IDR-induced CAR condensation improves the cytotoxicity of CAR-Ts against low-antigen cancers.

Promoter usage regulating the surface density of CAR molecules may modulate the kinetics of CAR-T cells in vivo

Acute Kidney Injury After the CAR-T Therapy Tisagenlecleucel

Disclosure: W.J. Kulakowska: None. R.A. Taylor: None. W. Ranasinghe: None. P.K. Darcy: None. J.A. Trapani: None. G.P. Risbridger: None. Background and aims While CAR (Chimeric Antigen Receptor) T cell therapies targeting PSMA and PSCA receptors are currently in phase 1/2 studies in metastatic castrate-resistant prostate cancer (mCRPC), antigen targets for neuroendocrine prostate cancer (NEPC) remain limited. Lewis Y antigen (LeY) is an oncofetal antigen expressed only in adult tumour cells and is considered a target for CAR T cell therapy in haematological, lung, ovarian and colon cancer. We previously found that LeY is a promising target for CAR T cell therapy in prostate cancer and that preconditioning LeY-directed CAR T cells with carboplatin further enhances tumour responses in mCRPC. We investigated if LeY can be used as a novel CAR T cell target for NEPC. Methods: The expression of the Lewis Y antigen was assessed in our patient-derived prostate cancer xenograft (PDX) collection using immunohistochemistry (IHC). To assess the efficacy of LeY-CAR T cells, we used specimens established from a patient (Patient 508) with NEPC who had incurable metastatic disease and failed standard treatments. We generated CAR T cells from his peripheral blood mononuclear cells (PBMCs) to test their specificity and efficacy in vitro. Results: In our MURAL PDX collection of 59 prostate tumours, 81% (9/11) NEPCs expressed LeY antigen on IHC, including Patient 508. We successfully produced anti-LeY CAR T cells from the patient’s PBMCs using retroviral transduction. Generated CAR T cells presented a nearly 1:1 CD4+:CD8+ T cell ratio (51.7% and 41.4% of total T cells, respectively), which according to existing literature yields better treatment outcomes. Next, we used an immunobead assay to investigate CAR-T specificity in releasing proinflammatory cytokines. Significant cytokine production was observed after a co-culture with LeY+ human ovarian cancer cell line, OVCAR3, vs co-culture with control LeY- MDA-MB435 cells (p=0.0004), demonstrating the specificity of the CAR T cells. Lastly, we showed that patient CAR T cells effectively killed LeY+ cells (OVCAR3), but not control LeY- cells (MDA-MB435; p<0.0001) in a 16-hour chromium release assay. CAR T cells killed LeY+ target cells more effectively than their non-transduced equivalents at a range of effector-to-target ratios, with up to 15-fold increase in cell death. Conclusion: Our preclinical studies show that most NEPC tumors express the LeY antigen and that patient-derived CAR T cells can specifically kill those cells. These findings identified a potential CAR T treatment target for NEPC for a phase 1/2 study. Presentation: Monday, July 14, 2025

Chimeric Antigen Receptor T Cell Therapies Clinical Trials in Pediatric Oncology: A Retrospective Analysis from Clinicaltrials.Gov

