Dual ON/OFF-switch chimeric antigen receptor controlled by two clinically approved drugs

Proceedings of the National Academy of Sciences, Год журнала: 2024, Номер 121(44)

Опубликована: Окт. 25, 2024

The ability to remotely control the activity of chimeric antigen receptors (CARs) with small molecules can improve safety and efficacy gene-modified T cells. Split ON- or OFF-switch CARs involve dissociation tumor-antigen binding from cell activation (i.e., CD3ζ) on receptor (R-) signaling (S-) chains, respectively, that either associate are disrupted in presence a molecule. Here, we have developed an inducible (i)ON-CAR comprising anti-apoptotic B lymphoma protein 2 ectodomain both chains which venetoclax. We showed ON (iON)-CAR cells respond target tumors venetoclax BH3 mimetic navitoclax dose-dependent manner, while there is no impact drugs equivalent second generation-CAR Within 48 h withdrawal, iON-CAR lose tumor vitro as evaluated by Interferon-gamma (IFNγ) production, they reliant upon for vivo activity. Finally, fusing degron sequence endodomain S-chain generated all-in-one ON/OFF-switch CAR, iONØ-CAR, down-regulated lenalidomide within 4 6 functionally inactive (no IFNγ production) 24 h. propose our remote-control CAR designs reduce toxicity clinic. Moreover, periodic rest iON iONØ-CAR may alleviate exhaustion hence augment persistence long-term patients.

Язык: Английский

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Drug-controlled CAR-T cells through the regulation of cell-cell interactions

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Авг. 6, 2024

CAR T-cell therapy is constrained by on-target, off-tumor toxicities as well cellular exhaustion due to chronic antigen exposure. CARs comprising small-molecule controlled switches can enhance both safety and therapeutic efficacy but are limited the scarcity of non-immunogenic protein elements responsive non-immunosuppressive, clinically approved drugs with favorable pharmacodynamics. Here, we combine rational design library-based optimization a protein-protein interaction (PPI) human origin develop venetoclax-controlled Drug-Regulated Off-switch PPI (DROP)-CARs. DROP-CARs enable dose-dependent release tumor-targeting scFv consequent dissociation from target tumor cell. Additionally, present proof-of-concept for dual DROP-CAR different small molecules, logic-gated synthetic receptors enabling STAT3 signaling. We demonstrate in vitro vivo function T cells conclude that approach holds important promise clinical application.

Язык: Английский

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Pyroptosis in Lung Cancer: The Emerging Role of Non-Coding RNAs

Pathology - Research and Practice, Год журнала: 2024, Номер 263, С. 155619 - 155619

Опубликована: Сен. 28, 2024

Язык: Английский

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Preclinical model for evaluating human TCRs against chimeric syngeneic tumors

Journal for ImmunoTherapy of Cancer, Год журнала: 2024, Номер 12(12), С. e009504 - e009504

Опубликована: Дек. 1, 2024

Background The adoptive cell transfer (ACT) of T receptor (TCR)-engineered cells targeting the HLA-A2-restricted epitope NY-ESO-1 157-165 (A2/NY) has yielded important clinical responses against several cancers. A variety approaches are being taken to augment tumor control by ACT including TCR affinity-optimization and T-cell coengineering strategies address suppressive microenvironment (TME). Most TCRs interest evaluated in immunocompromised mice enable human engraftment do not recapitulate dynamic interplay that occurs with endogenous immunity a treated patient. humanized mouse models have been described but they limitations immune reconstitution technically challenging implement. Here, we developed chimeric syngeneic model which A2Kb transgenic C57BL/6 engrafted B16 expressing A2Kb:NY as single chain trimer (SCT) murine A2/NY comprising variable fused constant regions. Methods We compared function supraphysiological affinity (wtc51m), computationally designed an optimal range (DMβ), near non-binding (V49I), engineered both primary lentiviral retroviral transduction, respectively. stably express on surface lines melanoma, ultimately achieving success SCT β2m GS linkers NY-peptide α1 HLA complex. studies were performed B16-A2Kb:NY tumor-bearing, non-preconditioned immune-competent HLA-A*0201/H-2Kb (A2Kb) tumors characterized post-transfer. Results observed significantly improved DMβ-T well superior infiltration upon TCR-T cells. Moreover, our model, able track favorable changes TME transfer. Conclusions robust, simple, inexpensive preclinical strategy for evaluating context fully competent system can aid development coengineered combination treatments translated clinic.

