Publications by authors named "José Cervera"

131 Publications

A Single-Run Next-Generation Sequencing (NGS) Assay for the Simultaneous Detection of Both Gene Mutations and Large Chromosomal Abnormalities in Patients with Myelodysplastic Syndromes (MDS) and Related Myeloid Neoplasms.

Cancers (Basel) 2021 Apr 18;13(8). Epub 2021 Apr 18.

Hematology Research Group, Instituto de Investigación Sanitaria La Fe, 46026 Valencia, Spain.

Myelodysplastic syndromes (MDS) and myelodysplastic/myeloproliferative neoplasms are clonal disorders that share most of their cytogenetic and molecular alterations. Despite the increased knowledge of the prognostic importance of genetics in these malignancies, next-generation sequencing (NGS) has not been incorporated into clinical practice in a validated manner, and the conventional karyotype remains mandatory in the evaluation of suspected cases. However, non-informative cytogenetics might lead to an inadequate estimation of the prognostic risk. Here, we present a novel targeted NGS-based assay for the simultaneous detection of all the clinically relevant genetic alterations associated with these disorders. We validated this platform in a large cohort of patients by performing a one-to-one comparison with the lesions from karyotype and single-nucleotide polymorphism (SNP) arrays. Our strategy demonstrated an approximately 97% concordance with standard clinical assays, showing sensitivity at least equivalent to that of SNP arrays and higher than that of conventional cytogenetics. In addition, this NGS assay was able to identify both copy-neutral loss of heterozygosity events distributed genome-wide and copy number alterations, as well as somatic mutations within significant driver genes. In summary, we show a novel NGS platform that represents a significant improvement to current strategies in defining diagnosis and risk stratification of patients with MDS and myeloid-related disorders.
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http://dx.doi.org/10.3390/cancers13081947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8072643PMC
April 2021

Case Report: Partial Uniparental Disomy Unmasks a Novel Recessive Mutation in the Gene in a Patient With a Severe Phenotype of Chédiak-Higashi Syndrome.

Front Immunol 2021 31;12:625591. Epub 2021 Mar 31.

Department of Hematology, Hospital Universitario y Politécnico La Fe, Barcelona, Spain.

Chédiak-Higashi syndrome (CHS) is a rare autosomal recessive (AR) immune disorder that has usually been associated to missense, nonsense or indels mutations in the gene. In this study, we describe for the first time the case of a CHS patient carrying a homozygous mutation in the gene inherited as a result of a partial uniparental isodisomy (UPiD) of maternal origin. Sanger sequencing of the cDNA and single nucleotide polymorphism (SNP)-arrays were performed to identify the causative mutation and to explain the molecular mechanism of inheritance, respectively. Partial-UPiD leads to a copy neutral loss of heterozygosity (CN-LOH) of the telomeric region of chromosome 1 (1q41q44), unmasking the potential effect of the mutation detected. The mutation (c.8380dupT) is an insertion located in exon 32 of the gene resulting in a premature stop codon and leading to the loss of all the conserved domains at the C-terminal of the LYST protein. This would account for the severe phenotype observed. We also reviewed the only two previously reported cases of CHS as a result of a uniparental disomy. In this study, we show that the combination of different strategies, including the use of SNP-arrays, is pivotal to fine-tune the diagnosis of rare AR disorders, such as CHS. Moreover, this case highlights the relevance of uniparental disomy as a potential mechanism of CHS expression in non-consanguineous families.
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http://dx.doi.org/10.3389/fimmu.2021.625591DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8044466PMC
March 2021

Influence of polymorphisms in anthracyclines metabolism genes in the standard induction chemotherapy of acute myeloid leukemia.

Pharmacogenet Genomics 2021 Mar 5. Epub 2021 Mar 5.

Grupo de Farmacogenética, Instituto Investigación Sanitaria La Fe and Área del Medicamento Servicio de Farmacia, Área del Medicamento Servicio de Hematología y Hemoterapia, Hospital Universitari i Politècnic La Fe. Av. Fernando Abril Martorell, Valencia CIBERONC, Instituto Carlos III, Madrid Departamento Farmacología, Facultad de Medicina, Universidad de Valencia, Av. Blasco Ibáñez 15 Unidad de Farmacología Clínica, Área del Medicamento, Hospital Universitari I Politècnic La Fe, Av. Fernando Abril Martorell, Valencia, Spain.

