Search

Esta depositada la versión del artículo enviada a la revista, A series of blue and blue-green emitters based on neutral bis- and tris-cyclometalated Ir(III) complexes with 1-benzyl-4-(2,6-difluorophenyl)-1H-1,2,3-triazole (dfptrBn) as cyclometalating ligand is reported. The bis-cyclometalated complexes of the type [Ir(dfptrBn)(2)((LX)-X-boolean AND)] with different ancillary ligands, (LX)-X-boolean AND = picolinate (pic) (2) or 2-(5-(perfluorophenyl)-2H-1,2,4-triazol-3-yl)pyridine (pytrF(5)) (3), are described and their photophysical properties compared with the analogous complexes containing the archetypal 2-(2,4-difluorophenyl)-pyridinato (dfppy) as cyclometaled ligand ((CN)-N-boolean AND). Complex 2 exhibits a marked solvatochromic behavior, from 475 nm in toluene to 534 nm in formamide, due to the strong MLCT character of its emissive excited state. Complex 3 displays a true-blue emission, narrower in the visible part than FIrpic. In addition, the homoleptic complex [Ir(dfprBn)(3)] (4) and the heteroleptic compounds with mixed arylpyridine/aryltriazole ligands, [Ir(dfptrBn)(2)((CN)-N-boolean AND)] ((CN)-N-boolean AND = 2-phenylpyridinato (ppy) (5) or dfppy (6)), have been synthesized and fully characterized. The facial (fac) complex fac-4 is emissive at 77 K showing a deep-blue emission, but it is not luminescent in solution at room temperature similarly to their phenylpyrazole counterparts. However, the fac isomers, fac-5 and fac-6, are highly emissive in solution and thin films, reaching emission quantum yields of 76%, with emission colors in the blue to blue-green region. The photophysical properties for all complexes have been rationalized by means of quantum-chemical calculations. In addition, we constructed electroluminescent devices, organic light-emitting diodes (OLEDs) by sublimation of fac-6, and by solution processed polymer-based devices (PLEDs) using complexes fac-5 or fac-6 as dopants., Fundacion Seneca, Instituto de Salud Carlos III, Ministry of Science and Technology, Taiwan, Deutscher Akademischer Austausch Dienst (DAAD), Fonds de la Recherche Scientifique - FNRS, Ministerio de Ciencia e Innovación (España), Depto. de Física de Materiales, Fac. de Ciencias Físicas, TRUE, pub

Diagnosis and treatment of hematological cancers is usually provided in many healthcare facilities including large but also middle size centers.1 Providing cancer care in local institutions might be advantageous for patients and caregivers in terms of financial burden and quality of life. However, it might carry potential risks derived of the limited experience of smaller centers and differences in accessibility to complex therapies including allogeneic hematopoietic cell transplantation (allo-HCT) and chimeric antigen receptor (CAR) T-cells. These risks might be especially relevant in infrequent cancers as adult acute lymphoblastic leukemia (ALL).

13q14 deletion is the most recurrent chromosomal aberration reported in B-CLL, having a favorable prognostic significance when occurring as the sole cytogenetic alteration. However, its clinical outcome is also related to the deletion size and number of cells with the del(13)(q14) deletion. In 10% of cases, 13q14 deletion arises following a translocation event with multiple partner chromosomes, whose oncogenic impact has not been investigated so far due to the assumption of a possible role as a passenger mutation. Here, we describe a t(4;13)(q21;q14) translocation occurring in a B-CLL case from the diagnosis to spontaneous regression. FISH and SNP-array analyses revealed a heterozygous deletion at 4q21, leading to the loss of the Rho GTPase Activating Protein 24 (ARHGAP24) tumor suppressor gene, down-regulated in the patient RNA, in addition to the homozygous deletion at 13q14 involving DLEU2/miR15a/miR16–1 genes. Interestingly, targeted Next Generation Sequencing analysis of 54 genes related to B-CLL indicated no additional somatic mutation in the patient, underlining the relevance of this t(4;13)(q21;q14) aberration in the leukemogenic process. In all tested RNA samples, RT-qPCR experiments assessed the downregulation of the PCNA, MKI67, and TOP2A proliferation factor genes, and the BCL2 anti-apoptotic gene as well as the up-regulation of TP53 and CDKN1A tumor suppressors, indicating a low proliferation potential of the cells harboring the aberration. In addition, RNA-seq analyses identified four chimeric transcripts (ATG4B::PTMA, OAZ1::PTMA, ZFP36::PTMA, and PIM3::BRD1), two of which (ATG4B::PTMA and ZFP36::PTMA) failed to be detected at the remission, suggesting a possible transcriptional remodeling during the disease course. Overall, our results indicate a favorable prognostic impact of the described chromosomal aberration, as it arises a permissive molecular landscape to the spontaneous B-CLL regression in the patient, highlighting ARHGAP24 as a potentially re

