Chimeric Antigen Receptor T Therapy Research Articles

Change in Neurocognitive Function in Patients Who Receive CAR-T Cell Therapies: A Steep Hill to Climb.

Immunotherapy with chimeric antigen receptor T (CAR-T) cell therapies has brought substantial improvement in clinical outcomes in patients with relapsed/refractory B cell neoplasms. However, complications such as cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS) limit the therapeutic efficacy of this treatment approach. ICANS can have a broad range of clinical manifestations, while various scoring systems have been developed for its grading. Cognitive decline is prevalent in CAR-T therapy recipients including impaired attention, difficulty in item naming, and writing, agraphia, and executive dysfunction. In this review, we aim to present the diagnostic methods and tests that have been used for the recognition of cognitive impairment in these patients. Moreover, up-to-date data about the duration of cognitive impairment symptoms after the infusion are presented. More research on the risk factors, pathogenesis, preventive measures, and therapy of neurocognitive impairment is crucial for better outcomes for our patients.

Abstract 4008: DASH CAR-T: accelerating CD276-targeted immunotherapy for pancreatic cancer

Abstract B7 homolog 3 protein (B7-H3), recognized as CD276, is a pivotal immune checkpoint molecule extensively expressed in cancer cells, presenting a limited presence in normal tissues. Immunohistochemical analysis of 150 pancreatic cancer patients revealed positive CD276 staining in 66% of cases, with elevated expression correlating to diminished disease-free survival. Additional findings from an independent study on 59 patients highlighted significantly elevated CD276 levels, particularly in lymph node metastasis and advanced pathology. This study introduces an innovative Chimeric Antigen Receptor T (CAR-T) therapy targeting CD276, leveraging our DASH CAR-T manufacturing platform. Significantly, our approach reduces the ex vivo cell expansion time from 10 days to 1-2 days. T cells were enriched, activated, and transduced with a CD276 CAR-encoding retroviral vector, achieving the final product in 2-3 days. In preclinical models, our shorten-time CAR-T against CD276 exhibited superior expansion and heightened tumor-killing efficacy. Remarkably, DASH CAR-T demonstrated superior tumor control compared to conventional CAR-T, requiring only one-tenth of the dosage. This research advances our understanding of CD276 in pancreatic cancer and presents a transformative approach in CAR-T therapy manufacturing. The shortened production timeline and enhanced therapeutic outcomes underscore the potential clinical impact of our novel CAR-T strategy, offering a more efficient and potent treatment option for pancreatic cancer. Citation Format: Haiying Wang, Tian Deng, Hao Li, Yue Yao, Yuan Tian, Hongwei Wang, Pengfei Jiang. DASH CAR-T: accelerating CD276-targeted immunotherapy for pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4008.

Multi-centers experience using therapeutic plasma exchange for corticosteroid/tocilizumab-refractory cytokine release syndrome following CAR-T therapy

The chimeric antigen receptor T (CAR-T) cell therapy significantly enhances the prognosis of various hematologic malignancies; however, the systemic expansion of CAR-T cells also gives rise to severe cytokine release syndrome (CRS), and immune effector cell-associated neurotoxicity syndrome (ICANS). Despite the successful application of corticosteroids and tocilizumab in alleviating severe CRS in most patients, there are still individuals who experience life-threatening CRS without responding to the aforementioned therapies. In our retrospective cohort, we conducted an analysis of clinical and laboratory parameters, including inflammatory cytokines, in 17 patients from three centers who underwent therapeutic plasma exchange (TPE) for refractory CRS with or without ICANS following CAR-T products treatment. Our findings demonstrate a significant improvement in both clinical symptoms and laboratory parameters subsequent to TPE treatment. The rapid decrease in temperature and levels of inflammatory indexes indicates the remarkable scavenging efficacy of TPE against cytokine storm following CAR-T therapy. In conclusion, TPE may serve as a valuable and safe adjunct to corticosteroids and tocilizumab in the management of severe CRS resulting from CAR-T cell infusion. We eagerly await further prospective studies to validate this finding.

Targeting autophagy overcomes cancer-intrinsic resistance to CAR-T immunotherapy in B-cell malignancies.