Objectives: Genetic variability in anthracycline metabolism could modify the response and safety of acute myeloid leukemia (AML) induction.

Methods: Polymorphisms in genes that encodes enzymes of anthracyclines metabolic pathway (CBR3: rs1056892, rs8133052, NQO1: rs1800566, NQO2: rs1143684, NOS3: rs1799983, rs2070744) were evaluated in 225 adult de novo AML patients.

Results: The variant CBR3 rs8133052 was associated with lower hepatotoxicity (P = 0.028). Wild-type genotype of NQO2 rs1143684 was related to higher complete remission (P = 0.014), and the variant allele with greater gastrointestinal toxicity (P = 0.024). However, the variant genotype of NQO1 rs1800566 was associated with mucositis (P = 0.018), but heterozygous genotype showed less gastrointestinal toxicity (P = 0.028) and thrombocytopenia (P = 0.009). Protective effects against nephrotoxicity and thrombocytopenia were reported with variant NOS3 rs1799983 (P = 0.006, P = 0.014), whereas carriers of NOS3 rs2070744 showed higher hepatotoxicity and thrombocytopenia (P = 0.017, P = 0.013).

Conclusions: This study supports the influence of genetic variability of idarubicin metabolizing could be critical in predicting anthracycline-induced toxicities.
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http://dx.doi.org/10.1097/FPC.0000000000000431DOI Listing
March 2021

Healthcare resource utilization in adult patients with relapsed/refractory FLT3 mutated acute myeloid leukemia: A retrospective chart review from Spain.

Eur J Haematol 2021 May 5;106(5):724-733. Epub 2021 Mar 5.

Instituto de Investigación Sanitaria La Fe (IISLAFE), Valencia, Spain.

Background: Information regarding impact on healthcare systems of relapsed or refractory (R/R) FLT3 mutated (FLT3mut) acute myeloid leukemia (AML) is scarce.

Objective: To assess the time and reimbursement associated with hospitalizations of patients with R/R FLT3mut AML in a tertiary Spanish hospital.

Methods: Retrospective review of medical charts identified patients aged ≥ 18 years with R/R FLT3mut AML between 1998 and 2018. Data were collected from the date of first diagnosis of R/R FLT3mut AML (index) until death or loss to follow-up. The primary end point was duration and frequency of hospitalization, use of outpatient resources and transfusion burden. Reimbursement associated with hospitalizations (including associated chemotherapy) was also assessed.

Results: Thirty-eight patients were eligible for inclusion. Their median age was 52 years, and 30 (79%) received intensive salvage chemotherapy; FLAG-IDA-based regimens were the most frequent (24 patients, 63%). Overall, there were 150 hospitalizations (mean 3.9/patient; mean duration 21 days). Patients spent a mean of 24% of the study period in hospital. Total mean reimbursement was €108 293 per patient; the majority (€89 834) attributable to inpatient stays (€22 576 /hospitalization). During chemotherapy period (prior to first alloHSCT), there were 73 hospitalizations (mean duration 22 days); mean reimbursement was €19 776 per hospitalization and €49 819 per patient. AlloHSCT (n = 16) involved 77 hospitalizations (mean duration 21 days), mean reimbursement €25 231/hospitalization and €131 515 per patient.

Conclusion: Data from this study suggest that there is a substantial healthcare resource utilization and cost burden on R/R FLT3mut AML patients in Spain receiving active treatments.
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http://dx.doi.org/10.1111/ejh.13604DOI Listing
May 2021

Helpful Criteria When Implementing NGS Panels in Childhood Lymphoblastic Leukemia.

J Pers Med 2020 Nov 26;10(4). Epub 2020 Nov 26.

Hematology Laboratory, Hospital Sant Joan de Déu Barcelona, Passeig Sant Joan de Déu 2, 08950 Esplugues de Llobregat, Barcelona, Spain.

