CD19 CAR-T expressing PD-1/CD28 chimeric switch receptor as a salvage therapy for DLBCL patients treated with different CD19-directed CAR T-cell therapies

CD19-targeted chimeric antigen receptor T (CAR T) cell therapy is a promising option to treat relapsed/refractory diffuse large B-cell lymphoma (R/R DLBCL). However, the majority of CAR T-treated patients will eventually progress and require salvage treatment, for which there is no current standard. In this study, we analyzed data from 6 patients with R/R DLBCL who experienced progression following CD19-CAR T therapy, and then received CD19-specific CAR T cells that express a PD-1/CD28 chimeric switch-receptor (CD19-PD-1/CD28-CAR T) as salvage therapy at our institution. After the second infusion of CAR T cells, 3 of 6 patients achieved complete remissions and the duration of the response of responsive patients ranged from 8 to 25 months. One patient showed a stable disease. In contrast, 2/6 patients died on 60 days because of progression disease. Importantly, no severe neurologic toxicity or cytokine release syndrome was observed. These data suggest that CD19-PD-1/CD28-CAR-T cells, a novel anti-CD19 CAR-T cell therapy, elicit a potent and durable anticancer response, and can be used in the post-CD19-CAR T failure setting. Supplementary Information The online version contains supplementary material available at 10.1186/s13045-021-01044-y.


To the Editor:
CD19-specific CAR T cell therapy has significantly improved the outcome of patients with R/R DLBCL, resulting in durable remissions in approximately 40% of heavily pretreated patients. Despite these encouraging results, nearly half of the patients could not achieve durable response after CD19-CAR T therapy and a significant proportion of patients will eventually relapse and develop treatment-refractory, fatal disease [1][2][3][4]. Recently, Spiegel et al. [5] reported on outcomes of large B-cell lymphoma patients who experienced progression following CD19-CAR T (Axicabtagene ciloleucel, axicel). The results showed that median overall survival (OS) from date of progression disease was 180 days (95% CI 105-242). Until now, there is no recommended therapeutic schedule for this fatal disease.
Six patients who relapsed or were refractory to CD19-CAR T therapy have been treated with CD19-PD-1/ CD28-CAR-T, a novel anti-CD19 CAR-T cell therapy [6], on compassionate-use basis at our institution between January 2018 and August 2019. This retrospective study was approved by the Ethics Committee of the Second Open Access *Correspondence: lab7182@tongji.edu.cn; ken.young@duke.edu; qianwb@zju.edu.cn Affiliated Hospital, Zhejiang University (Hangzhou, China) and conducted in accordance with the principles of the Declaration of Helsinki. As of October 1, 2020, the median follow-up time was 14 months. Six consecutive patients were enrolled. Patients ranged from 47 to 63 years of age and had received prior CD19-CAR T therapy with CD28-based or 4-1BB-based CAR T cells. Three of the six patients were refractory to the first CAR T treatment, and two patients had partial remission (PR) with response duration of 3 and 4 months, respectively.
