bims-carter 2025-10-26 papers

bims-carter

Biomed News

on CAR-T Therapies

Issue of 2025–10–26
forty-one papers selected by
Luca Bolliger, lxBio

Genome-edited allogeneic CAR-T cells: the next generation of cancer immunotherapies.
Lipid nanoparticles for engineering next generation CAR T cell immunotherapy.
Chimeric switch and inverted cytokine receptors in T cell therapy: reprogramming T cells to overcome immune suppression in the solid tumor microenvironment.
The potential of CAR T cells in autoimmune diseases.
Associations Between Pre-Existing Autoimmunity and Chimeric Antigen Receptor T Cell Therapy Toxicity for Cancer Treatment.
Development of antigen multimers for detection and evaluation of CAR T cells.
Preventing secondary primary malignancies (SPMs) in CAR-T cell therapy through site-specific transgene integration into genomic safe harbors (GSHs).
Infectious Complications of Chimeric Antigen Receptor T Cell Therapy and Bispecific Antibodies - A Review.
Current Role of CAR-T Therapy in Haematological Care.
Commercialization of Cell Therapies: A CDMO Perspective.
Engineered natural killer cells for cancer therapy.
Innovation to Integration: The Translation of hPSC based cell therapies from Academic Research to Industry Standards.
CCR5-targeted allogeneic gamma-delta CD19 chimeric antigen receptor T cells for HIV-associated B cell-malignancy immunotherapy.
Chimeric antigen receptor cells as a tool for localized delivery of TNFα in solid cancer treatment.
Production of functional CD19 CAR T cells under hypoxic manufacturing conditions.
Chimeric Antigen Receptor-Engineered Natural Killer (CAR-NK) Cell Therapy in Acute Myeloid Leukemia: A Systematic Review of the Literature.
CAR NK cell therapy for solid tumors: potential and challenges.
Challenges in the Patent and Legal Environment for Cell Therapies.
GMP Manufacturing of Allogenic iPSC-NK Cells for Immunotherapy.
Autologous stem cell transplantation meets CAR-T therapy: A synergistic strategy for B-cell lymphoma.
CAR-T and CAR-NK cell therapies in AML: breaking barriers and charting the future.
Engineering T cells with a membrane-tethered version of SLP-76 overcomes antigen-low resistance to CAR T cell therapy.
Raw Material Considerations for the Manufacture of Cell-Based Therapies.
Nanobody-enhanced T cell cancer immunotherapy: Advancing precision medicine.
Emerging Plant-Derived Exosome-Like Particles Reveal Key Therapeutic Benefits. A Comprehensive Review of Evidence.
Barriers to Effective Cryopreservation of Cell Therapies: Challenges and Emerging Solution.
Emerging macrophage-based therapies for cancer: a review of preclinical and clinical advances.
The Developing Standards Landscape for Cell and Gene Therapies.
Decoding signaling architectures: CAR versus TCR dynamics in solid tumor immunotherapy.
Acquisition of an immunosuppressive microenvironment after anti-CD19 CAR T-cell therapy is associated with T-cell dysfunction and resistance.
Peripheral T cell lymphoma following CD19-targeted chimeric antigen receptor T cell therapy.
Generation of the augmented IL-15-secreting anti-HER2 chimeric antigen receptor (CAR)-NK cells: an encouraging immunotherapeutic tool.
Development of Automated Systems for Cell Therapy.
Neoantigen identification and TCR-T therapy development for solid tumors: current advances and future perspectives.
Development of Stem Cell Data Systems and Associated Data Standards for Cell Therapy.
Cold Chain Delivery in Cell Therapies.
Advancing a US Single-Payer Model for Cell and Gene Therapy.
Standardised Matrices for Stem Cells.
The French Experience of Pharmacists and CAR T-Cells: A Study of the French Society of Oncology Pharmacy (SFPO).
Co-expression of an adapter CAR retains efficacy of CAR T cells after single and dual antigen loss in lymphoma.
Facts and Hopes of Chimeric Antigen Receptor-Redirected Natural Killer T cells.

J Hematol Oncol. 2025 Oct 24. 18(1): 90

Genome-edited allogeneic CAR-T cells: the next generation of cancer immunotherapies.

Jingchao Su, Yifei Zeng, Zhuojin Song, Yinglu Liu, Kaixin Ou, Yuhan Wu, Minhong Huang, Yuhua Li, Sanfang Tu.

  Chimeric Antigen Receptor T (CAR-T) cell therapy has revolutionized cancer immunotherapy, particularly in hematological malignancies. However, the clinical application of autologous CAR-T cells faces significant high cost and manufacturing challenges. Universal allogeneic CAR-T cells, derived from healthy donors, represent a promising solution to these obstacles. These "off-the-shelf" therapies aim to reduce the complexity and cost of CAR-T production. Despite exciting advancements in genome-editing technologies and promising clinical trial data, significant challenges remain, including graft-versus-host disease (GVHD), Host-versus-graft reaction (HVGR), off-target effects, genotoxicity, and manufacturing scalability. To address these concerns, genome-editing technologies such as ZFNs, TALENs, Meganucleases, CRISPR systems, base editing, and prime editing are being employed. This review summarizes the progress of universal allogeneic CAR-T cell therapies, addresses the critical challenges, and discusses the future directions for their clinical implementation.

Keywords:  Allogeneic CAR-T cells; Chimeric antigen receptor (CAR); Gene editing; Graft-versus-host disease (GVHD); Haematological malignancies; Host-versus-graft reaction (HVGR); Solid tumours; T cell receptor; Universal CAR-T cells

DOI:  https://doi.org/10.1186/s13045-025-01745-8
Nanoscale Horiz. 2025 Oct 20.

Lipid nanoparticles for engineering next generation CAR T cell immunotherapy.

Melgious Jin Yan Ang, Ann E Metzloff, Ajay S Thatte, Michael J Mitchell.

Lipid nanoparticles are a burgeoning technology which has vast potential to improve chimeric antigen receptor (CAR) T cell immunotherapy. This focused review provides an overview of CAR T cell therapy - highlighting its promises, limitations, and challenges - and describes ways in which lipid nanoparticles (LNPs) can be rationally designed to circumvent some of the challenges. Of particular note are antigen presenting cell-mimetic LNPs, which have the potential to streamline the CAR T cell production process by activating T cells and delivering the CAR transgene in a single step. Although the current clinical standard is ex vivo CAR T cell production, in vivo CAR T cell production represents a potentially transformative alternative. Recent innovations in each production method are described, with a particular emphasis on ways in which LNPs may enable in vivo CAR T cell production. The review concludes with a discussion of safety, immunogenicity, scalability, manufacturing, and regulatory factors which will be essential as LNP-based CAR T cell immunotherapies move toward clinical translation.

DOI: https://doi.org/10.1039/d5nh00432b
Front Immunol. 2025 ;16 1662238

Chimeric switch and inverted cytokine receptors in T cell therapy: reprogramming T cells to overcome immune suppression in the solid tumor microenvironment.

Riley Rane, Fengqiao Li, Alexis Williams, Avaneesh Jayadev, Nhan L Tran, Jeffrey A Winkles, Gloria B Kim.

  Adoptive T cell therapy has transformed cancer treatment, with chimeric antigen receptor (CAR) T cell therapy demonstrating remarkable clinical success in hematological malignancies. By genetically engineering a patient's own T cells to recognize and attack cancer cells, CAR T therapy has achieved durable remissions in several blood cancers. However, its efficacy in solid tumors remains limited, largely due to the immunosuppressive tumor microenvironment (TME), which impairs T cell infiltration, persistence, and function. To address these challenges, innovative strategies are being developed to reprogram T cell signaling within the hostile TME. One promising class involves chimeric non-antigen receptors (CNARs), which modulate T cell activity independently of direct antigen recognition. Among these, chimeric switch receptors (CSRs) convert inhibitory checkpoint signals into activating cues, while inverted cytokine receptors (ICRs) redirect suppressive cytokine signals to promote T cell activation. In this review, we provide a focused overview of the design principles, mechanistic functions, and therapeutic potentials of CSRs and ICRs as adjuncts to CAR T therapy in solid tumors. We also discuss key considerations regarding safety, specificity, and clinical translation to inform future advancements in engineered receptor strategies for cancer immunotherapy.

Keywords:  CAR-T; chimeric switch receptors; immune suppression; immunotherapy; inverted cytokine receptors; solid cancers; synthetic biology

DOI:  https://doi.org/10.3389/fimmu.2025.1662238
Immunotherapy. 2025 Oct 23. 1-16

The potential of CAR T cells in autoimmune diseases.

Johanna Richter, Martin Krusche, Isabell Haase, Ina Kötter, Nicolaus Kröger.

