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PURPOSE: Biallelic variants in TARS2, encoding the mitochondrial threonyl-tRNA-synthetase, have been reported in a small group of individuals displaying a neurodevelopmental phenotype but with limited neuroradiological data and insufficient evidence for causality of the variants. METHODS: Exome or genome sequencing was carried out in 15 families. Clinical and neuroradiological evaluation was performed for all affected individuals, including review of 10 previously reported individuals. The pathogenicity of TARS2 variants was evaluated using in vitro assays and a zebrafish model. RESULTS: We report 18 new individuals harboring biallelic TARS2 variants. Phenotypically, these individuals show developmental delay/intellectual disability, regression, cerebellar and cerebral atrophy, basal ganglia signal alterations, hypotonia, cerebellar signs, and increased blood lactate. In vitro studies showed that variants within the TARS2301-381 region had decreased binding to Rag GTPases, likely impairing mTORC1 activity. The zebrafish model recapitulated key features of the human phenotype and unraveled dysregulation of downstream targets of mTORC1 signaling. Functional testing of the variants confirmed the pathogenicity in a zebrafish model. CONCLUSION: We define the clinico-radiological spectrum of TARS2-related mitochondrial disease, unveil the likely involvement of the mTORC1 signaling pathway as a distinct molecular mechanism, and establish a TARS2 zebrafish model as an important tool to study variant pathogenicity.
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ARN de Transferencia , Pez Cebra , Animales , Humanos , Mutación , Pez Cebra/genética , Diana Mecanicista del Complejo 1 de la Rapamicina , Ligasas , FenotipoRESUMEN
We describe two female infants at the age of five and six months with urinary tract infections presenting with vomiting and reduced drinking behavior. On laboratory analysis, severe hyponatremia (106â mmol/L and 109â mmol/L) was seen with hyperkalemia and compensated metabolic acidosis. Endocrinological analyses revealed massively increased levels of aldosterone and renin, leading to the diagnosis of type III pseudohypoaldosteronism (PHA). A review of the current literature 2013-2023 revealed 26 type III PHA cases aged up to ten months with reduced drinking behavior, weight loss and/or failure to thrive being the most common clinical presentations. Given the severe presentation of PHA electrolyte measurements in infants with urinary tract infections and/or in infants with congenital anomalies of the kidney and urinary tract (CAKUT) are strongly recommended.
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At the fundamental level, messenger RNA (mRNA)-based therapeutics involves the delivery of in vitro-transcribed (IVT) mRNA into the cytoplasm of a target cell, where it is translated into the desired protein. IVT mRNA presents various advantages compared to DNA and recombinant protein-based approaches that make it ideal for a broad range of therapeutic applications. IVT mRNA, which is translated in the cytoplasm after transfection into cells, can encode virtually any target protein. Notably, it does not enter the nucleus, which avoids its integration into the genome and the risk of insertional mutagenesis. The large-scale production of IVT mRNA is less complex than production of recombinant proteins, and Good Manufacturing Practice-compliant mRNA production is easily scalable, ideally poising mRNA for not only off-the-shelf, but more personalized treatment approaches. IVT mRNA's safety profile, pharmacokinetics, and pharmacodynamics, including its inherent immunostimulatory capacity, can be optimized for different therapeutic applications by harnessing a wide array of optimized sequence elements, chemical modifications, purification techniques, and delivery methods. The value of IVT mRNA was recently proved during the COVID-19 pandemic when mRNA-based vaccines outperformed the efficacy of established technologies, and millions of doses were rapidly deployed. In this review, we will discuss chemical modifications of IVT mRNA and highlight numerous preclinical and clinical applications including vaccines for cancer and infectious diseases, cancer immunotherapy, protein replacement, gene editing, and cell reprogramming.
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COVID-19 , Neoplasias , COVID-19/terapia , Humanos , Inmunoterapia , Pandemias , Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
After one year of absence, the 18th Annual Meeting of the Association for Cancer Immunotherapy (CIMT), Europe's cancer immunotherapy meeting, took place virtually from 10 to 12 May 2021. Over 850 academic and clinical professionals from 30 countries met to discuss the recent advancements in cancer immunotherapy and the current progress in COVID19-related research. This meeting report summarizes the highlights of CIMT2021.
