Efficacy Of CAR-T Cell Therapy Research Articles

A Review of Current and Pipeline Drugs for Treatment of Melanoma.

Malignant melanoma is the most aggressive form of skin cancer. Standard treatment options include surgery, radiation therapy, systemic chemotherapy, targeted therapy, and immunotherapy. Combining these modalities often yields better responses. Surgery is suitable for localized cases, sometimes involving lymph node dissection and biopsy, to assess the spread of the disease. Radiation therapy may be sometimes used as a standalone treatment or following surgical excision. Systemic chemotherapy, while having low response rates, is utilized as part of combination treatments or when other methods fail. The development of resistance to systemic chemotherapies and associated side effects have prompted further research and clinical trials for novel approaches. In the case of advanced-stage melanoma, a comprehensive approach may be necessary, incorporating targeted therapies and immunotherapies that demonstrate significant antitumor activity. Targeted therapies, including inhibitors targeting BRAF, MEK, c-KIT, and NRAS, are designed to block the specific molecules responsible for tumor growth. These therapies show promise, particularly in patients with corresponding mutations. Combination therapy, including BRAF and MEK inhibitors, has been evidenced to improve progression-free survival; however, concerns about resistance and cutaneous toxicities highlight the need for close monitoring. Immunotherapies, leveraging tumor-infiltrating lymphocytes and CAR T cells, enhance immune responses. Lifileucel, an FDA-approved tumor-infiltrating lymphocyte therapy, has demonstrated improved response rates in advanced-stage melanoma. Ongoing trials continue to explore the efficacy of CAR T-cell therapy for advanced melanoma. Checkpoint inhibitors targeting CTLA-4 and PD-1 have enhanced outcomes. Emerging IL-2 therapies boost dendritic cells, enhancing anticancer immunity. Oncolytic virus therapy, approved for advanced melanoma, augments treatment efficacy in combination approaches. While immunotherapy has significantly advanced melanoma treatment, its success varies, prompting research into new drugs and factors influencing outcomes. This review provides insights into current melanoma treatments and recent therapeutic advances.

Enhancing the Efficacy of CAR-T Cell Therapy: A Comprehensive Exploration of Cellular Strategies and Molecular Dynamics.

The emergence of chimeric antigen receptor T cell (CAR-T cell) therapy has revolutionized cancer treatment, particularly for hematologic malignancies. This commentary discusses developments in CAR-T cell therapy, focusing on the molecular mechanisms governing T cell fate and differentiation. Transcriptional and epigenetic factors play a pivotal role in determining the specificity, effectiveness, and durability of CAR-T cell therapy. Understanding these mechanisms is crucial to improve the efficacy and decrease the adverse events associated with CAR-T cell therapies, unlocking the full potential of these approaches. T cell differentiation in CAR-T cell product manufacturing plays an important role in clinical outcomes. A positive correlation exists between the clinical efficacy of CAR-T cell therapy and signatures of memory, whereas a negative correlation has been observed with signatures of effector function or exhaustion. The effectiveness of CAR-T cell products is likely influenced by T-cell frequency and by their ability to proliferate, which is closely linked to early T cell differentiation. The differentiation process involving distinct T memory cell subsets is initiated upon antigen elimination, indicating infection resolution. In chronic infections or cancer, T cells may undergo exhaustion, marked by continuous inhibitory receptor expression, decreased cytokine production, and diminished proliferative capacity. Other cell subsets, such as CD4+ T cells, innate-like T lymphocytes, NKT cells, and cord blood-derived hematopoietic stem cells, offer unique advantages in developing the next-generation CAR-T cell-based therapies. Future research should focus on optimizing T-cell-enhancing approaches and developing strategies to potentially cure patients with hematological diseases and solid tumors.

Germline Determinants of Toxicity and Efficacy in Patients with Large B-Cell Lymphoma Treated with Anti-CD19 Autologous CAR T-Cell Therapy

Background. Increasing data have demonstrated that acquired genetic aberrations, including TP53 mutations in the tumor cells and clonal hematopoiesis, can impact the efficacy and/or toxicity of anti-CD19 autologous CAR T-cell therapy (CART) in patients with large B-cell lymphoma (LBCL). Recent data have suggested that germline genetic aberrations, including polymorphisms in CD19, ABCB1, MISP and CPVL (Strati P et al, Leukemia 2022) and deleterious germline variants in STXBP2, can also affect outcomes in these patients. However, a comprehensive analysis of germline determinants of response and toxicity after CART in this patient population has not yet been described. Methods. Genome-wide genotyping using the Illumina Genome Screening Array (GSAv2.0) was performed in 170 patients with relapsed or refractory LBCL treated with standard of care axicabtagene ciloleucel in third line and beyond. Response was assessed according to 2014 Lugano criteria. Cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS) were graded according to ASTCT criteria, and cytopenia according to CTCAE v5. Polygenic risk score (PRS) instruments for blood cell traits and inflammatory markers were obtained from the PGS Catalog and analyzed using PRSice. Risk of poor response and toxicity was estimated using logistic regression models with standardized PRS scores as a continuous variable adjusting for age at diagnosis, gender, and bridging therapy (yes/no). Exploratory genome-wide association study (GWAS) analyses were also performed. Results. The median age was 58 years, 116 (68%) were male, and 91 (54%) received bridging therapy. Interestingly, increasing PRS for baseline IL-6 levels was associated with decreased risk of CRS grade (G) 2-4 (β: -0.15, 95% CI: -0.30 to -0.013, p=0.030) and ICANS G3-4 (β: -0.21, 95% CI: -0.37 to -0.066, p=0.005). Similarly, increased genetically-predicted monocyte levels decreased risk of ICANS G3-4 (β: -0.16, 95% CI: -0.31 to -0.018, p=0.027). Baseline genetically-predicted monocyte levels were also associated with increased probability of a CR at D90 (β: 0.19, 95% CI: 0.041 to 0.34, p=0.011). For prolonged cytopenia (defined as G3-4 cytopenia ongoing at day 30), risk was increased with increasing PRS for reticulocyte levels (β: 0.29, 95% CI: 0.10 to 0.48, p=0.002). No genome-wide significant (p<5x10 -8) variants were identified in the exploratory GWAS analyses, however, several loci at p=10 -5 were located within/near potential biologically-relevant candidate genes. This included an association with prolonged cytopenia for rs111940551 located upstream of IKZF1, a regulator of lymphocyte differentiation, and serving as a putative expression quantitative trait loci (eQTL) for this gene. An intragenic variant identified for ICANS G3-4 (rs1149927) was in high linkage disequilibrium (D'=0.97) with a variant previously shown to be associated with brain morphology. Conclusion. PRS representing baseline levels of several key blood cell traits, particularly monocytes, and inflammatory markers may impact toxicities and efficacy of CART in patients with LBCL. The intriguing finding of an inverse association for IL-6 with both high-grade CRS and ICANS was unexpected and needs further investigation. Larger studies are warranted to confirm these observations and better understand how relevant germline genetic determinants may influence efficacy and toxicity after CART therapy. Elucidating such determinants may help in improving patient selection and in developing strategies to enhance the therapeutic index of CART.

