RESUMEN
Gene-editing technologies have made it feasible to create nonhuman primate models for human genetic disorders. Here, we report detailed genotypes and phenotypes of TALEN-edited MECP2 mutant cynomolgus monkeys serving as a model for a neurodevelopmental disorder, Rett syndrome (RTT), which is caused by loss-of-function mutations in the human MECP2 gene. Male mutant monkeys were embryonic lethal, reiterating that RTT is a disease of females. Through a battery of behavioral analyses, including primate-unique eye-tracking tests, in combination with brain imaging via MRI, we found a series of physiological, behavioral, and structural abnormalities resembling clinical manifestations of RTT. Moreover, blood transcriptome profiling revealed that mutant monkeys resembled RTT patients in immune gene dysregulation. Taken together, the stark similarity in phenotype and/or endophenotype between monkeys and patients suggested that gene-edited RTT founder monkeys would be of value for disease mechanistic studies as well as development of potential therapeutic interventions for RTT.
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Proteína 2 de Unión a Metil-CpG/genética , Síndrome de Rett/genética , Animales , Encéfalo/fisiología , Cromosomas Humanos X , Ritmo Circadiano , Modelos Animales de Enfermedad , Electrocardiografía , Femenino , Edición Génica , Humanos , Macaca fascicularis , Imagen por Resonancia Magnética , Masculino , Mutación , Dolor , Síndrome de Rett/fisiopatología , Sueño , Nucleasas de los Efectores Tipo Activadores de la Transcripción/metabolismo , TranscriptomaRESUMEN
BACKGROUND: Tocilizumab is commonly used for the management of chimeric antigen receptor (CAR) T-cell therapy-associated cytokine release syndrome (CRS). However, it remains unknown whether tocilizumab or its dosage affects the efficacy and safety of CAR T-cell therapy. The objective of this multicenter retrospective study was to explore the impact of tocilizumab on CAR T-cell therapy. METHODS: In total, 93 patients with B-cell acute lymphoblastic leukemia (B-ALL) receiving humanized anti-CD19 CAR T cells were recruited from May 2016 to November 2022. Forty-five patients received tocilizumab (tocilizumab group), whereas 48 patients did not (nontocilizumab group). Thirteen patients received >1 dose of tocilizumab. The primary end point was the effect of tocilizumab on the efficacy and safety of CAR T cells. Additionally, proliferation, killing, and cytokine assays of CAR T cells were performed in vitro in the presence of tocilizumab. RESULTS: The median age of the patients was 33 years, with 47 males and 46 females. Patients in the tocilizumab group showed similar complete response (CR) rate, overall survival (OS), and event-free survival (EFS) compared with the nontocilizumab group. Compared with patients who received ≤1 dose of tocilizumab, receiving >1 dose of tocilizumab did not affect their CR rate, OS, or EFS. In the tocilizumab group, all patients experienced CRS and 26.7% experienced immune effector cell-associated neurotoxicity syndrome (ICANS). In the nontocilizumab group, 64.6% of patients experienced CRS and 8.3% experienced ICANS. Up to 75% of ICANS and 87.5% of grade ≥3 ICANS occurred in the tocilizumab group. In vitro, tocilizumab did not impair the proliferation and killing effects of CAR T cells. CONCLUSIONS: Tocilizumab does not affect the efficacy of CAR T cells but may increase the likelihood of ICANS.
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Anticuerpos Monoclonales Humanizados , Antígenos CD19 , Síndrome de Liberación de Citoquinas , Inmunoterapia Adoptiva , Receptores Quiméricos de Antígenos , Humanos , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales Humanizados/efectos adversos , Masculino , Femenino , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Adulto , Antígenos CD19/inmunología , Estudios Retrospectivos , Persona de Mediana Edad , Adolescente , Adulto Joven , Síndrome de Liberación de Citoquinas/etiología , Receptores Quiméricos de Antígenos/inmunología , Niño , Leucemia-Linfoma Linfoblástico de Células Precursoras B/terapia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/tratamiento farmacológicoRESUMEN
Few studies have described chimeric antigen receptor (CAR) T-cell therapy for patients with B-cell acute lymphoblastic leukemia (B-ALL) with central nervous system leukemia (CNSL) because of concerns regarding poor response and treatment-related neurotoxicity. Our study included 48 patients with relapsed/refractory B-ALL with CNSL to evaluate the efficacy and safety of CD19-specific CAR T cell-based therapy. The infusion resulted in an overall response rate of 87.5% (95% confidence interval [CI], 75.3-94.1) in bone marrow (BM) disease and remission rate of 85.4% (95% CI, 72.8-92.8) in CNSL. With a median follow-up of 11.5 months (range, 1.3-33.3), the median event-free survival was 8.7 months (95% CI, 3.7-18.8), and the median overall survival was 16.0 months (95% CI, 13.5-20.1). The cumulative incidences of relapse in BM and CNS diseases were 31.1% and 11.3%, respectively, at 12 months (P = .040). The treatment was generally well tolerated, with 9 patients (18.8%) experiencing grade ≥3 cytokine release syndrome. Grade 3 to 4 neurotoxic events, which developed in 11 patients (22.9%), were associated with a higher preinfusion disease burden in CNS and were effectively controlled under intensive management. Our results suggest that CD19-specific CAR T cell-based therapy can induce similar high response rates in both BM and CNS diseases. The duration of remission in CNSL was longer than that in BM disease. CD19 CAR T-cell therapy may provide a potential treatment option for previously excluded patients with CNSL, with manageable neurotoxicity. The clinical trials were registered at www.clinicaltrials.gov as #NCT02782351 and www.chictr.org.cn as #ChiCTR-OPN-16008526.