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. This collaborative study among Spanish centers was aimed to assess the feasibility, pros, and cons of our customized panel and other commercial alternatives of NGS-targeted approaches. The custom panel was tested in three different sequencing centers. We used the same samples to assess other commercial panels (Oncomine Childhood Cancer Research Assay; ArcherFusionPlex ALL, and Human Comprehensive Cancer Panel GeneRead Panel v2). Overall, the panels showed a good performance in different centers and platforms, but each NGS approach presented some issues, as well as pros and cons. Moreover, a previous consensus on the analysis and reporting following international guidelines would be preferable to improve the concordance in results among centers. Our study shows the challenges posed by NGS methodology and the need to consider several aspects of the chosen NGS-targeted approach and reach a consensus before implementing it in daily practice.
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http://dx.doi.org/10.3390/jpm10040244DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7711852PMC
November 2020

Genotype FBN1/phenotype relationship in a cohort of patients with Marfan syndrome.

Clin Genet 2021 Feb 23;99(2):269-280. Epub 2020 Nov 23.

Unidad de Regeneración y Trasplante cardíaco, Instituto de Investigación Sanitaria La Fe, Valencia, España, Spain.

Marfan syndrome (MFS) is a systemic connective tissue disorder caused by mutations in the fibrillin-1 (FBN1) gene, and cardiovascular involvement is the leading cause of mortality. We sought to examine the genotype/phenotype realtionship in 61 consecutive patients with a phenotype and genotype compatible with MFS. The FBN1 gene was analyzed by massive sequencing using a hybridization capture-based target enrichment custom panel. Forty-three different variants of FBN1 were identified, of which 17 have not been previously reported. The causal variants of MFS were grouped into mutations resulting in haploinsufficiency (HI group; 23 patients) and mutations producing a dominant-negative effect (DN group; 38 patients). Patient information was collected from electronic medical records and clinical evaluation. While no significant differences were found between the two groups, the HI group included more cases with aortic dissection and occurring at a younger age that the DN group (34.7% vs. 15.8%; p = 0.160). Irrespective of the mutation group, males presented with a higher probability of aortic involvement (4-fold higher risk than females) and aortic dissections events occurred at younger ages. Patients with DN variants carrying a cysteine substitution had a higher incidence of ectopia lentis.
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http://dx.doi.org/10.1111/cge.13879DOI Listing
February 2021

Impact of combinations of single-nucleotide polymorphisms of anthracycline transporter genes upon the efficacy and toxicity of induction chemotherapy in acute myeloid leukemia.

Leuk Lymphoma 2021 03 2;62(3):659-668. Epub 2020 Nov 2.

Servicio de Hematología y Hemoterapia. Hospital Universitari i Politècnic, Valencia, Spain.

Anthracycline uptake could be affected by influx and efflux transporters in acute myeloid leukemia (AML). Combinations of single-nucleotide polymorphisms (SNPs) of wild-type genotype of influx transporters () and homozygous variant genotypes of polymorphisms (, , ) were evaluated in 225 adult AML patients. No differences in complete remission were reported, but higher induction death was observed with combinations of rs4149056 and (triple variant haplotype, rs1128503), previously associated with and SNPs. Several combinations of and with SNPs were associated with higher toxicities, including nephrotoxicity and hepatotoxicity, neutropenia, previously related to , and a novel correlation with mucositis. Combination of rs714368 and rs2231142 was related to cardiac toxicity, reproducing previous correlations with . This study shows the impact of transporter polymorphisms in AML chemotherapy safety. Further prospective studies with larger populations are needed to validate these associations.
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http://dx.doi.org/10.1080/10428194.2020.1839650DOI Listing
March 2021

Implications of TP53 allelic state for genome stability, clinical presentation and outcomes in myelodysplastic syndromes.

Nat Med 2020 10 3;26(10):1549-1556. Epub 2020 Aug 3.

Department of Hematology, Democritus University of Thrace Medical School, Alexandroupolis, Greece.

Tumor protein p53 (TP53) is the most frequently mutated gene in cancer. In patients with myelodysplastic syndromes (MDS), TP53 mutations are associated with high-risk disease, rapid transformation to acute myeloid leukemia (AML), resistance to conventional therapies and dismal outcomes. Consistent with the tumor-suppressive role of TP53, patients harbor both mono- and biallelic mutations. However, the biological and clinical implications of TP53 allelic state have not been fully investigated in MDS or any other cancer type. We analyzed 3,324 patients with MDS for TP53 mutations and allelic imbalances and delineated two subsets of patients with distinct phenotypes and outcomes. One-third of TP53-mutated patients had monoallelic mutations whereas two-thirds had multiple hits (multi-hit) consistent with biallelic targeting. Established associations with complex karyotype, few co-occurring mutations, high-risk presentation and poor outcomes were specific to multi-hit patients only. TP53 multi-hit state predicted risk of death and leukemic transformation independently of the Revised International Prognostic Scoring System (IPSS-R). Surprisingly, monoallelic patients did not differ from TP53 wild-type patients in outcomes and response to therapy. This study shows that consideration of TP53 allelic state is critical for diagnostic and prognostic precision in MDS as well as in future correlative studies of treatment response.
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http://dx.doi.org/10.1038/s41591-020-1008-zDOI Listing
October 2020

Analysis of SNP Array Abnormalities in Patients with DE NOVO Acute Myeloid Leukemia with Normal Karyotype.

