The introduction of Novartis CD19 CAR-T with T-chargeTM process

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The introduction of Novartis CD19 CAR-T with T-chargeTM process

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The introduction of Novartis CD19 CAR-T with T-chargeTM process.

Introduction of T-charge rapid production process

Prolonged T cell culture in vitro depletes the CAR-T end product of naive and stem cell memory T cell (Tscm) subsets associated with enhanced antitumor efficacy.

YTB323 is a CD19-directed autologous CAR-T cell therapy with a greatly simplified manufacturing process that eliminates complexities such as long culture cycles.

This improved T-Charge™ process preserves T cell stemness, an important feature closely related to therapeutic potential, resulting in enhanced CAR-T cell expansion and enhanced antitumor activity.

T-charge rapid production process method

The new T-Charge ™ manufacturing platform was evaluated in a preclinical setting, which reduced ex vivo culture time to approximately 24 hours, with the final product being manufactured in less than 2 days.

T cells were enriched from healthy donor leukocytes, then activated and transduced with a lentiviral vector encoding the same CAR used for tisagenlecleucel.

After approximately 24 hours in culture, cells (YTB323) were harvested, washed and formulated. Meanwhile, CAR-T cells (CTL*019) were generated from the same healthy donor T cells and the same lentiviral vector using a traditional in vitro expanded CAR-T manufacturing protocol (TM process).

After manufacture, the CAR-T product was evaluated in in vitro and in vivo T cell function assays in immunodeficient NSG mice (NOD-scid IL2Rg-null) vaccinated with pre-B-ALL cells (NALM6) or DLBCL cells line (TMD-8) to evaluate antitumor activity and CAR-T amplification.

Test results of T-charge rapid production

The YTB323 CAR-T product, produced by this novel scale-free manufacturing process, preserves the immunophenotype of input leukapheresis; specifically, preserves naive/Tscm cells (CD45RO-/CCR7+) as shown by flow cytometry . In contrast, the process of in vitro expansion yielded a final product consisting primarily of central memory T cells (Tcm) (CD45RO+/CCR7+) (Panel A).

Further evidence supporting retention of the initial phenotype was illustrated by bulk and single-cell RNA sequencing experiments comparing leukapheresis and final product of CAR-T generated using the T-Charge™ protocol.

YTB323 CAR-T cell potency was assessed in vitro using a cytokine secretion assay and a tumor repeat stimulation assay designed to test the persistence and depletion of the cellular product. Compared with CTL*019, YTB323 T cells secreted 10- to 17-fold higher levels of IL-2 and IFN-γ after CD19-specific activation.

Furthermore, YTB323 cells were able to control tumors with a 30-fold lower effector:tumor cell ratio and were stimulated at least 7 more times in repeated stimulation experiments. Both assays clearly demonstrated the enhanced potency of YTB323 CAR-T cells in vitro.

The final preclinical assessment of YTB323 cell efficacy was in vivo expansion and antitumor efficacy against B cell tumors in a multi-dose immunodeficient NSG mouse model by comparison with CTL*019.

The expansion of CD3+/CAR+ T cells in blood was analyzed weekly by flow cytometry for up to 4 weeks. Dose-dependent expansion (Cmax and AUC 0-21d) was observed for both YTB323 and CTL*019.

Compared with multiple doses of CTL*019, the Cmax of YTB323 was ≈40 times higher and the AUC0-21d was ≈33 times higher.

Peak expansion (Tmax) was observed to be delayed by at least 1 week in YTB323 compared to CTL*019, supporting that the increase in expansion is driven by the less differentiated T cell phenotype of YTB323.

YTB323 controlled NALM6 B-ALL tumor growth at a lower dose of 0.1 × 10 CAR+ cells compared to 0.5 × 10 CAR+ cells required for CTL*019 (Panel B).

In the DLBCL model TMD-8, only YTB323 was able to control tumors, while CTL*019 led to tumor progression at various doses. This ability of YTB323 cells to control tumors at lower doses confirms their robustness and efficacy.

The new manufacturing platform T-Charge™ for the YTB323 is simplified, shortened and does not require expansion. Therefore, it preserves T cell stemness, which is associated with improved in vivo CAR-T expansion and antitumor efficacy.

Compared with approved CAR-T therapies, YTB323 has the potential to achieve higher clinical efficacy at respective lower doses. T-Charge™ is designed to revolutionize CAR-T manufacturing while increasing the likelihood of long-term deep response.

