Leukemia-intrinsic determinants of CAR-T response revealed by in vivo genome-wide CRISPR screening [scRNA-seq]
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE196142 | GEO | 2023/10/31
REPOSITORIES: GEO
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Project description:CAR-T cells are a promising new treatment for B cell malignancies. However, the majority of patients inevitably go on to experience disease relapse through largely unknown means. To investigate leukemia-intrinsic CAR-T resistance mechanisms, we performed genome-wide CRISPR-Cas9 loss-of-function screening in an immunocompetent murine model of B cell acute lymphoblastic leukemia (B-ALL). We identified IFN /JAK/STAT signaling and components of the antigen processing and presentation pathway as key mediators of resistance to anti-CD19 CAR-T therapy in vivo, but not in vitro. Transcriptional characterization of this model demonstrated an upregulation of these pathways in CAR-T treated relapsed tumors, and examination of data from CAR-T treated B-ALL patients revealed an association between poor outcomes and increased expression of JAK/STAT/MHC-I in leukemia cells. Together, our data identify an unexpected mechanism of resistance to immunotherapy, in which tumor cell interaction with IFN -secreting CAR-T cells in vivo activates the expression of an MHC-I inhibitory T cell program.
2023-10-31 | GSE196135 | GEO
Project description:Combined gene expression and DNA occupancy profiling identifies JAK/STAT signaling as a valid therapeutic target of t(8;21) AML t(8;21) is commonly associated with acute myeloid leukemia (AML). The resulting AML1-ETO fusion proteins are involved in the pathogenesis of AML. To identify novel molecular and therapeutic targets, we performed combined gene expression and promoter occupancy profiling using a primary leukemia initiating cell-enriched population induced by AML1-ETO9a (AE9a). CD45, a negative regulator of cytokine/growth factor receptor and JAK/STAT signaling, is greatly downregulated; furthermore JAK1 and JAK2 are upregulated in these leukemia cells. Consequently, JAK/STAT signaling is enhanced in the AE9a leukemia cells. Importantly, AE9a leukemia cells are highly susceptible to perturbation of JAK/STAT signaling, and a JAK2-selective inhibitor, TG101209, effectively targets these leukemia cells in vivo, suggesting the potential efficacy of JAK2 inhibitors in treating t(8;21) AML. Wild-type or AE9a leukemic samples in triplicate.
2012-09-04 | E-GEOD-15195 | biostudies-arrayexpress
Project description:The efficacy of chimeric antigen receptor (CAR)-engineered T cell therapy is suboptimal in most of the cancer types , necessitating further improvement in their therapeutic actions. However, enhancing antitumor T cell response inevitably confers an increased risk of cytokine release syndrome associated with monocyte-derived IL-6 secretion. Thus, a novel approach to simultaneously enhance therapeutic efficacy and safety is warranted. In this study, we developed a chimeric cytokine receptor composed of the extracellular domains of GP130 and IL6RA linked to the transmembrane and cytoplasmic domain of IL-7R mutant that constitutively activate JAK-STAT pathway (G6/7R). CAR-T cells with G6/7R efficiently absorbed and degraded monocyte-derived IL-6 both in vitro and in vivo. The G6/7R-expressing CAR-T cells showed superior expansion and persistence in vivo, which resulted in durable antitumor response in multple tumor models. Our strategy can be widely applicable to CAR-T cell therapy to enhance its efficacy and safety, irrespective of the target antigen.
2024-03-24 | GSE216734 | GEO
Project description:Maier2022 - Stochastic Dynamics of Type I Interferon Responses
Our study aims to determine whether and how biochemical noise affects the information transduced in the JAK-STAT signaling pathway and investigate the transition between basal and activated state. To this end, we studied the stochastic responses of MxA and IFIT1 expression in Huh7.5 cells stimulated with IFN-$\alpha$. Using fluorescent reporters under the control of the authentic promoter/enhancer region of IFIT1 and MxA we collected data displaying the differences between expressing and non-expressing cells for the marker genes in a time-course experiment. We hypothesize that the JAK-STAT signaling pathway efficiently transmits information under stochastic environments. To test our working hypothesis, we developed a detailed mathematical model using the obtained time-resolved flow cytometry data to describe the elements in the JAK-STAT signaling pathway at single-cell resolution. This model allowed us to systematically test the influence of intrinsic and extrinsic noise in the IFN response.