Язык: Английский

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DNA-Dependent Protein Kinase Inhibitors PI-103 and Samotolisib Augment CRISPR/Cas9 Knockin Efficiency in Human T Cells

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Окт. 18, 2024

1. Abstract The adoptive cell transfer of ex vivo expanded tumor infiltrating lymphocytes (i.e., TIL therapy) is a promising clinical strategy and recently FDA approved for melanoma but has major limitations including that not all tumors are inflamed. Moreover, tumor-specific clones can be rare in an exhausted state due to the suppressive microenvironment. These obstacles overcome by engineering autologous peripheral blood T cells with pre-selected receptors (TCRs) viral vector-mediated gene insertion. While transduction highly efficient, insertional site specific persistence oftentimes limited. In contrast, site-specific integration TCR into α chain ( TRAC ) locus CRISPR/Cas9 been shown enable more consistent physiological levels exogenous expression coupled superior control preclinical studies. Here, we sought improve efficiency mediated knockin (KI) primary human cells. addition previously reported DNA-dependent protein kinase inhibitor M3814, demonstrate PI-103 samotolisib markedly increase KI process GMP-compatible, while CC-115 had variable effect. Importantly, do negatively impact viability, fold-expansion nor phenotype conclude they suitable generation gene-modified use.

Язык: Английский

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Golden age of immunoengineering

Immunological Reviews, Год журнала: 2023, Номер 320(1), С. 4 - 9

Опубликована: Окт. 23, 2023

Immunology has long been the source of many significant medical breakthroughs, from vaccines for infections to therapeutics cancer, autoimmunity, and transplant rejection. Indeed, only diseases we have successfully eradicated, example, smallpox polio, were achieved through our understanding immune system. Furthermore, system often plays an unexpected role in outcome a treatment not designed engage For instance, chemotherapy or radiation therapies initially target cancer cells directly. However, subsequent investigations uncovered critical these stimulating As deepens, its involvement homeostasis surveillance almost every human organ becomes more apparent. bidirectional response between central nervous systems now recognized as major determinant some neurodegenerative (e.g., Parkinson's disease) psychiatric disorders. Other chronic diseases, such heart disease diabetes, are influenced by such, study mechanisms would be deemed incomplete without considering dynamic interaction aliment with This recognition poses challenge especially humans, because studying interest is no longer sufficient get whole picture. Due importance, developed modulate various diseases. A balance activation suppression maintaining healthy, functional uncontrolled overactive responses can lead autoimmunity tissue damage. Yet hyporesponsive render patient vulnerable infection development. Many current either enhance restrain systemic modulation tends generate severe side effects. Therefore, precise spatiotemporal control become focus next generation immunotherapy. In this issue, 13 reviews prepared most innovative research groups describing development tools strategies harness therapeutic applications. issue will comprehensive overview immunoengineering. Instead, it on applying protein genetic engineering improve safety, specificity, efficacy immunotherapies. system's direct all organs provides intriguing opportunity intervention. The system, while highly complex, also very accessible. One collect genetically modify primary cells, essentially converting them into smart drug delivery devices cell-killing machines active homing capability that remotely controlled small molecules, light, ultrasound. These attributes being explored treat immune-mediated (PMID: 37495877). Cytokines soluble proteins regulate exploited types. prominent examples interleukin- (IL) 2 interferon-gamma, which clinically approved. due their ability widespread inflammation, toxicity limited utility clinics. Much effort devoted improving performance cytokine therapy, yet success clinics marginal. Santonelli Wittrup1 elegantly summarize field discuss challenges developing therapy. They provide rationales based principles clinical data conventions highlight promising developments. Heller Spangler's team2 dive deeply biology IL-4/IL-13, specific class type inflammatory response. well-known protecting against multicellular pathogens involved responding allergens. addition providing biology, they thoroughly discussed diagnostic opportunities IL-4/IL-13 pathway synthetic gene circuits allergy, other Besides cytokines, molecules derived enormous biotechnological potential. Antibodies B encourage readers peruse excellent latest Recently, new molecule T-cell receptors (TCR) emerged compelling modality therapeutics. While less versatile than antibodies, TCR bind intracellular-derived peptides, thus accessing mutated intracellular undruggable targets. Dao, Scheinberg, team3 potential compare modalities, antibodies chimeric antigen (CAR). Interestingly, combined together create novel (Figure 1). Small biologics dominant form advances cell enabled modality—immune receptor (CAR) six FDA approvals hematological malignancies since 2017. Several cover foundation CAR Roybal his team outline considerations needed effective therapies.4 Maus colleagues specifically preclinical results using T digestive tract cancers hurdles technology cancer.5 Even impressive number short time, currently approved treatments represent possibilities. challenging part treating always killing enough sparing healthy tissues. typically requires recruitment cytotoxic agents tumor, fundamentally difficult address alone. Immune well suited tackle challenge, integrating complex precision. Hernandez-Lopez team6 summarized developments circuit design specificity safety controls 2A). Designing optimal crucial consideration ensuring appropriate function cellular therapies. An example where importance solid tumors, means suppressing evading suffer exhaustion stimulation cells. Multiple available issues related potency, one way engineer promote survival. Wang, Xu, recently discovered electrostatic signaling domain CARs. Their review7 discusses recent findings knowledge charge-based designing better CARs comprised several domains impact physiology different ways. Comprehensively exploring possible combinations too time-consuming. incorporating literature reduce space, efficient approach needed. Daniels Capponi8 artificial intelligence (AI) machine learning (ML) adoptive Specifically, work constructing library AI facilitate understand designs persistence large-scale undoubtedly sources therapy receptor, host Irving concept “coengineering”,9 highlighting benefit introducing multiple features besides tumor-targeting 2B). checkpoint inhibitors) cytokines inducibly overexpressed proliferation survival within tumor microenvironment. overexpression, directly modifying genome important coengineering approach. CRISPR/Cas essential tool engineering. Chen colleagues10 experimental unbiased screens creating potent CRISPR ever-expanding profoundly proven vehicle shortcomings. main TCR, leads graft vs. if allogeneically sourced. use source. drastically increases complexity cost manufacturing process. It widely should allogeneic situ engineered ensure access commercial viability. Deleting approaches intriguing, complexity. types do express vehicles CAR. Natural killer (NK) alternative player cancer. foundational works NK review Rezvani colleagues,11 Recent trials demonstrated exciting fewer effects inherently shorter lifespan Further activity could pave off-the-shelf, comparable but 3A). macrophages under investigation medium Macrophages naturally migrate accumulate routinely observed tumors. modified expressed induce antigen-dependent phagocytosis properties make attractive Our macrophage context Gill colleagues12 summarizing natural abundance tumor-homing over types, unlocking advantage. Most applications leveraged Autoimmunity rejection caused attacking patient's tissues allografts. Only subset auto- alloreactive. Systemic immunosuppression standard care, accompanied complications reduced capacity fight infections. Using approach, eradicate autoreactive alloreactive unwanted immunosuppression. identifying autoimmune cases remains challenging. leverage regulatory (Treg) CD4 suppress when activated 3B). Unsurprisingly, CAR-expressing Treg suppression. Levings colleagues13 detailed designer Treg's past achievements status. ongoing studies We golden age immunoengineering special Immunological Reviews illustrate immense disease-modifying, maybe even curative, untreatable are, part, accelerated convergence fields immunology. RNA powers unprecedented COVID situ, completely circumventing need cumbersome ex vivo processing. investigated encapsulate messenger (mRNA) encoding CAR) lipid nanoparticles deliver mRNA inside patient. mRNA/LNP-based was cardiac fibrotic mice 34990237). conventional aim type. how environment learned both CD8 Various combination utilizing inhibitors parts elicit efficacious anticipate purposeful aspects further unlock addition, carefully orchestrated seen patients eliminate before administering anti-cancer efficacy. Thus, temporal order multi-pronged regulation dimension accelerate progress thank Sarah Adams Arun Nambiar reading manuscript. W.W.W acknowledges support U01CA265713, R01EB029483, R01DK132576, R01EB031904. W.A.L. NIH grants U54CA244438, U01CA265697, RO1CA249018, R01CA258789 NSF grant 14-600. W.W.W. holds equity Senti Biosciences 4Immune Therapeutics. Gilead Sciences Intellia Therapeutics adviser Allogene Note applicable.