Low Target Antigen Expression Mediates Resistance to BCMA CAR T Cell Therapy

Pediatric cancer poses a major health challenge globally, especially in low-middle-income countries like Indonesia. The survival rate of pediatric cancer in many high-income countries (HICs) reaches 90%, while it only ranges from 5 to 60% in LMICs. Over 80% of children with cancer live in low-middle-income countries, indicating the urgency to improve the survival rate of pediatric cancer in LMICs [1]. In Indonesia, the prevalence of pediatric cancer was 43.5% in 2020, making it the highest among Southeast Asian countries [2]. According to Dharmais Cancer Hospital (2024), the national cancer referral center for all of Indonesia, the 5-year survival rate of high-risk pediatric acute lymphoblastic leukemia is only 48.8% (unpublished data).One key factor contributing to the low survival rate of pediatric cancer in Indonesia is the lack of effective therapy options, especially for high-risk and relapsed or refractory patients. Several therapeutic approaches, such as immunotherapy, have been widely used in HICs but are still not very popular in Indonesia. CAR (Chimeric Antigen Receptor) T-cell therapy is one of the most promising immunotherapeutic approaches to treat pediatric cancer. Implementing CAR T Cell therapy in Indonesia offers promising prospects for improving the survival rates of pediatric cancer patients.CAR T cell therapy utilizes the body's immune system to specifically target and eliminate cancer cells. This innovative therapy entails extracting a patient's T cells, genetically modifying them to express chimeric antigen receptors specific to tumor-associated antigens, and then reinfusing them into the patient. Once infused, these engineered T cells recognize and eliminate cancer cells bearing the targeted antigen, thereby offering a highly targeted and potentially curative treatment option [3]. This innovative therapy has demonstrated remarkable success in treating certain hematologic malignancies, including pediatric leukemia. The most extensively studied case in childhood patients involves CAR T cells that target CD19, a B cell surface receptor [4].CAR T cell therapy holds great promise for improving survival rates among pediatric cancer patients in Indonesia. Children with refractory or relapsed leukemia, such as B-cell acute lymphoblastic leukemia (B-ALL), who have exhausted standard treatment options, can benefit from CAR T cell therapy. Most relapsed or refractory pediatric cancer patients in Indonesia do not have effective therapy options to treat the disease. CAR T cell therapy emerges as a novel therapy that can significantly improve the survival of this subset of patients. Numerous studies have documented high remission rates (ranging from 70% to 90%) in adults and children diagnosed with refractory B-ALL [4]. A study by Maude et al. [5] reported high remission rates and durable responses in young adults and children with refractory or relapsed B-ALL treated with CAR T cells. Similarly, Park et al. [6] demonstrated long-term remissions and improved survival in pediatric leukemia patients receiving CAR T cell therapy. Several groups also have observed the persistence of CAR T cells and sustained remission lasting over six months in the majority of patients examined [4]. Efforts have been made to implement CAR T cells in Indonesia. Dharmais Cancer Hospital, as a National Cancer Center in Indonesia, has initiated this effort by collaborating with iCarTAB Biomed Inc., a China-based CAR T cell manufacturer with one of its manufacturing sites located in Malaysia. However, this approach involves sending patients' blood samples that have been processed through leukapheresis to Malaysia for CAR T cell manufacturing, followed by the shipment of the manufactured cells back to Indonesia for administration to patients. This process is impractical and incurs intangible costs such as transportation and cryopreservation, ultimately making it more expensive for patients. Regulatory issues related to the shipment of cells across borders in the region and early preparation of patients for CAR T cell therapy soon after relapse before they succumb to treatment-related mortality or relapse-related complications are also challenges that need to be addressed [7]. Reflecting on the abovementioned issue, CAR T cell therapy adoption in Indonesia faces significant challenges. Limited healthcare infrastructure, including specialized facilities for cell therapy manufacturing and administration, poses logistical hurdles. Moreover, cost remains a major barrier, as CAR T cell therapy is often expensive and inaccessible to many patients in Indonesia. Furthermore, the lack of local expertise in cellular immunotherapy may impede the successful implementation of CAR T cell therapy programs.Efforts to address these challenges and maximize the potential of CAR T cell therapy in Indonesia are essential. This requires a multi-faceted approach involving investment in healthcare infrastructure, including establishing specialized centers equipped for CAR T cell therapy manufacturing and administration. Two alternative models have been proposed for manufacturing CAR-T cell therapy: centralized and de-centralized models [8]. In the centralized manufacturing model, point of manufacturing and point of care are located in different geographical areas, while decentralized manufacturing focuses on establishing point of care and manufacturing in close proximity. A decentralized manufacturing model might be the best approach to be implemented in LMICs like Indonesia. Building hospital-based cellular therapy manufacturing reduces the need for transportation and cryopreservation. The decentralized system's geographic proximity improves communication between manufacturing and treatment teams, facilitating the creation of customized products based on a patient's phenotype. This setup also reduces administration time and the risk of delays and mix-ups compared to centralized manufacturing, making hospital-based cellular therapy manufacturing a potentially more cost-effective option [8].In addition, initiatives to reduce the cost of therapy through partnerships with pharmaceutical companies, government subsidies, or philanthropic endeavors can improve affordability and access. Furthermore, capacity-building initiatives aimed at training local healthcare professionals in cellular immunotherapy techniques are essential for ensuring the successful implementation and sustainability of CAR T cell therapy programs in Indonesia. Collaboration between local institutions, international organizations, and industry stakeholders can facilitate knowledge transfer and technology transfer, fostering indigenous expertise in this cutting-edge treatment modality.CAR T cell therapy represents a transformative approach to improving survival rates among pediatric cancer patients in Indonesia. By harnessing the power of immunotherapy, specifically tailored to target cancer cells, CAR T cell therapy offers hope for children with refractory or relapsed leukemia who have limited treatment options. Through continued research, collaboration, and investment in healthcare infrastructure, CAR T cell therapy potentially could greatly improve the prognosis and quality of life for pediatric cancer patients in Indonesia.