The developed model consists of 42 species and 62 reactions (reactions m1 to m62). To name the variables in the model we used the following conventions: 1) variables referring to mRNA are denoted by $m$ prefix. 2) Variables in phosphorylated use $p$ as prefix. 3) Gene promoters are represented by the gene's name in lowercase. 4) R1, R2, IR, AR and RC, represent the IFN receptor subunits, inactive, active and complex forms, respectively. 5) The compartment is superscripted to the species if the species exist in multiple compartments. A graphical representation of the interaction between variables in the model is given in Maier et. al 2022 (Fig. 1) and all reactions are listed in the Supplementary Information, Section S2.1.
Note that we publish the SBML model without the observables (e.g. exp_IRF9_n) as the change in the number of particles is defined in relation to the starting value in the COPASI model which is not supported in SBML format. In order to reproduce the parameter estimation without any extra worj, we recommend the direct download of the provided copasi model linked in the article.
This model is described in the article:
Stochastic Dynamics of Type I Interferon Responses
Benjamin D. Maier(*), Luis U. Aguilera(*), Sven Sahle, Pascal Mutz, Priyata Kalra, Christopher Dächert, Ralf Bartenschlager, Marco Binder, Ursula Kummer
PLOS Computational Biology, 2022
(*) Equally contributing authors
Abstract:
Interferon (IFN) activates the transcription of several hundred of IFN stimulated genes (ISGs) that constitute a highly effective antiviral defense program. Cell-to-cell variability in the induction of ISGs is well documented, but its source and effects are not completely understood. The molecular mechanisms behind this heterogeneity have been related to randomness in molecular events taking place during the JAK-STAT signaling pathway. Here, we study the sources of variability in the induction of the IFN-alpha response by using MxA and IFIT1 activation as read-out. To this end, we integrate time-resolved flow cytometry data and stochastic modeling of the JAK-STAT signaling pathway. The complexity of the IFN response was matched by fitting probability distributions to time-course flow cytometry snapshots. Both, experimental data and simulations confirmed that the MxA and IFIT1 induction circuits generate graded responses rather than all-or-none responses. Subsequently, we quantify the size of the intrinsic variability at different steps in the pathway. We found that stochastic effects are transiently strong during the ligand-receptor activation steps and the formation of the ISGF3 complex, but negligible for the final induction of the studied ISGs. We conclude that the JAK-STAT signaling pathway is a robust biological circuit that efficiently transmits information under stochastic environments.
2022-10-25 | MODEL2210070001 | BioModels
Project description:Combined gene expression and DNA occupancy profiling identifies JAK/STAT signaling as a valid therapeutic target of t(8;21) AML t(8;21) is commonly associated with acute myeloid leukemia (AML). The resulting AML1-ETO fusion proteins are involved in the pathogenesis of AML. To identify novel molecular and therapeutic targets, we performed combined gene expression and promoter occupancy profiling using a primary leukemia initiating cell-enriched population induced by AML1-ETO9a (AE9a). CD45, a negative regulator of cytokine/growth factor receptor and JAK/STAT signaling, is greatly downregulated; furthermore JAK1 and JAK2 are upregulated in these leukemia cells. Consequently, JAK/STAT signaling is enhanced in the AE9a leukemia cells. Importantly, AE9a leukemia cells are highly susceptible to perturbation of JAK/STAT signaling, and a JAK2-selective inhibitor, TG101209, effectively targets these leukemia cells in vivo, suggesting the potential efficacy of JAK2 inhibitors in treating t(8;21) AML.