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COVID-19 , Neoplasias , Humanos , Inmunoterapia , Neoplasias/terapia , Microambiente TumoralRESUMEN
BACKGROUND: The global COVID-19 pandemic has had a significant impact on the physical and mental health of people everywhere. The aim of the study is to understand how people living in 15 countries around the globe experience an unexpected crisis which threatens their health and that of loved ones, and how they make meaning of this disruption in their narratives. METHODS: Data were collected through an anonymous online survey during May-September 2020, which was during or just after the first wave of the COVID-19 pandemic, depending on the country. The questionnaire included demographic and three open-ended questions as prompts for stories about experiences during the initial months of the pandemic. The text was analyzed through inductive thematic content analysis and quantified for full sample description, demographic and subsequently international comparisons. RESULTS: The final qualitative dataset included stories from n = 1685 respondents. The sample was 73.6% women and 26.4% men. The mean age of participants was 39.55 years (SD = 14.71). The identified four groups of overarching themes were: The presence and absence of others; Rediscovering oneself; The meaning of daily life; Rethinking societal and environmental values. We discuss the prevalence of each theme for the sample as a whole and differences by demographic groups. The most prevalent theme referred to disruptions in interpersonal contacts, made meaningful by the increased appreciation of the value of relationships, present in (45.6%) of stories. It was more prevalent in the stories of women compared to men (χ² = 24.88, p = .001). CONCLUSIONS: The paper provides a detailed overview of the methodology, the main themes identified inductively in the stories and differences according to select demographic variables. We identify several major ways of making meaning of the pandemic. The pandemic has impacted many aspects of people's lives which give it meaning, no matter where they live.
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BACKGROUND: Charcot-Marie-Tooth disease type 4J (CMT4J) originates from mutations in the FIG4 gene and leads to distal muscle weakness. Two null alleles of FIG4 cause Yunis Varón syndrome with severe central nervous system involvement, cleidocranial dysmorphism, absent thumbs and halluces and early death. OBJECTIVES: To analyse the phenotypic spectrum of FIG4-related disease and explore effects of residual FIG4 protein. METHODS: Phenotyping of five new patients with FIG4-related disease. Western Blot analyses of FIG4 from patient fibroblasts. RESULTS: Next generation sequencing revealed compound heterozygous variants in FIG4 in five patients. All five patients presented with peripheral neuropathy, various degree of dysmorphism and a central nervous system involvement comprising Parkinsonism in 3/5 patients, cerebellar ataxia (1/5), spasticity of lower limbs (1/5), epilepsy (1/5) and/or cognitive deficits (2/5). Onset varied between the first and the seventh decade. There was no residual FIG4 protein detectable in fibroblasts of the four analysed patients. CONCLUSION: This study extends the phenotypic spectrum of FIG4-related disease to Parkinsonism as a feature and demonstrates new phenotypes on a continuum between CMT4J and Yunis Varón syndrome.
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Enfermedad de Charcot-Marie-Tooth/genética , Displasia Cleidocraneal/genética , Displasia Ectodérmica/genética , Flavoproteínas/genética , Deformidades Congénitas de las Extremidades/genética , Micrognatismo/genética , Trastornos Parkinsonianos/genética , Monoéster Fosfórico Hidrolasas/genética , Adulto , Anciano , Enfermedad de Charcot-Marie-Tooth/fisiopatología , Displasia Cleidocraneal/fisiopatología , Displasia Ectodérmica/fisiopatología , Femenino , Fibroblastos , Humanos , Deformidades Congénitas de las Extremidades/fisiopatología , Masculino , Micrognatismo/fisiopatología , Persona de Mediana Edad , Mutación , Trastornos Parkinsonianos/fisiopatología , Linaje , Fenotipo , Adulto JovenRESUMEN
α-Linolenic acid (ALA) is well-known for its anti-inflammatory activity. In contrast, the influence of an ALA-rich diet on intestinal microbiota composition and its impact on small intestine morphology are not fully understood. In the current study, we kept adult C57BL/6J mice for 4 weeks on an ALA-rich or control diet. Characterization of the microbial composition of the small intestine revealed that the ALA diet was associated with an enrichment in Prevotella and Parabacteroides. In contrast, taxa belonging to the Firmicutes phylum, including Lactobacillus, Clostridium cluster XIVa, Lachnospiraceae and Streptococcus, had significantly lower abundance compared to control diet. Metagenome prediction indicated an enrichment in functional pathways such as bacterial secretion system in the ALA group, whereas the two-component system and ALA metabolism pathways were downregulated. We also observed increased levels of ALA and its metabolites eicosapentanoic and docosahexanoic acid, but reduced levels of arachidonic acid in the intestinal tissue of ALA-fed mice. Furthermore, intestinal morphology in the ALA group was characterized by elongated villus structures with increased counts of epithelial cells and reduced epithelial proliferation rate. Interestingly, the ALA diet reduced relative goblet and Paneth cell counts. Of note, high-fat Western-type diet feeding resulted in a comparable adaptation of the small intestine. Collectively, our study demonstrates the impact of ALA on the gut microbiome and reveals the nutritional regulation of gut morphology.