Sci Rep 2020 04 3;10(1):5904. Epub 2020 Apr 3.

Hematology Service, Hospital Universitario y Politécnico La Fe, Valencia, Spain.

Nearly 50% of patients with de novo acute myeloid leukemia (AML) harbor an apparently normal karyotype (NK) by conventional cytogenetic techniques showing a very heterogeneous prognosis. This could be related to the presence of cryptic cytogenetic abnormalities (CCA) not detectable by conventional methods. The study of copy number alterations (CNA) and loss of heterozygozity (LOH) in hematological malignancies is possible using a high resolution SNP-array. Recently, in clinical practice the karyotype study has been complemented with the identification of point mutations in an increasing number of genes. We analyzed 252 de novo NK-AML patients from Hospital La Fe (n = 44) and from previously reported cohorts (n = 208) to identify CCA by SNP-array, and to integrate the analysis of CCA with molecular alterations detected by Next-Generation-sequencing. CCA were detected in 58% of patients. In addition, 49% of them harbored CNA or LOH and point mutations, simultaneously. Patients were grouped in 3 sets by their abnormalities: patients carrying several CCA simultaneously, patients with mutations in FLT3, NPM1 and/or DNMT3A and patients with an amalgam of mutations. We found a negative correlation between the number of CCA and the outcome of the patients. This study outlines that CCA are present in up to 50% of NK-AML patients and have a negative impact on the outcome. CCA may contribute to the heterogeneous prognosis.
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http://dx.doi.org/10.1038/s41598-020-61589-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7125150PMC
April 2020

Acute Promyelocytic Leukemia: A Constellation of Molecular Events around a Single Fusion Gene.

Cancers (Basel) 2020 Mar 8;12(3). Epub 2020 Mar 8.

Department of Hematology, Hospital Universitario y Politécnico La Fe, 46026 Valencia, Spain.

Although acute promyelocytic leukemia (APL) is one of the most characterized forms of acute myeloid leukemia (AML), the molecular mechanisms involved in the development and progression of this disease are still a matter of study. APL is defined by the rearrangement as a consequence of the translocation t(15;17)(q24;q21). However, this abnormality alone is not able to trigger the whole leukemic phenotype and secondary cooperating events might contribute to APL pathogenesis. Additional somatic mutations are known to occur recurrently in several genes, such as , , and , whereas mutations in other common AML genes are rarely detected, resulting in a different molecular profile compared to other AML subtypes. How this mutational spectrum, including point mutations in the fusion gene, could contribute to the 10%-15% of relapsed or resistant APL patients is still unknown. Moreover, due to the uncertain impact of additional mutations on prognosis, the identification of the APL-specific genetic lesion is still the only method recommended in the routine evaluation/screening at diagnosis and for minimal residual disease (MRD) assessment. However, the gene expression profile of genes, such as , and once combined with the molecular events, might improve future prognostic models, allowing us to predict clinical outcomes and to categorize APL patients in different risk subsets, as recently reported. In this review, we will focus on the molecular characterization of APL patients at diagnosis, relapse and resistance, in both children and adults. We will also describe different standardized molecular approaches to study MRD, including those recently developed. Finally, we will discuss how novel molecular findings can improve the management of this disease.
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http://dx.doi.org/10.3390/cancers12030624DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7139833PMC
March 2020

Impact of somatic mutations in myelodysplastic patients with isolated partial or total loss of chromosome 7.

Leukemia 2020 09 17;34(9):2441-2450. Epub 2020 Feb 17.

Department of Haematological Medicine, King's College Hospital, NHS Foundation Trust, London, UK.