2012-09-04 | GSE15195 | GEO
Project description:Chimeric antigen receptor (CAR)-expressing T-cells induce durable remissions in patients with relapsed/refractory B-cell malignancies. CARs are artificial constructs introduced into mature T-cells conferring a second, non-MHC restricted specificity in addition to the endogenous T-cell receptor (TCR). The impact of TCR activation on CAR T-cell efficacy in vivo has important implications for clinical optimization of CAR T-cell therapy, but cannot be systematically evaluated in xenograft models. Using an immunocompetent, syngeneic murine model of CD19-targeted CAR T-cell therapy for pre-B cell ALL, we demonstrate loss of CD8 CAR T-cell mediated clearance of leukemia associated with T-cell exhaustion and apoptosis when TCR antigen is present. CD4 CAR T-cells demonstrate equivalent cytotoxicity, as compared to CD8 CAR T-cells, and in contrast, retain in vivo efficacy in the presence of TCR stimulation. Gene expression profiles confirm increased exhaustion and apoptosis of CAR8 upon dual receptor stimulation compared to CAR4, and indicate inherent differences in T-cell pathways. Chimeric antigen receptor (CAR) T cells express two activating receptors, the CAR and the endogenous T cell receptor (TCR). CAR T cells can be derived from either CD8 or CD4 T cells to generate CAR8 and CAR4 cells, respectively. In vivo, CAR8 and CAR4 cells respond differently when simultaneously stimulated through the CAR and TCR.
2017-12-31 | GSE103632 | GEO
Project description:Organ transplant recipients (OTRs) on Cyclosporine A (CSA) are prone to catastrophic cutaneous squamous cell carcinoma (SCC). Allograft-sparing, cancer-targeting systemic treatments are unavailable. We have shown increased risk for catastrophic SCC in OTRs via CSA-mediated induction of Interleukin-22 (IL-22). Herein, we found CSA drives SCC proliferation and tumor growth through IL-22 and JAK/STAT pathway induction. We in turn inhibited SCC growth with an FDA-approved JAK 1/2 inhibitor, Ruxolitinib. In human SCC cells, greatest proliferative response to IL-22 and CSA treatment occurred in non-metastasizing lines. IL-22 treatment upregulated JAK1 and STAT1/3 in A431 SCC cells. JAK/STAT pathway genes were highly expressed in tumors from a cohort of CSA-exposed OTRs, and in SCC with high risk for metastasis. Compared to immunocompetent SCC, genes associated with innate immunity, response to DNA damage and p53 regulation were differentially expressed in SCC from OTRs. In nude mice engrafted with human A431 cells, IL-22 and CSA treatment increased tumor growth and upregulated IL-22 receptor, JAK1 and STAT 1/3 expression. Ruxolitinib treatment significantly reduced tumor volume and reversed the accelerated tumor growth. CSA and IL-22 exacerbate aggressive behavior in SCC. Targeting the IL-22 axis via selective JAK/STAT inhibition may reduce the progression of aggressive SCC in OTRs, without compromising immunosuppression. In this study, microarray data was used to compare normal skin to immunocompetent SCC, to transplant associated SCC (TSCC)
2018-07-18 | GSE117247 | GEO
Project description:Activation of the immune system is a way for host tissue to defend itself against tumor growth. Hence, treatment strategies that are based on immunomodulation are on the rise. Conventional cytostatic drugs such as the anthracycline doxorubicin can also activate immune cell functions of macrophages and natural killer cells. In addition, cytotoxicity of doxorubicin can be enhanced by combining this drug with the cytokine IFN-alpha. Although doxorubicin is one of the most applied cytostatics, the molecular mechanisms of its immunomodulation ability are not investigated thoroughly. In microarray analyses of HeLa cells, a set of 19 genes related to interferon signaling was significantly overrepresented among genes regulated by doxorubicin exposure including STAT-1, -2, IRF9, NMI, and caspase 1. Regulation of these genes by doxorubicin was verified with Real-Time PCR and immunoblotting. An enhanced secretion of IFN-alpha was observed when HeLa cells were exposed to doxorubicin as compared to untreated cells. IFN-alpha neutralizing antibodies and inhibitors of JAK-STAT signaling (ATA and AG490) significantly abolished doxorubicin-stimulated expression of interferon signaling-related genes. Furthermore, inhibition of JAK-STAT signaling significantly reduced doxorubicin induced caspase 3 activation and desensitized HeLa cells to doxorubicin cytotoxicity. In conclusion, we demonstrate that doxorubicin induces interferon-responsive genes via IFN-alpha-JAK-STAT1 signaling and that this pathway is relevant for doxorubicinM-bM-^@M-^Ys cytotoxicity in HeLa cells. As immunomodulation is a promising strategy in anticancer treatment, this novel mode of action of doxorubicin may help to further improve the use of this drug among different types of anticancer treatment strategies. One batch of HeLa Cell culture treated with doxorubicin and DMSO (control) were used for screening of global changes at the transcriptome level.