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Alimentación Animal , Biodiversidad , Microbioma Gastrointestinal , Mucosa Intestinal/microbiología , Intestino Delgado/metabolismo , Intestino Delgado/microbiología , Ácido alfa-Linolénico/metabolismo , Alimentación Animal/análisis , Animales , Ácidos Grasos/metabolismo , Heces/microbiología , Análisis de los Alimentos , Inmunohistoquímica , Mucosa Intestinal/citología , Metabolismo de los Lípidos , Masculino , Metagenoma , Metagenómica/métodos , Ratones , Ácido alfa-Linolénico/análisisRESUMEN
Superoxide dismutase 1 (SOD1) is the principal cytoplasmic superoxide dismutase in humans and plays a major role in redox potential regulation. It catalyses the transformation of the superoxide anion (O2â¢-) into hydrogen peroxide. Heterozygous variants in SOD1 are a common cause of familial amyotrophic lateral sclerosis. In this study we describe the homozygous truncating variant c.335dupG (p.C112Wfs*11) in SOD1 that leads to total absence of enzyme activity. The resulting phenotype is severe and marked by progressive loss of motor abilities, tetraspasticity with predominance in the lower extremities, mild cerebellar atrophy, and hyperekplexia-like symptoms. Heterozygous carriers have a markedly reduced enzyme activity when compared to wild-type controls but show no overt neurologic phenotype. These results are in contrast with the previously proposed theory that a loss of function is the underlying mechanism in SOD1-related motor neuron disease and should be considered before application of previously proposed SOD1 silencing as a treatment option for amyotrophic lateral sclerosis.
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Trastornos Heredodegenerativos del Sistema Nervioso/genética , Superóxido Dismutasa-1/deficiencia , Superóxido Dismutasa-1/genética , Esclerosis Amiotrófica Lateral , Niño , Preescolar , Mutación del Sistema de Lectura , Humanos , Masculino , Linaje , SíndromeRESUMEN
BACKGROUND AND AIM: Accurate assessment of structural and functional characteristics of the liver could improve the diagnosis and the clinical management of patients with chronic liver diseases. However, the structure-function relationship in the progression of chronic liver disease remains elusive. The aim of this study is the combined measurement of liver function by the 13 C-methacetin Liver MAximum capacity (LiMAx) test and tissue-structure related stiffness by 2D time-harmonic elastography for the assessment of liver disease progression. METHODS: LiMAx test and time-harmonic elastography were applied, and the serological scores fibrosis 4 index and aspartate aminotransferase to platelet ratio index were calculated in patients with chronic liver diseases (n = 75) and healthy control subjects (n = 22). In 47 patients who underwent surgery, fibrosis was graded by histological examination of the resected liver tissue. RESULTS: LiMAx values correlated negatively with liver stiffness (r = -0.747), aminotransferase to platelet ratio index (r = -0.604), and fibrosis 4 (r = -0.573). Median (interquartile range) LiMAx values decreased with fibrosis progression from 395 µg/kg/h (371-460 µg/kg/h) in participants with no fibrosis to 173 µg/kg/h (126-309 µg/kg/h) in patients with severe fibrosis. Median liver stiffness increased progressively with the stage of fibrosis from no fibrosis (1.56 m/s [1.52-1.63 m/s]) to moderate fibrosis (1.60 m/s [1.54-1.67 m/s]) to severe fibrosis (1.85 m/s [1.76-1.92 m/s]). CONCLUSION: Our findings show that structural changes in the liver due to progressing liver diseases and reflected by increased tissue stiffness correlate with a functional decline of the organ as reflected by a decreased metabolic capacity of the liver.