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Linfoma de Burkitt , Neoplasias del Sistema Nervioso Central , Leucemia-Linfoma Linfoblástico de Células Precursoras , Receptores Quiméricos de Antígenos , Enfermedad Aguda , Antígenos CD19 , Linfoma de Burkitt/tratamiento farmacológico , Neoplasias del Sistema Nervioso Central/tratamiento farmacológico , Síndrome de Liberación de Citoquinas , Humanos , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Receptores Quiméricos de Antígenos/uso terapéutico , Linfocitos TRESUMEN
CD19 CAR-T (chimeric antigen receptor-T) cell immunotherapy achieves a remission rate of approximately 70% in recurrent and refractory lymphoma treatment. However, the loss or reduction of CD19 antigen on the surface of lymphoma cells results in the escape of tumor cells from the immune killing of CD19 CAR-T cells (CAR19-T). Therefore, novel therapeutic strategies are urgently required. In this study, an anti-CD79b/CD3 bispecific antibody (BV28-OKT3) was constructed and combined with CAR19-T cells for B-cell lymphoma treatment. When the CD19 antigen was lost or reduced, BV28-OKT3 redirected CAR19-T cells to CD79b+ CD19- lymphoma cells; therefore, BV28-OKT3 overcomes the escape of CD79b+ CD19- lymphoma cells by the killing action of CAR19-T cells in vitro and in vivo. Furthermore, BV28-OKT3 triggered the antitumor function of CAR- T cells in the infusion product and boosted the antitumor immune response of bystander T cells, markedly improving the cytotoxicity of CAR19-T cells to lymphoma cells in vitro and in vivo. In addition, BV28-OKT3 elicited the cytotoxicity of donor-derived T cells toward lymphoma cells in vitro, which depended on the presence of tumor cells. Therefore, our findings provide a new clinical treatment strategy for recurrent and refractory B-cell lymphoma by combining CD79b/CD3 BsAb with CAR19-T cells.
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Anticuerpos Biespecíficos , Linfoma de Células B , Linfoma , Humanos , Linfocitos T , Antígenos CD19 , Muromonab-CD3 , Linfoma/tratamiento farmacológico , Inmunoterapia Adoptiva/métodosRESUMEN
Chimeric antigen receptor (CAR) T-cell therapy has emerged as a promising form of adoptive T-cell immunotherapy for selected hematologic malignancies including leukemia, lymphoma and multiple myeloma. China has become the country with the largest number of registered CAR T-cell trials. Despite the remarkable clinical outcomes achieved with CAR Tcell therapy, challenges such as disease relapse, the process of manufacturing the CAR T cells and safety have limited the therapeutic efficacy of CAR T cells in hematologic malignancies. In this period of innovation, several clinical trials have reported the design of CAR directed at new targets in hematologic malignancies. In this review, we comprehensively summarize the contemporary landscape and clinical development of CAR T-cell therapy in China. In addition, we present strategies for further improving the clinical utility of CAR T-cell therapy, such as increasing the efficacy and response duration, in hematologic malignancies.