A Compilation of Experiences in Utilizing CAR-T Cell Therapy for Richter's Transformation

Objective:Even with Bruton tyrosine kinase inhibitors (BTKis) and BCL2 inhibitors, the prognosis for Richter's Transformation (RT) remains dismal, with a median overall survival (OS) of only 9-12 months from initial diagnosis. Although there are reports on CAR-T therapy for RT, the sample size is small and the conclusions vary greatly. Methods: We conducted a retrospective investigation of the efficacy of CAR-T cell therapy in treating RT at our center. Each participant provided their informed consent to partake in the “Phase II Clinical Trial of Sequential CAR-T Cell Therapy Targeting Different B-Cell Epitopes for Recurrent, Refractory Aggressive B-Cell Lymphoma”(Protocol number: BR-XG-CART-II, ChiCTR2100055062, Ethical Approval Number 20191225-PJ-003). Before lymphocyte collection, chemotherapy regimen without fludarabine/ Bendamustine is first used to reduce peripheral blood tumor load if peripheral blood tumor cells are higher than 10%. Patients with extramedullary tumor loads of 5cm or larger were treated with tumor reduction chemotherapy between lymphocyte collection and CART cell transfusion. The pretreatment scheme based on Bendamustine or FC scheme was adopted before CAR-T cells were infused. The efficacy and safety of CART cells were observed. Results: Between Feb 1, 2020 and Feb 28, 2023, our center administered CAR-T cell therapy to 16 patients diagnosed with RT, all of whom developed into diffuse large B-cell lymphoma. The patient population had a median age of 59 years (range: 42-69 years), consisting of 62.5% (10/16) males and 37.5% (6/16) females. The ECOG scores varied between 0 and 3. The median number of prior treatment lines was 5 (2-7). 93.8% failed previous BTKis treatment and 75% failed BCL2 inhibitor treatment; 93.8% had undergone chemotherapy (median 6 courses). 1 patient had previously failed universal CAR-T cell therapy, and another relapsed after autologous transplantation. 90% had at least one TP53 abnormality detected by either FISH or second-generation gene sequencing. Extramedullary masses were found in 87.5% (14/16) of patients, with a median maximum diameter of 42.5mm (18-144mm). Masses larger than 5cm were found in 25% (4/16) of cases, marrow involvement was noted in 62.5% (10/16), and cerebrospinal fluid was affected in 18.75% (3/16). None of the patients had central masses observed in their MRIs. During lymphocyte collection, 58.3% of patients showed positivity for peripheral blood tumor cells, with a maximum tumor cell proportion of 5.69%. The median infused CAR-T cell dose was 1.37×10 6/kg. 12 of them were injected with mouse CD19CAR-T, 2 with human CD20CART, and 2 with human CD19CART. CRS occurred in 81.3%of patients, of which, 68.8%were grade 1-2 and 12.5%were grade 3. Central CRS occurred in 6.3% (1/16) of patients. The complete response (CR) rate at the 3-month mark stood at 56.3% (9/16), with an overall response rate (ORR) of 68.8% (11/16). As of July 6, 2023, the median follow-up time was 10.6 months (4.4-33.6m), the 1-year OS was 73.4%, and the 1-year PFS was 68.8%. Median OS and PFS were not reached. One of the five patients who did not respond later underwent allogeneic hematopoietic stem cell transplantation and is still alive. The other four, despite attempts at salvage treatment including second CAR-T cell therapy, all died, with a median OS of only 5.6 months (4.4-8.9m). None of the four patients with extramedullary masses larger than 5cm achieved ORR despite tumor-reducing chemotherapy prior to CAR-T infusion. Only 1 of 3 patients with central involvement achieved CR and experienced grade 3 CRS along the way, while the other 2 did not respond to CAR-T therapy. Among all patients, only one experienced central CRS, and his cerebrospinal fluid showed no signs of tumor cells prior to the administration of CAR-T treatment. A patient who was in CR succumbed to an acute coronary event 19 months following infusion, despite remaining in CR. Conclusion: Even in the presence of TP53 gene abnormalities, resistance to BTK/BCL2 inhibitors, and unsuccessful previous chemotherapy regiments, A single CAR T cell therapy can result in CR in more than half of RT patients. Nonetheless, the prognosis remains bleak for patients presenting with large tumor masses and central nervous system involvement, despite having received chemotherapy aimed at reducing tumor size. The outlook is particularly dismal for those patients who fail to respond to CAR-T cell therapy.

