to be silenced simultaneously, thus resisting T-cell exhaustion and improving CAR T-cell persistence.
| microRNAs | Targets |
|---|---|
| miR15a/b | ALDOA, PI3K, Glut3, PKM, Glut3, AKT1, AKT2, Drp1 |
| miR26a/b | Glut3, TIP1, GAPDH, PGK1, ACAT2, AMPK, MCT1, GAPDH |
| miR146a/b | Glut3, ALDOA, PGK1, AMPK, AKT1, MCT4 |
| miR101 | TIP1, mTOR, MCT, MFF |
| miR142 | Glut3, GPI, PFKM, ALDOA, PI3K, AKT2 |
| miR150 | Glut3, GAPDH, PKM, LDHA |
| miR16 | Glut4, HK1, ALDOA, PGK1, PKM, LDHA |
| miR29a | Glut3, GPI, ALDOA, PI3K, AKT2, MCT1 |
Recently, we generated a unique microRNA gene silencing modified CAR construct for the co-expression of a mini-CAR element and a microRNA gene silencing domain. We choose a three-generation anti-HER2 CAR with CD28, 4-1BB, as well as CD3z stimulatory domains. The sequence derived from clone trastuzumab biosimilar, a recombinant antibody produced by our company that can bind to HER2 antigen, has been used in CAR scFv design. Moreover, three popular inhibitory receptors, including PD-1, TIM3, and LAG3, have been subjected to sequence optimization and gene knockout. This chimeric anti-HER2 microRNA mini-CAR construct is then transferred to primary T cells via a lentiviral vector.
Fig.2 The MicroRNA Gene Silencing-Based Anti-HER2 CAR T-Cell Design.
Fig.3 and 4 The Preparation and Function Analysis of The Third-Generation Anti-HER2 MicroRNA CAR-T cells.
If you have any special needs in the design and generation of novel multiplex microRNA gene silencing technology or be interested in learning more about Creative Biolabs' microRNA gene silencing CAR services, please feel free to contact us for more details.
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