Summary
The landscape of medical sequencing has rapidly changed with the evolution of next generation sequencing (NGS). These technologies have contributed to the molecular characterization of the ...myelodysplastic syndromes (MDS) and chronic myelomonocytic leukaemia (CMML), through the identification of recurrent gene mutations, which are present in >80% of patients. These mutations contribute to a better classification and risk stratification of the patients. Currently, clinical laboratories include NGS genomic analyses in their routine clinical practice, in an effort to personalize the diagnosis, prognosis and treatment of MDS and CMML. NGS technologies have reduced the cost of large‐scale sequencing, but there are additional challenges involving the clinical validation of these technologies, as continuous advances are constantly being made. In this context, it is of major importance to standardize the generation, analysis, clinical interpretation and reporting of NGS data. To that end, the Spanish MDS Group (GESMD) has expanded the present set of guidelines, aiming to establish common quality standards for the adequate implementation of NGS and clinical interpretation of the results, hoping that this effort will ultimately contribute to the benefit of patients with myeloid malignancies.
Nicotinamide adenine dinucleotide (NADH) is an important enzyme cofactor with emissive properties that allow it to be used in fluorescence microscopies to study cell metabolism. Its oxidized form ...NAD+, on the other hand, is considered to produce negligible fluorescence. In this contribution, we describe the photophysics of the isolated nicotinamidic system in both its reduced and oxidized states. This was achieved through the study of model molecules that do not carry the adenine nucleotide since its absorbance would overlap with the absorption spectrum of the nicotinamidic chromophores. We studied three model molecules: nicotinamide (niacinamide, an oxidized form without nitrogen substitution), the oxidized chromophore 1-benzyl-3-carbamoyl-pyridinium bromide (NBzOx), and its reduced form 1-benzyl-1,4-dihydronicotinamide (NBz). For a full understanding of the dynamics, we performed both femtosecond-resolved emission and transient absorption experiments. The oxidized systems, nicotinamide and NBzOx, have similar photophysics, where the originally excited bright state decays on an ultrafast timescale of less than 400 fs. The depopulation of this state is followed by excited-state positive absorption signals, which evolve in two timescales: the first one is from 1 to a few picoseconds and is followed by a second decaying component of 480 ps for nicotinamide in water and of 80–90 ps for nicotinamide in methanol and NBzOx in aqueous solution. The long decay times are assigned as the S1 lifetimes populated from the original higher-lying bright singlet, where this state is nonemissive but can be detected by transient absorption. While for NBzOx in aqueous solution and for nicotinamide in methanol, the S1 signal decays to the solvent-only level, for the aqueous solutions of nicotinamide, a small transient absorption signal remains after the 480 ps decay. This residual signal was assigned to a small population of triplet states formed during the slower S1 decay for nicotinamide in water. The experimental results were complemented by XMS-CASPT2 calculations, which reveal that in the oxidized forms, the rapid evolution of the initial π–π* state is due to a direct crossing with lower-energy dark n–π* singlet states. This coincides with the experimental observation of long-lived nonemissive states (80 to 480 ps depending on the system). On the other hand, the reduced model compound NBz has a long-lived emissive π–π* S1 state, which decays with a 510 ps time constant, similarly to the parent compound NADH. This is consistent with the XMS-CASPT2 calculations, which show that for the reduced chromophore, the dark states lie at higher energies than the bright π–π* S1 state.
