Whole-genome sequencing reveals complex genomic features underlying anti-CD19 CAR T-cell treatment failure in lymphoma
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PROVIDER: PRJNA869173 | ENA |
REPOSITORIES: ENA
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Project description:Whole-genome sequencing reveals complex genomic features underlying anti-CD19 CAR T-cell treatment failure in lymphoma
| PRJNA869172 | ENA
Project description:Chimeric antigen receptor T-cell (CAR-T) therapy is at the forefront of cell immunotherapy. In this study, we generated an anti-CD19 CAR-Jurkat T cell line using a locally produced second-generation CD19 CAR construct, enabling us to analyze early proteomic changes crucial for understanding the signaling pathways and mechanisms of action of this CAR-T cell. SILAC-heavy tagged RAJI B-cells and anti-CD19 CAR-Jurkat T-cells were co-cultured for ten minutes. The proteomic profiles were obtained via DIA methodology on the Orbitrap Astral LC-MS/MS platform. The proteome was extensively covered, resulting in approximately 8800 protein identifications at 1% FDR. The effector CAR-Jurkat cells showed proteomic changes involving antigen presentation by CD74. The target RAJI B-cells exhibited more significant alterations, such as CD28, essential for T-cell survival and activation.
2025-07-30 | PXD062748 | Pride
Project description:Chimeric antigen receptor T cell (CAR-T) targeting the CD19 antigen represents an innovative therapeutic approach to improve the outcome of relapsed or refractory B-cell acute lymphoblastic leukemia (B-ALL). Yet, despite a high initial remission rate, CAR-T therapy ultimately fails for some patients. Notably, around half of relapsing patients develop CD19 negative (CD19neg) B-ALL allowing leukemic cells to evade CD19-targeted therapy. Herein, we investigate leukemic cells of a relapsing B-ALL patient, at two-time points: before (T1) and after (T2) anti-CD19 CAR-T treatment. We show that at T2, the B-ALL relapse is CD19 negative due to the expression of a non-functional CD19 transcript retaining intron 2. Then, using single-cell RNA sequencing (scRNAseq) approach, we demonstrate that CD19neg leukemic cells were present before CAR-T cell therapy and thus that the relapse results from the selection of these rare CD19neg B-ALL clones. In conclusion, our study shows that scRNAseq profiling can reveal pre-existing CD19neg subclones, raising the possibility to assess the risk of targeted therapy failure.
2020-12-11 | GSE153697 | GEO
Project description:Background and methods: CD19-directed CAR T-cell therapy (CAR-T) has emerged as a promising treatment modality for multiple B-cell malignancies. However, its utility is hampered by a unique toxicity profile that classically includes cytokine release syndrome (CRS) and neurotoxicity. Hematological toxicity represents a vexing and common side effect. Cytopenia can occur long after lymphodepleting chemotherapy and resolution of CRS and is often prolonged and biphasic in nature. Some patients have been described to develop severe bone marrow (BM) failure, which can predispose for severe infections and high non-relapse mortality. Overall, the underlying pathomechanisms of CAR-T-related hematotoxicity remain poorly understood. We performed flow cytometry, serum cytokine profiling, single cell transcriptome and TCR-sequencing of PBMCs from pre-/post-treatment sampels of a patient with DLBCL-type Richter Transformation receiving tisagenlecleucel in a standard-of-care setting and present one marrow failure. Results: The advent of chimeric antigen receptor (CAR) T-cell therapy has revolutionized the treatment landscape of refractory B-cell malignancies. Real-world evidence has highlighted the high incidence of hematological toxicity, including prolonged and profound cytopenia. Here, we present the case of bone marrow failure in a 57-year old man with DLBCL-type Richter Transformation receiving tisagenlecleucel in a standard-of-care setting. The clinical course was notable for underlying bone marrow infiltration, severe cytokine release syndrome and multiple infectious complications in the setting of prolonged, profound neutropenia. Cytokine profiling revealed a signature consistent with acquired aplastic anemia – including downregulation of CD40-L and EGF. Flow cytometric analyses demonstrated expansion of both CAR and non-CAR bearing CD8+ CD57+ T-cells in the bone marrow, an immunophenotype linked to oligoclonality in aplastic anemia. On a single-cell level, this was accompanied by T-cell receptor Vβ oligoclonal expansion and post-CAR clonal drift. Gene expression profiling revealed upregulation of exhaustion markers on (CAR) T-cells and decreased expression of genes involved in STAT signalling and inflammatory response. Conclusion: In conclusion, this case highlights the complex nature of CAR-T-related hematological toxicity and introduces oligoclonal (CAR) T-cell expansion as a potential contributing pathomechanism.
2023-01-29 | GSE201704 | GEO
Project description:To characterize transfer of molecules from target cells into CAR T cells via trogocytosis we cultured NALM-6 leukemia cell line expressing a CD19-mCherry fusion protein with CAR T cells. NALM6-CD19-mCherry were loaded with heavy amino acid and cocultured with CAR T cells for 1 hour. CAR T cells were next sorted into two fractions, mCherry-positive (TrogPos), and -negative (TrogNeg). Proteomics analysis revealed the presence of targeted antigen (CD19) in the TrogPos only.