Monosomy 7 [-7] and/or partial loss of chromosome 7 [del(7q)] are associated with poor and intermediate prognosis, respectively, in myelodysplastic syndromes (MDS), but somatic mutations may also play a key complementary role. We analyzed the impact on the outcomes of deep targeted mutational screening in 280 MDS patients with -7/del(7q) as isolated cytogenetic abnormality (86 with del(7q) and 194 with -7). Patients with del(7q) or -7 had similar demographic and disease-related characteristics. Somatic mutations were detected in 79% (93/117) of patients (82% in -7 and 73% in del(7q) group). Median number of mutations per patient was 2 (range 0-8). There was no difference in mutation frequency between the two groups. Patients harbouring ≥2 mutations had a worse outcome than patients with <2 or no mutations (leukaemic transformation at 24 months, 38% and 20%, respectively, p = 0.044). Untreated patients with del(7q) had better overall survival (OS) compared with -7 (median OS, 34 vs 17 months, p = 0.034). In multivariable analysis, blast count, TP53 mutations and number of mutations were independent predictors of OS, whereas the cytogenetic subgroups did not retain prognostic relevance. This study highlights the importance of mutational analysis in terms of prognosis in MDS patients with isolated -7 or del(7q).
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http://dx.doi.org/10.1038/s41375-020-0728-xDOI Listing
September 2020

Factor XIII deficiency in two Spanish families with a novel variant in gene F13A1 detected by next-generation sequencing; symptoms and clinical management.

J Thromb Thrombolysis 2020 Oct;50(3):686-688

Unidad de Hemostasia Y Trombosis, Servicio de Hematología, Hospital Universitario Y Politécnico La Fe, Valencia, Spain.

Coagulation factor XIII (FXIII) has a major role in coagulation stabilizing the haemostatic clot. FXIII deficiency is associated with an increased risk of bleeding. Severe phenotypes lead to spontaneous, traumatic and surgical bleeding. Umbilical cord bleeding is especially common, and intracranial bleeding may occur in up to one third of patients without prophylaxis. In this work, we used NGS for screening all the coding and intronic boundary regions of F13A1 and F13B genes in two families affected by severe FXIII deficiency. Outcome confirmation analysis and variant studies in related patients was done by Sanger sequencing. Two variants were found: c.34A > G (p.Arg12Gly; NM_00129.3) and c.514C > T (p.Arg172Ter; NM_00129.3), both located in the F13A1 gene. The variant p.Arg172Ter is already described in literature and was found in homozygosis in one family and in compound heterozygosis in the other family. The variant p.Arg12Gly variant has not been described previously. This variant is located in the activation peptide of the FXIII A-subunit which is highly conserved among FXIII homologs. Given the high risk of dangerous bleeding and early manifestation in severe FXIII-deficient patients, a prompt genetic confirmation is imperative. In this sense, NGS technology allows a rapid and simultaneous analysis of all regions of all the genes involved in the pathology.
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http://dx.doi.org/10.1007/s11239-020-02065-zDOI Listing
October 2020

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

Br J Haematol 2020 03 16;188(5):605-622. Epub 2019 Oct 16.

Department of Haematology, Hospital Universitari i Politècnic La Fe, València, Spain.

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.
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http://dx.doi.org/10.1111/bjh.16175DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7064979PMC
March 2020

Dexamethasone does not prevent malignant cell reintroduction in leukemia patients undergoing ovarian transplant: risk assessment of leukemic cell transmission by a xenograft model.

Hum Reprod 2019 08;34(8):1485-1493

Reproductive Medicine Research Group, Instituto de Investigación Sanitaria La Fe, Valencia, Spain.

Study Question: Does dexamethasone (DXM) incubation avoid the reintroduction of leukemic malignant cells after ovarian tissue retransplantation in vivo?

Summary Answer: DXM incubation prior to retransplantation of ovarian tissue does not prevent reintroduction of leukemic cells.

What Is Known Already: Retransplantation of cryopreserved ovarian cortex from patients diagnosed with acute lymphoblastic leukemia (ALL) involves a risk of reintroducing malignant cells. DXM treatment is effective at inducing leukemic cell death in vitro.

Study Design, Size, Duration: This was an experimental study where ovarian cortex fragments from patients with ALL were randomly allocated to incubation with or without DXM (n = 11/group) and grafted to 22 immunodeficient mice for 6 months. In a parallel experiment, 22 immunodeficient mice were injected i.p. with varying amounts of RCH-ACV ALL cells (human leukemia cell line) and maintained for 4 months.

