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Atypical acute promyelocytic leukemia (aAPL) presents a complex landscape of retinoic acid receptor (RAR) fusion genes beyond the well-known PML::RARA fusion. Among these, 31 individually rare RARA and RARG fusion genes have been documented, often reported in the canonical X::RAR bipartite fusion form. Intriguingly, some artificially mimicked bipartite X::RAR fusions respond well to all-trans retinoic acid (ATRA) in vitro, contrasting with the ATRA resistance observed in patients. To unravel the underlying mechanisms, we conducted a comprehensive molecular investigation into the fusion transcripts in 27 RARA fusion gene-positive aAPL (RARA-aAPL) and 21 RARG-aAPL cases. Our analysis revealed an unexpected novel form of X::RAR::X or X::RAR::Y-type tripartite fusions in certain RARA- and all RARG-aAPL cases, with shared features and notable differences between these two disease subgroups. In RARA-aAPL cases, the occurrence of RARA 3' splices was associated with their 5' fusion partner genes, mapping across the coding region of helix 11_12 (H11_12) within the ligand-binding domain (LBD), resulting in LBD-H12 or H11_12 truncation. In RARG-aAPL cases, RARG 3' splices were consistently localized to the terminus of exon 9, leading to LBD-H11_12 truncation. Significant differences were also observed between RARA and RARG 5' splice patterns. Our analysis also revealed extensive involvement of transposable elements in constructing RARA and RARG 3' fusions, suggesting transposition mechanisms for fusion gene ontogeny. Both protein structural analysis and experimental results highlighted the pivotal role of LBD-H11_12/H12 truncation in driving ATRA unresponsiveness and leukemogenesis in tripartite fusion-positive aAPL, through a protein allosteric dysfunction mechanism.
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BACKGROUND: Colorectal cancer is a malignant tumor of the digestive system originating from abnormal cell proliferation in the colon or rectum, often leading to gastrointestinal symptoms and severe health issues. Nucleotide metabolism, which encompasses the synthesis of DNA and RNA, is a pivotal cellular biochemical process that significantly impacts both the progression and therapeutic strategies of colorectal cancer METHODS: For single-cell RNA sequencing (scRNA-seq), five functions were employed to calculate scores related to nucleotide metabolism. Cell developmental trajectory analysis and intercellular interaction analysis were utilized to explore the metabolic characteristics and communication patterns of different epithelial cells. These findings were further validated using spatial transcriptome RNA sequencing (stRNA-seq). A risk model was constructed using expression profile data from TCGA and GEO cohorts to optimize clinical decision-making. Key nucleotide metabolism-related genes (NMRGs) were functionally validated by further in vitro experiments. RESULTS: In both scRNA-seq and stRNA-seq, colorectal cancer (CRC) exhibited unique cellular heterogeneity, with myeloid cells and epithelial cells in tumor samples displaying higher nucleotide metabolism scores. Analysis of intercellular communication revealed enhanced signaling pathways and ligand-receptor interactions between epithelial cells with high nucleotide metabolism and fibroblasts. Spatial transcriptome sequencing confirmed elevated nucleotide metabolism states in the core region of tumor tissue. After identifying differentially expressed NMRGs in epithelial cells, a risk prognostic model based on four genes effectively predicted overall survival and immunotherapy outcomes in patients. High-risk group patients exhibited an immunosuppressive microenvironment and relatively poorer prognosis and responses to chemotherapy and immunotherapy. Finally, based on data analysis and a series of cellular functional experiments, ACOX1 and CPT2 were identified as novel therapeutic targets for CRC. CONCLUSION: In this study, a comprehensive analysis of NMRGs in CRC was conducted using a combination of single-cell sequencing, spatial transcriptome sequencing, and high-throughput data. The prognostic model constructed with NMRGs shows potential as a standalone prognostic marker for colorectal cancer patients and may significantly influence the development of personalized treatment approaches for CRC.
Assuntos
Neoplasias Colorretais , MicroRNAs , Humanos , RNA-Seq , Nucleotídeos , Análise da Expressão Gênica de Célula Única , Transcriptoma , Redes e Vias Metabólicas , Neoplasias Colorretais/genética , Microambiente Tumoral/genéticaRESUMO
CD7-targeted chimeric antigen receptor T-cell (CAR-T) therapy has shown promising initial complete remission (CR) rates in patients with refractory or relapsed (r/r) T-cell acute lymphoblastic leukaemia and lymphoblastic lymphoma (T-ALL/LBL). To enhance the remission duration, consolidation with allogeneic haematopoietic stem cell transplantation (allo-HSCT) is considered. Our study delved into the outcomes of 34 patients with r/r T-ALL/LBL who underwent allo-HSCT after achieving CR with autologous CD7 CAR-T therapy. These were compared with 124 consecutive T-ALL/LBL patients who received allo-HSCT in CR following chemotherapy. The study revealed that both the CAR-T and chemotherapy cohorts exhibited comparable 2-year overall survival (OS) (61.9% [95% CI, 44.1-78.1] vs. 67.6% [95% CI, 57.5-76.9], p = 0.210), leukaemia-free survival (LFS) (62.3% [95% CI, 44.6-78.4] vs. 62.0% [95% CI, 51.8-71.7], p = 0.548), non-relapse mortality (NRM) rates (32.0% [95% CI, 19.0-54.0] vs. 25.3% [95% CI, 17.9-35.8], p = 0.288) and relapse incidence rates (8.8% [95% CI, 3.0-26.0] vs. 15.8% [95% CI, 9.8-25.2], p = 0.557). Patients aged ≤14 in the CD7 CAR-T group achieved high 2-year OS and LFS rates of 87.5%. Our study indicates that CD7 CAR-T therapy followed by allo-HSCT is not only effective and safe for r/r T-ALL/LBL patients but also on par with the outcomes of those achieving CR through chemotherapy, without increasing NRM.