Chimeric antigen receptor T (CAR-T) therapy has substantially revolutionized the clinical outcomes of patients with hematologic malignancies, but the cancer-intrinsic mechanisms underlying resistance to CAR-T cells remain yet to be fully understood. This study aims to explore the molecular determinants of cancer cell sensitivity to CAR-T cell-mediated killing and to provide a better understanding of the underlying mechanisms and potential modulation to improve clinical efficacy. The human whole-genome CRISPR/Cas9-based knockout screening was conducted to identify key genes that enable cancer cells to evade CD19 CAR-T-cell-mediated killing. The in vitro cytotoxicity assays and evaluation of tumor tissue and bone marrow specimens were further conducted to confirm the role of the key genes in cancer cell susceptibility to CAR-T cells. In addition, the specific mechanisms influencing CAR-T cell-mediated cancer clearance were elucidated in mouse and cellular models. The CRISPR/Cas9-based knockout screening showed that the enrichment of autophagy-related genes (ATG3, BECN1, and RB1CC1) provided protection of cancer cells from CD19 CAR-T cell-mediated cytotoxicity. These findings were further validated by in vitro cytotoxicity assays in cells with genetic and pharmacological inhibition of autophagy. Notably, higher expression of the three autophagy-related proteins in tumor samples was correlated with poorer responsiveness and worse survival in patients with relapsed/refractory B-cell lymphoma after CD19 CAR-T therapy. Bulk RNA sequencing analysis of bone marrow samples from B-cell leukemia patients also suggested the clinical relevance of autophagy to the therapeutic response and relapse after CD19 CAR-T cell therapy. Pharmacological inhibition of autophagy and knockout of RB1CC1 could dramatically sensitize tumor cells to CD19 CAR-T cell-mediated killing in mouse models of both B-cell leukemia and lymphoma. Moreover, our study revealed that cancer-intrinsic autophagy mediates evasion of CAR-T cells via the TNF-α-TNFR1 axis-mediated apoptosis and STAT1/IRF1-induced chemokine signaling activation. These findings confirm that autophagy signaling in B-cell malignancies is essential for the effective cytotoxic function of CAR-T cells and thereby pave the way for the development of autophagy-targeting strategies to improve the clinical efficacy of CAR-T cell immunotherapy.

Development and Validation of a Prediction Model of Outcome After B-Cell Maturation Antigen-Directed Chimeric Antigen Receptor T-Cell Therapy in Relapsed/Refractory Multiple Myeloma.

Although chimeric antigen receptor T therapy (CAR-T) cells are an established therapy for relapsed/refractory multiple myeloma (RRMM), there are no established models predicting outcome to identify patients who may benefit the most from CAR-T. This is an international retrospective observational study including patients with RRMM infused with currently available commercial or academically produced anti-B-cell maturation antigen (BCMA) CAR-T. We describe characteristics and outcomes in Europe (n = 136) and the United States (n = 133). Independent predictors of relapse/progression built a simple prediction model (Myeloma CAR-T Relapse [MyCARe] model) in the training cohort (Europe), which was externally validated (US cohort) and tested within patient- and treatment-specific subgroups. The overall response rate was 87% and comparable between both cohorts, and complete responses were seen in 48% (Europe) and 49% (the United States). The median time to relapse was 5 months, and early relapse <5 months from infusion showed poor survival across cohorts, with the 12-month overall survival of 30% (Europe) and 14% (the United States). The presence of extramedullary disease or plasma cell leukemia, lenalidomide-refractoriness, high-risk cytogenetics, and increased ferritin at the time of lymphodepletion were independent predictors of early relapse or progression. Each factor received one point, forming the three-tiered MyCARe model: scores 0-1 (low risk), scores 2-3 (intermediate risk), and a score of 4 (high risk). The MyCARe model was significantly associated with distinct 5-month incidence of relapse/progression (P < .001): 7% for low-risk, 27% for intermediate-risk, and 53% for high-risk groups. The model was validated in the US cohort and maintained prognostic utility for response, survival, and outcomes across subgroups. Outcomes of patients with RRMM after CAR-T are comparable between Europe and the United States. The MyCARe model may facilitate optimal timing of CAR-T cells in patient-specific subgroups.

An alternative fully human anti-BCMA CAR-T shows response for relapsed or refractory multiple myeloma with anti-BCMA CAR-T exposures previously

Chimeric antigen receptor T (CAR-T) cells therapy has made remarkable progress in relapsed/refractory multiple myeloma (R/R MM) treatment. Unfortunately, patients still eventually experience disease progression or relapse even after receiving anti-BCMA CAR-T therapy. At present, there are limited data on available treatment options for patients who have progressed on anti-BCMA CAR-T therapy. In this study, we evaluated the safety and efficacy of fully human anti-BCMA CAR-T (HRC0202) in seven R/R MM patients who were previously exposed to anti-BCMA CAR-T therapy. Three patients received 6.0 × 106 CAR+T cells/kg, one patient received 10.0 × 106 CAR+T cells/kg and three patients received 15.0 × 106 CAR+T cells/kg. Cytokine release syndrome (CRS) of grades 1–2 occurred in three patients (42.9%) and grade ≥3 in two patients (28.6%). Immune effector cell-associated neurotoxic syndrome (ICANS) was not observed in any of the patients. The best overall response rate (ORR) was 71.4% (5/7), with a stringent complete response/complete response (sCR/CR) achieved in three patients. The median progression-free survival (PFS) was 269 days, and median overall survival (OS) for all patients was not reached. The median peak concentration (Cmax) of HRC0202 was 30117.70 (range, 6084.35–147415.10) copies/μg DNA. This study indicated that fully human anti-BCMA CAR-T (HRC0202) is a promising treatment for R/R MM patients who relapsed or refractory from prior anti-BCMA CAR-T infusion.