  CAR T cell therapy represents an emerging, promising approach in the field of autoimmune diseases. Unlike monoclonal antibodies, CAR T cells offer a deeper B cell depletion than monoclonal antibodies through their ability to infiltrate tissues and their independence from effector mechanisms. Early data from case reports and phase I/II studies demonstrate encouraging clinical responses in patients with severely refractory autoimmune disorders. CD19, BCMA and dual-targeting CAR constructs were generally well tolerated with limited safety concerns. This promising data encourages further research investigating CAR T cell therapy in a broader range of autoimmune indications and to advance CAR T constructs to improve efficacy, safety, and clinical application. In this review, we explore the rationale of using CAR T cell therapy in autoimmune diseases, summarize relevant clinical data, and highlight future perspectives within the field.

Keywords:  CAR T cell therapy; Chimeric antigen receptor T cell; autoimmune diseases; autoreactive B cells; cell therapy

DOI:  https://doi.org/10.1080/1750743X.2025.2558348
ACR Open Rheumatol. 2025 Oct;7(10): e70112

Associations Between Pre-Existing Autoimmunity and Chimeric Antigen Receptor T Cell Therapy Toxicity for Cancer Treatment.

Gregory J Challener, Chen Wang, David Gritsch, Ilana M Usiskin, Zandra E Walton, Minna J Kohler, Hyon Choi, Jeffrey A Sparks.

OBJECTIVE: Chimeric antigen receptor (CAR) T cell therapy is being investigated to treat individuals with autoimmune diseases. Data regarding the safety of CAR T cell therapy in this population are limited. We aimed to assess whether pre-existing autoimmune disease was associated with an increase in major toxicity or hospital length of stay (LOS) among patients receiving CAR T cell therapy for the treatment of hematologic cancer.
METHODS: This retrospective cohort study used data from the National Inpatient Sample (2021-2022) to investigate in-hospital outcomes in adult patients receiving CAR T cell therapy. Comparisons were drawn between patients with and without pre-existing autoimmune disease. Multivariable regression models with adjustment for age, sex, race, cancer type, and comorbidities assessed associations with hospital LOS, major toxicity, and inpatient mortality.
RESULTS: Among 1,321 patients receiving CAR T cell therapy, 62 (4.7%) had diagnosed autoimmune disease. Patients with pre-existing autoimmunity had a significantly shorter hospital stay compared with patients without, with a mean reduction of 2.1 days (95% confidence interval [CI] 0.5-3.6). Major toxicity was less frequent in patients with pre-existing autoimmunity than patients without (67.7% vs 78.6%, respectively; adjusted odds ratio 0.55; 95% CI 0.31-0.99). Inpatient mortality did not differ significantly between groups.
CONCLUSION: Patients with pre-existing autoimmune disease had shorter hospital stays and lower risk of major toxicity following CAR T cell therapy for cancer treatment, offering reassurance as to its safety in this population while providing additional data for research studies of CAR T cell therapy to treat autoimmune diseases.

DOI: https://doi.org/10.1002/acr2.70112
Cancer Immunol Immunother. 2025 Oct 23. 74(11): 344

Development of antigen multimers for detection and evaluation of CAR T cells.

Rasmus U W Friis, Maria Ormhøj, Cecilie S Krüger-Jensen, Carlos Rodriguez Pardo, Hólmfridur Rósa Halldórsdóttir, Markus Barden, Nikolaj P Kristensen, Keerthana Ramanathan, Mikkel R Hansen, Hinrich Abken, Sine R Hadrup.

   BACKGROUND: Chimeric antigen receptor (CAR) T cell therapy has transformed the treatment landscape of hematologic cancers by engineering T cells to specifically target and destroy cancer cells. Monitoring CAR T cell activity and function is essential for optimizing therapeutic outcomes, but existing tools for CAR detection are often limited in specificity and functional assessment capability.
METHODS: We developed dextran multimers by conjugating multiple CAR-specific antigens to a dextran backbone. The multimers were compared to previously reported antigen tetramers for their ability to stain and detect CAR T cells. Because these multimers incorporate the CAR target antigen, they uniquely enable assessment of CAR T cell functionality. We tested the staining and functional properties of the multimers across a range of CAR constructs with different affinities, using flow cytometry and microscopy.
RESULTS: The dextran multimers demonstrated high specificity and sensitivity in staining CAR T cells, with adjustable antigen density to optimize binding. Dextran multimers also enabled effective clustering and subsequent activation of CARs, showing their utility as both a staining and functional assessment tool. The multimers revealed that CARs with different affinities and clustering tendencies displayed varied binding and activation in response to different antigen densities.
CONCLUSION: Dextran multimers offer a dual advantage as versatile reagents for both staining and functional analysis of CAR T cells. Their capacity to engage CARs with the specific antigen provides a valuable platform for evaluating CAR functionality, informing CAR design improvements, and enhancing therapeutic precision.

Keywords:  Affinity; Antigen multimer; Chimeric antigen receptor; Clustering; Detection; Dextran; Flow cytometry; Microscopy.; Stimulation; Tetramer

DOI:  https://doi.org/10.1007/s00262-025-04134-9
J Transl Med. 2025 Oct 21. 23(1): 1155

Preventing secondary primary malignancies (SPMs) in CAR-T cell therapy through site-specific transgene integration into genomic safe harbors (GSHs).

Pooria Safarzadeh Kozani, Pouya Safarzadeh Kozani.

  Chimeric antigen receptor (CAR)-T cell therapy has revolutionized oncology by achieving durable remissions in refractory hematologic malignancies. However, emerging reports link this therapy to second primary malignancies, including CAR+ lymphomas and leukemias, driven by insertional mutagenesis from semi-random viral vector integration near oncogenes or tumor suppressor loci. These rare but serious complications underscore the dual challenge of eradicating primary tumors while mitigating delayed genotoxic risks. Conventional CAR-T cell manufacturing, reliant on gamma-retroviral or lentiviral vectors, introduces genomic instability through integration into fragile sites or transcriptionally active regions. CRISPR/Cas9-mediated genome editing further amplifies risks via off-target double-strand breaks and chromosomal rearrangements. This review evaluates genomic safe harbors (GSHs)-such as AAVS1, TRAC, CCR5, ROSA26 and CLYBL-as loci validated for stable, high-level CAR transgene expression without oncogenic disruption. GSHs meet stringent criteria: distal from cancer-related genes, resistant to epigenetic silencing, and transcriptionally permissive. Preclinical studies demonstrate that site-directed CAR integration into GSHs preserves antitumor efficacy while eliminating malignant transformation risks. Challenges persist in optimizing homology-directed repair efficiency, mitigating residual dsDNA toxicity, and standardizing regulatory frameworks for long-term genomic surveillance. Emerging technologies-base/prime editing, hybrid nucleases, and rigorous monitoring-promise enhanced precision and safety. By reconciling therapeutic innovation with genomic integrity, GSH-engineered CAR-T cells herald a paradigm shift toward precision immunotherapies, offering curative potential while preempting secondary oncogenesis. Collaborative efforts to refine manufacturing, harmonize global standards, and prioritize patient-specific risk stratification will be critical to advancing this transformative approach.

Keywords:  Cancer immunotherapy; Chimeric antigen receptor; Genomic safe harbors; Insertional mutagenesis; Secondary primary malignancies

DOI:  https://doi.org/10.1186/s12967-025-07183-x
Am J Med Sci. 2025 Oct 18. pii: S0002-9629(25)01249-2. [Epub ahead of print]

Infectious Complications of Chimeric Antigen Receptor T Cell Therapy and Bispecific Antibodies - A Review.

Bradley Dye, Daniel Gromer, Bibi Maryam, Emily A Siegrist, Joseph Sassine.

  Chimeric antigen receptor (CAR) T cell therapy has shifted the therapeutic landscape of hematological malignancies in the last decade, and bispecific antibodies (BsAb) are an emerging treatment for this patient population. Immunosuppression associated with these therapies can lead to a variety of infectious complications. In this narrative review, we explore the epidemiology, risk factors and prevention of bacterial, viral and fungal infections after CAR T cell and BsAb therapy.

Keywords:  CAR T cell; bispecific antibody; infection

DOI:  https://doi.org/10.1016/j.amjms.2025.10.007
Adv Exp Med Biol. 2025 ;1486 193-216

Current Role of CAR-T Therapy in Haematological Care.

Miroslava Jandová, Miriam Lánská, Alice Sýkorová, Jiří Gregor, Petra Rozsívalová, Lenka Beková, Zuzana Woidigová Ducháčová, Jakub Radocha, Glyn N Stacey, Pavel Měřička, Barry Fuller.