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Acetamidas/administración & dosificación , Isótopos de Carbono/administración & dosificación , Diagnóstico por Imagen de Elasticidad , Cirrosis Hepática/diagnóstico por imagen , Cirrosis Hepática/metabolismo , Pruebas de Función Hepática , Hígado/diagnóstico por imagen , Hígado/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Aspartato Aminotransferasas/sangre , Estudios de Casos y Controles , Progresión de la Enfermedad , Femenino , Hepatectomía , Humanos , Hígado/cirugía , Cirrosis Hepática/sangre , Cirrosis Hepática/cirugía , Masculino , Persona de Mediana Edad , Recuento de Plaquetas , Valor Predictivo de las Pruebas , Reproducibilidad de los Resultados , Índice de Severidad de la Enfermedad , Adulto JovenRESUMEN
Single-nucleotide variants that abolish the stop codon ("nonstop" alterations) are a unique type of substitution in genomic DNA. Whether they confer instability of the mutant mRNA or result in expression of a C-terminally extended protein depends on the absence or presence of a downstream in-frame stop codon, respectively. Of the predicted protein extensions, only few have been functionally characterized. In a family with autosomal dominant Charcot-Marie-Tooth disease type 2, that is, an axonopathy affecting sensory neurons as well as lower motor neurons, we identified a heterozygous nonstop variant in REEP1. Mutations in this gene have classically been associated with the upper motor neuron disorder hereditary spastic paraplegia (HSP). We show that the C-terminal extension resulting from the nonstop variant triggers self-aggregation of REEP1 and of several reporters. Our findings support the recently proposed concept of 3'UTR-encoded "cryptic amyloidogenic elements." Together with a previous report on an aggregation-prone REEP1 deletion variant in distal hereditary motor neuropathy, they also suggest that toxic gain of REEP1 function, rather than loss-of-function as relevant for HSP, specifically affects lower motor neurons. A search for similar correlations between genotype, phenotype, and effect of mutant protein may help to explain the wide clinical spectra also in other genetically determined disorders.
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Regiones no Traducidas 3'/genética , Proteínas de Transporte de Membrana/genética , Enfermedades del Sistema Nervioso Periférico/genética , Enfermedad de Charcot-Marie-Tooth/genética , Femenino , Genotipo , Humanos , Masculino , Mutación/genética , Linaje , Fenotipo , Paraplejía Espástica Hereditaria/genéticaRESUMEN
Hereditary neuropathies comprise a wide variety of chronic diseases associated to more than 80 genes identified to date. We herein examined 612 index patients with either a Charcot-Marie-Tooth phenotype, hereditary sensory neuropathy, familial amyloid neuropathy, or small fiber neuropathy using a customized multigene panel based on the next generation sequencing technique. In 121 cases (19.8%), we identified at least one putative pathogenic mutation. Of these, 54.4% showed an autosomal dominant, 33.9% an autosomal recessive, and 11.6% an X-linked inheritance. The most frequently affected genes were PMP22 (16.4%), GJB1 (10.7%), MPZ, and SH3TC2 (both 9.9%), and MFN2 (8.3%). We further detected likely or known pathogenic variants in HINT1, HSPB1, NEFL, PRX, IGHMBP2, NDRG1, TTR, EGR2, FIG4, GDAP1, LMNA, LRSAM1, POLG, TRPV4, AARS, BIC2, DHTKD1, FGD4, HK1, INF2, KIF5A, PDK3, REEP1, SBF1, SBF2, SCN9A, and SPTLC2 with a declining frequency. Thirty-four novel variants were considered likely pathogenic not having previously been described in association with any disorder in the literature. In one patient, two homozygous mutations in HK1 were detected in the multigene panel, but not by whole exome sequencing. A novel missense mutation in KIF5A was considered pathogenic because of the highly compatible phenotype. In one patient, the plasma sphingolipid profile could functionally prove the pathogenicity of a mutation in SPTLC2. One pathogenic mutation in MPZ was identified after being previously missed by Sanger sequencing. We conclude that panel based next generation sequencing is a useful, time- and cost-effective approach to assist clinicians in identifying the correct diagnosis and enable causative treatment considerations.
