15 research outputs found
Epigenetic Profiling and Response to CD19 Chimeric Antigen Receptor T-Cell Therapy in B-Cell Malignancies
Background: Chimeric antigen receptor (CAR) T cells directed against CD19 (CART19) are effective in B-cell malignancies, but little is known about the molecular factors predicting clinical outcome of CART19 therapy. The increasingly recognized relevance of epigenetic changes in cancer immunology prompted us to determine the impact of the DNA methylation profiles of CART19 cells on the clinical course.
Methods: We recruited 114 patients with B-cell malignancies, comprising 77 patients with acute lymphoblastic leukemia and 37 patients with non-Hodgkin lymphoma who were treated with CART19 cells. Using a comprehensive DNA methylation microarray, we determined the epigenomic changes that occur in the patient T cells upon transduction of the CAR vector. The effects of the identified DNA methylation sites on clinical response, cytokine release syndrome, immune effector cell-associated neurotoxicity syndrome, event-free survival, and overall survival were assessed. All statistical tests were 2-sided.
Results: We identified 984 genomic sites with differential DNA methylation between CAR-untransduced and CAR-transduced T cells before infusion into the patient. Eighteen of these distinct epigenetic loci were associated with complete response (CR), adjusting by multiple testing. Using the sites linked to CR, an epigenetic signature, referred to hereafter as the EPICART signature, was established in the initial discovery cohort (n = 79), which was associated with CR (Fisher exact test, P < .001) and enhanced event-free survival (hazard ratio [HR] = 0.36; 95% confidence interval [CI] = 0.19 to 0.70; P = .002; log-rank P = .003) and overall survival (HR = 0.45; 95% CI = 0.20 to 0.99; P = .047; log-rank P = .04;). Most important, the EPICART profile maintained its clinical course predictive value in the validation cohort (n = 35), where it was associated with CR (Fisher exact test, P < .001) and enhanced overall survival (HR = 0.31; 95% CI = 0.11 to 0.84; P = .02; log-rank P = .02).
Conclusions: We show that the DNA methylation landscape of patient CART19 cells influences the efficacy of the cellular immunotherapy treatment in patients with B-cell malignancy.Supported by CERCA Programme/Generalitat de Catalunya, Health Department PERIS #SLT/002/16/00374, AGAUR-project #2017SGR1080; MCI/AEI/ERDF project #RTI2018-094049-B-I00; ERC EPIPHARM; Cellex Foundation; “la Caixa” Foundation (LCF/PR/GN18/51140001 and LCF/PR/GN18/50310007), RF-2016–02364388, Accelerator Award—Cancer Research UK/AIRC—INCAR Associazione Italiana Ricerca per la Ricerca sul Cancro (AIRC) Project 5 × 1000 no. 9962, AIRC IG 2018 id. 21724, AIRC MFAG id. 21769 and id. 20450; MIUR (Grant PRIN 2017); and RCR-2019–23669115
Enhanced detection of viral RNA species using fokI-assisted digestion of DNA duplexes and DNA/RNA hybrids
The accurate detection of nucleic acids from certain biological pathogens is critical for the diagnosis of human diseases. However, amplified detection of RNA molecules from a complex sample by direct detection of RNA/DNA hybrids remains a challenge. Here, we show that type IIS endonuclease FokI is able to digest DNA duplexes and DNA/RNA hybrids when assisted by a dumbbell-like fluorescent sensing oligonucleotide. As proof of concept, we designed a battery of sensing oligonucleotides against specific regions of the SARS-CoV-2 genome and interrogated the role of FokI relaxation as a potential nicking enzyme for fluorescence signal amplification. FokI-assisted digestion of SARS-CoV-2 probes increases the detection signal of ssDNA and RNA molecules and decreases the limit of detection more than 3.5-fold as compared to conventional molecular beacon approaches. This cleavage reaction is highly specific to its target molecules, and no detection of other highly related B-coronaviruses was observed in the presence of complex RNA mixtures. In addition, the FokI-assisted reaction has a high multiplexing potential, as the combined detection of different viral RNAs, including different SARS-CoV-2 variants, was achieved in the presence of multiple combinations of fluorophores and sensing oligonucleotides. When combined with isothermal rolling circle amplification technologies, FokI-assisted digestion reduced the detection time of SARS-CoV-2 in COVID-19-positive human samples with adequate sensitivity and specificity compared to conventional reverse transcription polymerase chain reaction approaches, highlighting the potential of FokI-assisted signal amplification as a valuable sensing mechanism for the detection of human pathogens.Funding was provided by the ISCIII (COV00624 to J.R.T. andM.F.F., PI18/01527 and PI21/01067 to M.F.F.), CSIC (202020E092 to M.F.F), the European Commission