Assuntos
Antígenos CD7 , Transplante de Células-Tronco Hematopoéticas , Imunoterapia Adotiva , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Indução de Remissão , Humanos , Masculino , Feminino , Transplante de Células-Tronco Hematopoéticas/métodos , Adulto , Adolescente , Pessoa de Meia-Idade , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Leucemia-Linfoma Linfoblástico de Células T Precursoras/terapia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/mortalidade , Adulto Jovem , Criança , Recidiva , Transplante Homólogo , Receptores de Antígenos Quiméricos/uso terapêutico , Resultado do Tratamento , Pré-Escolar , Taxa de SobrevidaRESUMO
Derivation of CD7-targeted chimeric antigen receptor (7CAR) T cells often requires genetic manipulations to ablate the CD7 gene or block CD7 cell surface expression. Our novel approach deriving naturally selected 7CAR (NS7CAR) T cells from bulk T cells was able to overcome major fratricide by minimizing accessible CD7 epitopes. The CD7 molecules of NS7CAR T cells were masked or sequestered by the CD7-targeting CAR. Compared with sorted CD7-negative 7CAR T cells and CD7 knocked-out 7CAR T cells, NS7CAR exhibited similar or superior therapeutic properties, including a greater percentage of CAR+ cells and a higher proportion of CD8+ central memory T cells. In our first-in-human phase 1 trial (NCT04572308), 20 patients with relapsed/refractory T-cell acute lymphoblastic leukemia (T-ALL) (n = 14) and T-cell lymphoblastic lymphoma (T-LBL) (n = 6) were treated with NS7CAR. Nineteen patients achieved minimal residual disease negative complete remission (CR) in the bone marrow (BM) by day 28, and 5 of 9 patients achieved extramedullary CR. With a median follow-up of 142.5 (32-311) days after infusion, 14 patients subsequently received allogeneic hematopoietic stem cell transplant (10 consolidative, 4 salvage) following NS7CAR infusion with no relapses to date. Of the 6 patients who did not receive a transplant, 4 remained in CR at a median time of 54 (32-180) days. Eighteen patients experienced mild cytokine release syndrome (CRS) (grade ≤2), 1 developed grade 3 CRS, and 2 had grade 1 neurotoxicity. These results indicate that NS7CAR-T therapy is a safe and highly effective treatment for T-ALL/LBL. More patients and longer follow-up are needed for validation.
Assuntos
Leucemia-Linfoma Linfoblástico de Células Precursoras , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Receptores de Antígenos Quiméricos , Antígenos CD19 , Humanos , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/uso terapêutico , Linfócitos TRESUMO
Two recent guidelines, the 5th edition of the World Health Organization Classification of Haematolymphoid Tumours (WHO-HAEM5) and the International Consensus Classification (ICC), were published to refine the diagnostic criteria of acute myeloid leukemia (AML). They both consider genomic features more extensively and expand molecularly defined AML subtypes. In this study, we compared the classifications of 1135 AML cases under both criteria. According to WHO-HAEM5 and ICC, the integration of whole transcriptome sequencing, targeted gene mutation screening, and conventional cytogenetic analysis identified defining genetic abnormalities in 89% and 90% of AML patients, respectively. The classifications displayed discrepancies in 16% of AML cases after being classified using the two guidelines, respectively. Both new criteria significantly reduce the number of cases defined by morphology and differentiation. However, their clinical implementation heavily relies on comprehensive and sophisticated genomic analysis, including genome and transcriptome levels, alongside the assessment of pathogenetic somatic and germline variations. Discrepancies between WHO-HAEM5 and ICC, such as the assignment of RUNX1 mutations, the rationality of designating AML with mutated TP53 as a unique entity, and the scope of rare genetic fusions, along with the priority of concurrent AML-defining genetic abnormalities, are still pending questions requiring further research for more elucidated insights.