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Predisposición Genética a la Enfermedad , Neuropatía Hereditaria Motora y Sensorial/genética , Mutación/genética , Enfermedades Raras/genética , Enfermedad de Charcot-Marie-Tooth/genética , Femenino , Proteínas de Choque Térmico HSP27/genética , Proteínas de Choque Térmico , Neuropatía Hereditaria Motora y Sensorial/diagnóstico , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Masculino , Chaperonas Moleculares , FenotipoRESUMEN
PURPOSE: Mutations in the RLBP1 gene encoding the cellular retinaldehyde-binding protein (CRALBP) cause autosomal recessive progressive retinopathy, such as retinitis punctata albescens (RPA), Bothnia-type dystrophy (BD), Newfoundland rod-cone dystrophy (NFRCD), retinitis pigmentosa (RP) and fundus albipunctatus (FA). We present the clinical heterogeneity and genetic findings of seven patients from five families with RLBP1 mutations, including three novel mutations. METHODS: Seven patients underwent complete ophthalmological examination including psychophysical tests (visual acuity, colour vision, visual field, dark adaptation) and electrophysiology (Ganzfeld and multifocal ERG). Additionally, fundus photography, autofluorescence (FAF) and spectral domain optical coherence tomography (OCT) recordings were performed. Genomic DNA was analysed by high-throughput sequencing for all RP-related genes in a diagnostic set-up. RESULTS: The patients presented with variable phenotypes, including RPA, BD, RP and a mild form of NFRCD. No detectable or severely depressed responses in electrophysiological examinations were seen in all cases. Visual field constriction was variable among individuals. Severely reduced visual acuity was only observed in the patient presenting with BD. The other patients retained mild to moderate reduction of visual function. Despite the morphological differences, central retinal thinning - as a common feature - could be observed. CONCLUSIONS: The fact that different mutations in RLBP1 are correlated with quite different morphological and functional characteristics outlines the complexity of the protein. Identifying new mutations and comparing the different phenotypes may help to better understand the function of the protein and the consequences in pathological changes that involve RPE and choroid.
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Proteínas Portadoras/genética , Mutación , Distrofias Retinianas/genética , Adulto , Anciano , Niño , Electrorretinografía , Femenino , Angiografía con Fluoresceína , Estudios de Asociación Genética , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Distrofias Retinianas/diagnóstico , Hermanos , Tomografía de Coherencia Óptica , Agudeza Visual/fisiología , Adulto JovenRESUMEN
Hereditary retinal dystrophies (RD) constitute a group of blinding diseases that are characterized by clinical variability and pronounced genetic heterogeneity. The different forms of RD can be caused by mutations in >100 genes, including >1600 exons. Consequently, next generation sequencing (NGS) technologies are among the most promising approaches to identify mutations in RD. So far, NGS is not routinely used in gene diagnostics. We developed a diagnostic NGS pipeline to identify mutations in 170 genetically and clinically unselected RD patients. NGS was applied to 105 RD-associated genes. Underrepresented regions were examined by Sanger sequencing. The NGS approach was successfully established using cases with known sequence alterations. Depending on the initial clinical diagnosis, we identified likely causative mutations in 55% of retinitis pigmentosa and 80% of Bardet-Biedl or Usher syndrome cases. Seventy-one novel mutations in 40 genes were newly associated with RD. The genes USH2A, EYS, ABCA4, and RHO were more frequently affected than others. Occasionally, cases carried mutations in more than one RD-associated gene. In addition, we found possible dominant de-novo mutations in cases with sporadic RD, which implies consequences for counseling of patients and families. NGS-based mutation analyses are reliable and cost-efficient approaches in gene diagnostics of genetically heterogeneous diseases like RD.