2019-03-29 | PXD012833 | Pride
Project description:Comparation of the anti-CD19 CAR-T and the NMN treated anti-CD19 CAR-T
| PRJNA824334 | ENA
Project description:Long-lived, self-renewing, multipotent T memory stem cells (TSCM) can trigger profound and sustained tumor regression but their rareness poses a major hurdle to their clinical application. Presently, clinically compliant procedures to generate relevant numbers of this T cell population are undefined. Here, we provide a strategy for deriving large numbers of clinical grade tumor-redirected TSCM cells starting from naïve precursors. CD8+CD62L+CD45RA+ naïve T cells enriched by streptamer-based serial positive selection were activated by CD3/CD28 engagement in the presence of IL-7, IL-21 and the glycogen synthase-3β inhibitor TWS119, and genetically engineered to express a CD19-specific chimeric antigen receptor (CD19-CAR). These conditions allowed for the generation of CD19-CAR modified TSCM cells that were phenotypically, functionally and transcriptomically equivalent to their naturally occurring counterpart. Compared with T cell products currently under clinical investigation, CD19-CAR modified TSCM cells exhibit enhanced metabolic fitness, persistence and anti-tumor activity against systemic acute lymphoblastic leukemia xenografts. Based on these findings, we have initiated a phase 1 clinical study to evaluate the activity of CD19-CAR modified TSCM in patients with B-cell malignancies refractory to prior allogeneic hematopoietic stem cell transplantation. Three healthy human blood donors provided lymphocyte-enriched apheresis blood for this study after informed consent. From all samples, total RNA was isolated using an miRNeasy Mini Kit (Qiagen), processed by Ambionâ??s WT expression kit, fragmented and labeled with a WT Terminal Labeling Kit (Affymetrix), hybridized to WT Human Gene 1.0 ST arrays (Affymetrix) and stained on a Genechip Fluidics Station 450 (Affymetrix), all according to the respective manufacturer's instructions. Samples represent exon-level and gene-level analyses.
2016-06-29 | E-GEOD-68003 | biostudies-arrayexpress
Project description:CAR T-cell therapy has led to tremendous successes in the treatment of B-cell malignancies. However, 30%-50% of treated patients relapse – often with reduced target antigen expression. We report that anti-CD19 CAR T-cells cause a rapid reduction of CD19 expression within hours in CAR-T exposed CD19+ B-ALL cells. Initially, anti-CD19 CAR T-cells cause CD19 clusters at the T-cell – leukemia cell interface followed by CD19 internalization and decreased CD19 surface expression. Subsequently, CD19 expression is repressed by transcriptional rewiring. Using single-cell RNA-seq and single-cell ATAC-seq we demonstrate that a subset of CD19low cells that are refractory to CAR T-cell killing employ transcriptional programs of physiological B-cell activation and germinal center reaction in order to sustain decreased CD19 expression. Inhibiting B-cell activation programs with the BTK inhibitor ibrutinib increased the cytotoxic efficacy of anti-CD19 CAR T-cells without effecting CAR T-cell viability. These results demonstrate transcriptional plasticity as an underlying mechanism of CAR T-resistance and highlight the importance of combining CAR T-cell therapy with targeted therapies that aim to overcome this plasticity.
2022-06-09 | GSE176418 | GEO
Project description:<p>Anti-CD19 chimeric antigen receptor (CAR) T-cell therapy for relapsed or refractory (r/r) large B-cell lymphoma (LBCL) results in durable response in only a subset of patients. MYC overexpression in LBCL tumors is associated with poor response to treatment. We tested whether a MYC-driven polyamine signature, as a liquid biopsy, is predictive of response to anti-CD19 CAR-T therapy in patients with r/r LBCL. Elevated plasma acetylated polyamines were associated with non-durable response. Concordantly, increased expression of spermidine synthase, a key enzyme which regulates levels of acetylated spermidine, was prognostic for survival in r/r LBCL. A broad metabolite screen identified additional markers which resulted in a 6-marker panel (6MetP) consisting of acetylspermidine, diacetylspermidine and lysophospholipids which was validated in an independent set from another institution as predictive of non-durable response to CAR T therapy. A polyamine centric metabolomics liquid biopsy panel has predictive value for response to CAR-T therapy in r/r LBCL. </p>
2022-10-31 | MTBLS5381 | MetaboLights
Project description:The phase 3 ZUMA-7 trial in second-line large B-cell lymphoma demonstrated superiority of anti-CD19 CAR T-cell therapy (axicabtagene ciloleucel; axi-cel) over standard of care (SOC; salvage chemotherapy followed by hematopoietic transplantation). Here, we present a prespecified exploratory analysis examining the association between pretreatment tumor characteristics and the efficacy of axi-cel versus SOC. B-cell gene expression signature (GES) and CD19 expression significantly associated with improved event-free survival (EFS) for axi-cel (P=.0002 for B-cell GES; P=.0165 for CD19 expression) but not SOC (P=.9374 for B-cell GES; P=.5526 for CD19 expression). Axi-cel showed superior EFS over SOC irrespective of B-cell GES and CD19 expression (P=8.56e–9 for B-cell GES high; P=.0019 for B-cell GES low; P=3.85e–9 for CD19 gene high; P=.0017 for CD19 gene low). Low CD19 expression in malignant cells correlated with a tumor GES consisting of immune suppressive stromal and myeloid genes, highlighting the inter-relation between malignant cell features and immune contexture substantially impacting axi-cel outcomes. Tumor burden, lactate dehydrogenase, and cell-of-origin impacted SOC more than axi-cel outcomes. T-cell activation and B-cell GES, which are associated with improved axi-cel outcome, decreased with increasing lines of therapy. These data highlight differences in resistance mechanisms to axi-cel and SOC and support earlier intervention with axi-cel.
2024-02-26 | GSE248835 | GEO