The Impact of Pre-Existing Autoimmune Disease on the Safety and Efficacy of CAR T-Cell Therapy for Non-Hodgkin Lymphoma

Introduction: Chimeric antigen receptor T-cell therapy (CAR-T) has revolutionized the treatment of non-Hodgkin lymphoma (NHL). Despite having an increased risk of NHL, patients with pre-existing autoimmune disease (AID) were excluded from pivotal CAR-T trials due to concerns that CAR-T may exacerbate AID. As a result, there are limited data on the safety and efficacy of CAR-T for NHL in patients with AID. Interestingly, there is also growing evidence that CAR-T can be therapeutically effective for AID, such as in the treatment of systemic lupus erythematosus (SLE). As CAR-T utilization for NHL increases, it is essential to characterize the safety and efficacy of this potentially curative therapy in patients with AID. This study aimed to evaluate the impact of AID on outcomes in patients treated with commercial CD19-targeted CAR-T for NHL. Methods: Following Institutional Review Board approval, we conducted a retrospective analysis of all patients who were treated with commercial CD19-targeted CAR-T for NHL and had a pre-existing AID across the Mayo Clinic Enterprise (Arizona, Florida, and Minnesota) between January 2018 and May 2023. Variables collected included age, gender, NHL type, AID type, and CAR-T product. Efficacy outcomes, including response rates at 6 and 12 months after CAR-T, were collected. Toxicities, including cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS), were collected and analyzed. Our chart review for AID included SLE, sarcoidosis, ulcerative colitis, celiac disease, pernicious anemia, Crohn's disease, psoriasis, Sjogren's syndrome, hypothyroidism (Hashimoto's disease), hyperthyroidism (Graves' disease), rheumatoid arthritis (RA), vasculitis, scleroderma, and mixed connective tissue disease (MCTD). Results: A total of 235 patients who received commercial CD19-targeted CAR-T for NHL across the Mayo Clinic Enterprise were identified (Table 1). Of these 235 patients, 27 had a pre-existing AID and 208 did not. The median age was 66 years (range 39-81) for patients with AID and 59 years (range 18-88) for patients without AID. Over half of the patients were male (59% of patients with AID and 51% of patients without AID). The most common NHL type was diffuse large B cell lymphoma (41% of patients with AID and 56% of patients without AID). Most patients received axicabtagene ciloleucel (85% of patients with AID and 84% of patients without AID). The most common AID was hypothyroidism (81%, n=22), although other conditions were also identified, including RA (11%, n=3), SLE (4%, n=1), pulmonary sarcoidosis (4%, n=1), psoriasis (4%, n=1), and MCTD (4%, n=1). Two patients had more than one AID. Median follow-up was 18 months (range 1-36) for patients with AID and 14 months (range 2-49) for patients without AID. Six months after CAR-T, the complete response (CR) rate was 77.2% and 45.1% in patients with AID and without AID, respectively (p=0.0042). Twelve months after CAR-T, the CR rate was 50% and 31.4% in patients with AID and without AID, respectively (p=0.0801). CRS of any grade occurred in 78% and 88% of patients with and without AID, respectively. ICANS of any grade occurred in 41% and 54% of patients with and without AID, respectively. Conclusion: AID did not appear to adversely impact the safety or efficacy of CAR-T for NHL. Limitations of our study include the small number of patients with AID and the fact that the most common AID was hypothyroidism. Larger studies are warranted to confirm these findings. Additional data are needed to characterize the impact of CAR-T for NHL on AID outcomes and to describe the effect of disease-modifying antirheumatic drugs on NHL outcomes after CAR-T.

18F-FDG PET/CT Imaging Biomarkers for CAR-T Cell Therapy Outcomes in Large-B-Cell Lymphoma