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Neoplasias Hematológicas , Receptores Quiméricos de Antígenos , Humanos , Inmunoterapia Adoptiva/efectos adversos , Receptores Quiméricos de Antígenos/genética , Receptores de Antígenos de Linfocitos T/genética , Neoplasias Hematológicas/patología , Tratamiento Basado en Trasplante de Células y TejidosRESUMEN
DNA damage is assumed to accumulate in stem cells over time and their ability to withstand this damage and maintain tissue homeostasis is the key determinant of aging. Nonetheless, relatively few studies have investigated whether DNA damage does indeed accumulate in stem cells and whether this contributes to stem cell aging and functional decline. Here, we found that, compared with young mice, DNA double-strand breaks (DSBs) are reduced in the subventricular zone (SVZ)-derived neural stem cells (NSCs) of aged mice, which was achieved partly through the adaptive upregulation of Sirt1 expression and non-homologous end joining (NHEJ)-mediated DNA repair. Sirt1 deficiency abolished this effect, leading to stem cell exhaustion, olfactory memory decline, and accelerated aging. The reduced DSBs and the upregulation of Sirt1 expression in SVZ-derived NSCs with age may represent a compensatory mechanism that evolved to protect stem cells from excessive DNA damage, as well as mitigate memory loss and other stresses during aging.
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Ventrículos Laterales , Células-Madre Neurales , Sirtuina 1 , Envejecimiento/genética , Animales , ADN/metabolismo , Roturas del ADN de Doble Cadena , Reparación del ADN por Unión de Extremidades , Reparación del ADN , Ventrículos Laterales/metabolismo , Ratones , Células-Madre Neurales/metabolismo , Sirtuina 1/genética , Sirtuina 1/metabolismoRESUMEN
BACKGROUND AIMS: Chimeric antigen receptor (CAR) T-cell therapy is a breakthrough treatment for patients with relapsed or refractory diffuse large B-cell lymphoma. However, many patients do not achieve remission or relapse after remission. Previous studies have demonstrated that eosinophils have synergistic anti-tumor effects with CD8+T cells and pre-CAR T-eosinophil counts are associated with the efficacy of CAR T cells. METHODS: We retrospectively analyzed the eosinophil counts of patients with diffuse large B-cell lymphoma and found it changed remarkably pre- and post-CAR T-cell therapy. RESULTS: Patients who achieved complete remission after CAR T-cell infusion had greater post-CAR T-eosinophil counts than those who did not. Kaplan-Meier curves showed that patients with greater eosinophil counts during the second month after CAR T-cell infusion had superior progression-free survival and overall survival compared with those with lower eosinophil counts. CONCLUSIONS: For patients who responded to CAR T-cell therapy, eosinophil counts also can be used to predict 6-month duration of response. In conclusion, the post-CAR T-eosinophil count is associated with the prognosis of patients treated with CAR T-cell therapy and can be used to clinically identify patients who can achieve longer remission after CAR T-cell infusion.
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Linfoma de Células B Grandes Difuso , Receptores Quiméricos de Antígenos , Humanos , Inmunoterapia Adoptiva/efectos adversos , Receptores Quiméricos de Antígenos/genética , Eosinófilos/patología , Pronóstico , Estudios Retrospectivos , Recurrencia Local de Neoplasia/patología , Linfoma de Células B Grandes Difuso/terapia , Linfoma de Células B Grandes Difuso/etiología , Tratamiento Basado en Trasplante de Células y Tejidos , Antígenos CD19 , Receptores de Antígenos de Linfocitos TRESUMEN
Internal tandem duplication (ITD) mutations within the FMS-like receptor tyrosine kinase-3 (FLT3) can be found in up to 25% to 30% of acute myeloid leukemia (AML) patients and confer a poor prognosis. Although FLT3 tyrosine kinase inhibitors (TKIs) have shown clinical responses, they cannot eliminate primitive FLT3-ITD+ AML cells, which are potential sources of relapse. Therefore, elucidating the mechanisms underlying FLT3-ITD+ AML maintenance and drug resistance is essential to develop novel effective treatment strategies. Here, we demonstrate that FLT3 inhibition induces histone deacetylase 8 (HDAC8) upregulation through FOXO1- and FOXO3-mediated transactivation in FLT3-ITD+ AML cells. Upregulated HDAC8 deacetylates and inactivates p53, leading to leukemia maintenance and drug resistance upon TKI treatment. Genetic or pharmacological inhibition of HDAC8 reactivates p53, abrogates leukemia maintenance, and significantly enhances TKI-mediated elimination of FLT3-ITD+ AML cells. Importantly, in FLT3-ITD+ AML patient-derived xenograft models, the combination of FLT3 TKI (AC220) and an HDAC8 inhibitor (22d) significantly inhibits leukemia progression and effectively reduces primitive FLT3-ITD+ AML cells. Moreover, we extend these findings to an AML subtype harboring another tyrosine kinase-activating mutation. In conclusion, our study demonstrates that HDAC8 upregulation is an important mechanism to resist TKIs and promote leukemia maintenance and suggests that combining HDAC8 inhibition with TKI treatment could be a promising strategy to treat FLT3-ITD+ AML and other tyrosine kinase mutation-harboring leukemias.