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Leucemia Mieloide Aguda , Humanos , Consenso , Mutação , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Genômica , Organização Mundial da SaúdeRESUMO
The adaptor proteins NCK1 and NCK2 are well-established signalling nodes that regulate diverse biological processes including cell proliferation and actin dynamics in many tissue types. Here we have investigated the distribution and function of Nck1 and Nck2 in the developing mouse mammary gland. Using publicly available single-cell RNA sequencing data, we uncovered distinct expression profiles between the two paralogs. Nck1 showed widespread expression in luminal, basal, stromal and endothelial cells, while Nck2 was restricted to luminal and basal cells, with prominent enrichment in hormone-sensing luminal subtypes. Next, using mice with global knockout of Nck1 or Nck2, we assessed mammary gland development during and after puberty (5, 8 and 12 weeks of age). Mice lacking Nck1 or Nck2 displayed significant defects in ductal outgrowth and branching at 5 weeks compared to controls, and the defects persisted in Nck2 knockout mice at 8 weeks before normalizing at 12 weeks. These defects were accompanied by an increase in epithelial cell proliferation at 5 weeks and a decrease at 8 weeks in both Nck1 and Nck2 knockout mice. We also profiled expression of several key genes associated with mammary gland development at these timepoints and detected temporal changes in transcript levels of hormone receptors as well as effectors of cell proliferation and migration in Nck1 and Nck2 knockout mice, in line with the distinct phenotypes observed at 5 and 8 weeks. Together these studies reveal a requirement for NCK proteins in mammary gland morphogenesis, and suggest that deregulation of Nck expression could drive breast cancer progression and metastasis.
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Proteínas Adaptadoras de Transdução de Sinal , Glândulas Mamárias Animais , Animais , Camundongos , Camundongos Knockout , Camundongos Endogâmicos C57BL , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/crescimento & desenvolvimento , Glândulas Mamárias Animais/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proliferação de Células , Células Epiteliais/citologia , Expressão GênicaRESUMO
This study aimed to investigate the relationship between anomalous DNA nucleotidylexotransferase (DNTT) activation and the mutagenesis of gene length mutations (LMs) in acute myeloid leukemia (AML), and the relevance of their prognosis in antithymocyte globulin (ATG)-based regimen allogeneic hematopoietic stem cell transplantation (allo-HSCT). A cohort of 578 AML cases was enrolled. Next-generation sequencing was performed to screen mutations of 86 leukemia driver genes. RNA-seq was used to analyze gene expression. Prognostic analysis was investigated in 239 AML cases who underwent ATG-based regimen allo-HSCT. We report a refined subtyping algorithm of LMs (type I-IV) based on sequence anatomy considering the TdT-aided mutagenesis mechanism. GC content adjacent to LM junctions, inserted nontemplate nucleotide bases, and DNTT expression analysis supported the DNTT activation and TdT-aided mutagenesis in type II/III LMs in the total AML cohort. Both single-variate and multivariate analyses showed a better overall survival of FLT3 type III compared to type I in a subset of ATG-based regimen allo-HSCT cases. The novel LM subtyping algorithm not only deciphers the etiology of the mutagenesis of LMs but also helps to fine-tune prognosis differentiation in AML. The possible prognostic versatility of this novel LM subtyping algorithm in terms of chemotherapy, targeted therapy, and allo-HSCT merits further investigation.
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Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Humanos , DNA Nucleotidilexotransferase/genética , Soro Antilinfocitário/genética , Soro Antilinfocitário/uso terapêutico , Leucemia Mieloide Aguda/terapia , Leucemia Mieloide Aguda/tratamento farmacológico , Mutação , Estudos RetrospectivosRESUMO
While the use of chimeric antigen receptor-T (CAR-T) therapy for T-cell malignancies is in the early stage of clinical trials, it exhibits substantial potential to offer long-term remission for patients with refractory/relapsed (R/R) T-cell malignancies. In our phase I/II clinical trials, 65 pediatric and adult patients with R/R T-cell acute lymphoblastic leukemia and lymphoblastic lymphoma (T-ALL/LBL) were enrolled (NCT04572308 and NCT04916860). Of these, 60 participants (T-ALL 35, T-LBL 25) received a single dose of naturally selected anti-CD7 CAR (NS7CAR) T cells at three levels: a low dose (5 × 105 /kg), a medium dose (1 to 1.5 × 106 /kg), and a high dose (2 × 106 /kg). On day 28, 94.4% of patients achieved deep complete remission (CR) in bone marrow. Among the 32 patients with extramedullary disease, 78.1% showed response, with 56.3% in CR and 21.9% in partial remission. The 2-year overall survival and progression-free survival (PFS) were 63.5% (95% CI 47.7-79.4) and 53.7% (95% CI, 38.9-68.6), with no difference between pediatric and adult patients. PFS was significantly higher among the 37 CR patients who proceeded with consolidation transplant than the 10 patients who did not with 1-year PFS 67.2% (95% CI 51.9-82.4) versus 15.0% (95% CI 0-40.2), p < .0001. Of the 10 CR patients without transplants, eight relapsed, while two sustained CR on day 128, and day 180, respectively. Cytokine release syndrome occurred in 91.7% of patients (grade 1/2 in 80.0%, grade 3/4 in 11.7%) and 5% of patients had neurotoxicity. NS7CAR-T therapy is effective in treating R/R T-ALL/LBL patients with promising PFS while maintaining a manageable safety profile.