Participants/materials, Setting, Methods: Cryopreserved ovarian fragments from patients with ALL were exposed in vitro to 0.4 μM DXM or basal media (control) prior to xenograft into ovariectomized severe combined immunodeficiency (SCID) mice (experiment 1). After 6 months of monitoring, leukemia cell contamination was assessed in ovarian grafts and mouse organs by histology, PCR (presence of mouse mtDNA and absence of p53 were together considered a negative result for the presence of human cells) and detection of immunoglobulin monoclonality and specific ALL markers if present in the patient.In experiment 2, a series of 22 immunodeficient female mice was injected with specific doses of the leukemia cell line RCH-ACV (103 - 5 × 106, n = 4/group) to assess the engraftment competence of the SCID model.

Main Results And The Role Of Chance: ALL metastatic cells were detected, by PCR, in five DXM-treated and one control human ovarian tissue graft as well as in a control mouse liver, although malignant cell infiltration was not detected by histology in any sample after 6 months. In total, minimal residual disease was present in three DXM-treated and three control mice.RCH-ACV cells were detected in liver and spleen samples after the injection of as little as 103 cells, although only animals receiving 5 × 106 cells developed clinical signs of disease and metastases.

Limitations, Reasons For Caution: This is an experimental study where the malignant potential of leukemic cells contained in human ovarian tissues has been assessed in immunodeficient mice.

Wider Implications Of The Findings: These results indicate that DXM incubation prior to retransplantation of ovarian tissue does not prevent reintroduction of leukemic cells. Therefore, caution should be taken in retransplanting ovarian tissue from patients with leukemia until safer systems are developed, as leukemic cells present in ovarian grafts were able to survive, proliferate and migrate after cryopreservation and xenograft.

Study Funding/competing Interest(s): Funded by the Regional Valencian Ministry of Education (PROMETEO/2018/137) and by the Spanish Ministry of Economy and Competitiveness (PI16/FIS PI16/01664 and PTQ-16-08222 for S.H. participation). There are no competing interests.
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http://dx.doi.org/10.1093/humrep/dez115DOI Listing
August 2019

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

Ann Hematol 2019 Sep 16;98(9):2151-2162. Epub 2019 Jul 16.

Hematology Department, University Hospital of Salamanca, IBSAL Institute for Biomedical Research of Salamanca, Paseo de San Vicente, 139, 37007, Salamanca, Spain.

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. We found that patients with a higher number of mutated genes (more than 2) had a worse outcome (2 years overall survival [OS] 54.8% vs. 31.1%, p = 0.035). The only two significant variables in the multivariate analysis for OS were TET2 mutations (p = 0.046) and the development of cGVHD, considered as a time-dependent variable (p < 0.001), correlated with a worse and a better outcome, respectively. TP53 mutations also demonstrated impact on the cumulative incidence of relapse (CIR) (1 year CIR 47.1% vs. 9.8%, p = 0.006) and were related with complex karyotype (p = 0.003). cGVHD improved the outcome even among patients with more than 2 mutated genes (1-year OS 88.9% at 1 year vs. 31.3%, p = 0.02) and patients with TP53 mutations (1-year CIR 20% vs. 42.9%, p = 0.553). These results confirm that cGVHD could ameliorate the adverse impact of somatic mutations in patients with MDS with HSCT.
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http://dx.doi.org/10.1007/s00277-019-03751-6DOI Listing
September 2019

Clinical and molecular characterization by next generation sequencing of Spanish patients affected by congenital deficiencies of fibrinogen.

Thromb Res 2019 Aug 25;180:115-117. Epub 2019 Jun 25.

Unidad de Hemostasia y Trombosis, Servicio de Hematología, Hospital Universitario y Politécnico La Fe, Valencia, Spain.

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http://dx.doi.org/10.1016/j.thromres.2019.06.015DOI Listing
August 2019

Incidence and outcome of invasive fungal disease after front-line intensive chemotherapy in patients with acute myeloid leukemia: impact of antifungal prophylaxis.

Ann Hematol 2019 Sep 25;98(9):2081-2088. Epub 2019 Jun 25.

Department of Hematology, Hospital Universitari i Politècnic La Fe, Avda. Fernando Abril Martorell 106, CP: 46026, Valencia, Spain.