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Biomarcadores de Tumor/metabolismo , Resistencia a Antineoplásicos , Proteína Forkhead Box O1/metabolismo , Histona Desacetilasas/metabolismo , Leucemia Mieloide Aguda/patología , Proteínas Represoras/metabolismo , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , Tirosina Quinasa 3 Similar a fms/antagonistas & inhibidores , Animales , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Proteína Forkhead Box O1/genética , Regulación Neoplásica de la Expresión Génica , Histona Desacetilasas/genética , Humanos , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Ratones , Ratones Endogámicos NOD , Ratones SCID , Mutación , Pronóstico , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Represoras/genética , Secuencias Repetidas en Tándem , Células Tumorales Cultivadas , Regulación hacia Arriba , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Site-specific C>T DNA base editing has been achieved by recruiting cytidine deaminases to the target C using catalytically impaired Cas proteins; the target C is typically located within 5-nt editing window specified by the guide RNAs. The prototypical cytidine base editor BE3, comprising rat APOBEC1 (rA1) fused to nCas9, can indiscriminately deaminate multiple C's within the editing window and also create substantial off-target edits on the transcriptome. A powerful countermeasure for the DNA off-target editing is to replace rA1 with APOBEC proteins which selectively edit C's in the context of specific motifs, as illustrated in eA3A-BE3 which targets TC. However, analogous editors selective for other motifs have not been described. In particular, it has been challenging to target a particular C in C-rich sequences. Here, we sought to confront this challenge and also to overcome the RNA off-target effects seen in BE3. RESULTS: By replacing rA1 with an optimized human A3G (oA3G), we developed oA3G-BE3, which selectively targets CC and CCC and is also free of global off-target effects on the transcriptome. Furthermore, we created oA3G-BE4max, an upgraded version of oA3G-BE3 with robust on-target editing. Finally, we showed that oA3G-BE4max has negligible Cas9-independent off-target effects at the genome. CONCLUSIONS: oA3G-BE4max can edit C(C)C with high efficiency and selectivity, which complements eA3A-editors to broaden the collective editing scope of motif selective editors, thus filling a void in the base editing tool box.
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Desaminasa APOBEC-3G/genética , Sistemas CRISPR-Cas , Citidina Desaminasa/metabolismo , Edición Génica , ARN Guía de KinetoplastidaRESUMEN
BACKGROUND: Despite of the frequently reported Dnmt3a abormality in classical myeloproliferative neoplasms (cMPNs) patients, few research explores how the Dnmt3a is regulated by Jak2V617F mutation. In this study, we have investigated how the Dnmt3a is regulated by Jak2V617F mutation and its effects on downstream signaling pathways in cMPNs. METHODS: Specimens of Jak2V617F positive cMPN patients and normal controls were collected. Murine BaF3 cell line was used to construct cell models. Dual-Glo luciferase assays and chromatin immunoprecipitation (ChIP)-qPCR were performed to detect the impact of Stat5a on transcription activity of Dnmt3a. Soft agar colony formation assay and cell counting assay were performed to detect cell proliferation. BrdU staining and flow cytometry were used to investigate cell cycle distribution. Western blotting and quantitative reverse-transcription PCR (qPCR) were performed to detect the expression levels of genes. RESULTS: Firstly, the results of western blotting and qPCR revealed that compared with the control samples, Dnmt3a is downregulated in Jak2V617F positive samples. Then we explored the mechanism behind it and found that Dnmt3a is a downstream target of Stat5a, the transcription and translation of Dnmt3a is suppressed by the binding of aberrantly activated Stat5a with Dnmt3a promoter in Jak2V617F positive samples. We further revealed the region approximately 800 bp upstream of the first exon of the Dnmt3a promoter, which includes a gamma-activated sequence (GAS) motif of Stat5a, is the specific site that Stat5a binds to. Soft agar colony formation assay, cell counting assay, and BrdU staining and flow cytometry assay found that Dnmt3a in Jak2V617F-BaF3 cells significantly affected the cell proliferation capacity and cell cycle distribution by suppressing Cdkn1a via miR-17-5p/Cdkn1a axis and mediated G0/G1 arrest. CONCLUSIONS: Transcription and translation of Dnmt3a is downregulated by the binding of Stat5a with Dnmt3a promoter in Jak2V617F cells. The GAS motif at promoter of Dnmt3a is the exact site where the Stat5a binds to. Dnmt3a conducted G0/G1 arrest through regulating miR-17-5p/Cdkn1a axis. The axis of Stat5a/Dnmt3a/miR-17-5p/Cdkn1a potentially provides a treatment target for cMPNs.