  CAR-T therapy based on the genetic modification of T-lymphocyte receptors is currently a rapidly expanding modern method of treatment of hematological malignancies. In this chapter, the authors review the history of the development of this kind of therapy, principles of the CAR-T therapy product manufacturing, and indications for the use of registered CAR-T therapy products. They also review the perspectives for the expansion of indications using this treatment on the basis of analysis of approved clinical trials. The need for the establishment of effective logistics pathways and the benefits of cryopreservation at different manufacturing steps are reviewed as well. During their 2-year experience, the authors established a system of fluent cooperation between the local licensed Tissue Establishment (TE), CAR-T therapy product manufacturers, the Hospital Pharmacy, and a certified clinical CAR-T therapy centre. The leukapheresis of starting material, its processing, storage, and release for the manufacture took place in the authorized TE. The starting material is usually fresh mature peripheral blood mononuclear cell concentrate, which is sent to the manufacturing site in a chilled (or sometimes cryopreserved) state and the final registered product is sent back in a frozen state. Individual manufacturers use different cold chains. Sometimes the starting material is frozen by the manufacturer and the final manufacture is carried out before the actual administration to the patient as a fresh suspension. However, the most common variant used in registered products is the supply of the final product to the place of the administration in the cryopreserved state.Receipt of final products in the hospital cryobank attached to the TE takes place in cooperation between the TE staff with representatives of the Hospital Pharmacy and is followed by storage in a vapour phase of liquid nitrogen in a separate GMP-compliant container used exclusively for the storage of registered and investigational CAR-T therapy products at a temperature below -150 °C. Before the transport to the Clinical Department, the chain of recipient identity is checked, then the product is transported to the Clinical Department in a dry shipper at temperature below -150 °C. After the second check in the presence of the clinical hematologists, the product is thawed and immediately infused. In the case of the use of investigational products, special attention is paid to meeting specific genetic safety rules in the regimen of genetically modified organisms.

Keywords:  CD-19; Cell expansion; Cell processing; Chimeric antigen receptor; Cryopreservation; Cytokine release syndrome; Cytokines; Gene editing; Good Manufacturing Practice; Immunotherapy; Quality control; Regulatory compliance; Sterility assurance; T-cell isolation; Viral vector

DOI:  https://doi.org/10.1007/978-3-031-97297-3_16
Adv Exp Med Biol. 2025 ;1486 137-145

Commercialization of Cell Therapies: A CDMO Perspective.

Behnam Ahmadian Baghbaderani.

  Cell and gene therapies (CGT) hold tremendous curative potential for the treatment of devastating diseases, such as Parkinson's disease, Type I diabetes, and various types of cancer. Besides demonstrating the safety and efficacy of these therapies, it is important to address several challenges in early development and clinical manufacturing to commercialize cell and gene therapies and meet the demand to treat many patients. The source and intensity of challenges can vary as these therapies move from early clinical to late-stage clinical and commercialization readiness. Here, we highlight some of the main challenges the CGT industry faces from the contract development and manufacturing organization (CDMO) perspective and some of the bioprocessing best practices required to de-risk these challenges in the path to commercial launch.The key to commercializing cell and gene therapies is to establish a step-by-step, phase-appropriate, risk-assessment-based approach to meet the product lifecycle requirements from early development to clinical and late-stage commercial readiness campaigns. In this journey, it would be critical to cover key aspects of chemistry, manufacturing, and controls (CMC), including materials, manufacturing process, analytics, sterility assurance, equipment, and technology, as well as tissue acquisition, depending on the type of application. Appropriate process improvement studies may be needed, including implementing innovative technologies with proper in-process control and monitoring analytical methods to ensure the robustness and reproducibility of the manufacturing process. In particular, it may be necessary to eliminate open and variable manual unit operations while incorporating comprehensive characterization assays to verify the long-term stability of the products. Implementing an automated, scalable, computer-controlled 3D bioreactor in the process, with appropriate downstream and cell-processing technologies, would be critical to minimizing commercialization risks and producing high-quality cell therapy products. Finally, in preparation for commercial launch, the commercialization readiness campaign should be based on a Failure Mode and Effects Analysis (FMEA) to identify modes of failure, risks, and mitigations and design appropriate process characterization studies prior to process validation.

Keywords:  Automation; Bioreactor; Cell and gene therapies; Current good manufacturing practice; Process analytics; Process characterization

DOI:  https://doi.org/10.1007/978-3-031-97297-3_11
Cancer Cell. 2025 Oct 23. pii: S1535-6108(25)00405-2. [Epub ahead of print]

Engineered natural killer cells for cancer therapy.

Alexander Biederstädt, Katayoun Rezvani.

  Allogeneic natural killer (NK) cell immunotherapy is emerging as a promising and scalable, off-the-shelf platform for treating relapsed and refractory cancers. Early-phase clinical trials have demonstrated remarkable safety and encouraging therapeutic efficacy of chimeric antigen receptor (CAR)-NK cells in heavily pretreated patients with lymphoid malignancies. Current efforts are expanding these therapies to solid tumors, with translational research increasingly leveraging precision gene editing to enhance effector function, persistence, and resistance to the immunosuppressive tumor microenvironment. In this review, we summarize findings from early-phase clinical trials and discuss emerging synthetic biology and engineering approaches to improve NK cell potency. We also highlight advances in high-throughput discovery platforms that have identified actionable gene targets for NK cell reprogramming, offering a path to design multi-engineered CAR-NK cells to overcome the challenges of solid tumors. Together, these translational innovations define the trajectory of next-generation NK cell therapies and their integration into the broader cancer immunotherapy landscape.

Keywords:  CRISPR screening; adoptive cell therapy; cancer immunotherapy; cellular engineering; chimeric antigen receptor; natural killer cells; perturbomics; precision gene editing; solid tumors; tumor microenvironment

DOI:  https://doi.org/10.1016/j.ccell.2025.09.013
Adv Exp Med Biol. 2025 ;1486 147-156

Innovation to Integration: The Translation of hPSC based cell therapies from Academic Research to Industry Standards.

Ida Kjær, Maria Rathmann Sørensen, Sofia Håkansson Buch, Arvind Pradip.

  The trajectory from breakthrough discovery to scalable cell therapy can be as daunting as it is thrilling. While the cell therapy field is rapidly developing, early scientific research needs to be rapidly adapted and integrated into CMC development and subsequently commercial manufacturing. During the transfer of promising new therapies from academia researchers into pharmaceutical industry partners often face a huge conundrum: materials and technologies transfer from laboratory setting to manufacturer for clinical trials or marketing applications often are challenged by regulatory agencies based on their country or regional pharmaceutical regulations. However, this pathway is navigable with the right guidance. This chapter provides recommendations and discusses strategies on starting materials, raw materials, process development, and adequate documentation of early development activities aiming to aid researchers in refining and adapting the product development that meets with country or regional regulations and assures patient safety. This is needed not only to obtain early-stage regulatory approval but also to achieve consistent product quality and efficiency of the cell therapy and set the stage for the transition into late-stage development and manufacturing.

Keywords:  CMC; Cell therapy; Documentation; Process development; Raw material

DOI:  https://doi.org/10.1007/978-3-031-97297-3_12
Nat Biomed Eng. 2025 Oct 21.

CCR5-targeted allogeneic gamma-delta CD19 chimeric antigen receptor T cells for HIV-associated B cell-malignancy immunotherapy.

Ángel Ramírez-Fernández, Alexander J Dimitri, Fang Chen, Robert Bartoszek, Gregory M Chen, Laura Córdoba-Espejo, Yuqi Zhou, Yun-Hsin Tang, Chien-Ting Lin, Reyes Acosta, John Scholler, Guido Ghilardi, Patrizia Porazzi, Mireia Pellicer, Núria Profitós-Pelejà, Stefan K Barta, Anne Chew, Julie K Jadlowsky, Vanessa E Gonzalez, Donald L Siegel, Bruce L Levine, Gaël Roué, Marco Ruella, Michael T Lotze, Carl H June, James L Riley, Joseph A Fraietta.

Immune-based cell therapy offers a promising approach to cancer treatment. While autologous chimeric antigen receptor (CAR) T cells have shown success, production is time-consuming, costly and patient specific. Gamma-delta (γδ) T cells are promising for 'off-the-shelf' CAR T cell therapy. However, clinical translation of γδ CAR T cells is hampered by low frequency, resistance to genetic manipulation and advanced differentiation after expansion, limiting therapeutic feasibility. Here we demonstrate a method for in vitro activation and expansion of peripheral blood γδ T cells, facilitating high rates of gene editing and efficient CAR integration. Using artificial antigen-presenting cells, we produce minimally differentiated, highly functional γδ CAR T cells. By targeting a US Food and Drug Administration-approved CD19 CAR to the CCR5 locus, we generate CCR5-deficient γδ CD19 CAR T cells (γδ CCR5KI-CAR19), which demonstrated resistance to HIV-mediated depletion and robust antitumour responses against B cell lymphoma and leukaemia. γδ CCR5KI-CAR19 T cells enable the immunotherapy of HIV-associated B cell malignancies. These studies provide preclinical evidence supporting large-scale development of potent allogeneic γδ CAR T cells for diverse immunotherapies.