Assuntos
Linfoma de Células T Periférico , Leucemia-Linfoma Linfoblástico de Células Precursoras , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Receptores de Antígenos Quiméricos , Adulto , Humanos , Criança , Receptores de Antígenos Quiméricos/uso terapêutico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Imunoterapia Adotiva/efeitos adversos , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Linfócitos T , Linfoma de Células T Periférico/tratamento farmacológico , Recidiva , Terapia Baseada em Transplante de Células e Tecidos , Antígenos CD19RESUMO
BACKGROUND: Maintenance of the kidney filtration barrier requires coordinated interactions between podocytes and the underlying glomerular basement membrane (GBM). GBM ligands bind podocyte integrins, which triggers actin-based signaling events critical for adhesion. Nck1/2 adaptors have emerged as essential regulators of podocyte cytoskeletal dynamics. However, the precise signaling mechanisms mediated by Nck1/2 adaptors in podocytes remain to be fully elucidated. METHODS: We generated podocytes deficient in Nck1 and Nck2 and used transcriptomic approaches to profile expression differences. Proteomic techniques identified specific binding partners for Nck1 and Nck2 in podocytes. We used cultured podocytes and mice deficient in Nck1 and/or Nck2, along with podocyte injury models, to comprehensively verify our findings. RESULTS: Compound loss of Nck1/2 altered expression of genes involved in actin binding, cell adhesion, and extracellular matrix composition. Accordingly, Nck1/2-deficient podocytes showed defects in actin organization and cell adhesion in vitro, with podocyte detachment and altered GBM morphology present in vivo. We identified distinct interactomes for Nck1 and Nck2 and uncovered a mechanism by which Nck1 and Nck2 cooperate to regulate actin bundling at focal adhesions via α actinin-4. Furthermore, loss of Nck1 or Nck2 resulted in increased matrix deposition in vivo, with more prominent defects in Nck2-deficient mice, consistent with enhanced susceptibility to podocyte injury. CONCLUSION: These findings reveal distinct, yet complementary, roles for Nck proteins in regulating podocyte adhesion, controlling GBM composition, and sustaining filtration barrier integrity.
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Podócitos , Actinina/metabolismo , Actinas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Membrana Basal Glomerular/metabolismo , Camundongos , Proteínas Oncogênicas/metabolismo , Podócitos/metabolismo , ProteômicaRESUMO
Haematopoietic cell transplantation (HCT) is the only curative treatment for haematological complications in patients with Fanconi anaemia (FA). Haploidentical (haplo-) HCT is a promising alternative for FA. We aimed to analyse the outcomes of unmanipulated haplo-HCT in patients with FA with radiation-free conditioning. A total of 56 patients from 11 centres between 2013 and 2021 in China were retrospectively analysed. The mean (SD) cumulative incidence was 96.4% (0.08%) for 30-day neutrophil engraftment and 85.5% (0.24%) for 100-day platelet engraftment. With a median (range) follow-up of 2.4 (0.2-5.8) years, favourable mean (SD) overall survival of 80.9% (5.5%) and event-free survival of 79.3% (5.6%) were achieved. The mean (SD) incidences of acute graft-versus-host disease (aGvHD) Grade II-IV and Grade III-IV were 55.4% (0.45%) and 42.9 (0.45%) respectively. The mean (SD) cumulative incidence of 3-year chronic graft-versus-host disease (cGvHD) was 34.7% (0.86%) and that of moderate-to-severe cGvHD was 9.0% (0.19%). Our data demonstrate that in unmanipulated haplo-HCT for patients with FA, radiation-free regimens based on fludarabine and low-dose cyclophosphamide ± busulfan achieved favourable engraftment and survival with relatively high incidences of aGvHD and cGvHD. These results prompt the use of low-intensity conditioning without radiation and intensive GvHD prophylaxis when considering unmanipulated haplo-HCT in patients with FA.
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Anemia de Fanconi , Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Humanos , Doença Enxerto-Hospedeiro/etiologia , Doença Enxerto-Hospedeiro/prevenção & controle , Bussulfano/uso terapêutico , Estudos Retrospectivos , Condicionamento Pré-Transplante/efeitos adversos , Condicionamento Pré-Transplante/métodos , Anemia de Fanconi/terapia , Anemia de Fanconi/complicações , Medula Óssea , Recidiva Local de Neoplasia/tratamento farmacológico , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Transplante de Células-Tronco Hematopoéticas/métodos , Ciclofosfamida/uso terapêutico , Sistema de RegistrosRESUMO
CD19-targeted chimeric antigen receptor (CAR) T cell therapy has demonstrated striking responses among B cell acute lymphoblastic leukemia (B-ALL), but analyses of potential factors associated with poor response and relapse are lacking. Here, we summarize the long-term follow-up of 254 B-ALL treated with CD19 CAR-T cells from 5 clinical trials (NCT03173417, NCT02546739, and NCT03671460 retrospectively registered on May 23, 2017, March 1, 2018, and September 7, 2018, respectively, at www.clinicaltrials.gov ; ChiCTR-ONC-17012829, and ChiCTR1800016541 retrospectively registered on September 28, 2017, and June 7, 2018, at www.chictr.org.cn ). Our data showed that TP53 mutation, bone marrow blasts > 20%, prior CAR-T/blinatumomab treatment, and severe cytokine release syndrome (CRS) were associated with a lower complete remission (CR) rate while age, extramedullary disease, complex cytogenetics, history of prior transplant, prior courses of chemotherapy, CAR-T cell dose, and manufacturing source of the cellular product did not affect patients' CR rate. Risk factors related to leukemia-free survival (LFS) and overall survival (OS) were history of prior transplant, complex cytogenetics, TP53 mutation, severe CRS, neurotoxicity, and CAR-T therapy without consolidative allogeneic hematopoietic stem cell transplantation (allo-HSCT). Age and CAR-T cell dose did not influence LFS and OS. Patients with consolidative allo-HSCT after CAR-T therapy had a superior OS and LFS compared to those who did not. This benefit was also observed in both pediatric and adult patients as well as in patients either in high- or low-risk groups. This large study to identify risk factors of CR, LFS, and OS may help to maximize clinical outcomes of CAR-T therapy. Précis TP53 mutation and BM blasts > 20% are two independent factors associated with the CR rate. Patients with high tumor burden as well as those with bone marrow blasts < 5% can benefit from consolidative allo-HSCT post-CAR-T therapy.
