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1.
Immunity ; 56(7): 1631-1648.e10, 2023 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-37392737

RESUMO

CD137 (4-1BB)-activating receptor represents a promising cancer immunotherapeutic target. Yet, the cellular program driven by CD137 and its role in cancer immune surveillance remain unresolved. Using T cell-specific deletion and agonist antibodies, we found that CD137 modulates tumor infiltration of CD8+-exhausted T (Tex) cells expressing PD1, Lag-3, and Tim-3 inhibitory receptors. T cell-intrinsic, TCR-independent CD137 signaling stimulated the proliferation and the terminal differentiation of Tex precursor cells through a mechanism involving the RelA and cRel canonical NF-κB subunits and Tox-dependent chromatin remodeling. While Tex cell accumulation induced by prophylactic CD137 agonists favored tumor growth, anti-PD1 efficacy was improved with subsequent CD137 stimulation in pre-clinical mouse models. Better understanding of T cell exhaustion has crucial implications for the treatment of cancer and infectious diseases. Our results identify CD137 as a critical regulator of Tex cell expansion and differentiation that holds potential for broad therapeutic applications.


Assuntos
Linfócitos T CD8-Positivos , Neoplasias , Camundongos , Animais , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral , Diferenciação Celular , Proliferação de Células , Receptores de Antígenos de Linfócitos T
2.
Immunity ; 53(4): 824-839.e10, 2020 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-33053331

RESUMO

CD8+ T cells within the tumor microenvironment (TME) are exposed to various signals that ultimately determine functional outcomes. Here, we examined the role of the co-activating receptor CD226 (DNAM-1) in CD8+ T cell function. The absence of CD226 expression identified a subset of dysfunctional CD8+ T cells present in peripheral blood of healthy individuals. These cells exhibited reduced LFA-1 activation, altered TCR signaling, and a distinct transcriptomic program upon stimulation. CD226neg CD8+ T cells accumulated in human and mouse tumors of diverse origin through an antigen-specific mechanism involving the transcriptional regulator Eomesodermin (Eomes). Despite similar expression of co-inhibitory receptors, CD8+ tumor-infiltrating lymphocyte failed to respond to anti-PD-1 in the absence of CD226. Immune checkpoint blockade efficacy was hampered in Cd226-/- mice. Anti-CD137 (4-1BB) agonists also stimulated Eomes-dependent CD226 loss that limited the anti-tumor efficacy of this treatment. Thus, CD226 loss restrains CD8+ T cell function and limits the efficacy of cancer immunotherapy.


Assuntos
Antígenos de Diferenciação de Linfócitos T/imunologia , Linfócitos T CD8-Positivos/imunologia , Neoplasias/imunologia , Proteínas com Domínio T/imunologia , Animais , Humanos , Inibidores de Checkpoint Imunológico/imunologia , Imunoterapia/métodos , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/terapia , Receptor de Morte Celular Programada 1/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Transdução de Sinais/imunologia , Transcriptoma/imunologia , Microambiente Tumoral/imunologia , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/imunologia
3.
Blood ; 141(11): 1308-1315, 2023 03 16.
Artigo em Inglês | MEDLINE | ID: mdl-36375118

RESUMO

Cytogenetic abnormalities (CAs) are known to be the preponderant prognostic factor in multiple myeloma. Our team has recently developed a prognostic score based on 6 CAs, with which del(1p32) appears to be the second worst abnormality after del(17p). This study aimed to confirm the adverse effect of 1p32 deletion in patients with newly diagnosed multiple myeloma (NDMM). Among 2551 patients with newly diagnosed multiple myeloma, 11% were harboring del(1p32). Their overall survival (OS) was significantly inferior compared with patients without del(1p32) (median OS: 49 months vs 124 months). Likewise, progression-free survival was significantly shorter. More importantly, biallelic del(1p32) conferred a dramatically poorer prognosis than a monoallelic del(1p32) (median OS: 25 months vs 60 months). As expected, the OS of patients with del(1p32) significantly decreased when this abnormality was associated with other high-risk CAs [del(17p), t(4;14), or gain(1q)]. In the multivariate analysis, del(1p32) appeared as a negative prognostic factor; after adjustment for age and treatment, the risk of progression was 1.3 times higher among patients harboring del(1p32), and the risk of death was 1.9 times higher. At the dawn of risk-adapted treatment strategies, we have confirmed the adverse effect of del(1p32) in multiple myeloma and the relevance of its assessment at diagnosis.