Introduction: CAR-T cell therapy has transformed the care of patients with large-B-cell lymphoma (LBCL), but disease relapse and toxicities remain clinical challenges, while effective quantitative biomarkers are lacking. Our aim was to determine the role of 18F-FDG PET/CT imaging features as biomarkers in LBCL treated with CD19 CAR-T cell therapy. Methods: 18F-FDG PET/CT scans performed before CAR-T for LBCL were analyzed retrospectively by a board certified radiologist. Metabolic tumor volume (MTV), SUVmax, total lesion glycolysis (TLG), presence of bulky disease, and 106 radiomic features per lesion were recorded using the PET/CT and PyRadiomics plugins for FIJI. Laboratory values including IL-6, IL-10, TNF-α, LDH, ferritin, fibrinogen, and D-dimer pre CAR-T infusion were recorded. Response to CAR-T cell therapy was defined according to Lugano criteria. Cytokine release syndrome (CRS) and neurotoxicity were graded according to ASTCT guidelines. Progression-free survival (PFS) and overall survival (OS) were measured from time of CAR-T cell infusion; PFS events were death, relapse, and disease progression. Association between PET features, markers of inflammation and outcomes were studied with Spearman rank correlations, Cox, and logistic regression models. Multivariable generalized estimating equation models were applied to evaluate differential expression of radiomic features between responders and non-responders, accounting for the dependency structure among ROIs from the same individual. Results: A total of 180 LBCL patients (median age 66 years) treated with autologous CD19 CAR-T (93 axi-cel, 52 tisa-cel, 35 liso-cel) were included. Two time points were considered: 1) pre-apheresis scans (n=161; median SUVmax 19, MTV 56 mL, TLG 427 SUV*mL), and 2) pre-infusion, i.e., latest disease evaluation before CAR-T infusion including post-apheresis/post-bridging, as well as pre-apheresis scans for patients without additional imaging afterwards (n=180; median SUVmax 15, MTV 44 mL, TLG 338 SUV*mL), with a total of 341 PET/CT scans evaluated. At lymphodepletion, MTV was significantly correlated with multiple elevated inflammatory laboratory markers; at latest assessment, all PET and inflammatory markers showed significant correlations. MTV at last assessment before infusion was associated with increased risk of grade ≥ 2 CRS (OR 1.08 [95%CI 1.01-1.20], P=0.031), but not ICANS, in a multivariable logistic regression model adjusted for age, pre-lymphodepletion LDH, CAR-T product, bridging and SUVmax. A higher SUVmax at latest assessment was associated with a lower likelihood of non-complete response (CR) (OR 0.58 [0.41-0.80], P<0.001), while MTV lacked such an association after multivariable model. Increased MTV both before apheresis and at last disease assessment before infusion, but not SUVmax, were associated with decreased PFS (HR 1.11 [1.05-1.17] and HR 1.04 [1.02-1.07], respectively), in a multivariable adjustment. A similar pattern was observed for OS. Notably, the combination of MTV and LDH enabled clear separation of high and low PFS risk groups (P<0.001, Figure 1). Finally, when considering over 100 granular pre-infusion radiomic features ( Figure 2), which are not routinely evaluated in CAR-T patients, multiple novel determinants of CR were discovered, including gray level dependence, run length matrix, and shape-based features (P<0.05). Conclusions: To the best of our knowledge, this represents the most extensive analysis of quantitative 18F-FDG PET/CT features within an LBCL cohort undergoing CAR-T treatment. Our findings indicate that standard PET features (MTV and SUVmax) predict both toxicity and efficacy of CAR-T cell therapy, while also showing correlations with systemic inflammation. Additionally, we present, for the first time, the relevance of granular radiomic features in assessing the probability of treatment success. If confirmed, PET features could serve as effective biomarkers in CAR-T patients and have the potential to guide interventions in high-risk populations.

IMMU-04. LOW-INTENSITY PULSED ULTRASOUND ENHANCES CAR T CELL THERAPY BY INDUCING TWO-WAY IMMUNE CELL TRAFFICKING – T CELL INTO TUMOR AND MYELOID CELL INTO PERIPHERY IN A SYNGENEIC MOUSE GLIOBLASTOMA MODEL

Abstract Chimeric Antigen Receptor (CAR) T-cell therapy has been successful against hematological malignancies. However, it has not established efficacy in glioblastoma (GBM) patients. Significant challenges currently limiting the effectiveness of CAR T-cell therapy for patients with GBM include poor CAR-T cell trafficking to GBM due to the blood-brain barrier, and the presence of highly immunosuppressive and pro-tumoral tumor-associated macrophages (TAMs) in the tumor microenvironment. Furthermore, the heterogeneous expression of target antigens in GBM can lead to antigen escape and therapeutic resistance. Low-intensity pulsed ultrasound with microbubbles (LIPU/MB) has been shown to enhance CAR T-cell trafficking and improve survival in immunocompromised xenograft mice GBM models. In this study, we investigated whether multiple rounds of LIPU/MB can improve not only CAR T-cell but also endogenous T-cell infiltration into the tumors and induce myeloid cell trafficking into lymph nodes (LNs), thus enhancing CAR T-cell therapy against GBM in the immunocompetent syngeneic mice model. We recently developed a novel immunocompetent transgenic (Tg) mouse that expresses an anti-EGFRvIII CAR on T-cells and enables us to investigate the interaction between CAR T-cells and the endogenous immune environment. C57BL6/J mice bearing intracerebral EGFRvIII+ GBM received an intravenous administration of anti-EGFRvIII CAR T-cells derived from Tg-mice. The mice were stratified to receive single-LIPU (1 min before CAR T-cells administration) or triple-LIPU/MB (1 min before CAR T-cells administration, then two and four days later). Triple-LIPU/MB significantly suppressed tumor growth and improved survival and was associated with increased CAR T-cells and endogenous T-cells infiltration into the tumors compared with single-LIPU/MB or without LIPU. Furthermore, draining cervical LNs showed increased macrophages trafficking after triple-LIPU/MB. Moreover, TAMs isolated from triple-LIPU/MB mice brains showed higher levels of MHC-II expression. These results suggest that LIPU/MB may promote the tumor antigen presentation to endogenous T-cells and enhance the efficacy of CAR T-cell therapy for GBM.

Temporal Dynamics and Efficacy of CAR-T Cell Therapy in B-ALL with Extramedullary Cutaneous Involvement: A Case Report and Discussion

Temporal Dynamics and Efficacy of CAR-T Cell Therapy in B-ALL with Extramedullary Cutaneous Involvement: A Case Report and Discussion

Establishment of a Dynamic Ctdna Monitoring System to Predict the Prognosis of CAR-T Cell Therapy in R/R B-NHL Patients