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Antígenos CD19 , Imunoterapia Adotiva , Leucemia-Linfoma Linfoblástico de Células Precursoras B/terapia , Receptores de Antígenos Quiméricos , Adolescente , Adulto , Antígenos CD19/imunologia , Biomarcadores Tumorais/genética , Criança , Pré-Escolar , Síndrome da Liberação de Citocina/etiologia , Gerenciamento Clínico , Feminino , Humanos , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Lactente , Masculino , Pessoa de Meia-Idade , Mutação , Doenças do Sistema Nervoso/etiologia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/complicações , Leucemia-Linfoma Linfoblástico de Células Precursoras B/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras B/etiologia , Prognóstico , Receptores de Antígenos de Linfócitos T , Receptores de Antígenos Quiméricos/imunologia , Adulto JovemRESUMO
Cytomegalovirus (CMV) infection remains a major cause of mortality after hematopoietic stem cell transplantation (HSCT). Current treatments, including antiviral drugs and adoptive cell therapy with CMV-specific cytotoxic T lymphocytes (CTLs), only show limited benefits in patients. T-cell receptor (TCR)-T cell therapy offers a promising option to treat CMV infections. Here, using tetramer-based screening and single-cell TCR cloning technologies, we identified various CMV antigen-specific TCRs from healthy donors, and generated TCR-T cells targeting multiple pp65 epitopes corresponding to three major HLA-A alleles. The TCR-T cells showed efficient cytotoxicity toward epitope-expressing target cells in vitro. After transfer into immune-deficient mice bearing pp65+ HLA+ tumor cells, TCR-T cells induced dramatic tumor regression and exhibited long-term persistence. In a phase I clinical trial (NCT04153279), CMV TCR-T cells were applied to treat patients with CMV reactivation after HSCT. Except one patient who withdrew at early treatment stage, all other six patients were well-tolerated and achieved complete response (CR), no more than grade 2 cytokine release syndrome (CRS) and other adverse events were observed. CMV TCR-T cells persisted up to 3 months. Among them, two patients have survived for more than 1 year. This study demonstrates the great potential in the treatment and prevention of CMV infection following HSCT or other organ transplantation.
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Infecções por Citomegalovirus , Transplante de Células-Tronco Hematopoéticas , Animais , Antivirais , Linfócitos T CD8-Positivos , Ensaios Clínicos Fase I como Assunto , Citomegalovirus , Infecções por Citomegalovirus/etiologia , Infecções por Citomegalovirus/terapia , Epitopos , Antígenos HLA-A , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Camundongos , Fosfoproteínas/genética , Receptores de Antígenos de Linfócitos T/genética , Proteínas da Matriz ViralRESUMO
We developed a T-cell-receptor (TCR) complex-based chimeric antigen receptor (CAR) named Synthetic TCR and Antigen Receptor (STAR). Here, we report pre-clinical and phase I clinical trial data (NCT03953599) of this T-cell therapy for refractory and relapsed (R/R) B-cell acute lymphoblastic leukemia (B-ALL) patients. STAR consists of two protein modules each containing an antibody light or heavy chain variable region and TCR α or ß chain constant region fused to the co-stimulatory domain of OX40. T-cells were transduced with a STAR-OX40 lentiviral vector. A leukemia xenograft mouse model was used to assess the STAR/STAR-OX40 T cell antitumor activity. Eighteen patients with R/R B-ALL were enrolled into the clinical trial. In a xenograft mouse model, STAR-T-cells exhibited superior tumor-specific cytotoxicity compared with conventional CAR-T cells. Incorporating OX40 into STAR further improved the proliferation and persistence of tumor-targeting T-cells. In our clinical trial, 100% of patients achieved complete remission 4 weeks post-STAR-OX40 T-cell infusion and 16/18 (88.9%) patients pursued consolidative allogeneic hematopoietic stem cell transplantation (allo-HSCT). Twelve of 16 patients (75%) remained leukemia-free after a median follow-up of 545 (433-665) days. The two patients without consolidative allo-HSCT relapsed on Day 58 and Day 186. Mild cytokine release syndrome occurred in 10/18 (55.6%) patients, and 2 patients experienced grade III neurotoxicity. Our preclinical studies demonstrate super anti-tumor potency of STAR-OX40 T-cells compared with conventional CAR-T cells. The first-in-human clinical trial shows that STAR-OX40 T-cells are tolerable and an effective therapeutic platform for treating R/R B-ALL.