Efficacy and Safety of the Second CAR-T Therapy in Patients with Refractory/Relapsed Acute B-Cell Lymphoblastic Leukemia

Background: Chimeric antigen receptor T (CAR-T) cells therapy has shown significant efficacy in relapsed or refractory B-cell acute lymphoblastic leukemia (R/R B-ALL), but a substantial proportion of patients still relapse after a period of remission. The efficacy of a second CAR-T cell infusion is uncertain. The aim of this study was to explore the efficacy and safety of the second CAR-T therapy in R/R B-ALL patients. Methods: Between August 2018 and October 2022, 35 patients with R/R B-ALL successfully received the second CAR-T treatment. Among them, 18 patients received single CD19 or CD22 CAR-T therapy, 14 patients received tandem CD19/CD22 CAR-T therapy, and 3 patients received sequential CD19 and CD22 CAR-T therapy. Results: The complete remission (CR) rates were 55.6% (10/18) in patients who received CD19 or CD22 CAR-T therapy, 85.7% (12/14) in patients who received tandem CD19/CD22 CAR-T therapy, and 33.3% (1/3) in patients who received sequential CD19 and CD22 CAR-T therapy (tandem CD19/CD22 vs. others, P=0.045). Tandem CD19/CD22 CAR-T therapy remained one of the significant favorable factors in multivariate logistic regression analysis of CR rate in all patients (hazard ratio, 0.128; 95% CI, 0.017-0.974). A total of 5 patients with severe cytokine release syndrome (Grade ≥ 3) were observed, including 2 patients in the single CD19 CAR-T group, 2 patients in the tandem CD19/CD22 CAR-T group, and 1 patient in the sequential CD19 and CD22 CAR-T group. Of the 23 patients who achieved CR, 4 patients bridged to allogeneic hematopoietic stem cell transplantation (allo-HSCT) and 3 patients received decitabine (DAC) consolidation.With the median follow-up of 25.5 months (range, 1.1 to 33.5), the 2-year overall survival, leukemia-free survival (LFS) and cumulative incidence of relapse rates were 23.7%, 30.5% and 69.5%, respectively. Multivariate Cox regression analyses showed that a better LFS related to the absence of high-risk cytogenetics and genetic characteristics, DAC combination with fludarabine and cyclophosphamide lymphodepletion regimen, and bridging allo-HSCT or DAC consolidation. Conclusions: Our study showed that the second infusion of tandem CD19/CD22 CAR-T therapy obtains a better response than other infusion strategies. Bridging allo-HSCT or DAC consolidation had a significant survival benefit in patients with CR after the second CAR-T therapy.

Multiplex Genome Editing of Human T Cells with Innovative Transformer Base Editor (tBE) for Construction of Next Generation CAR-T Therapies

Autologous chimeric antigen receptor T (CAR-T) cell therapy has achieved great success for antitumor treatments especially against hematological malignancies, and several autologous CAR-T therapies have been approved around the world. However, high manufacturing cost and long manufacturing process hinder the broader application of autologous CAR-T cell therapy. As a result, off-the-shelf universal CAR-T cell therapy with donor derived healthy T cells is of great interest and under development by many companies globally. Gene editing technologies including TALENs, CRISPR/Cas nucleases and base editors are of great potential in T cell engineering for allogeneic T cell therapy. The innovative transformer base editor (tBE) is a base editing system that avoids to cause DNA double strand breaks (DSBs) and exhibits higher editing efficiency, no detected off-target mutations and lower cytotoxicity. Due to its capability to avoid off-target mutations and DSB-caused chromosomal translocations, tBE is potentially one of the best gene editing tools to induce multiplex editing in cells. Here we show that the tBE was used to induce highly efficient single or multiplex gene editing in primary human T cells. With optimized electroporation conditions for tBE, TRAC, CD52 and PDCD1 could be knocked out simultaneously with editing efficiency up to ~90% for each target gene. tBE triggered no detected off-target mutation or chromosomal abnormalities, which were reported in the cases of CRISPR/Cas- or TALEN-mediated multiplex editing. CD19-targeted CAR-T cells with triplex base editing exhibits enhanced antitumor efficacy in vitro and in mouse models compared with unedited CAR-T cells. Our results demonstrated the application of tBE for construction of next generation universal or enhanced CAR-T therapies.

Determinants of Response to Anti CD-19 CAR-T Cells for Diffuse Large B-Cell Lymphoma in Pre-Treatment Peripheral Blood Mononuclear Cells Using Single-Cell RNA-Seq