DOI: https://doi.org/10.1038/s41551-025-01527-0
Hum Vaccin Immunother. 2025 Dec;21(1): 2576262

Chimeric antigen receptor cells as a tool for localized delivery of TNFα in solid cancer treatment.

Zhenqiang Yao, Chutamath Sittplangkoon.

  Chimeric antigen receptor (CAR)-T cells are effective in treating blood cancers but not solid cancers and can cause severe side effects, including cytokine release syndrome (CRS). One strategy to enhance the efficacy of CAR-T cells while avoiding CRS in cancer treatment is to deliver a single tumoricidal factor. TNFα is well known for triggering apoptosis signaling; however, it alone is not effective in treating cancers because it significantly increases the levels of inhibitor of apoptosis proteins (IAPs), a family of E3 ubiquitin ligases that block caspase-induced apoptosis. Thus, localized delivery of TNFα by targeting the tumors using the adoptive cells combined with an IAP antagonist, which degrades IAP proteins, could lead to improved outcomes in cancer treatment. This article reviews TNFα-induced apoptosis signaling pathway, outlines the principles for designing CAR-T cells, CAR-macrophages and CAR-dendritic cells expressing TNFα, used alone or in combination with IAP antagonists, and discusses the potential contraindications of IAP antagonists with several clinical used drugs for cancer treatment.

Keywords:  Cancer; SMAC memetic (IAP antagonist); T cell; chimeric antigen receptor (CAR) cell therapy; dendritic cell (DC); inhibitor of apoptosis proteins (IAPs); macrophage; tumor necrosis factor-α (TNFα)

DOI:  https://doi.org/10.1080/21645515.2025.2576262
Front Immunol. 2025 ;16 1675786

Production of functional CD19 CAR T cells under hypoxic manufacturing conditions.

Isabella Micallef Nilsson, Thomas Poiret, Jinhye Ryu, Mohammadali Mohammadpour, Johan Henriksson, Anders Österborg, Jonas Mattsson, Anna Schurich, Isabelle Magalhaes.

   Introduction: Chronic lymphocytic leukemia (CLL) has proven difficult to treat with chimeric antigen receptor (CAR) T cell therapy. CLL cells can negatively alter T cell fitness and induce a pseudohypoxic state. We hypothesized that production of CAR T cells under restricted oxygen conditions resembling physiological oxygen levels that can be encountered in tissues (i.e. 2% O2) could promote outgrowth of hypoxia-tolerant CAR T cells.
Methods: We performed in vitro phenotypic and functional assessments of CD19-directed CAR T cells produced in either 21% (NorCAR) or 2% (HypCAR) O2 derived from healthy donors (HDs) or patients with CLL.
Results: Production of HD-derived CAR T cells in 2% O2 promoted the enrichment of a naïve-like subset. HypCAR and NorCAR cells were functionally distinct; CD4+ HypCAR cells produced more IL-2 and tumor necrosis factor than CD4+ NorCAR cells. Production in 2% O2 was not detrimental to viability or proliferation upon cognate antigen-stimulation and led to increased activation. After chronic stimulation in hypoxia, HypCAR-product remained enriched in naïve-like cells, and demonstrated cytotoxic and cytokine production capacity. In CAR T cells derived from patients with CLL, NorCAR and HypCAR subsets were functionally and phenotypically comparable, but displayed different mitochondrial metabolism.
Discussion: We demonstrated that production in 2% O2 is not detrimental, confers subtle but lasting functional and phenotypic changes in CAR T cells warranting further research on the impact of hypoxic production on CAR T cell functionality in hypoxic tumor microenvironments.

Keywords:  CAR T cells; CLL (chronic lymphocytic leukemia); hypoxia; mitochondria; tumor microenvironment

DOI:  https://doi.org/10.3389/fimmu.2025.1675786
Cureus. 2025 Oct;17(10): e94807

Chimeric Antigen Receptor-Engineered Natural Killer (CAR-NK) Cell Therapy in Acute Myeloid Leukemia: A Systematic Review of the Literature.

Mayra Steffania Novoa Arias.

  Chimeric antigen receptor (CAR)-engineered natural killer (NK) cells are attracting considerable interest as a potential therapeutic option in acute myeloid leukemia (AML), driven by the search for safe, scalable, and effective alternatives. To provide an overview of the current state of the field, a systematic review was performed to evaluate the safety and efficacy of CAR-NK cells for AML. The findings suggest possible directions for future development and also highlight important obstacles that must be addressed before they can be used in clinical settings. Therefore, although CAR-NK therapy remains in its early stages, its eventual application in AML is possible if these barriers can be resolved.

Keywords:  acute myeloid leukemia; car; chimeric antigen receptor; immunotherapy; natural killer cells

DOI:  https://doi.org/10.7759/cureus.94807
Antib Ther. 2025 Oct;8(4): 275-289

CAR NK cell therapy for solid tumors: potential and challenges.

Yanlin Yu, Mitchell Ho.

  CAR-T cell therapy has shown promise but is constrained by side effects and limited efficacy in treating solid tumors. Compared to CAR-T cells, CAR natural killer (NK) cells derived from multiple versatile sources exhibit more favorable safety profiles and possess the unique ability to refine cytotoxic activity, serve as off-the-shelf options, and target a broad range of tumors. However, several challenges still impede the development and implementation of CAR NK cell therapy for solid tumors. This review article outlines the therapeutic strategies, advantages, limitations, and potential solutions, while providing insight into the future landscape by challenging current knowledge in the field. It also discusses optimizing CAR NK cell structure, addressing obstacles in the clinical utility of CAR NK cell therapy, and integrating it into standard cancer treatment regimens. Ultimately, we aim to navigate the crucial challenges in treating solid tumors and explore the future of this approach.

Keywords:  CAR NK cell therapy; cancer treatment; natural killer (NK) cell; solid tumor

DOI:  https://doi.org/10.1093/abt/tbaf019
Adv Exp Med Biol. 2025 ;1486 43-52

Challenges in the Patent and Legal Environment for Cell Therapies.

Sebastian Sethe.

  Beyond specific regulations relating to manufacturing and safety, broader legal issues accompany the entire 'journey' of a cell therapy product: from procurement of the starting material to efforts of testing and development and operating in a medical marketplace. This chapter charts some legal considerations including the USA, EU, and UK law relating to consent, privacy, preclinical and clinical research, placing on the market, intellectual property protection, and medical commence as they uniquely shape innovation trajectories in cell therapies. These factors need to be considered from the inception of cell therapy and addressed strategically throughout its lifecycle.

Keywords:  Consent; Data protection; Intellectual property; Law; Legal; Medical tourism; Placing on the market; Privacy; Regulation; Research; Stem cells; cell therapies

DOI:  https://doi.org/10.1007/978-3-031-97297-3_4
Adv Exp Med Biol. 2025 ;1486 179-192

GMP Manufacturing of Allogenic iPSC-NK Cells for Immunotherapy.

Hongliang Zong, Richard Wang.

  Human induced pluripotent stem cell (iPSC)-derived natural killer (NK) cells are the most suitable immune cell therapy types for allogeneic application, due to the TCR-independent innate immune activity of NKs devoid of GvHD side effects. In addition, the amenability of genetic engineering and the potential of scalable manufacturing of homogenic iNK from an established bank of iPSC clones make it an excellent starting material for cell therapies. However, as there is no approved iNK therapeutic product on the market yet, the extent of robustness and regulatory compliance of good manufacturing practice (GMP) is not established and proven, nor is it straightforward. We will review the current development and validation status of iPSC-NK manufacturing processes, along with quality controls, based on literature information and our own experience, towards the goal of manufacturing large-scale iNK products for regulatory submission and future commercialization.

Keywords:  3-tier Banking; Allogenic Therapy; Critical Process Parameters (CPP); Critical Quality Attributes (CQA); Cryopreservation; Feeder-free Suspension Culture; GMP compliance; Induced Pluripotent Stem Cell (iPSC); NK Activation; Natural Killers (NK) Cell; Release Testing; Scale-up Manufacturing; iNK Characterization

DOI:  https://doi.org/10.1007/978-3-031-97297-3_15
Cell Transplant. 2025 Jan-Dec;34:34 9636897251379571

Autologous stem cell transplantation meets CAR-T therapy: A synergistic strategy for B-cell lymphoma.

Xumeng Zhao, Jiaying Wu, Xiaojian Zhu, Yi Xiao.