Introduction: The objective of this study is to establish a prediction system using circulating tumor DNA (ctDNA) to determine the effectiveness of CAR-T cell therapy in patients with relapsed or refractory B-cell non-Hodgkin lymphoma (R/R B-NHL). Furthermore, the study aims to compare the mutation characteristics between different therapeutic groups to provide guidance for the prevention and follow-up treatment of patients who fail CAR-T cell therapy. Methods: In this part of the study, a total of 79 (192 serum samples) R/R B-NHL patients with CAR-T cell therapy were collected for ctDNA detection (187 lymphoma-related gene panel), including 62 samples before treatment (T0), 69 samples at 1 week (T1) and 62 samples at 4 weeks (T2) after reinfusion. Differences of categorical variables were tested using the chi-square test or Fisher's exact test, and differences in survival events between different groups were compared using Kaplan-Meier analysis and log-rank test. Results: The results of Kaplan-Meier analysis showed that patients with ctDNA mutation genes >10 before CAR-T cell therapy had poorer OS (1-year OS rate: 0% vs 74.9%, 2-year OS rate: 0% vs 68%, P<0.001) and PFS (1-year PFS rate: 0% vs 52.5%, 2-year PFS rate: 0% vs 36.6%, P=0.0056). Patients with MYD88, FAT1 and BTG2 mutation before CAR-T cell therapy had poorer OS, while patients with MUC16 mutation had better OS. The CR rate in patients with TP53 mutation before CAR-T cell treatment was significantly lower than that of patients without TP53 mutation (33.3% vs 68.1%, P=0.02). However, the TP53 mutation status before treatment did not have an impact on the long-term survival of patients. All patients with TP53 mutations at 4 weeks after CAR-T cell therapy failed to achieve CR, and OS was poorer (1-year OS rate: 37.5% vs 66.4%; 2-year OS rate: 12.5% vs 56.3%, P=0.0023). For CR patients, patients with BCR mutation at 4 weeks after treatment had poorer OS (2-year OS rate: 40.9% vs 76.1%, P=0.035). At one week after CAR-T cell therapy(Figure 1), patients without mutations of CDKN2A, CBLB, APC, SPEN, KMT2D, CARD11, FOXO1 and PDGFRB, were more likely to achieve CR (76.6% vs 28.6%, P<0.001), and had better OS (1-year OS rate: 81.5% vs 38.9%, 2-year OS rate: 62.2% vs 5%, P<0.001) and PFS (1-year PFS rate: 67.2% vs 0%, P<0.001). Conclusions: In this study, we evaluated the characteristics of gene mutation between different therapeutic groups, and a gene set was screened to predict the efficacy of CAR-T cell therapy in R/R B-NHL patients, helping clinicians accurately evaluate the efficacy and assisting in decision-making of treatment options.

Intravenous IL-1 Receptor Antagonist for Prevention of Severe Immune Effector Cell-Associated Neurotoxicity Syndrome

Background and Significance: The management of toxicities from autologous CD19-directed chimeric antigen receptor (CAR) T cell therapy have evolved and the rates of severe cytokine release syndrome (CRS) have declined from nearly 50% in early trials to 8% (CRS grade 3 or higher) from real-world retrospective data with axicabtagene ciloleucel (axi-cel) (Jacobson et al. TCT 2022). However, the rate of severe immune effector cell-associated neurotoxicity (ICANS) has only marginally improved since CAR-T cell therapy has become commercially available. With expanding indications for CAR-T cell therapy, the prevention and management of ICANS remain an unmet need. We previously conducted a pilot, investigator-sponsored phase II study using Anakinra, an IL-1 receptor antagonist, administered subcutaneously at the onset of grade 2 CRS or any grade ICANS (Oliai et al. ASCO 2021) which demonstrated that anakinra is safe and feasible. There are several prospective studies that are currently open to accrual that incorporate subcutaneous Anakinra for prevention and treatment of severe ICANS (NCT04359784, NCT04148430, NCT04150913, and NCT04432506). Anakinra crosses the blood brain barrier and has been shown to be safe and efficacious in rheumatologic conditions. Relatively high doses of IV Anakinra have been shown to be safe in severe sepsis and in subarachnoid hemorrhage. We hypothesize that IV Anakinra is safe and effective in preventing severe ICANS in patients undergoing standard of care axi-cel therapy for large B cell lymphoma. Study Design and Methods: This investigator-sponsored trial (NCT4205838) is a phase Ib/II study of adults eligible for standard of care axi-cel for large B cell lymphoma. Anakinra will be administered intravenously using a standard 3+3 design with cohort 1 receiving 200mg IV loading dose, followed by 100mg IV q6h maintenance dose and cohort 2 receiving 400mg IV loading dose, followed by 200mg IV q6h. The trigger to initiate Anakinra is any grade ICANS or grade ≥2 CRS in the absence of ICANS, based on ASTCT 2019 criteria. In addition to Anakinra, participants will receive standard of care interventions for CRS and ICANS. Subjects with secondary CNS involvement are included. Key exclusion criteria are primary CNS lymphoma, Burkitt's lymphoma, transformed DLBCL from CLL, prior allogeneic stem cell transplantation, < 30 days of exposure to immune check point inhibitor therapy. The study is open to accrual at University of California, Los Angeles (UCLA) with 2 additional sites within the University of California Hematologic Malignancies Consortium (UCHMC). Objectives: The primary objective is to evaluate the safety and tolerability of Anakinra given intravenously in patients undergoing CAR-T cell therapy and to estimate the efficacy of Anakinra in prevention of severe ICANS (grade ≥3). Secondary objectives include the impact of Anakinra on the efficacy of CAR-T cell therapy for large B cell lymphoma, the duration of ICANS, and the pharmacokinetics of Anakinra. Exploratory objectives focus on changes in inflammatory cytokines during ICANS in the peripheral blood. Statistics and Current Status: Using historical control from the real-world experience using axi-cel for relapsed/refractory large B-cell lymphoma (38%, Jacobson et al. ASH 2018), a sample size of 36 patients achieves 80% power to detect a 50% reduction in the rate of severe ICANS when comparing to the historical control using a one-sample exact binomial test at the two-sided type I error rate of 0.10. This study has been open at UCLA since February 2023. Two of 4 enrolled participants met criteria to receive IV Anakinra. Conclusions: The safety and tolerability of IV Anakinra is being evaluated in patients with large B cell lymphoma undergoing treatment with axi-cel in order to prevent severe ICANS.