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Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , ADN Metiltransferasa 3A/metabolismo , Janus Quinasa 2/metabolismo , MicroARNs/metabolismo , Trastornos Mieloproliferativos/metabolismo , Factor de Transcripción STAT5/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Aminopiridinas/farmacología , Animales , Sitios de Unión , Western Blotting , Estudios de Casos y Controles , Recuento de Células , Línea Celular , Línea Celular Tumoral , Proliferación Celular , ADN Metiltransferasa 3A/genética , Regulación hacia Abajo , Exones , Puntos de Control de la Fase G1 del Ciclo Celular , Humanos , Imidazoles/farmacología , Janus Quinasa 2/antagonistas & inhibidores , Janus Quinasa 2/genética , Células K562 , Ratones , Monocitos/metabolismo , Mutación , Trastornos Mieloproliferativos/genética , Trastornos Mieloproliferativos/patología , Regiones Promotoras Genéticas , Pirazoles/farmacología , Piridazinas/farmacología , Transducción de Señal , Transcripción Genética , Ensayo de Tumor de Célula Madre , Células U937RESUMEN
In humans, hematopoietic stem cells (HSCs) adopt unique responsive pathways counteracting with the DNA-damaging assaults to weigh the balance between the maintenance of normal stem cell poor for whole-life blood regeneration and the transformation to leukemia stem cells (LSCs) for leukemia initiation. LSCs also take actions of combating with the attack launched by externally therapeutic drugs that can kill most leukemic cells, to avoid extermination and promote disease relapse. Therefore, the collection of knowledge about all these underlined mechanisms would present a preponderance for later studies. In this chapter, the universal DNA damage response (DDR) mechanisms were firstly introduced, and then DDR of HSCs were presented focusing on the DNA double-strand breaks in the quiescent state of HSCs, which poses a big advantage in promoting its transformation into preleukemic HSCs. Lastly, the DDR of LSCs were summarized based on the major outcomes triggered by different pathways in specific leukemia, upon which some aspects for future investigations were envisioned under our currently limited scope of knowledge.
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Daño del ADN , Células Madre Hematopoyéticas , Leucemia Mieloide Aguda , Células Madre Neoplásicas , División Celular , Células Madre Hematopoyéticas/patología , Humanos , Leucemia Mieloide Aguda/patología , Células Madre Neoplásicas/patologíaRESUMEN
BACKGROUND/AIMS: T-Cell Acute Lymphoblastic Leukemia (T-ALL) [corrected] is an aggressive disease which is highly resistant to chemotherapy. Studies show that enhanced ability of DNA damage repair (DDR) in cancer cells plays a key role in chemotherapy resistance. Here, we suggest that defect in DDR related genes might be a promising target to destroy the genome stability of tumor cells. METHODS: Since KU70 is highly expressed in Jurkat cells, one of the most representative cell lines of ATL, we knocked down KU70 by shRNA and analyzed the impact of KU70 deficiency in Jurkat cells as well as in NOD-SCID animal models by western blot, immunofluorescence, flow cytometry and measuring DNA repair efficiency. RESULTS: It is observed that silencing of KU70 resulted in accumulated DNA damage and impaired DDR in Jurkat cells, resulting in more apoptosis, decreased cell proliferation and cell cycle arrest. DNA damage leads to DNA double-strand breaks (DSBs), which are processed by either non-homologous end joining(NHEJ) or homologous recombination(HR). In our study, both NHEJ and HR are impaired because of KU70 defect, accompanied with increased protein level of SHP-1, a dephosphorylation enzyme. In turn, SHP-1 led to dephosphorylation of SIRT1, which further impaired HR repair efficiency. Moreover, KU70 deficiency prolonged survival of Jurkat-xenografted mice. CONCLUSION: These findings suggest that targeting KU70 is a promising target for ATL and might overcome the existing difficulties in chemotherapy.