Background: Chimeric antigen receptor T (CAR-T) cell therapy has become the standard of care in patients with relapsed/refractory diffuse large cell lymphoma (R/R DLBCL), providing durable remission in 40% of patients. Lack of response is likely to be attributed to inter- and intra-patient variability, including PBMC heterogeneity and apheresis material composition, CAR-T infusion product, in-vivo expansion and activity of CAR-T cells and the development of tumor-intrinsic mechanisms. Current methods for evaluating CAR-T cell activation and potency have limited predictive value for clinical outcomes. To address this challenge, we performed single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) obtained from patients with R/R LBCL patients at the time of apheresis for the manufacturing of anti-CD19 CAR-T cell therapy. Methods: Between 10/2020 and 03/2022, 31 patients who underwent lymphopheresis were enrolled. Samples were obtained - 1) At day of lymphopheresis from peripheral blood; 2) On the day of infusion of CAR-T from the product; and 3) On day +7 after infusion from peripheral blood. All patients had day +30 PET-CT. In addition, PBMCs samples from healthy donors were used as control to analyze baseline characteristics compared to the DLBCL patients. PBMCs from patients of healthy donors were purified from frozen blood samples by density gradient separation, stained using human CD45 antibody and loaded onto a 10x Chromium system. libraries were generated using the 10X Genomics Chromium Single Cell 3' Kit. The data was processed and analyzed using the Seurat R package. The study was approved by the local Ethic committee. Results: Patient characteristics: 31 RR DLBCL patients (42% females; median age 72, range 47-81 years), all treated with CAR-T cells therapy as third line of therapy, participated in the study. All patients underwent lymphopheresis, followed by CAR-T production. Disease status at lymphodepletion was complete remission (CR, n=4, 13%), partial remission (n=6, 19%), and progressive disease (n=21, 68%) and patients received either tisagenlecleucel (n=20, 65%) or axicabtagene ciloleucel (n=11, 35%) . PET scan performed on day 30 post treatment revealed that 68% and 32% obtained CR, and PD, respectively. scRNA analysis revealed differences in PBMC composition between healthy donors and LBCL patients (Figure 1A), with lower levels of naïve and memory T cells and elevated levels of activated monocytes in LBCL patients, suggesting a shift towards an inflamed and pre-dysfunctional immune state. Categorization of LBCL patients into responders (PR/CR on day +30 PET-CT) and non-responders (PD on day +30 PET-CT) revealed differences in immune cell subsets between responders and non-responders (Figure 2A), with enriched inflammation-related pathways in non-responders, In contrast, responders exhibited a distinct healthy-like immune profiles, particularly high levels of naive T cells, similar to that obtained in their age matched healthy controls. Of note, non-responders showed alterations in immune cell subsets, including increased levels of T regulatory cells and effector NK cells. Furthermore, non-responders demonstrated a prominent inflammatory immune profile, characterized by gene pathways related to inflammation and activation, such as type I and III IFN-G, and TNFb response (Figure 1B). Notably, an IFN-G score was significantly higher in non-responder's monocytes subpopulations, compared to responders. Conclusions: These findings indicate that the success of CAR-T therapy could be determined by the molecular characteristics and subpopulation distribution of each patient's PBMC. The diverse immune profiles of DLBCL patients might impact the effectiveness of their immune cells in generating successful CAR-T products or providing a conducive environment for CAR T cells to combat cancer. Furthermore, these findings emphasize the potential of scRNA-seq in evaluating PBMC from CAR-T patients prior to treatment, allowing the potential calculation of an “immune potency score” associated with the probability of a positive outcome from CAR-T treatment of DLBCL patients. This score could potentially be used by oncologists in decision-making processes to determine the appropriate candidates for CAR-T therapy.

Prediction Modeling of CAR-T Cell Therapy for Diffuse Large B-Cell Lymphoma Using Artificial Neural Networks on Tumor Vascular Phenotype

Chimeric antigen receptor T (CAR-T) cell therapy targeting CD19 only results in complete remission in approximately 30-40% of relapsed/refractory disease patients with diffuse large B-cell lymphoma (DLBCL). Failure may result from lack of CAR-T entry into the tumor. Endogenous T cell homing is initiated by binding to post-capillary venular endothelial cells (ECs). Naïve and central memory T cells use L-selectin binding to peripheral node addressin (PNad) on high endothelial venules (HEVs) of the lymph node while circulating effector memory T cells, lacking L-selectin, use other molecules to bind cytokine-induced endothelial proteins E-selectin, ICAM-1 and VCAM-1 on flat venules of inflamed tissues. Both of these venular EC types are present in inflamed lymph nodes. We hypothesized that the expression patterns of adhesion molecules on venular ECs in pre-treatment tumor biopsies could predict CAR-T efficacy. To test this, we applied cyclic immunofluorescence to 18 archived pre-treatment DLBCL patient FFPE biopsies (8 responders and 10 non-responders, defined by relapse at 12 months). We detected venular ECs using a combination of anti-CD31 and UEA lectin on larger diameter vessels and assessed expression for PNad, E-selectin, ICAM-1, VCAM-1. We detected endogenous CD8+ T cells and evaluated their expression of L-selectin, alpha 4 integrin and PSGL-1. See table 1. DAPI nuclear staining was included in each cycle. Images of each cycle were combined in ImageJ and imported into QuPath for analysis. Using Warpy, affine transformations were applied to DAPI staining to create overlays between imaging cycles. After alignment, representative images of several samples were randomly chosen for image training using a combination of Artificial Neural Networks (ANNs) and classifiers to perform identification and mask tissue regions of vessels and T-cells within the tissue. Positive and negative cells for each marker of interest (EC PNad, ICAM-1, E-Selectin, VCAM-1, and T cell alpha 4 integrin, L-Selectin, and PSGL-1) were used to create classification algorithms to determine marker expression for ECs and T-cells within the tissue. Results were analyzed by unpaired t-test and one-way ANOVA conducted in Graph-Pad Prism 9 software. Preliminary results from quantitative analysis of pre-treatment biopsies show a significant increase in CD31+/PNAD- venular ECs compared to non-responders that were double positive for ICAM1 and VCAM-1 (p=0.0273). This analysis also revealed a significant increase in CD31+/PNAD+ HEV ECs from responders that were triple positive for E-Selectin, ICAM-1, and VCAM-1 (p=0.0242). See figure 1. Our initial semi-quantitative analysis showed a trend toward elevated numbers of tissue-infiltrating CD3 + CD8 + T cells in the pre-treatment biopsies of responders. T cells expressing PSGL-1, a glycoprotein that may carry carbohydrate moieties recognized by E-Selectin and linked to T-cell exhaustion, was increased in the non-responder cohort. These data suggest an increased extent and activation state of intra-tumoral venular ECs and increased endogenous T-cell infiltrates, which may predict the outcome of CAR-T therapy. Analysis of T-cells in the pre-treatment biopsies leveraging machine learning algorithms is in progress and both complete quantitative findings and prognostic value of this approach will be presented. While our findings require further validation thru additional biopsy analysis, they provide emerging evidence for a rationale involving vascular readiness in DLBCL responder differences to CART therapy.

