Research Breaks Through Bottleneck in T-cell Leukemia Treatment

T-cell acute lymphoblastic leukemia (T-ALL) is a highly aggressive hematologic malignancy, accounting for approximately 15% of childhood acute lymphoblastic leukemia (ALL) and 25% of adult ALL. While cure rates for pediatric patients can reach 80%, the long-term survival rate for adult patients remains below 40%. More concerningly, more than half of patients relapse after treatment or fail to respond to standard therapy, with the median overall survival for relapsed/refractory T-ALL being only approximately eight months. Current treatment options primarily rely on intensive chemotherapy and allogeneic hematopoietic stem cell transplantation (alloHSCT). However, these treatments are associated with significant toxicity and high failure rates, necessitating an urgent need for safer and more effective targeted therapy strategies.

Compared to B-cell malignancies, immunotherapy for T-ALL has progressed more slowly. Because malignant T cells closely resemble healthy T cells in surface antigen expression, traditional CAR-T cell therapies face critical challenges such as fratricide and T cell aplasia. In recent years, researchers have attempted to utilize gene editing to knock out pan-T antigens such as CD7. Despite some progress, the technology remains complex, regulatory challenges remain, and salvage therapy still relies on alloHSCT. Therefore, identifying safe, specific, and non-pan-T antigen targets has become a key breakthrough in T-ALL immunotherapy.

Recently, a research report titled "CAR-T cells targeting CCR9 and CD1a for the treatment of T cell acute lymphoblastic leukemia" published in the international journal of Hematology & Oncology reported that scientists focused on two non-pan-T antigens—CCR9 and CD1a—to simultaneously develop and validate a dual-target CAR-T cell therapy strategy. This strategy aims to overcome antigen escape, manufacturing difficulties, and immunotoxicity issues in T-ALL treatment. This research may have significant clinical translational value.

In the article, researchers first performed immunophenotyping on samples from 180 T-ALL patients. The results revealed that CCR9 was highly expressed in 73% of patients, with a particularly high expression rate of 92% in relapsed samples. More importantly, CCR9 is barely expressed in normal T cells, hematopoietic stem cells, and other tissues, and is only weakly expressed in some thymocytes and intestinal lymphocytes, demonstrating a favorable safety profile. CD1a, on the other hand, is primarily expressed in cortical T-ALL, which accounts for approximately 30% of all T-ALL. The combination of these two drugs can cover 86% of patients and significantly expand the treatment population.

In the experimental design, the researchers used hybridoma technology to screen for high-affinity CCR9 monoclonal antibodies and constructed murine and humanized single-chain Fvs (scFvs). These scFvs were then embedded into second-generation CAR constructs to generate three types of CCR9-CAR-T cells (murine M, humanized H1, and H2). In vitro cytotoxicity assays revealed that H2 CAR-T cells exhibited the strongest activity and specificity against CCR9-positive T-ALL cell lines (such as MOLT4) and patient-derived xenograft (PDX) models, without significant fratricidal activity.

Figure 1. CCR9 is a safe and specific target for T-ALL. (Tirado N, et al., 2025)

To further validate its in vivo efficacy, researchers established a T-ALL PDX model in NSG mice. CCR9-positive leukemia cells were injected and then treated with different CAR-T cell therapies. Results showed that CCR9 H2 CAR-T cells significantly inhibited tumor progression, with 80% of mice achieving complete remission, surpassing other treatment groups. Building on this success, the researchers further developed dual-target CAR-T cells that simultaneously target both CCR9 and CD1a. Three strategies, co-transduction, tandem CAR, and bicistronic CAR, were employed for construction and comparison. Ultimately, the co-transduction strategy (individual transduction of CCR9 and CD1a CARs) demonstrated the best killing efficiency and antigen coverage. The results showed that dual-target CAR-T cells effectively eliminated leukemia cells expressing either antigen and avoided antigen escape associated with single-target therapy.

In addition, the researchers mimicked the antigenic heterogeneity commonly seen in T-ALL by using CRISPR/Cas9 technology to construct cell lines with varying levels of CCR9 and CD1a expression and then intermixing them into heterogeneous populations. Results showed that dual-targeted CAR-T cells were still able to effectively eliminate all subsets, while single-targeted CAR-T cells exhibited significant antigen escape. Finally, the researchers successfully generated CCR9/CD1a dual-targeted CAR-T cells from T-ALL patient samples. They demonstrated their potent cytotoxicity against autologous leukemia cells in vitro, further demonstrating the clinical feasibility of this strategy.

In summary, this study systematically validated the safety and efficacy of CCR9 as a therapeutic target for T-ALL and innovatively proposed a CCR9/CD1a dual-targeted CAR-T cell therapy strategy. This strategy successfully addresses three core challenges in T-ALL immunotherapy: antigen escape, manufacturing complexity, and immunotoxicity. The high expression rate and favorable safety profile of CCR9 make it an ideal CAR-T target, while the combined use of CD1a further expands therapeutic coverage. This dual-targeting strategy not only improves treatment efficiency but also effectively mitigates the risk of relapse due to antigen downregulation.

Reference

Tirado N, et al. CAR-T cells targeting CCR9 and CD1a for the treatment of T cell acute lymphoblastic leukemia. Journal of Hematology & Oncology, 2025, 18(1): 69.