NextGenerationEU, through CSIC’s Global Health Platform (PTI Salud Global) and the Spanish Ministry of Science and Innovation through the Recovery, Transformation and
Resilience Plan (GL2021-03-39 and GL2021-03-040), the PCTI from the Asturias Government, co-funded by 2018−2022/FEDER (IDI/2018/146 to M.F.F.), the AECC
(PROYE18061FERN to M.F.F), ISPA-Janssen (048-Intramural Nov-Tevar to J.R.T.) and the IUOPA. J.R.T is supported by a JdC fellowship from the Spanish Ministry of Science and Innovation (IJC2018-36825-I). R.F.P. and P.S.O. are supported by the Severo Ochoa program (BP17-114 and BP17-165). A.P. is supported by the PFIS program (ISCIII, FI19/
00085). J.J.A.L. is supported by the AECC fellowship. C.M. and V.L. are supported by IUOPA, and R.G.U. is supported by CIBERER.Peer reviewe
H3K4me1 marks DNA regions hypomethylated during aging in human stem and differentiated cells
In differentiated cells, aging is associated with hypermethylation of DNA regions enriched in repressive histone post-translational modifications. However, the chromatin marks associated with changes in DNA methylation in adult stem cells during lifetime are still largely unknown. Here, DNA methylation profiling of mesenchymal stem cells (MSCs) obtained from individuals aged 2 to 92 yr identified 18,735 hypermethylated and 45,407 hypomethylated CpG sites associated with aging. As in differentiated cells, hypermethylated sequences were enriched in chromatin repressive marks. Most importantly, hypomethylated CpG sites were strongly enriched in the active chromatin mark H3K4me1 in stem and differentiated cells, suggesting this is a cell type-independent chromatin signature of DNA hypomethylation during aging. Analysis of scedasticity showed that interindividual variability of DNA methylation increased during aging in MSCs and differentiated cells, providing a new avenue for the identification of DNA methylation changes over time. DNA methylation profiling of genetically identical individuals showed that both the tendency of DNA methylation changes and scedasticity depended on nongenetic as well as genetic factors. Our results indicate that the dynamics of DNA methylation during aging depend on a complex mixture of factors that include the DNA sequence, cell type, and chromatin context involved and that, depending on the locus, the changes can be modulated by genetic and/or external factors
The outcome of boosting mitochondrial activity in alcohol-associated liver disease is organ-dependent.
BACKGROUND AND AIMS
Alcohol-associated liver disease (ALD) accounts for 70% of liver-related deaths in Europe, with no effective approved therapies. Although mitochondrial dysfunction is one of the earliest manifestations of alcohol-induced injury, restoring mitochondrial activity remains a problematic strategy due to oxidative stress. Here, we identify methylation-controlled J protein (MCJ) as a mediator for ALD progression and hypothesize that targeting MCJ may help in recovering mitochondrial fitness without collateral oxidative damage.
APPROACH AND RESULTS
C57BL/6 mice [wild-type (Wt)] Mcj knockout and Mcj liver-specific silencing (MCJ-LSS) underwent the NIAAA dietary protocol (Lieber-DeCarli diet containing 5% (vol/vol) ethanol for 10 days, plus a single binge ethanol feeding at day 11). To evaluate the impact of a restored mitochondrial activity in ALD, the liver, gut, and pancreas were characterized, focusing on lipid metabolism, glucose homeostasis, intestinal permeability, and microbiota composition. MCJ, a protein acting as an endogenous negative regulator of mitochondrial respiration, is downregulated in the early stages of ALD and increases with the severity of the disease. Whole-body deficiency of MCJ is detrimental during ALD because it exacerbates the systemic effects of alcohol abuse through altered intestinal permeability, increased endotoxemia, and dysregulation of pancreatic function, which overall worsens liver injury. On the other hand, liver-specific Mcj silencing prevents main ALD hallmarks, that is, mitochondrial dysfunction, steatosis, inflammation, and oxidative stress, as it restores the NAD + /NADH ratio and SIRT1 function, hence preventing de novo lipogenesis and improving lipid oxidation.
