From April 14th to 17th, 2024, the 50th Annual Meeting of the European Society for Blood and Marrow Transplantation (EBMT) was held in Glasgow, United Kingdom. The conference focused on the latest advances in stem cell transplantation and cell therapy, driving towards better clinical outcomes for patients with hematologic diseases and blood tumors. At this year’s conference, a study by Professor Hui Wang‘s team from Beijing Lu Daopei Hematology Research Institute was included for poster presentation, revealing the significance of cytokines and T cell subsets in the process of CD7 CAR-T therapy. This journal invited Professor Hui Wang to share the research findings and their clinical significance with colleagues.
Research Background and Objectives
In recent years, CD7 CAR-T therapy for T cell acute lymphoblastic leukemia (T-ALL) has received increasing attention and expectations. Therefore, exploring the changes in cytokines and T cell subsets during the CD7 CAR-T therapy process is of great significance for the prognosis of CAR-T therapy.
Research Methods
Peripheral blood samples from 70 patients who underwent CD7 CAR-T therapy at Hebei Yanda Lu Daopei Hospital from December 2020 to November 2022 were collected. Among them, there were 56 males and 14 females, aged 2 to 60 years, with a median age of 19 years. According to the CRS criteria, the 70 patients were divided into low-grade group (CRS 0-1, 56 out of 70 cases) and high-grade group (CRS 2-4, 14 out of 70 cases). Based on the evaluation results, 55 of the 70 patients were classified as complete remission group (CR/CRi), and 15 were classified as non-complete remission group.
A 2-laser 4-color Calibur flow cytometer was used to detect the expression levels of 24 cytokines (IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17A, IL-17F, IL-22, TNFα, TNFβ, IFN-γ, IL-2RA/sCD25, MIP-1α, MCP-1, GM-CSF, IL-15, REG3a, Elafin, ST-2, Granzyme B, TNFRI) with microbeads; a 3-laser 8-10-color Canto flow cytometer was used to detect the expression of CAR-T cells, target cells (CD7+ cells), and T cell subsets (CD3+, CD4+, CD8+, CD8+TCM, CD8+TEM, Treg). All changes at the cellular level were calculated using D0 value as the baseline, by comparing peak time values and baseline differences. Changes in values were observed at seven time points after CAR-T infusion: D0, D4, D7, D11, D14, D21, and D30. FCM data were analyzed using Diva software. Statistical analysis was performed using Prism 8.
Research Results
According to patient characteristics, the median age and transfection efficiency of the four groups of patients were similar.
Firstly, according to the CRS level criteria, patients were divided into low-grade and high-grade groups (Table 1). The high-grade group showed a significantly delayed peak time of CAR-T compared to the low-grade group (P=0.0274). Additionally, at the peak time point, the high-grade group exhibited a significantly higher expansion of CD8+TEM (effector memory T) cells compared to the low-grade group (P=0.0258). The clearance rate of target cells (CD7+) in the low-grade group was significantly higher than that in the high-grade CRS group (P=0.003).
Based on the levels of cytokine secretion, in the high-grade group, IL-4 (P=0.0183), IL-6 (P=0.0001), IL-8 (P=0.043), IL-10 (P=0.0001), IL-22 (P=0.0463), IFN-γ (P=0.0001), sCD25 (P=0.0001), MCP-1 (P=0.0092), IL-15 (P=0.0029), and Granzyme B (P=0.0027) levels were all significantly elevated (see Figure 1).
The median peak time results for these 10 cytokines showed that in the high-grade group, these cytokines reached their peak levels between the 10th and 15th day after CAR-T infusion. Regarding the time interval between CAR-T expansion and the secretion peak of these 10 cytokines, MCP-1 had the longest interval, with MCP-1 reaching its peak 9 days before CAR-T cells. The median peak time interval between cytokine levels and CAR-T cell expansion was 6 days .Top of Form
Furthermore, based on the criteria for evaluating CD7 CAR-T therapy results, the 70 patients were divided into complete remission and non-complete remission groups. There were no statistically significant differences in CAR-T expansion or T cell subset variation between the two groups. Only the clearance rate of target cells (CD7+) was significantly higher in the complete remission group compared to the non-complete remission group (Figure 2). There were no significant differences observed in the levels of the 24 cytokines measured in this study between the two groups.