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Reparación del ADN por Unión de Extremidades , Autoantígeno Ku/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 6/metabolismo , Reparación del ADN por Recombinación , Sirtuina 1/metabolismo , Animales , Apoptosis , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Puntos de Control del Ciclo Celular , Roturas del ADN de Doble Cadena , Humanos , Células Jurkat , Autoantígeno Ku/antagonistas & inhibidores , Autoantígeno Ku/genética , Leucemia-Linfoma de Células T del Adulto/tratamiento farmacológico , Leucemia-Linfoma de Células T del Adulto/metabolismo , Leucemia-Linfoma de Células T del Adulto/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Fosforilación , Proteína Tirosina Fosfatasa no Receptora Tipo 6/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 6/genética , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , ARN Interferente Pequeño/uso terapéutico , Recombinasa Rad51/metabolismoRESUMEN
BACKGROUND/AIMS: Alternative splicing and DNA damage exhibit cross-regulation, with not only DNA damage inducing changes in alternative splicing, but alternative splicing itself possibly modulating the DNA damage response (DDR). Sirt1, a prominent anti-aging player, plays pivotal roles in the DDR. However, few studies have examined alternative splicing with DNA damage in neural stem cells (NSCs) and, in essence, nothing is known about whether SIRT1 regulates alternative splicing. Hence, we investigated the potential involvement of Sirt1-mediated alternative splicing in the NSC DDR. METHODS: Genome-wide alternative splicing profiling was performed upon DNA damage induction and SIRT1 deletion. RESULTS: DNA damage caused genome-wide changes in alternative splicing in adult NSCs and Sirt1 deficiency dramatically altered DDR-related alternative splicing. In particular, extensive alternative splicing changes in DDR-related processes such as cell cycle control and DNA damage repair were observed; these processes were dramatically influenced by Sirt1 deficiency. Phenotypically, Sirt1 deficiency altered the proliferation and DNA repair of adult NSCs, possibly by regulating alternative splicing. CONCLUSION: SIRT1 helps to regulate alternative splicing, which itself affects the DDR of NSCs. Our findings provide novel insight into the mechanisms underlying the DDR in stem cells.
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Reparación del ADN , Sirtuina 1/genética , Empalme Alternativo/efectos de la radiación , Animales , Puntos de Control del Ciclo Celular , Proliferación Celular , Células Cultivadas , Daño del ADN/efectos de la radiación , Ventrículos Laterales/citología , Ratones , Ratones Transgénicos , Microscopía Fluorescente , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Células-Madre Neurales/efectos de la radiación , Radiación Ionizante , Sirtuina 1/deficiencia , Sirtuina 1/metabolismoRESUMEN
Treatment failure in cancer chemotherapy is largely due to the toxic effects of chemotherapeutic agents on normal cells/tissues. The proteasome inhibitor bortezomib has been successfully applied to treat multiple myeloma (MM), but there are some common adverse reactions in the clinic including peripheral neuropathy (PN). The TAK1 selective inhibitor 5Z-7-oxozeaenol has been widely studied in cancer therapy. Here, we investigated the potential synergy of bortezomib and 5Z-7-oxozeaenol in Burkitt's lymphoma (BL) cell lines. Cell viability assay showed that co-treatment of bortezomib at 8 nM, representing a one-eighth concentration for growth arrest, and 5Z-7-oxozeaenol at 2 µM, a dose that exhibited insignificant cytotoxic effects, synergistically induced apoptosis in the cell line Daudi. In parallel with the increasing dose of the bortezomib, and 5Z-7-oxozeaenol at 0.5 µM, lower colony formation efficiencies were seen in the cell line Daudi. Western blotting analysis verified that TAK1 inhibition by 5Z-7-oxozeaenol completely blocked JNK, p38, Erk, IKK, and IκB phosphorylation, which was almost instantly activated by TAK1 both directly or indirectly. Both agents synergistically prevented nuclear translocation of NF-κB, a characteristic of NF-κB inactivation. Moreover, a synergistic effect of bortezomib and 5Z-7-oxozeaenol on Western blotting analysis and flow cytometry was disclosed. Collectively, our results indicated that the proteasome inhibitor bortezomib and the TAK1 inhibitor 5Z-7-oxozeaenol displayed synergy on inhibiting BL cell apoptosis by inhibiting NF-κB activity.
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Antineoplásicos/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Apoptosis , Bortezomib/administración & dosificación , Linfoma de Burkitt/tratamiento farmacológico , Quinasas Quinasa Quinasa PAM/metabolismo , Zearalenona/análogos & derivados , Transporte Activo de Núcleo Celular , Animales , Línea Celular Tumoral , Supervivencia Celular , Citoplasma/metabolismo , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Inhibidores Enzimáticos/química , Humanos , FN-kappa B/metabolismo , Inhibidores de Proteasoma/administración & dosificación , Ratas , Zearalenona/administración & dosificaciónRESUMEN
Recently, numerous types of human dental tissue-derived mesenchymal stem cells (MSCs) have been isolated and characterized, including dental pulp stem cells, stem cells from exfoliated deciduous teeth, periodontal ligament stem cells, dental follicle progenitor cells, alveolar bone-derived MSCs, stem cells from apical papilla, tooth germ progenitor cells, and gingival MSCs. All these MSC-like cells exhibit self-renewal, multilineage differentiation potential, and immunomodulatory properties. Several studies have demonstrated the potential advantages of dental stem cell-based approaches for regenerative treatments and immunotherapies. This review outlines the properties of various dental MSC-like populations and the progress toward their use in regenerative therapy. Several dental stem cell banks worldwide are also introduced, with a view toward future clinical application.