A Multicenter Study of CAR-T Cell Therapy for Relapse of B-ALL Post Allogeneic Stem Cell Transplantation

BACKGROUND: Patients with acute B-cell lymphoblastic leukemia (B-ALL) relapsed after allogeneic stem cell transplantation (allo-HSCT) have very poor prognosis. Chimeric antigen receptor T (CAR-T) cells targeting CD19 have been developed as one of the effective immunotherapies for relapsed/refractory B-ALL. The safety and efficacy of CAR-T cells in patients relapsed after allo-HSCT remain unclear. METHODS: This study analyzed 60 B-ALL patients relapsed after allo-HSCT treated with CAR-T cells between October 2015 and February 2022 from four Chinese centers. Their pre-treatment patient characteristics, treatment regimens, efficacy, adverse effects, and follow-up data were comprehensively analyzed. RESULTS: One patient refused efficacy evaluation. 7 patients died before day 28 after CAR-T infusion. A total of 52 were assessed for efficacy within 28 days after CAR-T infusion.76.92% (40/52) patients achieved complete remission (CR) at Day 28 after CAR-T infusion, including 38 patients who achieved MRD-negative CR. 4 of the 40 patients relapsed at a median follow-up date of 12.1 months. The 1-year OS and EFS of all patients were 52.8% ± 7.0% and 51.5% ± 7.0%, respectively, and the 2-year OS and EFS were 40.1% ± 8.6% and 45.1% ± 8.6%, respectively. Multifactorial analysis showed that patients with severe CRS had worse OS and EFS, and other factors including age, gender, site of CAR-T tumor involvement, type of previous transplantation, previous transplantation type, CAR-T product and CRS grade, etc., did not affect the long-term survival of patients. The incidence of CRS was 80.00%, the incidence of grade ≥3 CRS was 20.83%, and the median time of CRS was 2 days after CAR-T cell infusion (range: 0-34 days). 3 patients (5.00%) died due to multiorgan failure caused by severe CRS, and the rest of the patients' CRS was controlled, and the grade of CRS was related to the proportion of blast cells in the bone marrow prior to CAR-T cell infusion., independent of the type of transplantation and the source of CAR-T cells. The incidence of ICANS was 5.00%. The incidence of aGvHD was 30.00%, and patients who had received prior haploidentical allogeneic HSCT were significantly more likely to develop aGvHD after CAR-T cell infusion than those who had received prior matched allogeneic HSCT. The occurrence of aGvHD after CAR-T treatment was not associated with the occurrence of CRS (p = 1), the presence of aGvHD after prior transplantation (p = 0.3484), the presence of cGvHD after prior transplantation (p = 0.3167), the presence of cGvHD at the time of CAR-T infusion (p = 0.5136), and the presence of prior donor lymphocyte infusion (p = 0.7404), and whether CAR-T cells were from autologous or donor (p = 0.4819). CONCLUSIONS: CAR-T cells are safe and effective in treating B-ALL patients with post-transplant relapse, and donor-derived CAR-T cells do not exacerbate CRS with aGVHD, and there is a high incidence of aGVHD after CAR-T therapy in patients with HLA-haplotype-matched transplantation.

Characteristics of Central Nervous System Relapse after CD19 CAR-T Cell Therapy in R/R NHL