Co-expression of IL-4/IL-15-based inverted cytokine receptor in CAR-T cells overcomes IL-4 signaling in immunosuppressive pancreatic tumor microenvironment

The efficacy of CAR-T cell therapy has been hindered by several factors that are intrinsic to the tumor microenvironment. Many strategies are being employed to overcome these barriers and improve immunotherapies efficacy. Interleukin (IL)− 4 is a cytokine released by tumor cells inside the tumor microenvironment and it can oppose T cell effector functions via engagement with the IL-4 receptor on the surface of T cells. To overcome IL-4-mediated immunosuppressive signals, we designed a novel inverted cytokine receptor (ICR). Our novel CAR construct (4/15NKG2D-CAR), consisted of an NKG2D-based chimeric antigen receptor, co-expressing IL-4R as an extracellular domain and IL-15R as a transmembrane and intracellular domain. In this way, IL-4R inhibitory signals were converted into IL-15R activation signals downstream. This strategy increased the efficacy of NKG2D-CAR-T cells in the pancreatic tumor microenvironment in vitro and in vivo. 4/15NKG2D-CAR-T cells exhibited increased activation, degranulation, cytokine release, and cytotoxic ability of NKG2D-CAR-T cells against IL-4+ pancreatic cell lines. Furthermore, 4/15NKG2D-CAR-T cells exhibited more expansion, less exhaustion, and an increased percentage of less differentiated T cell phenotypes in vitro when compared with NKG2D-CAR-T cells. That is why IL-4R/IL-15R-modified CAR-T cells eradicated more tumors in vivo and outperformed NKG2D-CAR-T cells. Thus, we report here a novel NKG2D-CAR-T cells that could overcome IL-4-mediated immunosuppression in solid tumors.

Tumor microenvironment and CAR-T cell immunotherapy in B-cell lymphoma.

Chimeric receptor antigen T cell (CAR-T cell) therapy has demonstrated effectiveness and therapeutic potential in the immunotherapy of hematological malignancies, representing a promising breakthrough in cancer treatment. Despite the efficacy of CAR-T cell therapy in B-cell lymphoma, response variability, resistance, and side effects remain persistent challenges. The tumor microenvironment (TME) plays an intricate role in CAR-T cell therapy of B-cell lymphoma. The TME is a complex and dynamic environment that includes various cell types, cytokines, and extracellular matrix components, all of which can influence CAR-T cell function and behavior. This review discusses the design principles of CAR-T cells, TME in B-cell lymphoma, and the mechanisms by which TME influences CAR-T cell function. We discuss emerging strategies aimed at modulating the TME, targeting immunosuppressive cells, overcoming inhibitory signaling, and improving CAR-T cell infiltration and persistence. Therefore, these processes enhance the efficacy of CAR-T cell therapy and improve patient outcomes in B-cell lymphoma. Further research will be needed to investigate the molecular and cellular events that occur post-infusion, including changes in TME composition, immune cell interactions, cytokine signaling, and potential resistance mechanisms. Understanding these processes will contribute to the development of more effective CAR-T cell therapies and strategies to mitigate treatment-related toxicities.

Central nervous system lymphomas-Assessment and treatment and prevention of central nervous system relapse.

In this review focused on lymphoma and the central nervous system (CNS), we summarize recent developments in the management of primary (PCNSL) and secondary CNS lymphoma (SCNSL), treatment of CNS lymphoma in the older population, the neuroradiological assessment of CNS lymphoma and finally highlight the ongoing debate on optimal CNS prophylaxis. The section on PCNSL focuses on the different approaches available for frontline treatment in Europe and the United States and discusses consolidation strategies. We then highlight available strategies to treat PCNSL in the elderly population, an area of unmet need. New therapies aiming at minimizing toxicity and prioritizing quality of life are emerging for these patients. Secondary CNS lymphoma, especially in the relapsed/refractory setting is another area of unmet need, and the efficacy of CAR-T cell therapy is being explored. We provide an overview of the imaging challenges in the neuroradiological assessment of CNS lymphoma. Finally, the section on CNS prophylaxis summarizes recent findings from large retrospective studies challenging the efficacy of present approaches to prophylaxis in higher-risk patients with lymphoma.

Self-delivery of TIGIT-blocking scFv enhances CAR-T immunotherapy in solid tumors.

Chimeric antigen receptor T cell therapy has become an important immunotherapeutic tool for overcoming cancers. However, the efficacy of CAR-T cell therapy in solid tumors is relatively poor due to the complexity of the tumor microenvironment and inhibitory immune checkpoints. TIGIT on the surface of T cells acts as an immune checkpoint by binding to CD155 on the tumor cells' surface, thereby inhibiting tumor cell killing. Blocking TIGIT/CD155 interactions is a promising approach in cancer immunotherapy. In this study, we generated anti-MLSN CAR-T cells in combination with anti-α-TIGIT for solid tumors treatment. The anti-α-TIGIT effectively enhanced the efficacy of anti-MLSN CAR-T cells on the killing of target cells in vitro. In addition, we genetically engineered anti-MSLN CAR-T cells with the capacity to constitutively produce TIGIT-blocking single-chain variable fragments. Our study demonstrated that blocking TIGIT significantly promoted cytokine release to augment the tumor-killing effect of MT CAR-T cells. Moreover, the self-delivery of TIGIT-blocking scFvs enhanced the infiltration and activation of MT CAR-T cells in the tumor microenvironments to achieve better tumor regression in vivo. These results suggest that blocking TIGIT effectively enhances the anti-tumor effect of CAR-T cells and suggest a promising strategy of combining CAR-T with immune checkpoints blockade in the treatment of solid tumors.