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Linfoma de Burkitt , Transplante de Células-Tronco Hematopoéticas , Leucemia-Linfoma Linfoblástico de Células Precursoras , Receptores de Antígenos Quiméricos , Doença Aguda , Animais , Antígenos CD19 , Humanos , Imunoterapia Adotiva , Camundongos , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos Quiméricos/genética , Linfócitos TRESUMO
Targeting autophagy might be a promising anticancer strategy; however, the dual roles of autophagy in cancer development and malignancy remain unclear. NSCLC (non-small cell lung cancer) cells harbour high levels of SQSTM1 (sequestosome 1), the autophagy receptor that is critical for the dual roles of autophagy. Therefore, mechanistic insights into SQSTM1 modulation may point towards better approaches to treat NSCLC. Herein, we used multiple autophagy flux models and autophagy readouts to show that aldo-keto reductase family 1 member C1 (AKR1C1), which is highly expressed in NSCLC, promotes autophagy by directly binding to SQSTM1 in a catalytic-independent manner. This interaction may be strengthened by reactive oxygen species (ROS), important autophagy inducers. Further mechanistic research demonstrated that AKR1C1 interacts with SQSTM1 to augment SQSTM1 oligomerization, contributing to the SQSTM1 affinity for binding cargo. Collectively, our data reveal a catalytic-independent role of AKR1C1 for interacting with SQSTM1 and promoting autophagy. All these findings not only reveal a novel functional role of AKR1C1 in the autophagy process but also indicate that modulation of the AKR1C1-SQSTM1 interaction may be a new strategy for targeting autophagy.
Assuntos
Aldo-Ceto Redutases/metabolismo , Autofagia/fisiologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Estresse Oxidativo/fisiologia , Proteína Sequestossoma-1/metabolismo , Linhagem Celular Tumoral , HumanosRESUMO
Bladder cancer (BC) is among the most common tumors with a high recurrence rate, necessitating noninvasive and sensitive diagnostic methods. Accurate detection of exfoliated tumor cells (ETCs) in urine is crucial for noninvasive BC diagnosis but suffers from limited sensitivity when ETCs are rare and confounded by reactive, regenerative, or reparative cells. Single-cell sequencing (SCS) enables accurate detection of ETCs by surveying oncogenic driver mutations or genome-wide copy number alternations. To overcome the low-throughput limitation of SCS, we report a SCS-validated cellular marker, hexokinase 2 (HK2), for high-throughput screening cells in urine and detecting ETCs engaging elevated glycolysis. In the SCS-based training set, a total of 385 cells from urine samples of eight urothelial carcinoma (UC) patients were sequenced to establish a HK2 threshold that achieved >90% specificity for ETC detection. This urine-based HK2 assay was tested with a blinded patient group (n = 384) including UC and benign genitourinary disorders as a validation cohort for prospectively evaluating diagnostic accuracy. The sensitivity, specificity, positive predictive value, and negative predictive value of the assay were 90, 88, 83, and 93%, respectively, which were superior to urinary cytology. For investigating the potential to be a screening test, the HK2 assay was tested with a group of healthy individuals (n = 846) and a 6-month follow-up. The specificity was 98.4% in this health group. Three participants were found to have >5 putative ETCs that were sequenced to exhibit recurrent copy number alternations characteristic of malignant cells, demonstrating early BC detection before current clinical methods.