One patient had a CR duration of 30 months before relapse (Table 1 and Fig. 1a). CD19-PD-1/CD28-CAR-T cells manufactured from the leukapheresed or cryopreservated autologous peripheral blood mononuclear cells were successful for all 6 patients. After failure of first CAR T therapy, patients received conditioning chemotherapy containing cyclophosphamide 500 mg/m 2 and fludarabine 30 mg/m 2 daily on days -5 to -3, and followed by a single intravenous infusion of CD19-PD-1/ CD28-CAR-T as a salvage treatment for refractory or relapsed disease. The therapeutic doses of CAR T range from 0.32 × 10 6 to 4 × 10 6 /kg of body weight. A total of 90 related adverse events occurring within 30 days of CAR-T infusion were recorded between grade 1 and 4 (Additional file 1: Table S1). Overall, 3/6 patients experienced grade 1 CRS, Patient 2 experienced grade 2 CRS, and Patient 4 and Patient 6 had both grade 2 CRS and grade 3 ICANS (immune effector cell-associated neurotoxicity syndrome). Serum cytokine levels were detected in all patients during the first month following second CAR-T therapy (Additional file 2: Figure S1). IL-6, IL-4, IL-2, and TNFα were elevated in Patient 1. Patient 4 who experienced both grade 2 CRS and grade 3 ICANS exhibited increased levels of IL-6, IL-4, IL-2, IL-17A, and IFNγ, but such increased levels were not observed in Patient 6. Four cases of CRS resolved fully by supportive treatment while the 2 patients suffered from both CRS and ICANS resolved completely after treatment with supportive care, tocilizumab and glucocorticoids. The response was evaluated with FDG-PET-CT at 3 months after infusion (Fig. 1b), according to the International Working Group Response Criteria for Malignant Lymphoma. As shown in Table 1 and Fig. 1, three of six patients achieved a CR, and one patient showed stable disease. In contrast, 2/6 patients died on 60 days because of progression disease. Two of three patients achieving CR maintained ongoing response on the date for the last visit. But, another one relapsed within 8 months and eventually died 12 months after CAR T treatment. The presence of CAR-T cells in patients' blood was monitored by qPCR. The number of blood CAR + cells peaked within 2 weeks after infusion. However, peak blood CAR + cell numbers did not differ significantly between patients with response and those without response. Interestingly, CAR + cell numbers dropped rapidly after peaking but increased significantly by day 540 after CAR T treatment in Patient 3 who achieved durable remission with long, treatment-free interval (Fig. 1c). Treatment options are limited for DLBCL patients when disease fails to respond to or relapses after CD19 CAR-T cell therapy. Novel therapies with different mechanisms of actions are critical to improve unmet clinical needs and the outcome of these patients. Alternative CD19-specific CAR-T product may be an active salvage therapy although no clinical trials have defined the optimal approach. A recent study evaluated the efficacy of axi-cel, a CD28 costimulatory-based CD19-CAR T, as salvage therapy after failure of tisagenlecleucel or investigational CD19 CAR-T with 4-1BB costimulation in patients with DLBCL. One of three patients achieved CR, but died 180 days after axi-cel treatment because of progressive disease while two of three patients were refractory [7]. In another clinical study, second infusion of CD19-CAR T was used as salvage treatment after failure of first CD19 CAR T-cell therapy. Of 21 Non-Hodgkin lymphoma (NHL) patients, the overall response rate after the second CAR T was 52% (CR, n = 4; PR, n = 7) [8]. In the present study, 3 of 6 patients achieved CRs, 2 of 3 CRs are ongoing, suggesting that CD19-PD-1/CD28-CAR-T elicit a potent and durable anticancer response, and can be used in the post-CD19 CAR T failure setting. However, we did not find an association between the costimulatory domain of CAR T and disease control (Table 1).
CAR T cell efficacy can be enhanced by using engineering strategies to address the challenge relating to T cell exhaustion induced by PD-1/PD-L1 pathway [9][10][11]. Until now, few cases report rare lymphoma patients who could obtain better efficacy by a combination of CAR-T cell therapy and PD-1 blockade [10,12]. Recently, we have reported that CD19-PD-1/CD28-CAR T cells exhibited a superior capability of killing PD-L1 + B-cell lymphoma cells in vitro and in vivo relative to the prototype, CD19-CAR T cells. We also demonstrated that this therapy had a favorable safety profile and induced durable clinical responses in the patients with PD-L1 + R/R DLBCL [6]. An interesting aspect of the current study was that CD19-PD-1/CD28-CAR T was generally well-tolerated and resulted in a high response rate that was durable in R/R large B-cell lymphoma after failure of CD19-CAR T therapy. In conclusion, our data demonstrate the ability to augment CAR T cells targeting CD19 + lymphoma by co-expressing a chimeric PD-1/CD28 switch-receptor, and that this therapy has potential as a salvage treatment when first CAR T proves ineffective or resistant.
Additional file 2: Figure S1. Kinetics of serum cytokines. A-H. Fold change of listed serum cytokines obtained from patients at the indicated times was calculated relative to the baseline.