Background CD19-targeted chimeric antigen receptor T (CAR-T) cell therapy has shown remarkable antitumor activity against relapsed/refractory (R/R) non-Hodgkin lymphoma (NHL). However, NHL with central nervous system (CNS) relapse carries a poor prognosis and high mortality. We reported an analysis of 80 patients with NHL who underwent CAR-T cell therapy, and described the characteristics including the international prognostic index (IPI) and other clinical variables to evaluate the outcomes of CNS relapse in NHL patients treated with CAR-T cell therapy. Methods A total of 80 patients with NHL treated with CD19 CAR-T cell therapy in our center between April 2017 and November 2022 was retrospectively analyzed. Patient characteristics, laboratory parameters, and clinical outcomes were collected. CNS relapse was diagnosed by cerebrospinal fluid cytology, MRI or PET-CT. Results Baseline characteristics of all 80 patients are summarized in Table 1. The median age was 58.5 (range, 10 to 79) years, and 57.5% of patients were male. Overall, 67 (83.8%) patients achieved complete remission (CR) or partial remission (PR) after CD19 CAR-T therapy. Seven (8.43%) patients developed isolated CNS relapse after CAR-T cell therapy, all of which were heavily pre-treated with multiple lines of chemotherapy prior to CAR-T cell therapy, and one of them had received autologous stem cell transplant before infusion of CAR-T cells. Only 4 of 7 patients with isolated CNS relapse achieved CR (n=4), and the median time to isolated CNS relapse from CAR-T cell infusion was 6.3(range, 1.53 to 25.8) months. Thirty-seven patients (46.2%) developed systemic recurrence without CNS involvement, with the median time from CAR-T to relapse of 3.2 (range, 0.467-30.8) months. The other 36 patients remained in remission at the last follow-up. During the median follow-up of 13.5 months of the whole cohort, the number of recorded deaths in isolated CNS relapse, systemic only recurrence, and no relapse patients was 5 (71.4%), 22 (59.4%), 1(2.78%), respectively. The 1-year OS and 1-year PFS of patients with isolated CNS recurrence was significantly lower compared to the no relapse group (OS, 35.7% [95%CI 12.1-100] vs 97.22% [95%CI 92-100], P&amp;lt;0.001; PFS, 28.6% [95%CI 8.86-92.2] vs 100% [95%CI 100-100], P&amp;lt;0.001), but showed no difference compared to the systemic only recurrence group(OS, 53.3% [95%CI 39.3-72.4], P=0.79; PFS, 13.0% [95%CI 5.53-30.5], P=0.17). Conclusion In conclusion, patients with CNS relapse showed significant difference in 1-year outcomes with no relapse group but not systemic relapse group in our study, which suggested the importance of effective prophylaxis against CNS relapse. Further investigation is necessary to better define the characteristics of patients who will benefit from prophylaxis. Acknowledgements We gratefully thank all the participation from patients enrolled in clinical trials (ChiCTR1800015575; NCT03118180; NCT04532281; NCT04532268; NCT04213469).

Is YouTube a reliable source of information on chimeric antigen receptor T-cell therapy? An analysis.

486 Background: YouTube is an open-access source of information with more than 2.5 billion users. It plays an important role in disseminating information especially related to health, including cancer. Chimeric antigen receptor-T (CAR-T) cell therapy has significantly improved outcomes for hematologic malignancies over recent years. We aim to assess the quality and engagement parameters of the 100 most viewed videos on YouTube on CAR-T cell therapy. Methods: A systematic search was conducted on YouTube using the search terms (“CAR-T Cell therapy” AND “Cancer”). One hundred most viewed videos in English were selected. We utilized the already validated tools PEMAT and DISCERN to assess the quality of these videos on CAR-T. Additionally, we looked at the number of views, comments, and likes. Descriptive statistics and Pearson correlation tests were performed using R studio v2022.12.0-353. Results: The videos in our study had up to 3.4 million views (median number of views:19901), and the overall quality of information was fair (50%) by the DISCERN score. These videos were categorized into different sources academic (60%), media (22%), and industry (18%). The mean DISCERN score (44.27± 10.8), mean PEMAT actionability (47% ± 38%), mean PEMAT understandability (75.6% ± 15.5%). While benefits were mentioned by 67%, only 29% discussed the risks of CAR-T Cell therapy; 37% of videos promoted shared decision-making, and only 10% summarized the information at the end. There was no significant correlation between DISCERN and PEMAT scores with video engagement parameters, including likes, views, and comments. Interestingly, only 30% of videos discussed other treatment options except for CAR-T. While 88% of videos used everyday language, medical terminologies were defined by 73% of videos; 32 % of videos were broken into short sections with informative headers. Conclusions: This is the first study to our knowledge evaluating the quality of information on CAR-T therapy on YouTube. Compared to historical studies on YouTube 1, this study shows the quality of the information provided in these videos is valid and reliable. They also prove to be actionable and understandable; these videos focus on the benefits of CAR-T cell therapy and are less likely to discuss risks or shared decision-making. Contrary to previous studies2, we believe YouTube videos on CAR-T Cell therapy may serve as a supplementary source of providing information to patients wishing to learn more on this topic.

Gastrodin synergistically increases migration of interleukin-13 receptor α2 chimeric antigen receptor T cell to the brain against glioblastoma multiforme: A preclinical study.