2012-05-22 | E-GEOD-30988 | biostudies-arrayexpress
Project description:CAR gene expression on precursor T cells (preTs) that have been generated in vitro from engineered hematopoietic stem cells may reduce the risk of leukemia relapse upon co-transplantation in an MHC-independent manner. Setting the expression of an anti-murine CD19 CAR under the control of an inducible promoter allowed time-dependent assessment of its impact on in vivo development. Adoptive transfer of engineered preTs along with T cell-depleted bone marrow resulted in potent anti-leukemia effects even in fully MHC-class I and II mismatched recipients. Anti-leukemia effects entirely relied on the in vivo development of CAR-expressing natural killer cells since NK cell depletion after transplantation resulted in a complete abrogation of anti-leukemic efficacy. Gene expression profiling revealed that enforced CAR expression on preTs led to an activated shift of transcriptional key factors to those being associated with NK cell development such as Nfil3, Id2, and Eomes. In contrast, CAR expression on preTs blocked thymic seeding completely and prevented further T cell development. The CAR-expressing NK cell progeny of transferred preTs persisted for up to 60 days post transplantation allowing for complete B-cell recovery thereafter. Taken together, we provide new functional insights for the use of CAR-engineered preTs allowing for potent anti-leukemia efficiency within a limited time window for on target/off tumor effects.
2017-10-03 | GSE104512 | GEO
Project description:Natural Killer/T-cell lymphoma (NKTL) is a rare type of aggressive and heterogeneous non-Hodgkin's lymphoma (NHL) with poor prognosis and limited therapeutic options. Here, we demonstrate that chidamide, a novel histone deacetylase (HDAC) inhibitor, is effective in treating relapsed/refractory NKTL patients, achieving an overall response rate of 39.3%, with 17.9% complete response rate. However, the clinical response to chidamide remains variable as more than half of the patients exhibit primary resistance, limiting its utility in NKTL treatment. To unravel the resistance mechanisms of chidamide, we performed integrative transcriptomic and chromatin profiling of sensitive and resistant NKTL cells. Our results revealed that aberrant JAK-STAT signaling remodels the chromatin and confers resistance to chidamide. Subsequently, inhibition of JAK-STAT activity could overcome resistance to chidamide by reprogramming the chromatin from a resistant to sensitive state, leading to synergistic anti-tumor effect. More importantly, our clinical data demonstrated that combinatorial therapy with chidamide and JAK inhibitor ruxolitinib is effective against chidamide-resistant NKTL. In addition, we identified TNFRSF8 (CD30), a downstream target of JAK-STAT pathway, as a potential biomarker that could predict NKTL sensitivity to chidamide. Collectively, our study suggests that chidamide, in combination with JAK-STAT inhibitors, can be a novel targeted therapy in the standard of care for NKTL.
2023-03-02 | GSE152185 | GEO