HBV reactivation in patients with chronic or resolved HBV infection following BCMA-targeted CAR-T cell therapy.

B-cell maturation antigen (BCMA)-targeted chimeric antigen receptor-T cell (CAR-T) therapy is used for refractory or relapsed multiple myeloma (r/r MM). Concern of the safety and efficacy of CAR-T cell therapy in patients with chronic or resolved HBV infection is raised. In this study, we retrospectively reviewed 99 patients with r/r MM treated with BCMA-targeted CAR-T cell therapy, of which 7 (7.1%) patients had chronic HBV infection, 43 (43.4%) with resolved HBV infection, and the remaining 49 (49.49%) HBV-uninfected. Patients' characteristics before CAR-T cell administration were comparable in different status of HBV infection. Patients' liver function, cytokine levels, CAR-T cell expansion and cytokine release syndrome (CRS) grade after CAR-T cell therapy did not differ in different HBV serologic status. Furthermore, chronic HBV infection or resolved HBV infection did not affect clinical response, progress-free survival (PFS), or overall survival (OS). Four (4.04%) patients experienced HBV reactivation, 3 (6.98%) with resolved HBV infection, and 1 (14.29%) chronic HBV infection. Of 4 patients with HBV reactivation, 2 cases (50%) of severe hepatitis were noted and reported. Drops of serum IgG and elevation of alanine aminotransferase (ALT), alanine aminotransferase (AST), total bilirubin (TB) were observed in all four patients around the date of HBV reactivation.

Engineering memory-rich CAR-T cells by a piggyBac transposon system

The clinical application of chimeric antigen receptor T-cell therapy (CAR-T therapy) has significantly altered the therapeutic strategy for B-cell tumors and is now being used to treat myeloid and solid tumors. Nonetheless, the efficacy of CAR-T cell therapy for myeloid and solid tumors has been limited, and several studies are being conducted to understand and overcome the underlying mechanisms. Recent research achievements have revealed that the properties of CAR-T cells, particularly their memory function, which can be continuously amplified in the body without exhaustion after administration, are closely related to CAR-T cell clinical efficacy. Furthermore, because the characteristics of CAR-T cells are greatly influenced by the quality of peripheral blood mononuclear cells, the raw material of CAR-T cells, and the T-cell used during the manufacturing process, attention has been drawn to the development of high-quality CAR-T cell manufacturing methods and combination therapies that maintain CAR-T cell memory function and suppress immune exhaustion. This article provides an overview of the current state of CAR-T cell development and clinical application to cancer, particularly emphasizing the development of manufacturing processes and efforts to improve CAR-T cell efficacy in combination therapy with molecular-targeting drugs.

Cancer cell targeting by CAR-T cells: A matter of stemness.

Chimeric antigen receptor (CAR)-T cell therapy represents one of the most innovative immunotherapy approaches. The encouraging results achieved by CAR-T cell therapy in hematological disorders paved the way for the employment of CAR engineered T cells in different types of solid tumors. This adoptive cell therapy represents a selective and efficacious approach to eradicate tumors through the recognition of tumor-associated antigens (TAAs). Binding of engineered CAR-T cells to TAAs provokes the release of several cytokines, granzyme, and perforin that ultimately lead to cancer cells elimination and patient's immune system boosting. Within the tumor mass a subpopulation of cancer cells, known as cancer stem cells (CSCs), plays a crucial role in drug resistance, tumor progression, and metastasis. CAR-T cell therapy has indeed been exploited to target CSCs specific antigens as an effective strategy for tumor heterogeneity disruption. Nevertheless, a barrier to the efficacy of CAR-T cell-based therapy is represented by the poor persistence of CAR-T cells into the hostile milieu of the CSCs niche, the development of resistance to single targeting antigen, changes in tumor and T cell metabolism, and the onset of severe adverse effects. CSCs resistance is corroborated by the presence of an immunosuppressive tumor microenvironment (TME), which includes stromal cells, cancer-associated fibroblasts (CAFs), tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs), and immune cells. The relationship between TME components and CSCs dampens the efficacy of CAR-T cell therapy. To overcome this challenge, the double strategy based on the use of CAR-T cell therapy in combination with chemotherapy could be crucial to evade immunosuppressive TME. Here, we summarize challenges and limitations of CAR-T cell therapy targeting CSCs, with particular emphasis on the role of TME and T cell metabolic demands.

CD44‐targeted chimeric antigen receptor‐T eradicates acute myeloid leukaemic stem cells

CD44‐targeted chimeric antigen receptor‐T eradicates acute myeloid leukaemic stem cells