Assuntos
Hexoquinase/genética , Hexoquinase/metabolismo , Programas de Rastreamento , Análise de Célula Única , Neoplasias da Bexiga Urinária/diagnóstico , Neoplasias da Bexiga Urinária/urina , Linhagem Celular Tumoral , Humanos , Valor Preditivo dos Testes , Análise de Sequência , Neoplasias da Bexiga Urinária/patologiaRESUMO
BACKGROUND AIMS: The efficacy of CD19-targeted chimeric antigen receptor T (CAR T) cells for treatment of relapsed B-cell malignancies after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and the long-term outcomes of these patients remain inconclusive. METHODS: The authors focused on the survival of 35 patients with B-cell acute lymphoblastic leukemia who relapsed after allo-HSCT and received CAR T cells. RESULTS: Of the 34 eligible patients, 30 achieved minimal residual disease-negative complete remission (CR), with a total CR rate of 85.7% (79.8-91.6%). There were 14 patients who received various forms of additional therapy after achieving CR. After a median follow-up of 20.7 months, it was noted that 17 patients had relapsed at a median of 4.5 months (2-34 months). The cumulative recurrence rate (RR) at 18 months was 68.3% (57.6-79.0%). Additional treatment did not reduce the RR but seemed to delay the time to relapse (mean: 5.9 months vs 13.1 months; P = 0.046). Patients with a lower tumor burden (≤10%) had a lower RR (25.0% vs 78.6% at 12 months; P = 0.006). The overall survival (OS) rate for the CR patients was 30.0% (20.3-29.7%) at 18 months, with a median OS of 12.7 months. CONCLUSIONS: The authors' study indicated that for patients who relapsed after HSCT, although a high CR rate was achieved after CAR T therapy, the long-term efficacy was unsatisfactory. It is necessary to optimize additional treatment, including a second HSCT, to further improve long-term efficacy after CAR T infusion.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Imunoterapia Adotiva , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Receptores de Antígenos de Linfócitos T/metabolismo , Adulto , Linfócitos B/imunologia , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Interleucina-2/metabolismo , Interleucinas/metabolismo , Masculino , Pessoa de Meia-Idade , Antígeno Prostático Específico/metabolismo , Recidiva , Indução de Remissão , Linfócitos T/imunologiaRESUMO
The development of hematopoietic stem cell transplantation (HSCT) programs can face significant challenges in most developing countries because such endeavors must compete with other government health care priorities, including the delivery of basic services. Although this is may be a limiting factor, these countries should prioritize development of the needed expertise to offer state-of-the-art treatments, including transplantation, by providing financial, technological, legal, ethical, and other needed support. This would prove beneficial in providing successful programs customized to the needs of their population and potentially provide long-term cost savings by circumventing the need for their citizens to seek care abroad. The costs of establishing an HSCT program and the costs of the HSCT procedure itself can be substantial barriers in developing countries. In addition, socioeconomic factors intrinsic to specific countries can influence access to HSCT, patient eligibility for HSCT, and timely utilization of HSCT center capabilities. This report describes recommendations from the Worldwide Network for Blood and Marrow Transplantation for establishing HSCT programs, with a specific focus on developing countries, and identifies challenges and opportunities for providing this specialized procedure in resource-constrained settings.
Assuntos
Países em Desenvolvimento , Transplante de Células-Tronco Hematopoéticas , Sociedades Médicas , Condicionamento Pré-Transplante , Humanos , Guias de Prática Clínica como Assunto , Fatores Socioeconômicos , Transplante Autólogo , Transplante HomólogoRESUMO
BACKGROUND: MET amplification is associated with acquired resistance to first-generation epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) in treating non-small-cell lung cancer (NSCLC); however, the therapeutic strategy in these patients is undefined. Herein we report the clinical outcomes of patients with c-MET amplification resistance to EGFR-TKIs treated with crizotinib. METHODS: We retrospectively analyzed advanced NSCLC patients from five sites who were diagnosed with EGFR-mutant NSCLC and received EGFR-TKI treatment. After disease progression, these patients were confirmed to have a MET-to-centromere ratio (MET:CEN) ≥ 1.8 based on fluorescence in situ hybridization (FISH) examination and without a T790M mutation. We assessed the efficacy and safety of crizotinib to overcome EGFR-TKI resistance in EGFR-activating mutations NSCLC with acquired MET amplification. RESULTS: Amplification of the acquired MET gene was identified in 18 patients with EGFR-mutant NSCLC. Fourteen patients received crizotinib treatment after acquired resistance to EGFR-TKIs. Among the 14 patients, 6 (42.9%) received crizotinib plus EGFR-TKI and 8 (57.1%) received crizotinib monotherapy. The overall objective response rate (ORR) and disease control rate (DCR) were 50.0% (7/14) and 85.7% (12/14), respectively. The median PFS (mPFS) of patients receiving crizotinib monotherapy and crizotinib plus EGFR-TKI was 6.0 and 12.6 months, respectively (P = 0.315). Notably, treatment efficacy was more pronounced in patients with crizotinib than patients with chemotherapy (24.0 months vs. 12.0 months, P = 0.046). The mOS for 8 of 14 patients receiving crizotinib monotherapy and 6 of 14 patients receiving crizotinib plus EGFR-TKI was 17.2 and 24.0 months, respectively (P = 0.862). Among the 14 patients, 1 who received crizotinib monotherapy (grade 3 nausea) and 2 who received crizotinib plus EGFR-TKI (grade 3 elevated liver aminotransferase levels) received reduced doses of crizotinib (200 mg twice daily) to better tolerate the dose. CONCLUSIONS: We observed the clinical evidence of efficacy generated by combination of crizotinib and previous EGFR-TKIs after the resistance to first-generation EGFR-TKIs. These results might increase evidence of more effective therapeutic strategies for NSCLC treatment. Combination therapy did not increase the frequency of adverse reactions.