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Pulpa Dental/citología , Saco Dental/citología , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/citología , Animales , Diferenciación Celular/fisiología , Humanos , Ingeniería de TejidosRESUMEN
BACKGROUND: Acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) presents great challenges in the treatment of non-small cell lung cancer (NSCLC) patients, while the mechanisms are still not well understood. The ß-catenin signaling pathway has been found to be associated with chemoresistance and can activate the EGFR and its downstream pathways. This study aimed to investigate the role of ß-catenin in acquired resistance to EGFR-TKIs in NSCLC cell lines. METHODS: The expression and transcriptional activity of ß-catenin were measured in both the NSCLC cell line PC9 and its sub-line PC9/AB(2) which has acquired resistance to gefitinib. Knockdown and overexpression of ß-catenin in the PC9/AB(2) and PC9 cells were performed. The cell survival rate and the activation of the EGFR and its downstream pathways were detected in the two cell lines after transfection. RESULTS: Nuclear translocation of ß-catenin was increased in the PC9/AB(2) cells and the baseline expression of members of the ß-catenin signaling pathway was also higher in the PC9/AB(2) cells. Knocking down the expression of ß-catenin increased the sensitivity of the PC9/AB(2) cells to gefitinib by blocking the activation of the EGFR downstream pathways, while ß-catenin overexpression improved PC9 cells resistance to gefitinib by enhancing the activation of the EGFR and its downstream signaling. CONCLUSION: ß-catenin plays an important role in acquired resistance to EGFR-TKIs in NSCLC cell lines and may be a potential therapeutic target for NSCLC patients who have failed to respond to targeted therapy.
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Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Quinazolinas/farmacología , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Resistencia a Antineoplásicos , Receptores ErbB/antagonistas & inhibidores , Gefitinib , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Transducción de Señal/efectos de los fármacos , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
DNA microarray data for thrombus-related leukocyte from patients with acute coronary syndrome (ACS) was analyzed to acquire key genes associated with ACS. Microarray data set GSE19339, including four ACS patients' samples and four normal samples, were downloaded from Gene Expression Omnibus database. Raw data was pre-processed and differentially expressed genes (DEGs) were identified by Affy packages of R. The interaction network was established with STRING. DrugBank was retrieved to obtain relevant small molecules. A total of 487 differentially expressed genes were identified as DEGs between normal and disease samples. Among which, ten up-regulated genes belonging to chemokine family (CCL2, CCR1, CXCL3, CXCL2, CCL8, CXCL11, CCL7, IL10, CCL22 and CCL20) were related to inflammatory response. In addition, two inhibitors of CCL2 (L-Mimosine) were retrieved from the DrugBank database. Considering the roles of inflammatory response in the progression of ACS and the functions of the ten up-regulated genes, we speculated that these genes might be related to ACS. Moreover, the inhibitors could provide guidelines for future drug design acting on these genes.
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Síndrome Coronario Agudo/genética , Quimiocinas/genética , Regulación de la Expresión Génica/inmunología , Síndrome Coronario Agudo/tratamiento farmacológico , Síndrome Coronario Agudo/inmunología , Síndrome Coronario Agudo/patología , Cardiotónicos/farmacología , Estudios de Casos y Controles , Quimiocinas/antagonistas & inhibidores , Quimiocinas/inmunología , Biología Computacional , Danazol/farmacología , Bases de Datos Genéticas , Bases de Datos Farmacéuticas , Perfilación de la Expresión Génica , Humanos , Mimosina/farmacología , Análisis de Secuencia por Matrices de Oligonucleótidos , Mapeo de Interacción de Proteínas , Bibliotecas de Moléculas Pequeñas/farmacologíaRESUMEN
The quiescence and activation of adult stem cells are regulated by many kinds of molecular mechanisms, and RNA alternative splicing participates in regulating many cellular processes. However, the relationship between stem cell quiescence and activation regulation and gene alternative splicing has yet to be studied. In this study, we aimed to elucidate the regulation of stem cell quiescence and activation by RNA alternative splicing. The upregulated genes in activated mouse neural stem cells (NSCs), muscle stem cells, and hematopoietic stem cells were collected for Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis. The genes from three tissue stem cells underwent Venn analysis. The mouse NSCs were used for quiescence and reactivation induction. The immunostaining of cell-specific markers was performed to identify cell properties. The reverse transcription-polymerase chain reaction and western blotting were used to detect the gene expression and protein expression, respectively. We found that the upregulated genes in activated stem cells from three tissues were all enriched in RNA splicing-related biological processes; the upregulated RNA splicing-related genes in activated stem cells displayed tissue differences; mouse NSCs were successfully induced into quiescence and reactivation in vitro without losing differentiation potential; serine and arginine-rich splicing factor 3 (Srsf3) was highly expressed in the activated mouse NSCs, and the overexpression of SRSF3 protein promoted the activation of quiescent mouse NSCs and increased the neural cell production. Our data indicate that the alternative splicing change may underline the transition of quiescence and activation of stem cells. The manipulation of the splicing factor may benefit tissue repair by promoting the activation of quiescent stem cells.