[Background]: The microbiota is increasingly recognized as a significant factor in the pathophysiology of many diseases, including cardiometabolic diseases, with lifestyles probably exerting the greatest influence on the composition of the human microbiome. The main objectives of the study are to analyze the association of lifestyles (diet, physical activity, tobacco, and alcohol) with the gut and oral microbiota, arterial aging, and cognitive function in subjects without cardiovascular disease in the Iberian Peninsula. In addition, the study will examine the mediating role of the microbiome in mediating the association between lifestyles and arterial aging as well as cognitive function. [Methods and analysis]: MIVAS III is a multicenter cross-sectional study that will take place in the Iberian Peninsula. One thousand subjects aged between 45 and 74 years without cardiovascular disease will be selected. The main variables are demographic information, anthropometric measurements, and habits (tobacco and alcohol). Dietary patterns will be assessed using a frequency consumption questionnaire (FFQ) and the Mediterranean diet adherence questionnaire. Physical activity levels will be evaluated using the International Physical Activity Questionnaire (IPAQ), Marshall Questionnaire, and an Accelerometer (Actigraph). Body composition will be measured using the Inbody 230 impedance meter. Arterial aging will be assessed through various means, including measuring medium intimate carotid thickness using the Sonosite Micromax, conducting analysis with pulse wave velocity (PWA), and measuring pulse wave velocity (cf-PWV) using the Sphygmocor System. Additional cardiovascular indicators such as Cardio Ankle Vascular Index (CAVI), ba-PWV, and ankle-brachial index (Vasera VS-2000®) will also be examined. The study will analyze the intestinal microbiota using the OMNIgene GUT kit (OMR−200) and profile the microbiome through massive sequencing of the 16S rRNA gene. Linear discriminant

Trisomy 8 (+8) is the most frequent trisomy in myelodysplastic syndromes (MDS) and is associated with clinical heterogeneity and intermediate cytogenetic risk when found in isolation. The presence of gene mutations in this group of patients and the prognostic significance has not been extensively analyzed. Targeted deep sequencing was performed in a cohort of 79 MDS patients showing isolated +8. The most frequently mutated genes were: TET2 (38%), STAG2 (34.2%), SRSF2 (29.1%) and RUNX1 (26.6%). The mutational profile identified a high-risk subgroup with mutations in STAG2, SRSF2 and/or RUNX1, resulting in shorter time to acute myeloid leukemia progression (14 months while not reached in patients without these mutations, p < 0.0001) and shorter overall survival (23.7 vs. 46.3 months, p = 0.001). Multivariate analyses revealed the presence of mutations in these genes as an independent prognostic factor in MDS showing +8 isolated (HR: 3.1; p < 0.01). Moreover, 39.5% and 15.4% of patients classified as low/intermediate risk by the IPSS-R and IPSS-M, respectively, were re-stratified as a high-risk subgroup based on the mutational status of STAG2, SRSF2 and RUNX1. Results were validated in an external cohort (n = 2494). In summary, this study validates the prognosis significance of somatic mutations shown in IPSS-M and adds STAG2 as an important mutated gene to consider in this specific subgroup of patients. The mutational profile in isolated +8 MDS patients could, therefore, offer new insights for the correct management of patients with a higher risk of leukemic transformation.

The promoter‐less CRISPR‐Trap system efficiently the specific integration of the gene trap donor in BCR/ABL but is not a robust reporter system to select corrected cells. As a first approach, we generate a dsDNA donor by PCR amplification containing a SAT2A‐ Venus interfering cassette (Figure S1A). The K562 cell line was divided into three experimental groups according to the conditions for their subsequent electroporation: a) with the donor dsDNA (donor), b) with the donor DNA and Cas9 nuclease without sgRNA (Cas9 + donor) and c) with the donor DNA, Cas9 nuclease and the specific BCR/ABL sgRNA (CRISPR/Cas9 + donor). Twenty‐four hours after electroporation, no fluorescent cells (Venus + cells) were observed in any of the three groups (Figure S1B). The result of the site‐unspecific PCR (In 5´F/Venus R, Table 1) which amplify a region contained entirely in the donor HDR dsDNA, showed a band of the expected size in the Cas9 + donor and CRISPR/Cas9 + donor groups, but not in the donor group (Figure S1C), implying that all donor HDR dsDNA not integrated in the genome was fully degraded. In contrast, the sitespecific PCR (Out 5´F/Venus R, Table 1) corroborated the correct insertion of the donor HDR dsDNA at the BCR/ABL target sequence only in the CRISPR/Cas9 + donor group, with no sitespecific integration detected in any of the controls (Figure S1C). These results were corroborated by Sanger sequencing of the PCR products, supporting the HDR‐mediated insertion of the donor dsDNA into the CRISPR/Cas9 + donor group.

[Background]: Rare inherited thrombocytopenias are caused by alterations in genes involved in megakaryopoiesis, thrombopoiesis and/or platelet release. Diagnosis is challenging due to poor specificity of platelet laboratory assays, large numbers of culprit genes, and difficult assessment of the pathogenicity of novel variants. [Objectives]: To characterize the clinical and laboratory phenotype, and identifying the underlying molecular alteration, in a pedigree with thrombocytopenia of uncertain etiology. [Patients/Methods]: Index case was enrolled in our Spanish multicentric project of inherited platelet disorders due to lifelong thrombocytopenia and bleeding. Bleeding score was recorded by ISTH‐BAT. Laboratory phenotyping consisted of blood cells count, blood film, platelet aggregation and flow cytometric analysis. Genotyping was made by whole‐exome sequencing (WES). Cytoskeleton proteins were analyzed in resting/spreading platelets by immunofluorescence and immunoblotting. [Results]: Five family members displayed lifelong mild thrombocytopenia with a high number of enlarged platelets in blood film, and mild bleeding tendency. Patient's platelets showed normal aggregation and granule secretion response to several agonists. WES revealed a novel nonsense variant (c.322C>T; p.Gln108*) in TPM4 (NM_003290.3), the gene encoding for tropomyosin‐4 (TPM4). This variant led to impairment of platelet spreading capacity after stimulation with TRAP‐6 and CRP, delocalization of TPM4 in activated platelets, and significantly reduced TPM4 levels in platelet lysates. Moreover, the index case displayed up‐regulation of TPM2 and TPM3 mRNA levels. [Conclusions]: This study identifies a novel TPM4 nonsense variant segregating with macrothrombocytopenia and impaired platelet cytoskeletal remodeling and spreading. These findings support the relevant role of TPM4 in thrombopoiesis and further expand our knowledge of TPM4‐related thrombocytopenia.

Context: B-cell precursor acute lymphoblastic leukemia (BCP ALL) is a genetically heterogeneous neoplasm with >20 biologic subtypes. Each subtype shows specific genetic traits that determine relapse risk and patients' survival. Objectives: To establish the genetic subtype (primary alteration) of adult BCP ALL patients enrolled in the PETHEMA LAL19 trial (NCT 04179929) and to correlate them with measurable residual disease (MRD) level and survival. Patients and Methods: In the LAL19 trial (NCT04179929), Ph-negative patients (18–65 y) with MRD≥0.01% at day+35 or high-risk genetics receive alloHSCT and MRD<0.01% patients with standard-risk genetics receive maintenance chemotherapy. The genetic analyses are centralized: FISH and NGS DNA panel (Hospital de Salamanca), RNAseq panel (Hospital 12 de Octubre), FISH panel (Hospital La Fe), and SNP array (Josep Carreras Institute/ICO-Hospital Germans Trias i Pujol). MRD determinations are centrally done by next-generation flow cytometry in the Cytometry Service, NUCLEUS, University of Salamanca. Results: The genetic subtype was identified in 54% (82/152) of patients. The most recurrent subtypes were KMT2Ar (11%), Ph-like (mostly CRLF2::IGH, 11%), low-hypodiploid (7%), PAX5 P80R (7%), high-hyperdiploid (6%), and t(1;19)/TCF3::PBX1 (6%). In addition, t(12;21)/ETV6::RUNX1, ZNF384r, and iAMP21 subtypes (1.5% each) and MEF2Dr, MYCr, IDH1 R132 subtypes (<1% each) were found. Regarding secondary alterations, NRAS (15%), TP53 (13%), PAX5 (13%), and KRAS (10%) mutations were the most frequently observed. Twelve patients were refractory (mainly low-hypodiploid, Ph-like, MYCr, and B-other/unclassified patients). Statistically significant differences were observed for day+35 MRD levels between genetic subtypes. Ph-like, low-hypodiploid, and KMT2Ar showed lower frequencies of MRD<0.01% (17%, 33%, and 57%, respectively) than patients with PAX5P80R (100%), t(1;19)/TCF3::PBX1 (83%), and high-hyperdiploid (75%) (P=0.006). Despite the short med

Interstitial 14q32 deletions involving IGH gene are infrequent events in chronic lymphocytic leukemia (CLL), affecting less than 5% of patients. To date, little is known about their clinical impact and molecular underpinnings, and its mutational landscape is currently unknown. In this work, a total of 871 CLLs were tested for the IGH break-apart probe, and 54 (6.2%) had a 300 kb deletion of 3′IGH (del-3′IGH CLLs), which contributed to a shorter time to first treatment (TFT). The mutational analysis by next-generation sequencing of 317 untreated CLLs (54 del-3′IGH and 263 as the control group) showed high mutational frequencies of NOTCH1 (30%), ATM (20%), genes involved in the RAS signaling pathway (BRAF, KRAS, NRAS, and MAP2K1) (15%), and TRAF3 (13%) within del-3′IGH CLLs. Notably, the incidence of TRAF3 mutations was significantly higher in del-3′IGH CLLs than in the control group (p < .001). Copy number analysis also revealed that TRAF3 loss was highly enriched in CLLs with 14q deletion (p < .001), indicating a complete biallelic inactivation of this gene through deletion and mutation. Interestingly, the presence of mutations in the aforementioned genes negatively refined the prognosis of del-3′IGH CLLs in terms of overall survival (NOTCH1, ATM, and RAS signaling pathway genes) and TFT (TRAF3). Furthermore, TRAF3 biallelic inactivation constituted an independent risk factor for TFT in the entire CLL cohort. Altogether, our work demonstrates the distinct genetic landscape of del-3′IGH CLL with multiple molecular pathways affected, characterized by a TRAF3 biallelic inactivation that contributes to a marked poor outcome in this subgroup of patients.

Inherited antithrombin deficiency, the most severe form of thrombophilia, is predominantly caused by variants in SERPINC1. Few causal structural variants have been described, usually detected by multiplex ligation-dependent probe amplification or cytogenetic arrays, which only define the gain or loss and the approximate size and location. This study has done a complete dissection of the structural variants affecting SERPINC1 of 39 unrelated patients with antithrombin deficiency using multiplex ligation-dependent probe amplification, comparative genome hybridization array, long-range PCR, and whole genome nanopore sequencing. Structural variants, in all cases only affecting one allele, were deleterious and caused a severe type I deficiency. Most defects were deletions affecting exons of SERPINC1 (82.1%), but the whole cohort was heterogeneous, as tandem duplications, deletion of introns, or retrotransposon insertions were also detected. Their size was also variable, ranging from 193 bp to 8 Mb, and in 54% of the cases involved neighboring genes. All but two structural variants had repetitive elements and/or microhomologies in their breakpoints, suggesting a common mechanism of formation. This study also suggested regions recurrently involved in structural variants causing antithrombin deficiency and found three structural variants with a founder effect: the insertion of a retrotransposon, duplication of exon 6, and a 20-gene deletion. Finally, nanopore sequencing was determined to be the most appropriate method to identify and characterize all structural variants at nucleotide level, independently of their size or type.

Chronic myeloid leukaemia (CML) is a haematological neoplasm driven by the BCR/ABL fusion oncogene. The monogenic aspect of the disease and the feasibility of ex vivo therapies in haematological disorders make CML an excellent candidate for gene therapy strategies. The ability to abolish any coding sequence by CRISPR-Cas9 nucleases offers a powerful therapeutic opportunity to CML patients. However, a definitive cure can only be achieved when only CRISPR-edited cells are selected. A gene-trapping approach combined with CRISPR technology would be an ideal approach to ensure this. Here, we developed a CRISPR-Trap strategy that efficiently inserts a donor gene trap (SA-CMV-Venus) cassette into the BCR/ABL-specific fusion point in the CML K562 human cell line. The trapping cassette interrupts the oncogene coding sequence and expresses a reporter gene that enables the selection of edited cells. Quantitative mRNA expression analyses showed significantly higher level of expression of the BCR/Venus allele coupled with a drastically lower level of BCR/ABL expression in Venus+ cell fractions. Functional in vitro experiments showed cell proliferation arrest and apoptosis in selected Venus+ cells. Finally, xenograft experiments with the selected Venus+ cells showed a large reduction in tumour growth, thereby demonstrating a therapeutic benefit in vivo. This study represents proof of concept for the therapeutic potential of a CRISPR-Trap system as a novel strategy for gene elimination in haematological neoplasms.

Los Síndromes Mielodisplásicos (SMD) son un grupo heterogéneo de enfermedades clonales que afectan a las células madre hematopoyéticas los cuales presentan un curso clínico muy variable, por lo que su diagnóstico, pronóstico y toma de decisiones terapéuticas son todo un reto. En los últimos años, el uso de técnicas de alto rendimiento, como la secuenciación masiva, ha revolucionado la investigación genómica, aumentando nuestro conocimiento sobre las alteraciones moleculares subyacentes a la patogénesis del SMD y mejorando nuestra compresión acerca de los mecanismos implicados en la progresión de la enfermedad. Sin embargo, la ausencia de una combinación eficiente entre genética e información clínica ha limitado la interpretación de los datos de secuenciación y, por consiguiente, la implantación de la secuenciación masiva en la práctica clínica habitual. Por tanto, el objetivo principal de esta tesis es caracterizar los mecanismos moleculares y la evolución clonal que están implicados en la patogénesis del SMD y en la progresión de los SMD a Leucemia Aguda Mieloblástica secundaria (LAMs), tan bien como utilizar los datos moleculares para alcanzar un diagnóstico integrado más fiable y una estratificación pronóstica más precisa. Para ello, la tesis se ha centrado en 3 estudios: - Caracterización del perfil mutacional de los pacientes con SMD que evolucionan a LAMs. - Estudio de la relevancia de las mutaciones de las cohesinas en pacientes con SMD. - Estudio de la significancia del perfil mutacional de la vía de señalización de Ras en la patogénesis del SMD.

[Introduction]: Intestinal microbiota is arising as a new element in the physiopathology of cardiovascular diseases. A healthy microbiota includes a balanced representation of bacteria with health promotion functions (symbiotes). The aim of this study is to analyse the relationship between intestinal microbiota composition and arterial stiffness. [Methods and analysis]: An observational case—control study will be developed. Cases will be defined by the presence of at least one of the following: carotid-femoral pulse wave velocity (cf-PWV), Cardio-Ankle Vascular Index (CAVI), brachial ankle pulse wave velocity (ba or ba-PWV) above the 90th percentile, for age and sex, of the reference population. Controls will be selected from the same population as cases. The study will be developed in Primary Healthcare Centres. We will select 500 subjects (250 cases and 250 controls), between 45 and 74 years of age. Cases will be selected from a database that combines data from EVA study (Spain) and Guimarães/Vizela study (Portugal). Measurements: cf-PWV will be measured using the SphygmoCor system, CAVI, ba-PWV and Ankle-Brachial Index will be determined using VaSera device. Gut microbiome composition in faecal samples will be determined by 16S ribosomal RNA sequencing. Lifestyle will be assessed by food frequency questionnaire, adherence to the Mediterranean diet and IPAQ (International Physical Activity Questionnaire). Body composition will be evaluated by bioimpedance. [Ethics and dissemination]: The study has been approved by ‘Committee of ethics of research with medicines of the health area of Salamanca’ on 14 December 2018 (cod. 2018-11-136) and the ’Ethics committee for health of Guimaraes’ (Portugal) on 15 October 2019 (ref: 67/2019).All study participants will sign an informed consent form agreeing to participate in the study, in compliance with the Declaration of Helsinki and the WHO standards for observational studies. The results of this study will allow a better desc

The knowledge of chronic lymphocytic leukemia (CLL) has progressively deepened during the last forty years. Research activities and clinical studies have been remarkably fruitful in novel findings elucidating multiple aspects of the pathogenesis of the disease, improving CLL diagnosis, prognosis and treatment. Whereas the diagnostic criteria for CLL have not substantially changed over time, prognostication has experienced an expansion with the identification of new biological and genetic biomarkers. Thanks to next-generation sequencing (NGS), an unprecedented number of gene mutations were identified with potential prognostic and predictive value in the 2010s, although significant work on their validation is still required before they can be used in a routine clinical setting. In terms of treatment, there has been an impressive explosion of new approaches based on targeted therapies for CLL patients during the last decade. In this current chemotherapy-free era, BCR and BCL2 inhibitors have changed the management of CLL patients and clearly improved their prognosis and quality of life. In this review, we provide an overview of these novel advances, as well as point out questions that should be further addressed to continue improving the outcomes of patients.

Recent genomic studies have identified a wide range of novel genetic alterations that have substantially increased our knowledge of the biology of B- and T-progenitor acute lymphoblastic leukaemia (B-ALL, T-ALL) and defined new subtypes with prognostic and therapeutic relevance.1-4 Thanks to the use of transcriptome sequencing approaches, new cryptic fusion transcripts have been described, such as the ATP5L-KMT2A gene fusion, described by Gestrich et al. in a 14-month-old patient with aggressive B-ALL.5 ATP5L or ATP5MG (ATP Synthase Membrane Subunit G) catalyzes ATP synthesis during oxidative phosphorylation.6 This protein has recently been reported to interact with a SARS-CoV-2 protein.7 The histone lysine [K]-methyl transferase 2A (KMT2A) gene is a transcriptional coactivator that plays an essential role in regulating gene expression during early development and haematopoiesis. It is frequently rearranged to over 135 translocation partner genes in acute leukaemias.8 ATP5L is a novel KMT2A fusion partner not detectable by fluorescent in situ hybridization (FISH) or karyotype, due to the closeness of the two genes on chromosome 11q23. The Cleveland Medical Centre team found a reciprocal out-of-frame ATP5L-KMT2A rearrangement that juxtaposes the ATP5L exon 1 to the KMT2A exon 2, with the insertion of an extra nucleotide (G) at the fusion site.5 We sequenced leukaemic cells from eight adult ALL patients (two T-ALL, five B-ALL Philadelphia negative (Ph−) and one B-ALL Ph+; Table I) by a 199 gene RNA-sequencing panel (RNA-seq; Pan-Heme FusionPlex, ArcherDx Inc., Boulder, CO, USA).

Chronic myeloid leukemia (CML) is a hematopoietic malignancy produced by a unique oncogenic event involving the constitutively active tyrosine-kinase (TK) BCR/ABL1. TK inhibitors (TKI) changed its prognosis and natural history. Unfortunately, ABL1 remains unaffected by TKIs. Leukemic stem cells (LSCs) remain, and resistant mutations arise during treatment. To address this problem, we have designed a therapeutic CRISPR-Cas9 deletion system targeting BCR/ABL1. The system was efficiently electroporated to cell lines, LSCs from a CML murine model, and LSCs from CML patients at diagnosis, generating a specific ABL1 null mutation at high efficiency and allowing the edited leukemic cells to be detected and tracked. The CRISPR-Cas9 deletion system triggered cell proliferation arrest and apoptosis in murine and human CML cell lines. Patient and murine-derived xenografts with CRISPR-edited LSCs in NOD SCID gamma niches revealed that normal multipotency and repopulation ability of CRISPR edited LSCs were fully restored. Normal hematopoiesis was restored, avoiding myeloid bias. To the best of our knowledge, we show for the first time how a CRISPR-Cas9 deletion system efficiently interrupts BCR/ABL1 oncogene in primary LSCs to bestow a therapeutic benefit. This study is a proof of concept for genome editing in all those diseases, like CML, sustained by a single oncogenic event, opening up new therapeutic opportunities.

The need for allogeneic hematopoietic stem cell transplantation (allo-HSCT) in adults with Philadelphia chromosome–negative (Ph−) acute lymphoblastic leukemia (ALL) with high-risk (HR) features and adequate measurable residual disease (MRD) clearance remains unclear. The aim of the ALL-HR-11 trial was to evaluate the outcomes of HR Ph− adult ALL patients following chemotherapy or allo-HSCT administered based on end-induction and consolidation MRD levels. Patients aged 15 to 60 years with HR-ALL in complete response (CR) and MRD levels (centrally assessed by 8-color flow cytometry) <0.1% after induction and <0.01% after early consolidation were assigned to receive delayed consolidation and maintenance therapy up to 2 years in CR. The remaining patients were allocated to allo-HSCT. CR was attained in 315/348 patients (91%), with MRD <0.1% after induction in 220/289 patients (76%). By intention-to-treat, 218 patients were assigned to chemotherapy and 106 to allo-HSCT. The 5-year (±95% confidence interval) cumulative incidence of relapse (CIR), overall survival (OS), and event-free survival probabilities for the whole series were 43% ± 7%, 49% ± 7%, and 40% ± 6%, respectively, with CIR and OS rates of 45% ± 8% and 59% ± 9% for patients assigned to chemotherapy and of 40% ± 12% and 38% ± 11% for those assigned to allo-HSCT, respectively. Our results show that avoiding allo-HSCT does not hamper the outcomes of HR Ph− adult ALL patients up to 60 years with adequate MRD response after induction and consolidation. Better postremission alternative therapies are especially needed for patients with poor MRD clearance.

SF3B1 is a highly mutated gene in myelodysplastic syndrome (MDS) patients, related to a specific subtype and parameters of good prognosis in MDS without excess blasts. More than 40% of MDS patients carry at least two myeloid-related gene mutations but little is known about the impact of concurrent mutations on the outcome of MDS patients. In applying next-generation sequencing (NGS) with a 117 myeloid gene custom panel, we analyzed the co-occurrence of SF3B1 with other mutations to reveal their clinical, biological, and prognostic implications in very low/low- and intermediate-risk MDS patients. Mutations in addition to those of SF3B1 were present in 80.4% of patients (median of 2 additional mutations/patient, range 0–5). The most frequently mutated genes were as follows: TET2 (39.2%), DNMT3A (25.5%), SRSF2 (10.8%), CDH23 (5.9%), and ASXL1, CUX1, and KMT2D (4.9% each). The presence of at least two mutations concomitant with that of SF3B1 had an adverse impact on survival compared with those with the SF3B1 mutation and fewer than two additional mutations (median of 54 vs. 87 months, respectively: p = 0.007). The co-occurrence of SF3B1 mutations with specific genes is also linked to a dismal prognosis: SRSF2 mutations were associated with shorter overall survival (OS) than SRSF2wt (median, 27 vs. 75 months, respectively; p = 0.001), concomitant IDH2 mutations (median OS, 11 [mut] vs. 75 [wt] months; p = 0.001), BCOR mutations (median OS, 11 [mut] vs. 71 [wt] months; p = 0.036), and NUP98 and STAG2 mutations (median OS, 27 and 11 vs. 71 months, respectively; p = 0.008 and p = 0.002). Mutations in CHIP genes (TET2, DNMT3A) did not significantly affect the clinical features or outcome. Our results suggest that a more comprehensive NGS study in low-risk MDS SF3B1mut patients is essential for a better prognostic evaluation.

NGS Establishment in Multidisciplinary Healthcare (NEMHESYS) is an Erasmus+ programme with the purpose of providing qualified staff with the essential technical and bioinformatic knowledge and skills on next-generation sequencing (NGS) to be able to carry out NGS studies and perform some of the most common types of analyses. The clinical application of NGS has become easier with advancements in technologies.1 However, the investment needed to bring NGS into medical practice remains significant, with the scale of knowledge required being unprecedented at most hospitals. In addition, these novel technologies bring new challenges in translating NGS to clinical practice, at both technical and regulatory level, in terms of data management, interpretation of the results, and genetic counseling.2,3 All these aspects justify the consideration of what will be the precise role of NGS in diagnosis, risk assessment, response prediction, and treatment monitoring, today and tomorrow. Thus, to evaluate the implementation of NGS in European healthcare/research centers, a mapping survey was carried out, based on previous NGS mapping studies.

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.

During platelet activation, arachidonic acid (AA) is released from membrane phospholipids and metabolized to thromboxane A2 (TXA2) through the actions of cyclooxygenase-1 (COX-1) and TXA2 synthase. Note, TXA2 binds to the platelet TXA2 receptor, causing shape change, secretion and platelet aggregation.1 Also, COX-1 (599aa; 70 kDa) has cyclooxygenase and peroxidase activities and it is functionally active as a homodimer, with each COX-1 monomer consisting of four highly conserved domains: an N-terminal signal peptide, a dimerization domain, a membrane-binding domain (MBD) and a large C-terminal catalytic domain2 (Figure 1A). Irreversible COX-1 inhibition by aspirin is a widely established anti-platelet therapy in cardiovascular disease.

The potential prognostic value of conventional karyotyping in adult T-cell acute lymphoblastic leukemia (T-ALL) remains an open question. We hypothesized that a modified cytogenetic classification, based on the number and type of cytogenetic abnormalities, would allow the identification of high-risk adult T-ALL patients. Complex karyotype defined by the presence of ≥3 cytogenetic alterations identified T-ALL patients with poor prognosis in this study. Karyotypes with ≥3 abnormalities accounted for 16 % (22/139) of all evaluable karyotypes, corresponding to the largest poor prognosis cytogenetic subgroup of T-ALL identified so far. Patients carrying karyotypes with ≥3 cytogenetic alterations showed a significantly inferior response to therapy, and a poor outcome in terms of event-free survival (EFS), overall survival (OS) and cumulative incidence of relapse (CIR), independently of other baseline characteristics and the end-induction minimal residual disease (MRD) level. Additional molecular analyses of patients carrying ≥3 cytogenetic alterations showed a unique molecular profile that could contribute to understand the underlying molecular mechanisms of resistance and to evaluate novel targeted therapies (e.g. IL7R directed) with potential impact on outcome of adult T-ALL patients.