CONCLUSIONS
Improving mitochondrial respiration by liver-specific Mcj silencing might become a novel therapeutic approach for treating ALD.This work was supported by grants from Ministerio de
Ciencia e Innovación, Programa Retos-Colaboración
RTC2019-007125-1 (for Jorge Simon and Maria Luz
Martinez-Chantar); Ministerio de Economía, Industria y
Competitividad, Retos a la Sociedad AGL2017-
86927R (for F.M.); Instituto de Salud Carlos III,
Proyectos de Investigación en Salud DTS20/00138
and DTS21/00094 (for Jorge Simon and Maria Luz
Martinez-Chantar, and Asis Palazon. respectively);
Instituto de Salud Carlos III, Fondo de Investigaciones
Sanitarias co-founded by European Regional
Development Fund/European Social Fund, “Investing
in your future” PI19/00819, “Una manera de
hacer Europa” FIS PI20/00765, and PI21/01067 (for
Jose J. G. Marin., Pau Sancho-Bru,. and Mario F.
Fraga respectively); Departamento de Industria del
Gobierno Vasco (for Maria Luz Martinez-Chantar);
Asturias Government (PCTI) co-funding 2018-2023/
FEDER IDI/2021/000077 (for Mario F. Fraga.);
Ministerio de Ciencia, Innovación y Universidades
MICINN: PID2020-117116RB-I00, CEX2021-001136-S
PID2020-117941RB-I00, PID2020-11827RB-I00 and
PID2019-107956RA-100 integrado en el Plan Estatal
de Investigación Científica y Técnica y Innovación,
cofinanciado con Fondos FEDER (for Maria Luz Martinez-Chantar, Francisco J Cubero., Yulia A Nevzorova
and Asis Palazon); Ayudas Ramón y Cajal de la Agencia
Estatal de Investigación RY2013-13666 and RYC2018-
024183-I (for Leticia Abecia and Asis Palazon); European Research Council Starting Grant 804236 NEXTGEN-IO (for Asis Palazon); The German Research
Foundation SFB/TRR57/P04, SFB1382-403224013/
A02 and DFG NE 2128/2-1 (for Francisco J Cubero
and Yulia A Nevzorova); National Institute of Health (NIH)/National Institute of Alcohol Abuse and Alcoholism
(NIAAA) 1U01AA026972-01 (For Pau Sancho-Bru);
Junta de Castilla y León SA074P20 (for Jose J. G.
Marin); Junta de Andalucía, Grupo PAIDI BIO311 (for
Franz Martin); CIBERER Acciones Cooperativas y
Complementarias Intramurales ACCI20-35 (for Mario F.
Fraga); Ministerio de Educación, Cultura y Deporte
FPU17/04992 (for Silvia Ariño); Fundació Marato TV3
201916-31 (for Jose J. G. Marin.); Ainize Pena-Cearra is
a fellow of the University of the Basque Country (UPV/
EHU); BIOEF (Basque Foundation for Innovation and
Health Research); Asociación Española contra el Cáncer
(Maria Luz Martinez-Chantar and Teresa C. Delgado.);
Fundación Científica de la Asociación Española Contra
el Cáncer (AECC Scientific Foundation) Rare Tumor
Calls 2017 (for Maria Luz Martinez-Chantar); La Caixa
Foundation Program (for Maria Luz Martinez-Chantar);
Proyecto Desarrollo Tecnologico CIBERehd (for Maria
Luz Martinez-Chantar); Ciberehd_ISCIII_MINECO is
funded by the Instituto de Salud Carlos III.S