  B-cell lymphoma, the most common subtype of non-Hodgkin lymphoma, presents major therapeutic challenges due to molecular heterogeneity and high relapse rates. While autologous hematopoietic stem cell transplantation (ASCT) has been a cornerstone for relapsed/refractory (R/R) B-cell lymphoma, its efficacy is often compromised by minimal residual disease (MRD) persistence and an immunosuppressive tumor microenvironment. Chimeric antigen receptor (CAR)-T cell therapy has transformed treatment paradigms but faces limited long-term durability due to antigen escape and T-cell exhaustion. The integration of ASCT with CAR-T therapy may offer a complementary approach to address these limitations, leveraging ASCT-induced immune reconstitution to enhance CAR-T-cell persistence and reprogram the tumor milieu. Emerging clinical evidence supports this approach, indicating improved disease control and progression-free survival. Although preliminary clinical outcomes are encouraging, unresolved challenges persist, particularly in terms of cumulative toxicity, optimal therapeutic sequencing, CAR-T-cell longevity, and financial feasibility associated with these advanced therapies. This review provides a comprehensive overview of mechanistic synergies between ASCT and CAR-T therapy, critically evaluates emerging clinical evidence on treatment sequencing, and explores innovative strategies to increase safety, efficacy, and accessibility.

Keywords:  B-cell lymphoma; autologous hematopoietic stem cell transplantation; chimeric antigen receptor T-cell therapy; minimal residual disease clearance; tumor microenvironment modulation

DOI:  https://doi.org/10.1177/09636897251379571
J Transl Med. 2025 Oct 23. 23(1): 1163

CAR-T and CAR-NK cell therapies in AML: breaking barriers and charting the future.

Huichao Wu, Fatemeh Sadat Shafiei, Zahra Taghinejad, Mohsen Maleknia, Hanieh Noormohamadi, Atieh Raoufi, Sina Nouri, Nazli Servatian, Hamed Soleimani Samarkhazan.

  Acute myeloid leukemia (AML), characterized by aggressive relapse and dismal survival, remains a formidable challenge despite conventional therapies. Chimeric antigen receptor (CAR)-engineered T and natural killer (NK) cells have emerged as groundbreaking immunotherapies, offering targeted eradication of leukemic stem cells (LSCs) and resistant blasts. CAR-T cells, leveraging antigens like CD123 and CD33, demonstrate early clinical success, with complete remission rates up to 66% in refractory/relapsed (R/R) AML. CAR-NK cells complement this approach through inherent tumor surveillance, reduced toxicity, and "off-the-shelf" feasibility. However, barriers such as antigen escape, heterogeneous immunosuppressive microenvironments (including intratumoral microbiota variations), and on-target/off-tumor toxicity persist, limiting durable responses. Innovations in dual-targeting CARs, cytokine-armored constructs, and CRISPR-edited universal cells aim to overcome these hurdles. Emerging strategies integrating checkpoint inhibitors, metabolic modulators, and AI-driven antigen selection promise to enhance efficacy and safety. This review synthesizes the evolving landscape of CAR-T/NK therapies, critically analyzing preclinical breakthroughs, clinical trial outcomes, and persisting challenges. By addressing manufacturing scalability, cost barriers, and long-term safety, cellular immunotherapy holds transformative potential to redefine AML management. As the field advances, interdisciplinary collaboration and biomarker-guided personalization will be pivotal in translating laboratory innovations into life-saving therapies for AML patients.

Keywords:  Acute myeloid leukemia; CAR-NK cell therapy; CAR-T-cell therapy; Immunotherapy; Tumor Microenvironment

DOI:  https://doi.org/10.1186/s12967-025-07151-5
Nat Cancer. 2025 Oct 23.

Engineering T cells with a membrane-tethered version of SLP-76 overcomes antigen-low resistance to CAR T cell therapy.

Maria Caterina Rotiroti, Aidan M Tousley, Hoyin Chu, Marco Herrera-Barrera, Antigoni Manousopoulou, Won-Ju Kim, Yajie Yin, Thomas Spencer Parish, Aniela Mitchell, Malcolm Holterhus, Lea Wenting Rysavy, Guillermo Nicolas Dalton, Katherine Ann Freitas, Gernot Kaber, Korbinian N Kropp, Christopher A Klebanoff, Ansuman T Satpathy, Leo D Wang, Caleb A Lareau, Robbie G Majzner.

Chimeric antigen receptor (CAR) T cells can mediate durable complete responses in individuals with certain hematologic malignancies, but antigen downregulation is a common mechanism of resistance. Although the native T cell receptor can respond to very low levels of antigen, engineered CARs cannot, likely due to inefficient recruitment of downstream proximal signaling molecules. We developed a platform that endows CAR T cells with the ability to kill antigen-low cancer cells consisting of a membrane-tethered version of the cytosolic signaling adaptor molecule SLP-76 (MT-SLP-76). MT-SLP-76 can be expressed alongside any CAR to lower its activation threshold, overcoming antigen-low escape in multiple xenograft models. Mechanistically, MT-SLP-76 amplifies CAR signaling through recruitment of ITK and PLCγ1. MT-SLP-76 was designed based on biologic principles to render CAR T cell therapies less susceptible to antigen downregulation and is poised for clinical development to overcome this common mechanism of resistance.

DOI: https://doi.org/10.1038/s43018-025-01056-4
Adv Exp Med Biol. 2025 ;1486 97-110

Raw Material Considerations for the Manufacture of Cell-Based Therapies.

Jennifer N Solomon, Mike Jones, Lynn Csontos.

  Cell-based therapies are rapidly advancing as a promising approach for treating blood-based cancers and other unmet clinical needs. As the pace of development and commercialization of the therapies increases, regulatory scrutiny on the materials and processes used in their manufacturing intensifies. This chapter focuses on raw materials-also known as ancillary materials-that come into direct contact with the cell product during manufacturing but are not present in the final therapeutic product. Examples include: cell culture media, supplements, enzymes, cryopreservation agents, buffers, and antibodies. The selection and qualification of these materials are critical to ensuring the quality, safety, consistency, and efficacy of the final cell product. Raw materials vary in grade and composition, and their suitability for use in cell therapy manufacturing must be rigorously evaluated and continuously reassessed through a risk-based approach throughout clinical development. This chapter also examines the regulatory landscape for raw materials, noting the lack of specific regulations but highlighting guidance from key regional and international organizations. Given the variability in raw material quality and labeling, developers and suppliers need to collaborate closely to qualify materials for their intended use. The chapter concludes by outlining the regulatory considerations and risks involved in raw material selection, providing case studies that offer practical insights from real-world experiences.

Keywords:  Ancillary materials; Cell therapy development; Cell therapy manufacturing; GMP; Raw materials; Reagents; Risk-based approach

DOI:  https://doi.org/10.1007/978-3-031-97297-3_8
Mol Ther. 2025 Oct 23. pii: S1525-0016(25)00864-0. [Epub ahead of print]

Nanobody-enhanced T cell cancer immunotherapy: Advancing precision medicine.

Remy Boisgard, Juliette Prothon, Celia Torrielli, Patrick Chames, Brigitte Kerfelec.

Due to their small size, high specificity, and modularity, nanobodies, or single-domain antibodies, have emerged as versatile tools in cancer immunotherapy. This review provides a comprehensive and up-to-date overview of nanobody-based strategies designed to enhance T cell-mediated anti-tumor responses. It first outlines the structural and functional characteristics of nanobodies, highlighting their tumor penetration and modular design potential. Then, three major nanobody-driven therapeutic approaches are discussed in detail: immune checkpoint inhibition, T cell engagement, and CAR-T cell engineering. For each strategy, current preclinical and clinical developments are reviewed. Challenges are explored, along with emerging strategies to overcome them and expand the therapeutic applicability of nanobody-based immunotherapy, including format choice, valency tuning, pharmacokinetic optimization, and safety enhancements.

DOI: https://doi.org/10.1016/j.ymthe.2025.10.043
Int J Nanomedicine. 2025 ;20 12393-12411

Emerging Plant-Derived Exosome-Like Particles Reveal Key Therapeutic Benefits. A Comprehensive Review of Evidence.

Jia Chi, Xubo Wang, Yanrui Song, Jiazhi Wang, Lin Guo, Rui Wang, Guangzhi Wang.

