RESUMO
Gross cytogenetic anomalies are traditionally being used as diagnostic, prognostic and therapeutic markers in the clinical management of cancer, including childhood acute lymphoblastic leukemia (ALL). Recently, it has become increasingly clear that genetic lesions driving tumorigenesis frequently occur at the submicroscopic level and, consequently, escape standard cytogenetic observations. Therefore, we profiled the genomes of 40 childhood ALLs at high resolution. We detected multiple de novo genetic lesions, including gross aneuploidies and segmental gains and losses, some of which were subtle and affected single genes. Many of these lesions involved recurrent (partially) overlapping deletions and duplications, containing various established leukemia-associated genes, such as ETV6, RUNX1 and MLL. Importantly, the most frequently affected genes were those controlling G1/S cell cycle progression (e.g. CDKN2A, CDKN1B and RB1), followed by genes associated with B-cell development. The latter group includes microdeletions of the B-lineage transcription factors PAX5, EBF, E2-2 and IKZF1 (Ikaros), as well as genes with other established roles in B-cell development, that is RAG1 and RAG2, FYN, PBEF1 or CBP/PAG. The fact that we frequently encountered multiple lesions affecting genes involved in cell cycle regulation and B-cell differentiation strongly suggests that both these processes need to be targeted independently and simultaneously to trigger ALL development.
Assuntos
Ciclo Celular/genética , Diferenciação Celular/genética , Genes Neoplásicos , Linfócitos/citologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Linfócitos B/citologia , Aberrações Cromossômicas , Feminino , Dosagem de Genes , Perfilação da Expressão Gênica/métodos , Genômica/métodos , Humanos , Masculino , Hibridização de Ácido Nucleico , Leucemia-Linfoma Linfoblástico de Células Precursoras/etiologia , Fatores de TranscriçãoRESUMO
The contribution of genetic predisposing factors to the development of pediatric acute lymphoblastic leukemia (ALL), the most frequently diagnosed cancer in childhood, has not been fully elucidated. Children presenting with multiple de novo leukemias are more likely to suffer from genetic predisposition. Here, we selected five of these patients and analyzed the mutational spectrum of normal and malignant tissues. In two patients, we identified germline mutations in TYK2, a member of the JAK tyrosine kinase family. These mutations were located in two adjacent codons of the pseudokinase domain (p.Pro760Leu and p.Gly761Val). In silico modeling revealed that both mutations affect the conformation of this autoregulatory domain. Consistent with this notion, both germline mutations promote TYK2 autophosphorylation and activate downstream STAT family members, which could be blocked with the JAK kinase inhibitor I. These data indicate that germline activating TYK2 mutations predispose to the development of ALL.
Assuntos
Mutação em Linhagem Germinativa , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , TYK2 Quinase/genética , Alelos , Substituição de Aminoácidos , Exoma , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Modelos Moleculares , Fosforilação , Polimorfismo de Nucleotídeo Único , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Fatores de Transcrição STAT/metabolismo , TYK2 Quinase/química , TYK2 Quinase/metabolismoRESUMO
Autologous skin grafts are the gold standard for the treatment of burn wounds. In a number of cases, treatment with autologous tissue is not possible and skin substitutes are used. The outcome, however, is not optimal and improvements are needed. Inspired by scarless healing in early embryonic development, we here set out a strategy for the design and construction of embryonic-like scaffolds for skin tissue engineering. This strategy may serve as a general approach in the construction of embryonic-like scaffolds for other tissues/organ. As a first step, key effector molecules upregulated during embryonic and neonatal skin formation were identified using a comparative gene expressing analysis. A set of 20 effector molecules was identified, from which insulin-like growth factor 2 (IGF2) and sonic hedgehog (SHH) were selected for incorporation into a type I collagen-heparin scaffold. Porous scaffolds were constructed using purified collagen fibrils and 6% covalently bound heparin (to bind and protect the growth factors), and IGF2 and SHH were incorporated either individually (~0.7 and 0.4 µg/mg scaffolds) or in combination (combined ~1.5 µg/mg scaffolds). In addition, scaffolds containing hyaluronan (up to 20 µg/mg scaffold) were prepared, based on the up- or downregulation of genes involved in hyaluronan synthesis/degradation and its suggested role in scarless healing. In conclusion, based on a comprehensive gene expression analysis, a set of effector molecules and matrix molecules was identified and incorporated into porous scaffolds. The scaffolds thus prepared may create an 'embryonic-like' environment for cells to recapitulate embryonic events and for new tissues/organs.