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Crizotinibe/uso terapêutico , Receptores ErbB/genética , Amplificação de Genes , Neoplasias Pulmonares/tratamento farmacológico , Mutação/genética , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-met/genética , Carcinoma Pulmonar de Células não Pequenas/diagnóstico por imagem , Carcinoma Pulmonar de Células não Pequenas/genética , Crizotinibe/efeitos adversos , Crizotinibe/farmacologia , Progressão da Doença , Intervalo Livre de Doença , Feminino , Humanos , Neoplasias Pulmonares/diagnóstico por imagem , Neoplasias Pulmonares/genética , Masculino , Pessoa de Meia-Idade , Inibidores de Proteínas Quinases/efeitos adversos , Proteínas Proto-Oncogênicas c-met/metabolismo , Estudos Retrospectivos , Tomografia Computadorizada por Raios X , Falha de TratamentoRESUMO
Proteins of the Src homology and collagen (Shc) family are typically involved in signal transduction events involving Ras/MAPK and PI3K/Akt pathways. In the nervous system, they function proximal to the neurotrophic factors that regulate cell survival, differentiation, and neuron-specific characteristics. The least characterized homolog, ShcD, is robustly expressed in the developing and mature nervous system, but its contributions to neural cell circuitry are largely uncharted. We now report that ShcD binds to active Ret, TrkA, and TrkB neurotrophic factor receptors predominantly via its phosphotyrosine-binding (PTB) domain. However, in contrast to the conventional Shc adaptors, ShcD suppresses distal phosphorylation of the Erk MAPK. Accordingly, genetic knock-out of mouse ShcD enhances Erk phosphorylation in the brain. In cultured cells, this capacity is tightly aligned to phosphorylation of ShcD CH1 region tyrosine motifs, which serve as docking platforms for signal transducers, such as Grb2. Erk suppression is relieved through independent mutagenesis of the PTB domain and the CH1 tyrosine residues, and successive substitution of these tyrosines breaks the interaction between ShcD and Grb2, thereby promoting TrkB-Grb2 association. Erk phosphorylation can also be restored in the presence of wild type ShcD through Grb2 overexpression. Conversely, mutation of the ShcD SH2 domain results in enhanced repression of Erk. Although the SH2 domain is a less common binding interface in Shc proteins, we demonstrate that it associates with the Ptpn11 (Shp2) phosphatase, which in turn regulates ShcD tyrosine phosphorylation. We therefore propose a model whereby ShcD competes with neurotrophic receptors for Grb2 binding and opposes activation of the MAPK cascade.
Assuntos
MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Glicoproteínas de Membrana/metabolismo , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-ret/metabolismo , Receptor trkA/metabolismo , Proteínas Adaptadoras da Sinalização Shc/metabolismo , Motivos de Aminoácidos , Linhagem Celular , MAP Quinases Reguladas por Sinal Extracelular/genética , Proteína Adaptadora GRB2/genética , Proteína Adaptadora GRB2/metabolismo , Humanos , Glicoproteínas de Membrana/genética , Fosforilação/fisiologia , Proteína Tirosina Fosfatase não Receptora Tipo 11/genética , Proteína Tirosina Fosfatase não Receptora Tipo 11/metabolismo , Proteínas Tirosina Quinases/genética , Proteínas Proto-Oncogênicas c-ret/genética , Receptor trkA/genética , Receptor trkB , Proteínas Adaptadoras da Sinalização Shc/genéticaRESUMO
BACKGROUND: Mutations in the colony-stimulating factor 3 receptor (CSF3R) gene occur frequently in chronic neutrophilic leukemia and are rare in de novo acute leukemia. The objective of this study was to assess the incidence of CSF3R mutations in acute leukemia and their association with other genetic abnormalities. METHODS: Amplicon-targeted, next-generation sequencing of 58 genes was performed retrospectively on 1152 patients (acute myeloid leukemia [AML], n = 587; acute lymphoid leukemia [ALL], n = 565). Reverse transcriptase-polymerase chain reaction analysis was used to detect 35 leukemia-specific gene fusions. RESULTS: CSF3R mutations (26 patients) were detected in 3.6% (13 of 364 patients), 4.6% (8 of 175 patients), and 8.3% (4 of 48 patients) of those with de novo, relapsed, and secondary AML, respectively, and in 0.2% (1 of 565 patients) of those with ALL. In total, 9 distinct CSF3R mutations were detected. Membrane-proximal missense mutations and cytoplasmic truncations were identified as mutually exclusive. The proportion of patients who had French-American-British subtypes M2 and M4 in the CSF3R-mutated group was significantly greater than that in the CSF3R wild-type group for both the de novo AML cohort (P = .001) and the relapsed AML cohort (P = .024). All de novo and relapsed AMLs with CSF3R mutations were associated with genetic alterations in transcription factors, including RUNX1-RUNX1T1, CBFB-MYH11, double-mutated CCAAT/enhancer binding protein α (CEBPAdm), and NPM1 mutations; and core-binding factor gene abnormalities and CEBPAdm accounted for 90.5% (19 of 21 patients). CONCLUSIONS: CSF3R mutations are uncommon in AML; however, when they occur, they are often associated with core-binding factor gene abnormalities and CEBPAdm. An in-depth understanding of the interaction between these genetic alterations could facilitate a clearer understanding of the role of CSF3R mutations in AML development and may be used for disease classification, prognosis, and the development of targeted therapy.