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Arginina , Células-Madre Neurales , Animales , Ratones , Células-Madre Neurales/metabolismo , ARN/metabolismo , Factores de Empalme de ARN/metabolismo , SerinaRESUMEN
Tumor-derived exosomes (TEXs) enriched in immune suppressive molecules predominantly drive T-cell dysfunction and impair antitumor immunity. Chimeric antigen receptor (CAR) T-cell therapy has emerged as a promising treatment for refractory and relapsed hematological malignancies, but whether lymphoma TEXs have the same impact on CAR T-cell remains unclear. Here, we demonstrated that B-cell lymphoma-derived exosomes induce the initial activation of CD19-CAR T-cells upon stimulation with exosomal CD19. However, lymphoma TEXs might subsequently induce CAR T-cell apoptosis and impair the tumor cytotoxicity of the cells because of the upregulated expression of the inhibitory receptors PD-1, TIM3, and LAG3 upon prolonged exposure. Similar results were observed in the CAR T-cells exposed to plasma exosomes from patients with lymphoma. More importantly, single-cell RNA sequencing revealed that CAR T-cells typically showed differentiated phenotypes and regulatory T-cell (Treg) phenotype conversion. By blocking transforming growth factor ß (TGF-ß)-Smad3 signaling with TGF-ß inhibitor LY2109761, the negative effects of TEXs on Treg conversion, terminal differentiation, and immune checkpoint expression were rescued. Collectively, although TEXs lead to the initial activation of CAR T-cells, the effect of TEXs suppressed CAR T-cells, which can be rescued by LY2109761. A treatment regimen combining CAR T-cell therapy and TGF-ß inhibitors might be a novel therapeutic strategy for refractory and relapsed B-cell lymphoma.
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INTRODUCTION: Chimeric antigen receptor (CAR) T cell therapy has markedly improved the prognosis of patients with diffuse large B-cell lymphoma (DLBCL). The relative positioning of tumor lesions in lymphoma varies among patients, manifesting as either aggregation (clumped together) or dissemination (spread throughout the body). Prognostic significance of factors indicating the relative positioning of tumor lesions in CAR T cell therapy remains underexplored. For aggregation, prior research proposed the tumor volume surface ratio (TVSR), linking it to prognosis in chemotherapy. Regarding dissemination, indicators such as disease stage or extranodal involvement, commonly used in clinical practice, have not demonstrated prognostic significance in CAR T cell therapy. This study aims to analyze current indicators of tumor aggregation or dissemination and introduce a novel indicator to assess the prognostic value of tumor lesions' relative positioning in DLBCL patients undergoing CAR T cell therapy. METHODS: This retrospective study included 42 patients receiving CAR T cell therapy. Lesion image information was obtained from the last PET/CT scan prior to CAR T cell infusion, including total metabolic tumor volume, total tumor surface, diameter of lymphoma masses, and the sites of tumor lesions. We evaluated TVSR and bulky disease as descriptors of tumor aggregation. We refined existing indicators, stage III&IV and >1 site extranodal involvement, to distill a new indicator, termed 'extra stage', to better represent tumor dissemination. The study examined the prognostic significance of tumor aggregation and dissemination. RESULTS: Our findings indicate that TVSR, while prognostically valuable in chemotherapy, lacks practical prognostic value in CAR T cell therapy. Conversely, bulky disease emerged as an optimal prognostic indicator of tumor aggregation. Both bulky disease and extra stage were associated with poor prognosis and exhibiting synergistic prognostic impact in CAR T cell therapy. CONCLUSIONS: Overall, the relative positioning of tumor lesions significantly influences the prognosis of patients with DLBCL receiving CAR T cell therapy. The ideal scenario involves tumors with minimal dissemination and no aggregation.