Therapy with chimeric antigen receptor T (CAR-T) cells involves using reformative T lymphocytes that have three domains, antigen recognition, transmembrane, and costimulating to achieve the therapeutic purpose. CAR-T therapy on malignant hematologic has been successful; however, its effectiveness in patients with solid tumors is still limited. Few studies exist confirming the efficacy of natural products on the function of CAR-T cells. The purpose of this study is to assess the effect of gastrodin (GAS) on CAR-T cells that target interleukin-13 receptor α2 antigen (IL-13Rα2 CAR-T) in the brain against glioblastoma multiforme. Migration of IL-13Rα2 CAR-T was evaluated using the Transwell assay. The effects of GAS on IL-13Rα2 CAR-T cells were assessed both in vitro and situ glioblastoma models. The cytoskeleton was stained with Fluorescein 5-isothiocyanate (FITC)-phalloidin. Cytokines expression in cells was determined by flow cytometry and ELISA assay. Western blotting was used to detect the S1P1 expression, and quantitative PCR assay was used to determine the IL-13Rα2 gene level. GAS increased the migratory and destructive capacity of IL-13Rα2 CAR-T cells with no effect on cytokine release. By increasing the expression of S1P1, GAS encouraged the entry of CAR-T cells into the brain and bone marrow. Transcriptomic analysis revealed that genes related to skeletal migration such as add2 and gng8 showed increased expression in GAS-treated CAR-T cells. We found that GAS synergistically improves the mobility of IL-13Rα2 CAR-T, enhancing their ability to recognize the tumor antigen of glioblastoma, which could be advantageous for the application of CAR-T for the treatment of solid tumors.

Preconditioning of radiotherapy enhances efficacy of B7-H3-CAR-T in treating solid tumor models

AimsLimited efficacy of chimeric antigen receptor T (CAR-T) cells in treating solid tumors is largely due to the antigen heterogeneity and immunosuppressive tumor microenvironment (TME). B7-H3 is over-expressed in most kind of solid tumors, making it a promising target for cancer treatment. This study aims to explore the effect of B7-H3-CAR-T therapy combined with radiotherapy in treating solid tumor models. MethodsIrradiated tumor cell lines were prepared and tested. A humanized B7-H3-CAR-T was constructed, and it was evaluated that B7-H3-CAR-T cytotoxicity against solid tumor models with preconditioning of radiotherapy in vitro and vivo. ResultsIrradiation was found to increase expression level of B7-H3 in pancreatic cancer (PANC-1), colorectal cancer (HCT-15, SW620), acute myelocytic leukemia (AML-5), epidermoid carcinoma (KB) and glioma (U87-MG) human cell lines significantly. 6Gy irradiation was also found to up-regulate tumor-infiltration molecule like intracellular adhesion molecule-1 ICAM-1 or FAS in HCT-15 cells, supporting a possible synergistic enhancement effect of radiotherapy. In vitro and in vivo experiments demonstrated that irradiation indeed significantly enhanced the ability of B7-H3-CAR-T to infiltrate and kill tumors. Interestingly in dual-tumor mouse model study, not only tumor cells on irradiation side were eradicated completely, irradiation also enhanced CAR-T tumor-killing ability on non-irradiated side, confirming the abscopal effect of irradiation existed with CAR-T therapy. ConclusionsOur results suggest that B7-H3-CAR-T therapy combined with radiotherapy may be a promising modality in treating solid tumors.

CRISPR/Cas9: A Powerful Strategy to Improve CAR-T Cell Persistence.

As an emerging treatment strategy for malignant tumors, chimeric antigen receptor T (CAR-T) cell therapy has been widely used in clinical practice, and its efficacy has been markedly improved in the past decade. However, the clinical effect of CAR-T therapy is not so satisfying, especially in solid tumors. Even in hematologic malignancies, a proportion of patients eventually relapse after receiving CAR-T cell infusions, owing to the poor expansion and persistence of CAR-T cells. Recently, CRISPR/Cas9 technology has provided an effective approach to promoting the proliferation and persistence of CAR-T cells in the body. This technology has been utilized in CAR-T cells to generate a memory phenotype, reduce exhaustion, and screen new targets to improve the anti-tumor potential. In this review, we aim to describe the major causes limiting the persistence of CAR-T cells in patients and discuss the application of CRISPR/Cas9 in promoting CAR-T cell persistence and its anti-tumor function. Finally, we investigate clinical trials for CRISPR/Cas9-engineered CAR-T cells for the treatment of cancer.

The nutritional impact of CD19-targeted CAR-T therapy versus BEAM chemotherapy for adult patients with lymphoma.

Chimeric antigen receptor T (CAR-T) cell therapy is a novel therapy demonstrating durable remissions in patients with refractory or relapsing non-Hodgkin's B-cell lymphoma. Maintaining a patient's nutritional status has been demonstrated to improve outcomes in cancer treatment. However, no studies have investigated how CAR-T therapy affects nutritional status, nor compared its impact with other cancer treatments for this patient group. The primary aim of the present study was to investigate the effect of CAR-T therapy on the prevalence of nutrition impact symptoms (NIS) and nutritional status within 30 days post-treatment of patients with lymphoma compared to a conditioning regimen for autologous haematopoetic stem cell transplant (carmustine/BCNU, Etoposide, cytarabine/Ara-C, Melphalan [BEAM] auto-haematopoetic stem cell transplant [HSCT]). Clinical notes of patients with lymphoma who underwent either CAR-T therapy or BEAM auto-HSCT between 2018 and 2021 were reviewed. Data extracted included body weight measurements and NIS, including decreased appetite, nausea, vomiting, diarrhoea, constipation, mucositis, cytokine release syndrome (CRS) and neurotoxicity at baseline and 30 ± 7 days post-treatment. In total, 129 adults with lymphoma (n = 88 CAR-T vs. n = 41 BEAM) were included. Nutritional status was assessed in both groups at baseline prior to treatment. Mean absolute weight change was significantly different between groups (3.05 kg in CAR-T, -5.9 kg in BEAM, p ≤ 0.001). This was also significant when weight loss was categorised into percentage weight loss (p = 0.01). CAR-T patients experienced a significantly lower prevalence of decreased appetite (52.3% vs. 97.6%) nausea (25% vs. 78%,) vomiting (10.2% vs. 53.7%), diarrhoea (43.2% vs. 96.7%) and mucositis (5.7% vs. 75.6%) combined across all levels of severity compared to BEAM chemotherapy (all p ≤ 0.01). CRS and neurotoxicity, which are specific side effects of CAR-T therapy, were moderately positively associated with weight loss. Weight loss, percentage weight loss and NIS were significantly reduced in CAR-T compared to BEAM treatment. However, patients who experienced neurotoxicity during treatment did have significant weight loss.

