Overcomes the Challenges of CAR-T Cell Therapy

The development of chimeric antigen receptor T (CAR-T) cell therapy is a major breakthrough in cancer therapy due to the remarkable clinical responses observed in certain hematological cancer patients infused with cancer-targeting CAR-T cells. However, the widespread application of CAR-T cell therapy for the treatment of cancer has encountered several challenges, including (1) the potential for on-target/off-tumor toxicity and/or cytokine release syndrome (CRS); (2) relapses due to: (i) antigen escape and/or antigen heterogeneity of solid tumor, resulting in outgrowth of non-targeted tumor cells and/or (ii) a lack of CAR-T cell persistence; (3) lack of potency for the treatment of solid tumors; and (4) the high cost associated with CAR-T cell manufacturing.

Therefore, the potency and persistence of CAR-T cells are key to a successful CAR-T cell therapy. TSCM are characterized by stem-like properties with high capacity for self-renewal, resulting in long-term persistence. Furthermore, TSCM cells have the highest therapeutic efficacy due to enhanced metabolic fitness and low level of senescence and exhaustion (Figure 1). Consistent with these desired therapeutic properties, it has been clinically demonstrated that high levels of CAR-TSCM cells lead to better clinical outcomes.

CAR-T cell therapy faces several challenges in the context of solid tumor, including (1) lack of CAR-T cell trafficking to the tumor; (2) antigen heterogeneity of solid tumors; and (3) immunosuppressive tumor microenvironment (TME) which can adversely affect T cell fitness (e.g. differentiation, exhaustion, senescence, and survival). A technological breakthrough needs to be developed to overcome these hurdles.

Viral vector has been widely used for the development of CAR-T cell therapy due to its high efficiency in gene delivery and integration, resulting in stable long-term gene expression. However, CAR-T cells generated virally have several limitations. These include (1) virus-associated safety concerns; (2) limited payload capacity of viral vectors; (3) low expansion capacity and low percentage of CAR+ T cells following transduction; (4) low CAR-T cell persistence; and (5) high cost of manufacturing virally-produced CAR-T cells. 

A virus-free system can overcome most if not all of these hurdles, but electroporation-associated damages in virus-free systems present a critical challenge. In order for a virus-free system to be effective, we need: (1) a robust gene delivery system; (2) an effective system for integrating multiple genes into CAR-T cells; and (3) a robust cell expansion system for producing clinical-scaled CAR-T cells.

GenomeFrontier has developed Quantum CART (qCART™), a virus-free Quantum Engine™ for developing CAR-T cell therapy that synergistically integrates four platforms:

(1) G-Tailor™: a rapid multiplex gene design, construction, and screening system for designing CAR-T cells with (i) the ability to target to multiple tumor antigens; (ii) modulators for efficient CAR-T cell trafficking and TME resistance; and (iii) a safety control to terminate treatment as needed.

(2) Quantum Nufect™: a robust gene delivery buffer system for introducing therapeutic genes into T cells, resulting in (i) reliable CAR-T cell production while (ii) preserving high cell viability.

(3) Quantum pBac: a virus-free vector system with (i) a large payload gene integration capacity and (ii) high preference for TSCM genome integration.

(4) iCellar™: a robust cell expansion system for producing clinical-scaled CAR-T cells with (i) high percentage of CAR+ TSCM cells and (ii) enhanced fitness.

Collectively, qCART™ enables timely (~10 days) and cost-effective manufacturing of clinical-scaled CAR-T cells (1-3.5x109/L) (Table 1, Figure 2 and Figure 3).

Virus-free qCART™ robustly produces clinical-scaled CAR-T cells comprised of high percentage of CAR-TSCM cells (Table 1) capable of mediating high tumor cytolytic activities (Figure 2).

Virus-free qCART™ reliably produces CAR-TSCM cells with high expansion capacity against different targets and with various transgene sizes (Table 2), thereby demonstrating the feasibility of multiplex CAR-T construct design.