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Predisposición Genética a la Enfermedad , Patología Molecular , Distrofias Retinianas/genética , Retinitis Pigmentosa/genética , Síndromes de Usher/genética , Transportadoras de Casetes de Unión a ATP/genética , Exones , Proteínas de la Matriz Extracelular/genética , Proteínas del Ojo/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Mutación , Linaje , Distrofias Retinianas/etiología , Distrofias Retinianas/patología , Retinitis Pigmentosa/etiología , Análisis de Secuencia de ADN , Síndromes de Usher/etiología , Síndromes de Usher/patologíaRESUMEN
Autosomal recessive polycystic kidney disease (ARPKD), although less frequent than the dominant form, is a common, inherited ciliopathy of childhood that is caused by mutations in the PKHD1-gene on chromosome 6. The characteristic dilatation of the renal collecting ducts starts in utero and can present at any stage from infancy to adulthood. Renal insufficiency may already begin in utero and may lead to early abortion or oligohydramnios and lung hypoplasia in the newborn. However, there are also affected children who have no evidence of renal dysfunction in utero and who are born with normal renal function. Up to 30 % of patients die in the perinatal period, and those surviving the neonatal period reach end stage renal disease (ESRD) in infancy, early childhood or adolescence. In contrast, some affected patients have been diagnosed as adults with renal function ranging from normal to moderate renal insufficiency to ESRD. The clinical spectrum of ARPKD is broader than previously recognized. While bilateral renal enlargement with microcystic dilatation is the predominant clinical feature, arterial hypertension, intrahepatic biliary dysgenesis remain important manifestations that affect approximately 45 % of infants. All patients with ARPKD develop clinical findings of congenital hepatic fibrosis (CHF); however, non-obstructive dilation of the intrahepatic bile ducts in the liver (Caroli's disease) is seen at the histological level in only a subset of patients. Cholangitis and variceal bleeding, sequelae of portal hypertension, are life-threatening complications that may occur more often in advanced cases of liver disease. In this review we focus on common and uncommon kidney-related and non-kidney-related phenotypes. Clinical management of ARPKD patients should include consideration of potential problems related to these manifestations.
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Riñón Poliquístico Autosómico Recesivo/patología , Humanos , Fenotipo , Riñón Poliquístico Autosómico Recesivo/complicacionesRESUMEN
BACKGROUND: Dialectical behavior therapy (DBT) has been proven to be an efficacious treatment for borderline personality disorder (BPD) in several randomized controlled trials (RCTs). However, generalizability of this outcome to the routine health care (effectiveness) has rarely been investigated to date. The aim of this study is to examine the effectiveness of DBT for BPD under the routine health care situation in Germany. METHODS: The study has a longitudinal design over a course of four years with six assessment points. In this paper, results for the first year of treatment are reported. Outcome was assessed at four times throughout an initial phase (of up to five therapy-sessions) and an additional 12 months of therapy. Overall, n =78 patients started the study, 47 patients completed one year of treatment. Dependent variables were number and duration of inpatient treatment stays, number of suicide attempts and non-suicidal self-injury, severity of borderline symptoms, depression, level of dissociation, and general psychopathology. RESULTS: Patients significantly improved regarding self-injurious behaviors, number of inpatient hospital stays, severity of borderline symptoms and psychopathology. At the end of the first treatment year, 77% of the patients no longer met criteria for BPD diagnosis. Fewer therapy discontinuations by patients were observed when therapists participated in consultation teams. CONCLUSIONS: Under routine mental health care conditions in Germany, outpatient DBT leads to positive results comparable to those reported in other effectiveness studies and in randomized controlled trials.
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Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , Síndrome de Cornelia de Lange/genética , Sitios de Empalme de ARN/genética , Niño , Síndrome de Cornelia de Lange/patología , Femenino , Humanos , Recién Nacido , Mutación , CohesinasRESUMEN
The DNA damage pathway plays a central role in chemoresistance in chronic lymphocytic leukemia (CLL), as indicated by the prognostic impact of TP53 and ATM loss/mutations. We investigated the function of the p53 axis in primary CLL samples by studying p53 and p21 responses to irradiation by FACS and RT-PCR. We observed a distinct response pattern for most cases with a 17p deletion (n = 16) or a sole TP53 mutation (n = 8), but not all cases with a p53 aberration were detected based on a number of different assays used. Samples with a small clone with a TP53 mutation remained undetected in all assays. Only 1 of 123 cases showed high expression of p53, which is suggestive of p53 aberration without proof of mutation of TP53. Samples with an 11q deletion showed a heterogeneous response, with only 13 of 30 showing an abnormal response based on cutoff. Nevertheless, the overall induction of p53 and p21 was impaired, suggesting a gene-dosage effect for ATM in the 11q-deleted samples. The detectability of p53 defects is influenced by clonal heterogeneity and sample purity. Functional assays of p53 defects will detect a small number of cases not detectable by FISH or TP53 mutational analysis. The clinical utility of functional p53 testing will need to be derived from clinical trials.