Clinical trials of cells made by T-Charge

This Phase I, multicenter, dose-escalation study (NCT03960840) is evaluating the safety and preliminary efficacy of YTB323 in patients with B-cell malignancies.

The results presented here focus on the DLBCL cohort. Eligible patients were adults with measurable disease, ECOG 0-1, and r/r DLBCL at enrollment with ≥2 lines of prior therapy, including after autologous hematopoietic stem cell transplantation (aHSCT).

Patients received a single dose of YTB323 at dose level 1 (DL1; 1-2.5×106 CAR+ cells), DL2 (5-12.5×106 CAR+ cells), or DL3 (25-40×106 CAR+ cells). Bridging therapy prior to YTB323 is optional.

The primary endpoint was the incidence of dose-limiting toxicity (DLT) in the first 28 days and the safety of determining the recommended phase II dose (RP2D).

Secondary endpoints were cellular kinetics, local investigator-assessed overall response rate (ORR), duration of response, and overall survival.

Clinical trial results of cells made by T-Charge

As of April 16, 2021, 15 patients with r/r DLBCL had been infused with YTB323: 4 at DL1, 10 at DL2, and 1 at DL3 (figure, n=14 for DL1 and DL2).

The median age was 65 years; most (60%) had received 2nd-line therapy and 4 (27%) had received aHSCT. All adverse events were reported regardless of study drug relationship.

Of the 15 patients evaluable for safety, 4 patients (27%) reported at least 1 grade 3 (Gr) AE, 6 (40%) reported at least 1 grade 4 AE, 2 (13%) ) reported at least 1 grade 5 AE.

The most frequently reported Gr3/4 AEs were thrombocytopenia (n=2, 13%), neutropenia (n=3, 20%), and neutropenia (n=3, 20%) .

Seven patients (47%) had neurological AEs, of which 2 events (13%) were considered severe – Gr3 peripheral neuropathy (unrelated) and Gr2 seizures (immune effector cell-associated neurotoxicity syndrome grade 3, and treatment).

Cytokine release syndrome (CRS) was experienced in 4 patients (27%), 3 (20%) Gr1/2 and 1 (7%) 4 (Lee et al, 2014) who met criteria for DLT. Two (13%) and one (7%) patients were treated with tocilizumab and corticosteroids for CRS, respectively.

There were 4 deaths in the trial, all unrelated to YTB323: 2 from disease progression and 2 from sepsis (1 at DL1 and 1 at DL3).

The median time to onset of CRS was 9 days (range, 9-9 days) for DL1 and 11 days (range, 8-17 days) for DL2. A preliminary dose-dependent response was observed.

At DL1, 4 patients were evaluable for efficacy at Mo3, with both ORR and CR rates of 25% (95% CI, 0.6%-80.6%).

At DL2, 8 patients were evaluable for efficacy at Mo 3, of which 2 were in CR prior to YTB323 infusion; both ORR and CR rates were 75% (95% CI, 34.9%-96.8%).

Dose-dependent expansion (Cmax and AUC 0-28d) was observed following infusion at DL2 of 13.6% CAR+ or 32,100 copies/µg DNA in CD3+ cells.

Although long-term persistence could not be assessed, 3 of 8 patients with DL2 had 3-month follow-up and 3 had CAR expression detected by flow cytometry (≥1%).

Peak expansion time (Tmax) coincided with peak cytokine levels (approximately 16 days after infusion). The novel manufacturing method of YTB323 allows preservation of CD4 and CD8 naive/Tscm cells in the final product, as determined by flow cytometry.

Batch and single-cell RNAseq analysis showed that YTB323 retains a naive stem cell-like phenotype ( clinical trial data: CYTB323A12101 ).

DL3 is undergoing YTB323 recruitment; RP2D remains to be determined. At DL2, YTB323 showed promising efficacy and favorable safety profile.

Current data supports continued development of YTB323 in r/r DLBCL pts.

references:

1、Preservation of T-Cell Stemness with a Novel Expansionless CAR-T Manufacturing Process, Which Reduces Manufacturing Time to Less Than Two Days, Drives Enhanced CAR-T Cell Efficacy

2、A First-in-Human Study of YTB323, a Novel, Autologous CD19-Directed CAR-T Cell Therapy Manufactured Using the Novel T-ChargeTM platform, for the Treatment of Patients (Pts) with Relapsed/Refractory (r/r) Diffuse Large B-Cell Lymphoma (DLBCL)

The introduction of Novartis CD19 CAR-T with T-chargeTM process

(source:internet, reference only)

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