Chimeric antigen receptor (CAR) T-cell therapy has been shown to produce profound results in the treatment of certain hematologic malignancies, however treatment of solid tumors with CAR T cells has not been as successful. Studies have suggested that T-cell exhaustion plays a role in limiting the ability of CAR T cells to eradicate solid tumors. Additionally, stem-like qualities of T cells have been associated with better outcomes in patients treated with cellular therapies, including CAR T cells. Therefore, maintaining stem-like qualities and overcoming T-cell exhaustion may be key to improving clinical efficacy of CAR T cells in patients with solid tumors. ROR1 is a cell surface antigen expressed in several solid tumor types and chronic lymphocytic leukemia (CLL). ROR1 expression has been reported in 57% of triple-negative breast cancer (TNBC), as well as 42% of adenocarcinoma and 12% of squamous cell carcinoma subtypes of non-small cell lung cancer (NSCLC). These expression data of ROR1 in TNBC and NSCLC provide support for anti-ROR1 agents as a therapeutic strategy for these cancers. LYL797 is a novel, ROR1-targeted chimeric antigen receptor (CAR) T-cell product that incorporates genetic and epigenetic reprogramming technologies, Gen-R and Epi-R, to overcome barriers of CAR T-cell therapies in solid tumors. The ROR1-specific CAR contains a single-chain variable fragment (scFv) derived from an R12 rabbit monoclonal antibody that recognizes and binds with high specificity to human ROR1. Gen-R is ex vivo genetic reprogramming technology that engineers CAR T cells to overexpress c-Jun. Dysregulation of activator protein 1 (AP-1) has been implicated in CAR T-cell exhaustion, and studies have demonstrated that overexpression of c-Jun renders CAR T cells less susceptible to exhaustion, enhancing both anti-tumor efficacy and persistence in preclinical models of hematologic and solid tumors. Epi-R is a proprietary optimized manufacturing process that results in maintenance of stem-like phenotype and function of T-cell products. In preclinical studies LYL797 cells reprogrammed with Gen-R and Epi-R led to improved functional activity in the presence of ROR1+ tumor cells compared to conventional ROR1 CAR T cells. Additional studies are underway to determine the mechanisms by which antitumor activity of LYL797 in ROR1-positive solid tumor xenograft models is enhanced. LYL797 is anticipated to enter into Phase 1 clinical trials for TNBC and NSCLC in 2022. Citation Format: Spencer Park, Courtney Simianer, Sydney Spadinger, Xiao Wang, Purnima Sundar, Shobha Potluri, Rachel Lynn, Bijan Boldajipour, Grace Wang, Neeraj Sharma, Hajime Hiraragi, Veena Krishnamoorthy, Suman Kumar Vodnala, E-Ching Ong, Chang-Chih Wu, Martin Wohlfahrt, Byoung Ryu, Lisa Song, Brian D. Weitzner, Howell Moffett, Marc Lajoie, Scott Boyken, Tamer Shabaneh, Shivani Srivastava, Tina Albertson, Blythe Sather. LYL797, a ROR1 CAR T-cell therapy with genetic and epigenetic reprogramming for solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2754.

Development and Evaluation of an Optimal Human Single-Chain Variable Fragment-Derived BCMA-Targeted CAR T Cell Vector.

DAP10 integration in CAR-T cells enhances the killing of heterogeneous tumors by harnessing endogenous NKG2D

CAR T-cell therapy for solid tumours

217 Background: Neuroendocrine prostate cancers (NEPC) currently have no effective therapies. While CAR (Chimeric Antigen Receptor) T cell therapies targeting PSMA and PSCA receptors are currently in phase 1/2 studies in metastatic castrate-resistant prostate cancer (mCRPC), antigen targets for NEPC remain limited. Lewis Y antigen (LeY) is an oncofetal antigen expressed only in adult tumour cells and is considered a target for CAR T cell therapy in haematological, lung, ovarian and colon cancer. We previously found that LeY is a promising target for CAR T cell therapy in prostate cancer and that preconditioning LeY-directed CAR T cells with carboplatin further enhances tumour responses in mCRPC. We investigated if LeY can be used as a novel CAR T cell target for NEPC. Methods: The expression of the Lewis Y antigen was assessed in our patient-derived prostate cancer xenograft collection using immunohistochemistry (IHC). To assess the efficacy of LeY-CAR T cells, we used specimens established from a patient (Patient 508) with NEPC who had incurable metastatic disease and failed standard treatments. We generated CAR T cells from his peripheral blood mononuclear cells (PBMCs) to test their specificity and efficacy in vitro against established cell lines. Results: In our MURAL PDX collection of 59 prostate tumours, 81% (9/11) NEPCs expressed LeY antigen on IHC. Next, we successfully produced anti-LeY CAR T cells from the patient’s PBMCs using retroviral transduction. Flow cytometry verified efficient transduction of the CAR constructs. Previous studies found that a 1:1 ratio of CD4 + :CD8 + T cells improved outcomes following CAR T cell therapy 5,6 , so we further phenotyped patient T cells by flow cytometry. The CD4 + :CD8 + T cell ratio was nearly 1:1 (51.7% and 41.4% of total T cells, respectively). Next, we used an immunobead assay to investigate CAR-T specificity in releasing proinflammatory cytokines. Significant cytokine production was observed after a 4-hour co-culture with LeY+ human ovarian cancer cell line, OVCAR3, vs co-culture with control LeY- MDA-MB435 cells (p=0.0004), demonstrating the specificity of the CAR T cells.Lastly, we showed that patient CAR T cells effectively killed LeY+ cells (OVCAR3), but not control LeY- cells (MDA-MB435; p<0.0001). Tumour cell death was measured by the amount of released chromium following 16-hour co-culture. CAR T cells killed LeY+ target cells more effectively than their non-transduced equivalents at a range of effector-to-target ratios, with up to 15-fold increase in cell death. Conclusions: Our preclinical studies show that most NEPC tumors express the LeY antigen and that patient-derived CAR T cells specifically kill those cells. These findings identified a potential CAR T treatment target for NEPC for a phase 1/2 study.

Decision letter: Chimeric antigen receptors that trigger phagocytosis

B7-H3 as a Novel CAR-T Therapeutic Target for Glioblastoma

CRG-023 Is a Novel Tri-Specific CAR T Product Candidate Engineered to Prevent Antigen Escape and Sustain Durable Anti-Tumor Functionality Against B-Cell Malignancies

While chimeric antigen receptor T (CART) cell therapy has shown remarkable success, the development of exhaustion limits durable response. We identified a role for interleukin (IL)-4 in the development of CART cell exhaustion through three independent approaches including: 1) a genome-wide CRISPR knockout screen using healthy donor CART cells in an in vitro model for exhaustion, 2) RNA and ATAC sequencing on freshly produced and chronically stimulated healthy donor CART cells, and 3) RNA and ATAC sequencing on pre-infusion CART cell products from responders and non-responders in the Zuma-1 clinical trial that led to the FDA approval of axi-cel. Further, in vitro validation studies revealed that CD19 directed CART (CART19) cells chronically stimulated in the presence of human recombinant IL-4 (hrIL-4) displayed signs of exhaustion such as 1) decreased proliferation (p = 0.01), 2) increased coexpression of inhibitory receptors (p = 0.01), and 3) decreased production of IL-2 and interferon (IFN)-γ (p= 0.02, p = 0.002). Encouragingly, CART19 cells combined with an IL-4 monoclonal antibody improved antitumor activity (p = 0.045) and expansion (p = 0.01) while also decreasing the co-expression of inhibitory receptors (p = 0.02) in a mantle cell lymphoma xenograft mouse model. Building on these results, we asked if IL-4 driven exhaustion results from a direct impact of IL-4 on CART cells. To test this, we used a tumor-free assay where CART19 cells were chronically stimulated with CD19 beads in the presence of hrIL-4 or diluent. CART cells treated with hrIL-4 displayed an exhausted phenotype characterized by increased co-expression of inhibitory receptors (p = 0.04) and decreased production of IL-2 (p = 0.01). Next, we asked if IL-4 driven CART cell exhaustion is dependent on the costimulatory domain. We tested the impact of IL-4 on both CD28ζ and 41BBζ costimulated CART19 cells. Similar to our previous studies with CART19-28ζ cells, chronic stimulation of CART19-BBζ cells in the presence of hrIL-4 enhanced the exhausted phenotype as seen by increased co-expression of inhibitory receptors (p = 0.04) and decreased production of IL-2 and IFN-γ (p = 0.08 and p = 0.007). Finally, we asked if IL-4 induces exhaustion independently of its classic role in Th2 polarization of CD4 CART cells. Following CART production, we isolated CD8 cells and chronically stimulated them in the presence of hrIL-4 or diluent. CD8 CART cells treated with hrIL-4 displayed an enhanced exhausted profile as seen by 1) decreased proliferative ability (p < 0.0001), 2) increased co-expression of inhibitory receptors (p = 0.01), and 3) decreased production of IL-2 and IFN-γ (p < 0.0001, p = 0.004). Together, our data indicates a novel role for IL-4 in the development of CART cell exhaustion that is independent of tumor cells, costimulatory domain, and CD4 cells. As such, we believe IL-4 neutralization may be a widely applicable and actionable approach to improve the durable response to CART cell therapy. Citation Format: Carli M. Stewart, Elizabeth L. Siegler, Truc N. Huynh, R. Leo Sakemura, Brooke Kimball, Long Mai, Kun Yun, James H. Girsch, Jennifer Feigin, Omar Gutierrez Ruiz, Makena Rodriguez, Ekene Ogbodo, Ismail Can, Claudia Manriquez Roman, Olivia Sirpilla, Hong Xia, Jenny Kim, Justin Budka, Mike Mattie, Nathalie Scholler, Simone Filosto, Saad S. Kenderian. IL-4 drives CART cell exhaustion in a CD4 independent manner [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 37.