Assuntos
Mieloma Múltiplo , Humanos , Mieloma Múltiplo/diagnóstico , Mieloma Múltiplo/genética , Prognóstico , Aberrações Cromossômicas , Hibridização in Situ Fluorescente , Intervalo Livre de Progressão
4.
Blood ; 139(17): 2666-2672, 2022 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-35171994

RESUMO

Primary plasma cell leukemia (pPCL) is an aggressive form of multiple myeloma (MM) that has not benefited from recent therapeutic advances in the field. Because it is very rare and heterogeneous, it remains poorly understood at the molecular level. To address this issue, we performed DNA and RNA sequencing of sorted plasma cells from a large cohort of 90 newly diagnosed pPCL and compared with MM. We observed that pPCL presents a specific genomic landscape with a high prevalence of t(11;14) (about half) and high-risk genomic features such as del(17p), gain 1q, and del(1p32). In addition, pPCL displays a specific transcriptome when compared with MM. We then wanted to characterize specifically pPCL with t(11;14). We observed that this subentity displayed significantly fewer adverse cytogenetic abnormalities. This translated into better overall survival when compared with pPCL without t(11;14) (39.2 months vs 17.9 months, P = .002). Finally, pPCL with t(11;14) displayed a specific transcriptome, including differential expression of BCL2 family members. This study is the largest series of patients with pPCL reported so far.


Assuntos
Leucemia Plasmocitária , Mieloma Múltiplo , Aberrações Cromossômicas , Genômica , Humanos , Leucemia Plasmocitária/diagnóstico , Mieloma Múltiplo/genética , Prognóstico , Transcriptoma
5.
Blood ; 132(23): 2456-2464, 2018 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-30249784

RESUMO

The introduction of novel agents has led to major improvements in clinical outcomes for patients with multiple myeloma. To shorten evaluation times for new treatments, health agencies are currently examining minimal residual disease (MRD) as a surrogate end point in clinical trials. We assessed the prognostic value of MRD, measured during maintenance therapy by next-generation sequencing (NGS). MRD negativity was defined as the absence of tumor plasma cell within 1 000 000 bone marrow cells (<10-6). Data were analyzed from a recent clinical trial that evaluated the role of transplantation in newly diagnosed myeloma patients treated with lenalidomide, bortezomib, and dexamethasone (RVD). MRD negativity was achieved at least once during maintenance in 127 patients (25%). At the start of maintenance therapy, MRD was a strong prognostic factor for both progression-free survival (adjusted hazard ratio, 0.22; 95% confidence interval, 0.15-0.34; P < .001) and overall survival (adjusted hazard ratio, 0.24; 95% confidence interval, 0.11-0.54; P = .001). Patients who were MRD negative had a higher probability of prolonged progression-free survival than patients with detectable residual disease, regardless of treatment group (RVD vs transplant), cytogenetic risk profile, or International Staging System disease stage at diagnosis. These results were similar after completion of maintenance therapy. Our findings confirm the value of MRD status, as determined by NGS, as a prognostic biomarker in multiple myeloma, and suggest that this approach could be used to adapt treatment strategies in future clinical trials.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Sequenciamento de Nucleotídeos em Larga Escala , Mieloma Múltiplo/metabolismo , Idoso , Medula Óssea/metabolismo , Medula Óssea/patologia , Bortezomib/administração & dosagem , Dexametasona/administração & dosagem , Intervalo Livre de Doença , Feminino , Seguimentos , Humanos , Lenalidomida/administração & dosagem , Quimioterapia de Manutenção , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/genética , Mieloma Múltiplo/mortalidade , Neoplasia Residual , Plasmócitos/metabolismo , Plasmócitos/patologia , Taxa de Sobrevida
7.
J Biotechnol ; 131(3): 223-30, 2007 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-17720272