Inherited platelet disorders are a heterogeneous group of rare diseases, caused by inherited defects in platelet production and/or function. Their genetic diagnosis would benefit clinical care, ...prognosis and preventative treatments. Until recently, this diagnosis has usually been performed
Sanger sequencing of a limited number of candidate genes. High-throughput sequencing is revolutionizing the genetic diagnosis of diseases, including bleeding disorders. We have designed a novel high-throughput sequencing platform to investigate the unknown molecular pathology in a cohort of 82 patients with inherited platelet disorders. Thirty-four (41.5%) patients presented with a phenotype strongly indicative of a particular type of platelet disorder. The other patients had clinical bleeding indicative of platelet dysfunction, but with no identifiable features. The high-throughput sequencing test enabled a molecular diagnosis in 70% of these patients. This sensitivity increased to 90% among patients suspected of having a defined platelet disorder. We found 57 different candidate variants in 28 genes, of which 70% had not previously been described. Following consensus guidelines, we qualified 68.4% and 26.3% of the candidate variants as being pathogenic and likely pathogenic, respectively. In addition to establishing definitive diagnoses of well-known inherited platelet disorders, high-throughput sequencing also identified rarer disorders such as sitosterolemia, filamin and actinin deficiencies, and G protein-coupled receptor defects. This included disease-causing variants in
(n=2) and
(n=3). Our study reinforces the feasibility of introducing high-throughput sequencing technology into the mainstream laboratory for the genetic diagnostic practice in inherited platelet disorders.
This research presents a virtual tour performed on the oppidum of Ulaca, one of the most relevant archaeological sites of the Iberian Peninsula during the Late Iron Age (ca. 400–50 BC). Beyond the ...clear benefits of the tool to the interpretation, dissemination, and knowledge of the mentioned archaeological site and its surroundings, the novelty of this research is the implementation of the platform in alternative scenarios and purposes. In this way, the present work verifies how the access to multi-source and spatially geolocated information in the same tool (working as a geospatial database) allows the promotion of cross-sectional investigations in which different specialists intervene. This peculiarity is also considered useful to promote tourism with an interest beyond the purely historical/archaeological side. Likewise, the possibility of storing and managing a large amount of information in different formats facilitates the investigation in the contexts of excavations and archaeological or environmental works. In this sense, the use of this kind of tool for the study of cultural landscapes is especially novel. In order to better contextualize the potential of the virtual tour presented here, an analysis about the challenges and possibilities of implementing this tool in environments such as the Ulaca oppidum is performed. The selected site stands out for: (i) being in a unique geological, environmental and ecological context, allowing us to appreciate how human beings have modified the landscape over time; (ii) presenting numerous visible archaeological remains with certain conservation problems; and (iii) not having easy access for visitors.
Reef fishes support important fisheries throughout the Caribbean, but a combination of factors in the tropics makes otolith microstructure difficult to interpret for age estimation. Therefore, ...validation of ageing methods, via application of Δ14C is a major research priority. Utilizing known-age otolith material from north Caribbean fishes, we determined that a distinct regional Δ14C chronology exists, differing from coral-based chronologies compiled for ageing validation from a wide-ranging area of the Atlantic and from an otolith-based chronology from the Gulf of Mexico. Our north Caribbean Δ14C chronology established a decline series with narrow prediction intervals that proved successful in ageing validation of three economically important reef fish species. In examining why our north Caribbean Δ14C chronology differed from some of the coral-based Δ14C data reported from the region, we determined differences among study objectives and research design impact Δ14C temporal relationships. This resulted in establishing the first of three important considerations relevant to applying Δ14C chronologies for ageing validation: 1) evaluation of the applicability of original goal/objectives and study design of potential Δ14C reference studies. Next, we determined differences between our Δ14C chronology and those from Florida and the Gulf of Mexico were explained by differences in regional patterns of oceanic upwelling, resulting in the second consideration for future validation work: 2) evaluation of the applicability of Δ14C reference data to the region/location where fish samples were obtained. Lastly, we emphasize the application of our north Caribbean Δ14C chronology should be limited to ageing validation studies of fishes from this region known to inhabit shallow water coral habitat as juveniles. Thus, we note the final consideration to strengthen findings of future age validation studies: 3) use of Δ14C analysis for age validation should be limited to species whose juvenile habitat is known to reflect the regional Δ14C reference chronology.
Summary
The introduction of Rituximab has improved the outcome and survival rates of Burkitt lymphoma (BL). However, early relapse and refractoriness are current limitations of BL treatment and new ...biological factors affecting the outcome of these patients have not been explored. This study aimed to identify the presence of genomic changes that could predict the response to new therapies in BL. Forty adolescent and adult BL patients treated with the Dose‐Intensive Chemotherapy Including Rituximab (Burkimab) protocol (Spanish Programme for the Study and Treatment of Haematological Malignancies; PETHEMA) were analysed using array‐based comparative genomic hybridization (CGH). In addition, the presence of TP53, TCF3 (E2A), ID3 and GNA13 mutations was assessed by next‐generation sequencing (NGS). Ninety‐seven per cent of the patients harboured genomic imbalances. Losses on 11q, 13q, 15q or 17p were associated with a poor response to Burkimab therapy (P = 0·038), shorter progression‐free survival (PFS; P = 0·007) and overall survival (OS; P = 0·009). The integrative analysis of array‐CGH and NGS showed that 26·3% (5/19) and 36·8% (7/19) of patients carried alterations in the TP53 and TCF3 genes, respectively. TP53 alterations were associated with shorter PFS (P = 0·011) while TCF3 alterations were associated with shorter OS (P = 0·032). Genetic studies could be used for risk stratification of BL patients treated with the Burkimab protocol.
Synthetic control of the mutual arrangement of the cyclometalated ligands (C^N) in Ir(III) dimers, Ir(C^N)2Cl2, and cationic bis-cyclometalated Ir(III) complexes, Ir(C^N)2(L^L)+ (L^L = neutral ...ligand), is described for the first time. Using 1-benzyl-4-(2,4-difluorophenyl)-1H-1,2,3-triazole (HdfptrBz) as a cyclometalating ligand, two different Ir(III) dimers, Ir(dfptrBz)2Cl2, are synthesized depending on the reaction conditions. At 80 °C, the dimer with an unusual mutual cis-C,C and cis-N,N configuration of the C^N ligands is isolated. In contrast, at higher temperature (140 °C), the geometrical isomer with the common cis-C,C and trans-N,N arrangement of the C^N ligand is obtained. In both cases, an asymmetric bridge, formed by a chloro ligand and two adjacent nitrogens of the triazole ring of one of the cyclometalated ligands, is observed. The dimers are cleaved in coordinating solvents to give the solvento complexes Ir(dfptrBz)2Cl(S) (S = DMSO or acetonitrile), which maintain the C^N arrangement of the parent dimers. Controlling the C^N ligand arrangement in the dimers allows for the preparation of the first example of geometrical isomers of a cationic bis-cyclometalated Ir(III) complex. Thus, N,N-trans-Ir(dfptrBz)2(dmbpy)+ (dmbpy = 4,4′-dimethyl-2,2′-bipyridine), with cis-C,C and trans-N,N arrangement of the C^N ligands, as well as N,N-cis-Ir(dfptrBz)2(dmbpy)+, with cis-C,C and cis-N,N C^N ligand orientation, are synthesized and characterized. Interestingly, both isomers show significantly different photophysical and electroluminescent properties, depending on the mutual arrangement of the C^N ligands. Furthermore, quantum chemical calculations give insight into the observed photophysical experimental data.
Myelodysplastic syndromes (MDS) are hematological disorders at high risk of progression to secondary acute myeloid leukemia (sAML). However, the mutational dynamics and clonal evolution underlying ...disease progression are poorly understood at present. To elucidate the mutational dynamics of pathways and genes occurring during the evolution to sAML, next generation sequencing was performed on 84 serially paired samples of MDS patients who developed sAML (discovery cohort) and 14 paired samples from MDS patients who did not progress to sAML during follow-up (control cohort). Results were validated in an independent series of 388 MDS patients (validation cohort). We used an integrative analysis to identify how mutations, alone or in combination, contribute to leukemic transformation. The study showed that MDS progression to sAML is characterized by greater genomic instability and the presence of several types of mutational dynamics, highlighting increasing (STAG2) and newly-acquired (NRAS and FLT3) mutations. Moreover, we observed cooperation between genes involved in the cohesin and Ras pathways in 15-20% of MDS patients who evolved to sAML, as well as a high proportion of newly acquired or increasing mutations in the chromatin-modifier genes in MDS patients receiving a disease-modifying therapy before their progression to sAML.
PDF