  Plant-derived exosome-like nanoparticles (PELNs) are lipid bilayer-enclosed nanoscale vesicles isolated from plant cells, harboring a diverse cargo such as RNAs, proteins, lipids, and biologically active constituents. Increasing evidence indicates that PELNs can efficiently enter mammalian cells through multiple uptake pathways, including phagocytosis, clathrin/caveolin-mediated endocytosis, and macropinocytosis. In recent years, they have emerged as highly promising nanocarriers for targeted drug delivery, disease diagnosis, and therapeutic intervention. This review provides a systematic overview of the therapeutic applications of PELNs across various diseases and the signaling mechanisms involved, while briefly outlining their isolation and characterization to provide essential research background. Despite remarkable advancements, the field still has several challenges, including protocol standardization, precise marker identification, biological stability, and refinement of targeted delivery strategies. Nevertheless, owing to their intrinsic properties, such as low cytotoxicity, high biocompatibility, inherent targeting capacity, minimal immunogenicity, and surface modifiability, PELNs hold considerable promise as next-generation delivery vectors. Future investigations will likely focus on refining manufacturing processes, elucidating PELN-associated molecular mechanisms, and engineering more advanced delivery systems designed for clinical translation.

Keywords:  drug delivery; extracellular vesicles; plant-derived exosome-like nanoparticles; signal transduction pathways; therapeutic applications

DOI:  https://doi.org/10.2147/IJN.S543947
Adv Exp Med Biol. 2025 ;1486 253-282

Barriers to Effective Cryopreservation of Cell Therapies: Challenges and Emerging Solution.

Charles J Hunt, Brian H Johnstone, Erik J Woods.

  Many cellular therapies require a cryopreservation step during their lifecycle. Cryopreservation offers many advantages such as extending product shelf life, decoupling manufacture from "just-in-time" delivery, and allowing pre-conditioning and other treatment interventions before administration. Nevertheless, it can also pose significant risks to the product and by extension the patient, if the process is non-optimized or misapplied. A clear understanding of the principles and their relationship to cell viability and functionality is therefore a pre-requisite for effective cryopreservation. The effect of cooling rate on cell survival, the need to avoid the formation of intracellular ice, and the action of cryoprotectants in improving cell recovery are all well recognized. Less so, the need to control cryoprotectant exposure times/temperature and their part in controlling cryoprotectant toxicity and the avoidance during addition/removal of damaging excursions in cell volume. However, there are other, newly emerging, factors that often go unrecognized. This chapter sets out to examine some that are potential impediments to the successful generation of a cell therapy product.Recently, a debate has opened up on the use of dimethyl sulphoxide as the cryoprotectant of choice in the context of cell therapies. Its use, together with that of bovine serum as an excipient, is being questioned and an examination of the advantages, disadvantages, and alternatives is explored herein. The control of ice nucleation/propagation and the phenomenon of supercooling which, if uncontrolled, can lead to excessive ice nucleation and variable outcomes, and the means to control this process and inhibit ice crystallization are also discussed.The effect on cell viability of transient warming events during routine low-temperature storage has, until recently, gone unrecognized. This is an emerging issue with both regulatory and commercial implications. Lastly, cryopreservation-induced delayed-onset cell death, its causes, detection and prevention, and the implications of such injury on demonstrating acceptable viability and functionality for regulatory purposes and routine quality-control monitoring are also examined.

Keywords:  Controlled-supercooling; Cryopreservation-induced delayed-onset cell death; Dimethyl sulfoxide debate; Serum starvation; Transient warming events

DOI:  https://doi.org/10.1007/978-3-031-97297-3_20
Front Immunol. 2025 ;16 1679271

Emerging macrophage-based therapies for cancer: a review of preclinical and clinical advances.

Jan Brancewicz, Paulina Kucharzewska.

  Macrophages, the most abundant immune cells in many solid tumors, are no longer viewed solely as accomplices of cancer but as powerful therapeutic allies. This review charts the rapid rise of macrophage-based immunotherapies, from CD47/SIRPα checkpoint blockade and CAR-macrophages to macrophage-drug conjugates (MDCs). We emphasize emerging frontiers - RNA-based reprogramming, epigenetic modulation, small activating RNA and circRNA approaches, and macrophage-derived extracellular vesicles - that are redefining how tumor-associated macrophages can be targeted or harnessed. Distinct from earlier TAM reviews, we integrate outcomes from ongoing and completed clinical trials, highlight therapeutic platforms beyond classical depletion and polarization, and frame macrophages not only as targets but also as delivery vehicles. By spotlighting both innovative strategies and the challenges of moving them into the clinic, we aim to provide a forward-looking guide for researchers and clinicians shaping the next generation of cancer immunotherapy.

Keywords:  cancer; cell therapy; immunotherapy; macrophages; tumor microenvironment

DOI:  https://doi.org/10.3389/fimmu.2025.1679271
Adv Exp Med Biol. 2025 ;1486 17-26

The Developing Standards Landscape for Cell and Gene Therapies.

Jonathan J Campbell, Julian H Braybrook.

  Effective standardization strategies are critical to the successful development and adoption of complex technological solutions, in many cases due to meeting requirements of interdisciplinary expertise, interconnected systems, or the production of comparable products at scale. Cell and gene therapies, as a new generation of living drugs are a prime example of such a technology, yet with the pace of advance, the challenge to develop appropriate solutions is an increasingly complex and resource-intensive activity, requiring careful prioritization and management. This chapter introduces and develops various drivers for standardization in this sector, the stakeholders involved in this work, and governance structures required for standards development and deployment, helping to overcome challenges associated with product and process characterization, consistent manufacture and regulatory approval.

Keywords:  Characterization; Critical process parameters; Critical quality attributes; Documentary standards; Harmonization; Metrology; Quality; Reference materials; Reference methods; Standardization

DOI:  https://doi.org/10.1007/978-3-031-97297-3_2
Acta Biochim Biophys Sin (Shanghai). 2025 Oct 23.

Decoding signaling architectures: CAR versus TCR dynamics in solid tumor immunotherapy.

Zui Chen, Xin Zhou.

  The T cell receptor (TCR) initiates signaling by specifically recognizing peptide-MHC complexes, triggering the phosphorylation of CD3 chain immunoreceptor tyrosine-based activation motifs (ITAMs). This recruits kinases such as ZAP70, triggering a tightly regulated signaling cascade that governs T cell activation, differentiation, and effector functions. In contrast, the chimeric antigen receptor (CAR) is a synthetic construct that bypasses MHC restriction by fusing an antigen-binding domain with intracellular signaling modules (usually CD3ζ and co-stimulatory domains) from the TCR complex and other receptors. CAR-T cell therapy has revolutionized the treatment of hematologic malignancies, resulting in durable remission of B-cell leukemia, lymphoma, and multiple myeloma. However, its efficacy in solid tumors is limited by intrinsic barriers: poor CAR-T-cell trafficking/infiltration into tumors, the immunosuppressive tumor microenvironment (TME), intratumoral metabolic competition, and tumor antigen heterogeneity/loss. To improve CAR-T-cell function in solid tumors, numerous studies have explored multiple strategies: engineering CARs to boost immune synapse formation via optimized receptor clustering, increasing the ITAM number/strength to amplify downstream signaling, and incorporating novel/multiple co-stimulatory domains to sustain T-cell activation and persistence. Additionally, approaches include the use of CAR-T cells that secrete pro-inflammatory cytokines, epigenetic reprogramming to preserve T-cell stemness and functionality, and the use of synthetic biology tools for tunable/logic-gated CAR activation. Here, we summarize the current understanding of CAR signaling dynamics and highlight recent breakthrough strategies designed to overcome these challenges in solid tumors. These advances narrow the liquid-solid tumor efficacy gap, holding promise for better clinical outcomes in patients with solid malignancies and a new era of personalized immunotherapy.

Keywords:  CAR-T; TCR; immunotherapy; solid tumor

DOI:  https://doi.org/10.3724/abbs.2025190
J Immunother Cancer. 2025 Oct 23. pii: e011768. [Epub ahead of print]13(10):

Acquisition of an immunosuppressive microenvironment after anti-CD19 CAR T-cell therapy is associated with T-cell dysfunction and resistance.

Marianna Ponzo, Lorenzo Drufuca, Chiara Buracchi, Marco M Sindoni, Silvia Nucera, Cristina Bugarin, Ramona Bason, Grazisa Rossetti, Raoul Bonnal, Cristian Meli, Benedetta Rambaldi, Federico Lussana, Silvia Ferrari, Alex Moretti, Giuseppe Dastoli, Giulia Risca, Christian Pellegrino, Markus G Manz, Stefania Galimberti, Alessandro Rambaldi, Giuseppe Gaipa, Andrea Biondi, Massimiliano Pagani, Chiara F Magnani.

   BACKGROUND: Chimeric antigen receptor (CAR) T cells targeting CD19 induce durable responses in B-cell acute lymphoblastic leukemia (B-ALL). However, the contribution of the tumor microenvironment to the therapeutic response after CAR T-cell treatment remains incompletely understood.
METHODS: We performed single-cell RNA sequencing and spectral flow cytometry-based analyses of bone marrow-resident immune cells from B-ALL patients before and after CAR T-cell treatment.
RESULTS: We observed profound changes in the microenvironment in response to CAR T-cell-mediated inflammation, including an increase in myeloid cells. Significant induction of the interferon response, hypoxia, and TGF-β signaling was accompanied by expansion of myeloid-derived suppressor cells (MDSCs) and endogenous exhausted CD8+ T cells. PD-1 expression in endogenous T cells post-treatment was associated with a lack of durable response in the cohort of patients analyzed. Further, we revealed that HIF-1α, VEGF, and TGFBR2 are key players in the intercellular communication between CAR T cells and the immune niche, potentially driving widespread T-cell dysfunction. Infusion of anti-CD19 CAR T cells led to increased accumulation of human MDSCs, exacerbation of a hypoxic environment and T-cell exhaustion in hematopoietic stem/progenitor cell-humanized mice bearing a human tumor.
CONCLUSIONS: In conclusion, CAR T-cell-mediated myeloid activation is associated with pathways of immune dysregulation that may antagonize the effects of therapy.

Keywords:  Adoptive cell therapy - ACT; Chimeric antigen receptor - CAR; Leukemia; Tumor microenvironment - TME

DOI:  https://doi.org/10.1136/jitc-2025-011768
Int J Hematol. 2025 Oct 21.

Peripheral T cell lymphoma following CD19-targeted chimeric antigen receptor T cell therapy.

Hiro Tatetsu, Koji Kato, Atsushi Wada, Taichi Hirano, Shikiko Ueno, Yuko Miyasato, Asami Yamada, Takafumi Shichijo, Yusuke Higuchi, Yujiro Ueda, Kisato Nosaka, Yoshiki Mikami, Kennosuke Karube, Masao Matsuoka, Jun-Ichirou Yasunaga.

  Chimeric antigen receptor (CAR) T cell therapy has significantly improved outcomes for patients with refractory B cell lymphoma. However, rare cases of secondary T cell lymphomas have raised safety concerns. Here, we present a case of peripheral T cell lymphoma not otherwise specified (PTCL-NOS) that developed eight months following CD19-directed CAR T cell therapy (lisocabtagene maraleucel) in a 66-year-old male patient with recurrent diffuse large B cell lymphoma. The patient initially achieved complete remission but later developed a subcutaneous mass and systemic lymphadenopathy. Histopathology and flow cytometry confirmed a diagnosis of PTCL-NOS with a CD3 + , CD8 + , and CD30 + phenotype, as well as clonal T cell receptor gene rearrangements. No immunoglobulin rearrangements were detected, ruling out a lineage switch. Furthermore, the CAR transgene was undetectable by RNA-in situ hybridization, and flow cytometry showed no CAR protein expression, suggesting that the lymphoma was not caused by CAR gene integration. This case highlights the importance of re-biopsy in cases of suspected relapse following CAR T cell therapy, and emphasizes the need for long-term monitoring. While a direct causal link remains unclear, industry-academia collaboration is crucial for investigating the mechanisms underlying secondary T cell malignancies and improving the safety of CAR T cell therapy.

Keywords:  CAR T cell; PTCL; Secondary malignancies

DOI:  https://doi.org/10.1007/s12185-025-04074-1
Mol Biol Rep. 2025 Oct 23. 53(1): 3

Generation of the augmented IL-15-secreting anti-HER2 chimeric antigen receptor (CAR)-NK cells: an encouraging immunotherapeutic tool.

Reza Darvishvand, Maryam Asadi, Zohreh Mostafavi-Pour, Amin Ramezani, Nasrollah Erfani.

   BACKGROUND AND OBJECTIVE: Given the undeniable and promising outcomes of chimeric antigen receptor (CAR) technology-based immunotherapy reported in recent years, this study aimed to develop anti-HER2 CAR NK cells as a novel therapeutic strategy for cancer immunotherapy.
MATERIALS AND METHODS: The NK-92 cell line was transduced with a recombinant lentiviral vector encoding an anti-HER2 construct, either with or without IL-15 co-expression. This yielded two distinct CAR NK cell populations: (1) anti-HER2 CAR NK cells and (2) IL-15-secreting anti-HER2 CAR NK cells. The cytotoxic effects of these engineered cells against the HER2-positive SK-BR-3 target cells were then evaluated using the PE-Annexin V and 7-AAD assays. Flow cytometry analyses were performed to assess CAR NK cell activity by measuring the expression of degranulation marker CD107a and intracellular levels of granzyme B and perforin, following surface and intracellular staining.
RESULTS: Our findings demonstrated that anti-HER2 CAR NK cells and IL-15-secreting anti-HER2 CAR NK cells induced significantly higher levels of total apoptosis in HER-positive SK-BR-3 cells compared to mock-transduced (control) and non-transduced NK cells (control). The mean percentage (± SD) of CD107a was significantly higher in CAR NK cells co-cultured with SK-BR-3 cells compared to both control groups. Moreover, the mean expression (based on MFI) of granzyme B and perforin was significantly elevated in both CAR NK cell types following co-culture with HER2-positive SK-BR-3 cells. Notably, IL-15-secreting anti-HER2 CAR NK cells exhibited superior cytotoxic potential compared to their non-secreting counterparts.
CONCLUSION: In summary, our findings demonstrate potent antitumor activity of anti-HER2 CAR NK cells. The promising results suggest that these engineered CAR NK cells, particularly those capable of IL-15 secretion, hold significant potential as a novel immunotherapeutic strategy for HER2-positive malignancies.

Keywords:  Anti-HER2 CAR NK cell; Cancer; Chimeric antigen receptor (CAR); HER2 molecule; Natural killer (NK) cells

DOI:  https://doi.org/10.1007/s11033-025-11163-1
Adv Exp Med Biol. 2025 ;1486 157-163

Development of Automated Systems for Cell Therapy.

Takako Yamamoto, Masaki Hosoya, Shin Kawamata.

  The introduction of automated systems that can increase manufacturing efficiency and eliminate human errors is indispensable for the dissemination of cell therapy. However, a robotic operation without process monitoring and manufacturing records needs a "verification" process at the end of manufacturing for each shipment, just like manual operation would have required, which is called the Pharma 3.0 manufacturing stage, and would not visualize the manufacturing process nor link with inventory control. A simple robotic system that can mimic human operation will not contribute to innovation in the cell manufacturing industry. Here we show the quality by design (QbD)-based automated closed-type cell manufacturing system with in-process monitoring that could eliminate a "verification" process prior to shipment and assure the quality of the product by manufacturing the product as designed, which will contribute to the industrialization of cell manufacturing that can support the dissemination of cell therapy.

Keywords:  2-AAA; Automation; CCPs; CQAs; Digitization; Electric batch record; Glycolytic pathway; In-process monitoring; Kynurenine; OXPHOS; Pharma 4.0; QTPP; QbD; Quality by design; Warburg effect; eBR

DOI:  https://doi.org/10.1007/978-3-031-97297-3_13
J Natl Cancer Cent. 2025 Oct;5(5): 429-440

Neoantigen identification and TCR-T therapy development for solid tumors: current advances and future perspectives.

Xinyao Zheng, Yahui Zhao, Zhihua Liu.

  Recently, T cells expressing engineered T cell receptor (TCR-T cells) have become recognized as a promising tumor cell therapy for solid tumors because of their ability to selectively kill tumor cells with less destruction of other cells and their high safety when used as autologous T cells. Several studies and clinical tests have been conducted to demonstrate its potential as a novel therapy. However, previous research has mainly focused on antigens; these common targets for TCR-T are tumor-associated antigens, which exhibit expression not only in tumor cells but also in normal cells, resulting in off-target risk and not considering the heterogeneity of different patients. In contrast, neoantigens offer superior specificity as they are uniquely expressed on tumor cells due to genomic alterations. Given the frequent occurrence and notable role of genetic mutations in tumorigenesis and tumor progression, identification and targeting of neoantigens is a valuable therapeutic direction. This perspective delves into various antigen classifications, including their characteristics and advantages, as well as strategies for identifying and validating neoantigens that have emerged from numerous research studies. These insights are crucial for guiding the search for new neoantigens. We also review significant and representative studies involving TCR-T and other immunotherapies that target neoantigens to assess the therapeutic effectiveness of TCR-T therapy. Moreover, we discuss the challenges and complexities inherent in TCR-T therapy and propose potential solutions for these issues. In this perspective, we aim to provide fresh perceptions and strategies for cancer treatment by highlighting the potential of TCR-T and exploring its challenges and future directions. It also seeks to propel the development of precision medicine and personalized therapy, offering hope for more effective and targeted cancer treatments in the future.