Research Conclusion:
The data from this study indicate that IL-4, IL-6, IL-8, IL-10, IL-15, IL-22, IFN-γ, sCD25, MCP-1, and Granzyme B are associated with the severity of CRS following CD7 CAR-T therapy. These 10 cytokines are positively correlated with CRS levels, and their peak levels precede CAR-T expansion, suggesting that they can serve as indicators for predicting the occurrence and severity of CRS. The peak time of CD7 CAR-T cell expansion coincides with the expansion of CD8+TEM cells and CRS. For all patients achieving complete remission or CRi, the CD7+ clearance rate is higher and statistically significant. Our results demonstrate that cytokine levels and T cell subsets are potential indicators for predicting and assessing the prognosis of CRS.
Researcher’s Statement:
With the support of recent clinical trial data and advancements in cell production technologies, CAR-T cell therapy has shown remarkable results not only in treating CD19+ B-ALL but also in various diseases. Currently, CAR-T therapy targeting CD7+ T-ALL has achieved promising results. Furthermore, with improvements in manufacturing processes and accumulated clinical experience, side effects such as CRS induced by CAR-T therapy have been effectively controlled. However, according to our observations, different CAR-T therapies targeting different antigens may have differences in the timing and severity of CAR-T expansion, T cell subset changes, and CRS occurrence during treatment. Therefore, this study analyzed and summarized the above points and obtained some preliminary correlations. More comprehensive and accurate results still need further validation with more clinical data.
Expert Profile
Hui Wang
Researcher, Lu Daopei Medical (Yanda Lu Daopei Hospital, Beijing Lu Daopei Hospital)
Deputy Dean, Lu Daopei Institute of Hematology
Deputy Director (Deputy Dean level), Department of Laboratory Medicine, Lu Daopei Hospital
Director, SINAK CAP Certified Laboratory
Main Academic Positions
Chairman of the Expert Committee on Flow Cytometry Diagnosis of the Chinese Association of Integrated Traditional Chinese and Western Medicine Laboratory Medicine Branch
Vice President, Beijing Medical Laboratory Association
Deputy Director, Flow Cytometry and Cell Identification Quality Management Group, Medical Laboratory Quality Management Committee of China Medical Quality Management Association
Standing Committee Member, Hematology Committee, China Association of Non-Public Medical Institutions
Standing Committee Member, Cell Analysis Special Committee, China Biological Engineering Society
Standing Committee Member, China Clinical Flow Cytometry Alliance
Editorial Board Member, Chinese Journal of Laboratory Medicine
Committee Member, Clinical Hematology and Body Fluid Group, 11th Committee of Laboratory Medicine Branch, Chinese Medical Association
Committee Member, Laboratory Medicine Branch, Chinese Association of Integrated Traditional Chinese and Western Medicine
Committee Member, Lymphoma Group, China Anti-Cancer Association
Main Research Directions
Clinical diagnosis of flow cytometry
First inventor of 8 national invention patents, 1 international invention patent in the United States, 8 Beijing new technology and new product certificates, 3 utility model patents, 8 design patents, 3 works registrations
First author and corresponding author of 4 expert consensus documents
Published more than 40 SCI and national core journal articles as first author and corresponding author
Dongchu Wang: Master of Science, Assistant Researcher. Committee Member of the Flow Cytometric Diagnosis Expert Committee of the Chinese Association of Integrated Traditional Chinese and Western Medicine, Youth Committee Member of the Beijing Association of Integrated Traditional Chinese and Western Medicine.