Assuntos
Embrião de Mamíferos/citologia , Pele/metabolismo , Engenharia Tecidual/métodos , Alicerces Teciduais/química , Animais , Western Blotting , Bovinos , Colágeno/farmacologia , Colágeno Tipo I/farmacologia , Embrião de Mamíferos/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Proteínas Hedgehog/metabolismo , Heparina/farmacologia , Ácido Hialurônico/farmacologia , Imuno-Histoquímica , Fator de Crescimento Insulin-Like II/farmacologia , Camundongos Endogâmicos C57BLRESUMO
Response to therapy as determined by minimal residual disease (MRD) is currently used for stratification in treatment protocols for pediatric acute lymphoblastic leukemia (ALL). However, the large MRD-based medium risk group (MRD-M; 50-60% of the patients) harbors many relapses. We analyzed MRD in 131 uniformly treated precursor-B-ALL patients and evaluated whether combined MRD and IKZF1 (Ikaros zinc finger-1) alteration status can improve risk stratification. We confirmed the strong prognostic significance of MRD classification, which was independent of IKZF1 alterations. Notably, 8 of the 11 relapsed cases in the large MRD-M group (n=81; 62%) harbored an IKZF1 alteration. Integration of both MRD and IKZF1 status resulted in a favorable outcome group (n=104; 5 relapses) and a poor outcome group (n=27; 19 relapses), and showed a stronger prognostic value than each of the established risk factors alone (hazard ratio (95%CI): 24.98 (8.29-75.31)). Importantly, whereas MRD and IKZF1 status alone identified only 46 and 54% of the relapses, respectively, their integrated use allowed prediction of 79% of all the relapses with 93% specificity. Because of the unprecedented sensitivity in upfront relapse prediction, the combined parameters have high potential for future risk stratification, particularly for patients originally classified as non-high risk, such as the large group of MRD-M patients.
Assuntos
Fator de Transcrição Ikaros/genética , Neoplasia Residual/patologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Valor Preditivo dos Testes , Criança , Rearranjo Gênico , Humanos , Estimativa de Kaplan-Meier , Mutação , Leucemia-Linfoma Linfoblástico de Células Precursoras/mortalidade , Recidiva , Medição de Risco , Sensibilidade e EspecificidadeRESUMO
Relapse is the most common cause of treatment failure in pediatric acute lymphoblastic leukemia (ALL) and is often difficult to predict. To explore the prognostic impact of recurrent DNA copy number abnormalities on relapse, we performed high-resolution genomic profiling of 34 paired diagnosis and relapse ALL samples. Recurrent lesions detected at diagnosis, including PAX5, CDKN2A and EBF1, were frequently absent at relapse, indicating that they represent secondary events that may be absent in the relapse-prone therapy-resistant progenitor cell. In contrast, deletions and nonsense mutations in IKZF1 (IKAROS) were highly enriched and consistently preserved at the time of relapse. A targeted copy number screen in an unselected cohort of 131 precursor B-ALL cases, enrolled in the dexamethasone-based Dutch Childhood Oncology Group treatment protocol ALL9, revealed that IKZF1 deletions are significantly associated with poor relapse-free and overall survival rates. Separate analysis of ALL9-treatment subgroups revealed that non-high-risk (NHR) patients with IKZF1 deletions exhibited a approximately 12-fold higher relative relapse rate than those without IKZF1 deletions. Consequently, IKZF1 deletion status allowed the prospective identification of 53% of the relapse-prone NHR-classified patients within this subgroup and, therefore, serves as one of the strongest predictors of relapse at the time of diagnosis with high potential for future risk stratification.