Valproic Acid Increased CAR T Cell Cytotoxicity Against Acute Myeloid Leukemia

Background The treatment paradigm of B cell malignancies has dramatically changed in recent years with the introduction of immunotherapy, especially CAR-T cell therapy. However, only limited efficacy was observed in AML with CAR-T cell therapy. Methods In this study, the molecular, clinical, and tumor immune microenvironments characteristics of AML patients in TCGA were performed on an online bio-information analysis website. The expressions of CD123 and CLL-1 on the AML cells from clinical samples were analyzed by flow cytometry. Then we constructed anti-CD123 CAR and CLL-1 CAR with different co-stimulation domains CD28 or 4-1BB and detected the anti-AML effects. To increase the efficacy of CAR-T cell therapy, we tested different combination strategies, such as checkpoint inhibitor PD-1/PD-L1 antibodies and histone deacetylase inhibitor (HDACi) in vivo and in vitro. RNA sequencing and western blot were performed to analyze the potential mechanism of valproic acid (VPA) in upregulating NKG2DL. Results The data suggested that CD123 and CLL-1 were highly expressed on AML leukemia cells (p=0.0031, p=0.0069). The proportion of various immune cells such as T cell subsets and NK cells in AML patients was significantly lower than that in healthy donors. The process of immune infiltration mainly stagnated in the recruitment and activation of immune cells or the tracking migration of immune cells. The immunosuppressive molecule PD-1 was negatively correlated with memory T cells, while the active molecules CD27, CD28, and NKG2D were positively correlated with CD8 + memory T cells. Both CD123 CAR and CLL-1 CAR displayed specific anti-AML effects. To improve the lysis effects of CAR-T cells, we combined CAR-T cell therapy with different agents. PD-1/PD-L1 antibody could only slightly improve the potency of CAR-T cell therapy (CD123 CAR-T 60.92±2.9087% VS 65.43±2.1893%, 60.92±2.9087% VS 67.43± 3.4973%, CLL-1 CAR-T 37.37±3.908% VS 41.89 ±5.1568%, 37.37±3.908% VS 42.84± 4.2635%). While, one of HDACi, valproic acid (VPA) could synergize with CAR-T cell therapy and significantly improve CAR-T cell potency against AML cells, especially in pre-treated with VPA for 48h ( CLL-1 CAR-T 34.97±0.3051% VS 88.167±1.5327% p<0.0001, CD123 CAR-T 26.87±2.7010% VS 82.56±3.086%, p<0.0001 in MV411, CLL-1 CAR-T 78.77±1.2061% VS 93.743±1.2333% p<0.0001, CD123 CAR-T 64.10±1.5130% VS 94.427±0.142%, p=0.0001 in THP-1). What's more, the anti-AML effects of combination therapy were also observed in the AML mice model. NCG mice were injected with 1×106 THP-1 cells that express firefly luciferase and GFP label (THP-1-Fluc) via tail vein, 1 week later mice were infused with VPA 0.5mM (iv, n=6), VPA 100mg/kg (ip, n=6), VPA 0.5mM (iv, twice a day, n=6) and PBS (iv, n=6), then CD123 CAR-T cells or untreated T cells were infused 48h later. The combination therapy prolonged the overall survival of mice when compared to single CAR-T cell therapy (median survival:180 days VS unfollowed). One of the possible mechanisms underlying we found was that NKG2D could upregulate on activated CD8+T cells, and VPA could upregulate NKG2D ligand expression in the AML cells which may contribute to NKG2D-mediated cytotoxicity of CAR-T cells against tumor cells. In addition, the expression of HDAC2 and H4 in THP-1 cells was significantly decreased after VPA treatment, suggesting that VPA may enhance the expression of NKG2DL by degrading HDAC2 expression. Conclusion CD123 and CLL-1 are promising targets for AML CAR-T cell therapy. A combination of VPA pre-treatment and CAR-T against AML exhibits synergic effects, which may provide a promising strategy in the clinical treatment of R/R AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Multi-Omics Reveal Plasma Molecular Signatures of B-ALL Patients during CAR-T Cell Therapy

Background: The Chimeric antigen receptor (CAR) T cell therapy for hematological malignancies has revolutionized anticancer therapy. However, CAR-T cells can cause immunity-related adverse events, such as cytokine release syndrome and neurotoxicity; the efficacy of CAR-T cell therapy also varies substantially with different patients. Poor understanding of plasma proteomic and metabolomic features during chimeric antigen receptor (CAR) T cell therapy may affect the exploitation of therapeutic potential in the management of hematological malignancies. Methods: We longitudinally performed proteomic and metabolomic profiling of plasma from 20 patients with relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL) on CAR-T cell therapy. Blood samples were collected from B-ALL patients at different time points, including before fludarabine/cyclophosphamide lymphodepletion (Pre-FC), Day 0 (D0), Day 7 (D7), Day 14 (D14), Day 28 (D28), and Peak-IL-6 during CAR-T cell therapy. The plasma samples from 22 age- and gender-matched healthy controls (HC) were used as controls. Results: Compared with HC, 239, 217, 282, 260 and 261 proteins were markedly different in terms of expression level in these five groups (Pre-FC, D0, D7, D14, D28 versus HC), respectively (Figure 1). The pathways of differentially expressed proteins (DEPs) were highly enriched in processes implicated in actin binding, cadherin binding, regulation of ion transport, platelet aggregation and regulation of actin cytoskeleton, with actin binding obtaining the most significant change (Figure 1). 104, 139, 175, 223 and 87 metabolites were significantly different in these five groups, respectively (Figure 1). Interestingly, the alterations of plasma metabolites became more extensive on the 14th day after CAR-T cell infusion, indicating that the change of metabolites may lag behind that of proteins. Furthermore, CAR-T cell therapy reshaped plasma proteome and metabolome with discrete kinetics, as represented by upregulated acute-phase response and triglycerides metabolism and downregulated actin cytoskeleton organization, retrograde endocannabinoid signaling and glycosylphosphatidylinositol (GPI)-anchor biosynthesis during the therapy. We also identified sets of covarying molecules that were involved in acute inflammatory response, epithelial mesenchymal transition, glutathione metabolism and tryptophan metabolism in concert with the dynamics of circulating IL-6 in patients with cytokines storm. Subgroup analysis further revealed numerous prognostic biomarkers, such as proteins related to allograft rejection, which were down-regulated in patients who achieved stable complete remission. Conclusions: This study provided a dynamic landscape of plasma proteome and metabolome in B-ALL patients during CAR-T cell therapy and offered novel insights into CAR-T cell therapy-related toxicities and prognostic heterogeneity. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal