Molecular profiling of pre- and post- 5-azacytidine myelodysplastic syndrome samples identifies predictors of response.

Treatment with the hypomethylating agent 5-azacytidine (AZA) increases survival in high-risk (HR) myelodysplastic syndrome (MDS) patients, but predicting patient response and overall survival remains challenging. To address these issues, we analyzed mutational and transcriptional profiles in CD34+ hematopoietic stem/progenitor cells (HSPCs) before and following AZA therapy in MDS patients. AZA treatment led to a greater reduction in the mutational burden in both blast and hematological responders than non-responders.

Outcomes and effect of somatic mutations after erythropoiesis stimulating agents in patients with lower-risk myelodysplastic syndromes.

Background: Erythropoiesis stimulating agents (ESAs) are the first-line therapy in patients with lower-risk myelodysplastic syndromes (LR-MDS). Some predictive factors for ESAs response have been identified. Type and number of somatic mutations have been associated with prognosis and response to therapies in MDS patients.

RNA sequencing identifies novel regulated IRE1-dependent decay targets that affect multiple myeloma survival and proliferation.

Background: IRE1 is an unfolded protein response (UPR) sensor with kinase and endonuclease activity. It plays a central role in the endoplasmic reticulum (ER) stress response through unconventional splicing of XBP1 mRNA and regulated IRE1-dependent decay (RIDD). Multiple myeloma (MM) cells are known to exhibit an elevated level of baseline ER stress due to immunoglobulin production, however RIDD activity has not been well studied in this disease.

Helpful Criteria When Implementing NGS Panels in Childhood Lymphoblastic Leukemia.

The development of Next-Generation Sequencing (NGS) has provided useful diagnostic, prognostic, and therapeutic strategies for individualized management of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) patients. Consequently, NGS is rapidly being established in clinical practice. However, the technology's complexity, bioinformatics analysis, and the different available options difficult a broad consensus between different laboratories in its daily routine introduction.

Comprehensive Custom NGS Panel Validation for the Improvement of the Stratification of B-Acute Lymphoblastic Leukemia Patients.

Background: B-acute lymphoblastic leukemia (B-ALL) is a hematological neoplasm of the stem lymphoid cell of the B lineage, characterized by the presence of genetic alterations closely related to the course of the disease. The number of alterations identified in these patients grows as studies of the disease progress, but in clinical practice, the conventional techniques frequently used are only capable of detecting the most common alterations. However, techniques, such as next-generation sequencing (NGS), are being implemented to detect a wide spectrum of new alterations that also include point mutations.

Co-occurrence of cohesin complex and Ras signaling mutations during progression from myelodysplastic syndromes to secondary acute myeloid leukemia.

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).

The combination of WGS and RNA-Seq is superior to conventional diagnostic tests in multiple myeloma: Ready for prime time?

The diagnosis and risk stratification of multiple myeloma (MM) is based on clinical and cytogenetic tests. Magnetic CD138 enrichment followed by interphase FISH (fluorescence in situ hybridisation) is the gold standard to identify prognostic translocations and copy number alterations (CNA). Although clinical implications of gene expression profiling (GEP) or panel based sequencing results are evident, those tests have not yet reached routine clinical application.

CRISPR/Cas9-generated models uncover therapeutic vulnerabilities of del(11q) CLL cells to dual BCR and PARP inhibition.

The deletion of 11q (del(11q)) invariably comprises ATM gene in chronic lymphocytic leukemia (CLL). Concomitant mutations in this gene in the remaining allele have been identified in 1/3 of CLL cases harboring del(11q), being the biallelic loss of ATM associated with adverse prognosis. Although the introduction of targeted BCR inhibition has significantly favored the outcomes of del(11q) patients, responses of patients harboring ATM functional loss through biallelic inactivation are unexplored, and the development of resistances to targeted therapies have been increasingly reported, urging the need to explore novel therapeutic approaches.

/ Fusion Gene Abrogation Decreases the Oncogenicity of Tumour Cells in a Preclinical Model of Acute Lymphoblastic Leukaemia.

Background: The t(12;21)(p13;q22), which fuses and genes, is the most common genetic abnormality in children with B-cell precursor acute lymphoblastic leukaemia. The implication of the fusion protein in leukemogenesis seems to be clear. However, its role in the maintenance of the disease continues to be controversial.

Transcriptomic analysis of patients with immune thrombocytopenia treated with eltrombopag.

In the last years, the use of thrombopoietin receptor agonists (TPO-RA), eltrombopag and romiplostim, has improved the management of immune thrombocytopenia (ITP). Moreover, eltrombopag is also active in patients with aplastic anemia and myelodysplastic syndrome. However, their mechanisms of action and signaling pathways still remain controversial.

Spanish Guidelines for the use of targeted deep sequencing in myelodysplastic syndromes and chronic myelomonocytic leukaemia.

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.

Chronic graft-versus-host disease could ameliorate the impact of adverse somatic mutations in patients with myelodysplastic syndromes and hematopoietic stem cell transplantation.

Somatic mutations in patients with myelodysplastic syndromes (MDS) undergoing allogeneic hematopoietic stem cell transplantation (HSTC) are associated with adverse outcome, but the role of chronic graft-versus-host disease (cGVHD) in this subset of patients remains unknown. We analyzed bone marrow samples from 115 patients with MDS collected prior to HSCT using next-generation sequencing. Seventy-one patients (61%) had at least one mutated gene.

Splice donor site sgRNAs enhance CRISPR/Cas9-mediated knockout efficiency.

CRISPR/Cas9 allows the generation of knockout cell lines and null zygotes by inducing site-specific double-stranded breaks. In most cases the DSB is repaired by non-homologous end joining, resulting in small nucleotide insertions or deletions that can be used to construct knockout alleles. However, these mutations do not produce the desired null result in all cases, but instead generate a similar, functionally active protein.

Molecular Diagnosis of Inherited Coagulation and Bleeding Disorders.

Diagnosis of inherited bleeding disorders (IBDs) remains challenging, especially in the case of inherited platelet disorders, due to the heterogeneity of the clinical and laboratory phenotype, the limited specificity of platelet function tests, and the large number of potential culprit genes. Unraveling the underlying molecular defect provides the definitive diagnosis of IBDs, facilitating prognosis and clinical care, which are especially important for severe clinical syndromes and those that may be associated with an increased risk of malignancy. Until recently, Sanger sequencing of candidate genes has been the only method of molecular diagnosis, but this approach is time-consuming and costly and requires phenotype-based identification of any obvious candidate gene(s).

DNA damage response-related alterations define the genetic background of patients with chronic lymphocytic leukemia and chromosomal gains.

The presence of chromosomal gains other than trisomy 12 suggesting a hyperdiploid karyotype is extremely rare in chronic lymphocytic leukemia (CLL) and is associated with a dismal prognosis. However, the genetic mechanisms and mutational background of these patients have not been fully explored. To improve our understanding of the genetic underpinnings of this subgroup of CLL, seven CLL patients with several chromosomal gains were sequenced using a next-generation sequencing (NGS)-targeted approach.

Hidden myelodysplastic syndrome (MDS): A prospective study to confirm or exclude MDS in patients with anemia of uncertain etiology.

Introduction: Diagnosis of myelodysplastic syndromes (MDSs) when anemia is the only abnormality can be complicated. The aim of our study was to investigate the primary causes of anemia and/or macrocytosis of uncertain etiology.

Methods: We conducted a multicenter, prospective study over 4 months in three hematology laboratories.

Introducing high-throughput sequencing into mainstream genetic diagnosis practice in inherited platelet disorders.

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.

Prospective randomized trial of 5 days azacitidine versus supportive care in patients with lower-risk myelodysplastic syndromes without 5q deletion and transfusion-dependent anemia.

In this prospective trial, the efficacy of azacitidine in lower-risk myelodysplastic syndromes (LR-SMD) lacking del(5q) was compared to best supportive care (BSC) at 1:1. The primary endpoint was the achievement of erythroid hematologic improvement (HI-E) after nine cycles. Thirty-six patients received at least ≥1 cycle.

FISHing in the dark: How the combination of FISH and conventional karyotyping improves the diagnostic yield in CpG-stimulated chronic lymphocytic leukemia.

Despite significant advances in molecular genetic approaches, fluorescence in situ hybridization (FISH) remains the gold standard for the diagnostic evaluation of genomic aberrations in patients with chronic lymphocytic leukemia (CLL). Efforts to improve the diagnostic utility of molecular cytogenetic testing have led to the expansion of the traditional 4-probe CLL FISH panel. Not only do these efforts increase the cost of testing, they remain hindered by the inherent limitations of FISH studies - namely the inability to evaluate genomic changes outside of the targeted loci.

The presence of genomic imbalances is associated with poor outcome in patients with burkitt lymphoma treated with dose-intensive chemotherapy including rituximab.

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.