Keywords:  CAR-T; Immunotherapy; Neoantigen; TCR-T

DOI:  https://doi.org/10.1016/j.jncc.2025.07.001
Adv Exp Med Biol. 2025 ;1486 363-374

Development of Stem Cell Data Systems and Associated Data Standards for Cell Therapy.

Yong Zhang, Andreas Kurtz, Hongling Zhao, Lei Wang, Jie Hao, Martin Golebiewski, Glyn N Stacey, Tongbiao Zhao.

  As cell-based therapies near widespread clinical use, robust data management systems are crucial for ensuring safety, efficacy, and regulatory compliance. This chapter reviews the ISO 8472 series, which focuses on data interoperability for stem cell applications, and highlights the complexities of managing clinical, manufacturing, and quality control data. It underscores the need for standardized frameworks to streamline data sharing, analysis, and validation. Additionally, emerging technologies like AI and blockchain are explored for their potential to revolutionize data management, enhance security, and improve automation. Furthermore, this chapter addresses challenges in implementing data standards in cell therapy manufacturing and ensuring transparency. It outlines strategies to foster collaboration among stakeholders, including regulatory bodies, biobanks, CROs, and manufacturers, to drive innovation and scalability. Establishing comprehensive standards and interoperable data systems is essential for building trust and facilitating the commercialization of stem cell therapies.

Keywords:  Cell therapy; Data interoperability; Standards; Stem cell data systems

DOI:  https://doi.org/10.1007/978-3-031-97297-3_28
Adv Exp Med Biol. 2025 ;1486 295-302

Cold Chain Delivery in Cell Therapies.

Peter Kilbride.

  Cold-chain shipment (i.e. ultra-low cryogenic shipment) is a critical step to enable widespread patient access to cell therapies. These therapies often require multiple cryogenic transport steps, from sourcing cellular starting materials, to manufacturing, and ultimately to the final clinical application. Unlike conventional cold chain logistics, cryopreserved cell therapy shipments typically required much lower temperatures, involve high-value shipments (both in terms of financial cost of the therapy and the high patient impact incorrect shipment may have), and require a strict chain of custody and traceability.This chapter is intended to act as a guide for those developing or working with cell therapies and the wider cold-chain. Pros and cons of different temperature shipments are considered, from unfrozen down to LN2 temperatures. Typical shipping processes are considered, with a focus on key steps that must be taken that are not typical with standard, non-cryopreserved shipments. Finally the practicalities of developing a custom shipping system are considered, along with thoughts regarding the future direction of cold-chain processes as they become critical for ever more therapies, and more common within the logistics space.

Keywords:  Cell therapies; Chain of custody; Clinical delivery; Cold-chain; Cryopreservation; Dry ice; Dry shipper; Logistics; Regenerative medicine; Shipping

DOI:  https://doi.org/10.1007/978-3-031-97297-3_22
JAMA Intern Med. 2025 Oct 20.

Advancing a US Single-Payer Model for Cell and Gene Therapy.

Audun Brendbekken, Stacie B Dusetzina.

DOI: https://doi.org/10.1001/jamainternmed.2025.5058
Adv Exp Med Biol. 2025 ;1486 3-16

Standardised Matrices for Stem Cells.

Nilofar Faruqui, Ibolya E Kepiro, Maxim G Ryadnov.

  The chapter reviews the state of the art in establishing reference extracellular matrices or scaffolds to support commercial materials developed to promote and monitor cell growth and tissue development in 3D. The need and value of reference scaffolds as well as the technical barriers and current progress towards them are discussed from the end user perspective and as informed by standardization requirements for cell therapy applications.

Keywords:  3D cell culture; Biometrology; Cell therapy; Extracellular matrix; Hydrogels; Pre-clinical testing; Reference materials; Regenerative medicine; Standardization

DOI:  https://doi.org/10.1007/978-3-031-97297-3_1
Hematol Oncol. 2025 Nov;43(6): e70144

The French Experience of Pharmacists and CAR T-Cells: A Study of the French Society of Oncology Pharmacy (SFPO).

Vérane Schwiertz, Romain de Jorna, Adeline Quintard, Marie-Antoinette Lester, Marine Pinturaud, Nicolas Cormier, Elise D'Huart, Emmanuelle Fougereau, Muriel Carvalho, Benjamin Sourisseau, Pauline Gueneau, Mathieu Wasiak, Alexia Jouvance, Muriel Paul, Régine Chevrier, Bertrand Pourroy, Jean-Louis Cazin, Florence Ranchon, Isabelle Madelaine-Chambrin, Catherine Rioufol.

  The aim of this study was to describe the initial 3-year experience in vein-to-vein time for axi-cel therapy and the role of pharmacists in the first recruiting French centers. Retrospective observational data were collected for vein-to-vein time for commercial axi-cel after ≥ 2 lines of systemic therapy between January 2019 and December 2021 in the first 12 authorized French centers. Hospital pharmacists used a circuit database to ensure the prospective traceability at all steps. Totally 501 of the 562 intention-to-treat registrations on the database for cytapheresis (89,1%) led to the infusion of axi-cel. Median vein-to-vein time was shortened by 4 days. This was mainly due to tightening the interval from apheresis to release. The 36-day median vein-to-vein time achieved after 3 years' experience should be compared to the 29-34 days reported in Canada, the USA and Israel, where manufacturing sites are geographically closer to hospital centers than they are in France. The top 5 recruiting centers had the shortest vein-to-vein times. This French experience may serve as a model for other European centers, notably as regards deployment of pharmacists to improve the patient pathway with CAR T-cells and other gene and cellular therapies.

Keywords:  CAR T‐cells; patient pathway; pharmaceutical circuit; vein‐to‐vein time

DOI:  https://doi.org/10.1002/hon.70144
Mol Ther. 2025 Oct 23. pii: S1525-0016(25)00867-6. [Epub ahead of print]

Co-expression of an adapter CAR retains efficacy of CAR T cells after single and dual antigen loss in lymphoma.

Zeno Riester, Marlena Surowka, Nicole Seifert, Diana Darowski, Arindam Ghosh, Peter Spieler, Justus Weber, Reto Gianotti, Fabian Freitag, Miriam Alb, Thomas Nerreter, Elena Gerhard-Hartmann, Andreas Rosenwald, Markus Sauer, Ekkehard Moessner, Hermann Einsele, Claudia Ferrara-Koller, Christian Klein, Michael Hudecek, Sophia Danhof.

CAR T cell therapy is effective in many patients suffering from B cell malignancies, yet antigen escape is a major resistance mechanism by which efficacy can be diminished or lost. To counter this, we enhance anti-lymphoma CAR T cells by generating conventional and adapter dual (ConvAD) CAR T cells, which co-express a conventional antigen-specific CAR and the P329G adapter CAR that leverages Fc-mutated antibodies for redirection. ConvAD CAR T cells display robust functionality against the primary lymphoma antigen of the conventional CAR while providing flexible redirection to additional targets via target-specific adapters in the event of antigen escape. We demonstrate bimodal activity of ConvAD CAR T cells acting through direct engagement of the conventional CAR as well as through binding of adapters, resulting in enhanced multispecific anti-lymphoma targeting. In vitro and in vivo, ConvAD CAR T cells targeting combinations of ROR1, CD19, and CD20 prevent lymphoma outgrowth across models of stable antigen expression as well as single- or dual antigen loss, and benchmark superior to both single-antigen specific and bispecific CAR T cells. The ConvAD CAR platform thus addresses a medical need by offering an effective strategy for multi-antigen targeting, counteracting antigen escape.

DOI: https://doi.org/10.1016/j.ymthe.2025.10.046
Clin Cancer Res. 2025 Oct 21.

Facts and Hopes of Chimeric Antigen Receptor-Redirected Natural Killer T cells.

Amy N Courtney, Xin Zhou, Gengwen Tian, Ying Wang, Leonid S Metelitsa, Gianpietro Dotti.

Chimeric antigen receptor (CAR)-engineered invariant natural killer T cells (CAR-NKTs) are a novel cell platform for cancer immunotherapy. Unlike conventional T cells, NKTs are characterized by innate antitumor properties, minimal alloreactivity, and a unique ability to modulate the tumor microenvironment (TME). This article provides a comprehensive overview of preclinical and early clinical studies evaluating CAR-NKTs in both autologous and allogeneic clinical settings. We discuss the contributions of CAR signaling domains, cytokine co-expression, and other functional measures that correlate with CAR-NKT persistence, function, and metabolic fitness. We also discuss the critical role of immunocompetent animal models in elucidating the interactions of CAR-NKTs with the TME and other components of the immune system. Finally, we review strategies that combine CAR-NKTs with other therapeutic approaches to promote potential synergistic benefits in cancer patients.

DOI: https://doi.org/10.1158/1078-0432.CCR-25-0197