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Cromosomas Humanos Par 17/genética , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Daño del ADN , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/patología , Proteína p53 Supresora de Tumor/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Western Blotting , Adhesión Celular , Ciclo Celular , Diferenciación Celular , Movimiento Celular , Proliferación Celular , Células Cultivadas , Deleción Cromosómica , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Análisis Mutacional de ADN , Femenino , Citometría de Flujo , Humanos , Hibridación Fluorescente in Situ , Linfocitos/metabolismo , Linfocitos/patología , Masculino , Persona de Mediana Edad , Mutación/genética , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transcripción GenéticaRESUMEN
BACKGROUND: Chronic lymphocytic leukemia has a variable clinical course. Genomic aberrations identify prognostic subgroups, pointing towards distinct underlying biological mechanisms that are poorly understood. In particular it remains unclear whether the prognostic subgroups of chronic lymphocytic leukemia are characterized by different levels of leukemogenic proteins. DESIGN AND METHODS: Expression of 23 proteins involved in apoptosis, proliferation, DNA damage, and signaling or whose genes map to chromosomal regions known to be critical in chronic lymphocytic leukemia was quantified in 185 cytogenetically well characterized cases of chronic lymphocytic leukemia using immunoblotting. Cases were categorized hierarchically into deletion(17p), deletion(11q), trisomy 12, deletion(13q) as sole abnormality or normal karyotype. Statistical analysis was performed for expression differences between these subgroups. In addition, the expression levels of CDK4, P27 and P53 were quantified over the clinical course and compared to levels in immunopurified B cells from healthy individuals. RESULTS: In subgroups with a good prognosis, differential expression was mainly seen for proteins that regulate apoptosis. In contrast, in cytogenetic subgroups with a worse prognosis, differential expression was mostly detected for proteins that control DNA damage and proliferation. Expression levels of CDK4, P27 and P53 were higher compared to those in B cells from healthy individuals and significantly correlated with increasing hierarchical risk. In addition, no significant longitudinal changes of expression levels of CDK4, P27 and P53 could be detected in chronic lymphocytic leukemia patients. CONCLUSIONS: Differences in expression levels of apoptosis- and proliferation-controlling proteins define distinct prognostic subgroups of chronic lymphocytic leukemia and uncover a correlation of levels of CDK4, P27 and P53 proteins with higher hierarchical risk.
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Aberraciones Cromosómicas , Quinasa 4 Dependiente de la Ciclina/biosíntesis , Regulación Leucémica de la Expresión Génica , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Leucemia Linfocítica Crónica de Células B/metabolismo , Proteína p53 Supresora de Tumor/biosíntesis , Apoptosis/genética , Linfocitos B/metabolismo , Línea Celular Transformada , Línea Celular Tumoral , Proliferación Celular , Quinasa 4 Dependiente de la Ciclina/genética , Inhibidor p27 de las Quinasas Dependientes de la Ciclina , Femenino , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Leucemia Linfocítica Crónica de Células B/diagnóstico , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/mortalidad , Masculino , Factores de Riesgo , Proteína p53 Supresora de Tumor/genéticaRESUMEN
The prognosis of fludarabine (F)-refractory chronic lymphocytic leukemia (CLL) is very poor, and underlying mechanisms are only partly understood. To assess the contribution of p53 abnormalities to F-refractory CLL, we studied TP53 mutations in the CLL2H trial (subcutaneous alemtuzumab; n = 99). We found TP53 mutations in 37% of patients. Twelve of 67 (18%) patients without the 17p deletion showed a TP53 mutation and 50% showed evidence of uniparental disomy. A total of 75% of cases with TP53 mutation (without 17p-) showed clonal evolution/expansion. TP53 mutations had no impact on overall survival (P = .48). CLL with the 17p deletion or TP53 mutation showed very low miR-34a expression. To investigate the mechanisms underlying refractory CLL beyond p53, we studied cases without 17p-/TP53 mutation in detail. In several paired samples before and after F-refractory disease, no change in p21/p53 induction was observed after DNA damage. Although TP53 mutations and 17p deletions are found in a high proportion of F-refractory CLL, more than half of the cases cannot be explained by p53 defects (deletion or mutation), and alternative mechanisms need to be investigated. Alemtuzumab is effective irrespective of genetic high-risk subgroups with TP53 mutations. These clinical trials are registered at www.clinicaltrials.gov as #NCT00274976.