RESUMO

Construction of synthetic genes is today the most elegant way to optimize the heterologous expression of a recombinant protein. However, the selection of positive clones that incorporate the correct synthetic DNA fragments is a bottleneck as current methods of gene synthesis introduce 3.5 nucleotide deletions per kb. Furthermore, even when all predictable optimizations for protein production have been introduced into the synthetic gene, production of the protein is often disappointing: protein is produced in too low amounts or end up in inclusion bodies. We propose a strategy to overcome these two problems simultaneously by cloning the synthetic gene upstream of a reporter gene. This permits the selection of clones devoid of frame-shift mutations. In addition, beside nucleotide deletion, an average of three non-neutral mutations per kb are introduced during gene synthesis. Using a reporter protein downstream of the synthetic gene, allows the selection of clones with random mutations improving the expression or the folding of the protein of interest. The problem of errors found in synthetic genes is then turned into an advantage since it provides polymorphism useful for molecular evolution. The use of synthetic genes appears as an alternative to the error-prone PCR strategy to generate the variations necessary in protein engineering experiments.


Assuntos
Células Clonais/classificação , Clonagem Molecular/métodos , Proteínas de Fluorescência Verde/genética , Reação em Cadeia da Polimerase/métodos , Engenharia de Proteínas/métodos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Genes Reporter/genética , Vetores Genéticos/genética , Proteínas Recombinantes de Fusão/isolamento & purificação
9.
PLoS One ; 6(10): e25510, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21991314

RESUMO

(S(C5'), R(P)) α,ß-D- Constrained Nucleic Acids (CNA) are dinucleotide building blocks that can feature either B-type torsional angle values or non-canonical values, depending on their 5'C and P absolute stereochemistry. These CNA are modified neither on the nucleobase nor on the sugar structure and therefore represent a new class of nucleotide with specific chemical and structural characteristics. They promote marked bending in a single stranded DNA so as to preorganize it into a loop-like structure, and they have been shown to induce rigidity within oligonucleotides. Following their synthesis, studies performed on CNA have only focused on the constraints that this family of nucleotides introduced into DNA. On the assumption that bending in a DNA template may produce a terminator structure, we investigated whether CNA could be used as a new strong terminator of polymerization in PCR. We therefore assessed the efficiency of CNA as a terminator in PCR, using triethylene glycol phosphate units as a control. Analyses were performed by denaturing gel electrophoresis and several PCR products were further analysed by sequencing. The results showed that the incorporation of only one CNA was always skipped by the polymerases tested. On the other hand, two CNA units always stopped proofreading polymerases, such as Pfu DNA polymerase, as expected for a strong replication terminator. Non-proofreading enzymes, e.g. Taq DNA polymerase, did not recognize this modification as a strong terminator although it was predominantly stopped by this structure. In conclusion, this first functional use of CNA units shows that these modified nucleotides can be used as novel polymerization terminators of proofreading polymerases. Furthermore, our results lead us to propose that CNA and their derivatives could be useful tools for investigating the behaviour of different classes of polymerases.


Assuntos
DNA Polimerase Dirigida por DNA/metabolismo , Ácidos Nucleicos/química , Ácidos Nucleicos/metabolismo , Reação em Cadeia da Polimerase/métodos , Temperatura , Sequência de Bases , Estabilidade Enzimática , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Polimerização , Alinhamento de Sequência
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