Are we ready for personalized CAR-T therapy?

The future of chimeric antigen receptor T (CAR-T) therapy remains unclear. New studies are constantly being published confirming the efficacy and favorable safety profile of its innovative enhancements. Currently approved CAR-T drugs are manufactured exclusively for a specific patient from the recipient's own cells. This does not close the door to further modifications with subsequent personalization and better adaptation to the individual needs. Bringing such a drug to market would involve raising the already high costs, so it is necessary to lower the existing ones. On the other hand, so-called universal CAR-T are also getting closer to the patient's bed, but its implementation may struggle with multiple challenges, including development of graft-versus-host disease (GvHD) and alloimmunity. However, that off-the-shelf therapy could prove useful as a quick solution for patients in very poor condition or excluded from current therapy due to manufacturing limitations. The introduction of currently tested solutions may undoubtedly change the current paradigm of treatment.

Effects of CAR-T Cell Therapy on Immune Cells and Related Toxic Side Effect Analysis in Patients with Refractory Acute Lymphoblastic Leukemia.

To observe the effects of chimeric antigen receptor T (CAR-T) cell immunotherapy on immune cells and related toxic side effects in patients with refractory acute lymphoblastic leukemia (ALL). A retrospective study was conducted in 35 patients with refractory ALL. The patients were treated with CAR-T cell therapy in our hospital from January 2020 to January 2021. The efficacy was evaluated at one and three months post treatments. The venous blood of the patients was collected before treatment, 1 month after treatment, and 3 months after treatment. The percentage of regulatory T cells (Treg cells), natural killer (NK) cells, and T lymphocyte subsets (CD3+, CD4+, and CD8+ T cells) was detected by flow cytometry. The ratio of CD4+/CD8+ was calculated. Patient's toxic side effects such as fever, chills, gastrointestinal bleeding, nervous system symptoms, digestive system symptoms, abnormal liver function, and blood coagulation dysfunction were monitored and recorded. The incidence of toxic and side effects was calculated, and the incidence of infection was recorded. After one month of CAR-T cell therapy in 35 patients with ALL, the efficacy evaluation showed that complete response (CR) patients accounted for 68.57%, CR with incomplete hematological recovery (CRi) patients accounted for 22.86%, and partial disease (PD) patients accounted for 8.57%, and the total effective rate was 91.43%. In addition, compared with that before treatment, the Treg cell level in CR+CRi patients treated for 1 month and 3 months decreased prominently, and the NK cell level increased dramatically (P < 0.05). Compared with that before treatment, the levels of CD3+, CD4+, and CD4+/CD8+ in patients with CR+CRi in the 1-month and 3-month groups were markedly higher, and the levels of CD4+/CD8+ in the 3-month group were memorably higher than those in the 1-month group (P < 0.05). During CAR-T cell therapy in 35 patients with ALL, fever accounted for 62.86%, chills for 20.00%, gastrointestinal bleeding for 8.57%, nervous system symptoms for 14.29%, digestive system symptoms for 28.57%, abnormal liver function for 11.43%, and coagulation dysfunction for 8.57%. These side effects were all relieved after symptomatic treatment. During the course of CAR-T therapy in 35 patients with ALL, 2 patients had biliary tract infection and 13 patients had lung infection. No correlations were found between the infection and age, gender, CRS grade, usage of glucocorticoids or tocilizumab, and laboratory indicators such as WBC, ANC, PLT, and Hb (P > 0.05). CAR-T cell therapy had a good effect on patients with refractory ALL by regulating the immune function of the body via mediating the content of immune cells. CAR-T cell therapy may have therapeutic effect on refractory ALL patients with mild side effects and high safety.

SA3 Characteristics of Patients Treated with CAR-T Therapies across Multiple Indications Utilizing the TriNetX Network

Six Chimeric Antigen Receptor-T (CAR-T) therapies have been approved as treatments in relapsed/refractory (R/R) B-cell malignancies since 2017. This study aims to examine patient counts and characteristics of patients on CAR-T therapies in healthcare organizations in the United States covered by the TriNetX network, an electronic medical records (EMR) database.