BACKGROUND: Currently approved CAR-T products were all derived from viral vector transfection, which increases a potential tumorigenesis risk for patients' long-term survival, and manufacturing costs as well. We developed a novel, non-viral, CRISPR/Cas9-mediated PD-1 locus specifically integrated anti-CD19 CAR-T cells (BRL-201) to address these limitations. In the first-in-human phase I study for the treatment of relapsed/refractory (r/r) non-Hodgkin's lymphoma (NHL) patients (NCT04213469), our preliminary results showed a promising efficacy with a pretty good safety profile. Here, we update the patients with long term sustained remission after at least 4-year follow up. A phase I/II of BRL-201 (NCT05741359) is ongoing in multi-center with a large sample size to further evaluate of BRL-201 antitumor activity in China.

AIM： The study aims to evaluate the tolerability and safety of escalated doses of BRL-201 in patients with r/r B-cell NHL.

METHODS: The eligible participants underwent leukapheresis followed by lymphodepletion regimen with cyclophosphamide (500mg/m², D-3 to -2) and fludarabine (30mg/m², D-4 to -2) before BRL-201 infusion. Three dose levels (2.0, 4.0, and 6.0 × 10⁶ CAR-positive cells/kg) were designed with a single infusion using a 3+3 dose-escalation schema. Additional non-standard doses (0.56-0.8 × 10⁶ CAR-positive cells/kg) were also administered to three subjects. The final recommended optimal dose was 2.0 × 10⁶ CAR-positive cells/kg in the extended patients. The primary and secondary endpoints were the incidence of dose-limiting toxicities (DLT) and overall objective response at 3 months, respectively. The phase I/II trial also employed the 3+3 escalation rule.

RESULTS: From May 3, 2020, to August 10, 2021, 25 patients with r/r B-NHL were enrolled; 21 received BRL-201, with a median age of 56 (range: 34-70) and median of 4 (range: 1-9) prior therapy lines. 17 patients were diagnosed with disease stages III or IV, and 13 patients were high-intermediate to high risk per IPI/aaIPI. Two had previous autologous HSCT, and one had primary refractory disease.

As of July 11, 2025, the median follow-up was 55.2 months (95%CI: 47.7-58.5). The best overall response rate was 100%, with an 85.7% (18/21) complete response (CR) rate. The estimated 5-year progression-free survival (PFS) and overall survival (OS) rates were 40.6% and 42.9% respectively, and the survival curves showed a gradually flattering trend over time. Patients with CR had longer PFS and OS, with 5-year rates of 47.4% and 50.0%, respectively. All the survival times were matured, median duration of response (DOR), median PFS, and median OS were 19.5 months (95%CI: 5.9-20.9), 20.4 months (95%CI: 6.8-21.8), and 41.5 months (95%CI: 12.6-45.3), respectively. As the cutoff date, nine patients (42.8%) survived; among them 7 patients (33.3%) remained disease-free remission. In the 17 patients with available PD-L1 expression, PD-L1-negative accounted for 16.7% (1/6) and 45.5% (5/11) between alive and death groups, respectively. The OS of the two groups were 45.3 months (95%CI: 12.6-45.3) versus 10.3 months (95%CI: 1.9-16.8), which had a significant statistic difference (P=0.008), Hazard ratios 0.1746 (95%CI: 0.03788-0.8045). For all treated patients, mild cytokine release syndrome (CRS) (grade 1/2) occurred in 66.7% (14/21) of patients, with one requiring tocilizumab. Grade 1/2 ICANS was observed in 19.0% (4/21). No new safety issues were identified during the extended follow-up period. In the ongoing phase I/II study, no DLTs, neurotoxicity, or serious adverse events (SAE) were observed within 28 days after receiving BRL-201 infusion, nor were there any cases of grade≥3 CRS.

CONCLUSIONS: With a median 55.2 month follow-up, BRL-201 demonstrated durable and deep responses. One-third of patients remained long-term survival for more than 4 years with disease-free remission. The estimated 5-year OS rate with CR patients was 50.0%. The expression of PD-L1 may have a favorable correlation with long-term survival of relapsed or refractory lymphoma patients, which highlights the potential efficient antitumor activity of BRL-201 product. Further investigation is ongoing to verify its clinical benefits in patients with r/r B-cell NHL.

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2025
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