Язык: Английский

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Distinct host preconditioning regimens differentially impact the antitumor potency of adoptively transferred Th17 cells.

Опубликована: Дек. 18, 2023

Abstract Background Mechanisms by which distinct methods of host preconditioning impact the efficacy adoptively transferred antitumor T helper cells is unknown. Methods CD4 + with a transgenic TCR that recognize TRP-1 melanoma antigen were polarized to 17 (Th17) phenotype and then into melanoma-bearing mice preconditioned either total body irradiation or chemotherapy. Results We found non-myeloablative dose (TBI 5 Gy) was more effective than using an equivalently dosed chemotherapy (CTX at 200 mg/kg) augmenting therapeutic activity anti-tumor Th17 cells. Anti-tumor engrafted better following TBI regressed large established in all animals. Conversely, only half survived long-term when CTX infused anti-melanoma IL-17 IFN-g produced cells, detected animals given preconditioning. Interestingly, inflammatory cytokines (G-CSF, IL-6, MCP-1, IL-5, KC) significantly elevated serum versus after therapy. Conclusions Our results indicate, for first time, response, persistence, cytokine profiles resulting from therapy are impacted specific regimen This work important understanding mechanisms promote long-lived responses ACT, particularly as based cell therapies now emerging clinic.

Язык: Английский

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