Few reports analyze the incidence and clinical outcome of invasive fungal disease (IFD) in patients with newly diagnosed acute myeloid leukemia (AML) undergoing intensive chemotherapy, and thus the impact of different antifungal prophylactic regimens remains unclear. We analyze the incidence and clinical outcome of IFD in a large series of adult AML patients undergoing front-line intensive induction and consolidation chemotherapy between 2004 and 2015 in a single institution. Three antifungal prophylaxis regimens were given (2004-2005 oral fluconazole, 2006-2012 intravenous itraconazole, and 2013-2015 voriconazole). Overall, 285 patients and 589 intensive chemotherapy episodes were assessed (47%) (induction courses 47% and consolidation 53%). The median age was 51 years (range, 17-65). We observed 56 (10%) episodes of IFD. According to the EORTC 2008 criteria, IFD was classified as possible (29, 52%), probable (17, 30%), and proven (10, 18%). Possible/probable/proven IFD rate was significantly lower during HiDAC consolidation as compared to any anthracycline-containing chemotherapy courses (2% vs. 11%, P = 0.001), and under voriconazole prophylaxis as compared to itraconazole and fluconazole (6% vs. 11% vs. 15%, P = 0.007), and the multivariate analysis showed that they were independent risk factors. Patients under voriconazole prophylaxis had shorter hospitalization duration and less frequent use of empirical or directed antifungal therapy. In conclusion, IFD was a frequent complication during upfront intensive chemotherapy courses for adult AML patients. This retrospective study shows that voriconazole prophylaxis was feasible and associated with a lower risk of IFD compared with intravenous itraconazole or oral fluconazole schedules.
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http://dx.doi.org/10.1007/s00277-019-03744-5DOI Listing
September 2019

RNA Sequencing Analysis for the Identification of a PCM1/PDGFRB Fusion Gene Responsive to Imatinib.

Acta Haematol 2019 14;142(2):92-97. Epub 2019 May 14.

Hematology Department, University Hospital La Fe, Valencia, Spain.

The platelet-derived growth factor receptor β (PDGFRB) gene translocations lead to a spectrum of chronic myeloid neoplasms, frequently associated with eosinophilia. Clinical heterogeneity is associated with a molecular one. Here, we report a novel case of a patient harboring a t(5;8)(q33;p22) translocation, resulting in the PCM1/PDGFRB fusion. Conventional cytogenetics and RNA sequencing were performed to identify the chromosomes and the genes involved in the rearrangement, respectively. This study shows that the combination of different strategies is pivotal to fine-tune the diagnosis and the clinical management of the patient. After 1 year of treatment with imatinib, the patient achieves hematological and molecular remission. We present an attractive strategy to identify novel and/or cryptic fusions, which will be relevant for clinicians dealing with the diagnosis of the patients with myelodysplastic syndrome/myeloproliferative diseases with atypical manifestations.
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http://dx.doi.org/10.1159/000497348DOI Listing
February 2020

CRISPR to fix bad blood: a new tool in basic and clinical hematology.

Haematologica 2019 05 28;104(5):881-893. Epub 2019 Mar 28.

Hematology Research Group, Instituto de Investigación Sanitaria La Fe, Valencia

Advances in genome engineering in the last decade, particularly in the development of programmable nucleases, have made it possible to edit the genomes of most cell types precisely and efficiently. Chief among these advances, the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is a novel, versatile and easy-to-use tool to edit genomes irrespective of their complexity, with multiple and broad applications in biomedicine. In this review, we focus on the use of CRISPR/Cas9 genome editing in the context of hematologic diseases and appraise the major achievements and challenges in this rapidly moving field to gain a clearer perspective on the potential of this technology to move from the laboratory to the clinic. Accordingly, we discuss data from studies editing hematopoietic cells to understand and model blood diseases, and to develop novel therapies for hematologic malignancies. We provide an overview of the applications of gene editing in experimental, preclinical and clinical hematology including interrogation of gene function, target identification and drug discovery and chimeric antigen receptor T-cell engineering. We also highlight current limitations of CRISPR/Cas9 and the possible strategies to overcome them. Finally, we consider what advances in CRISPR/Cas9 are needed to move the hematology field forward.
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http://dx.doi.org/10.3324/haematol.2018.211359DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6518885PMC
May 2019

Clinical Utility of a Next-Generation Sequencing Panel for Acute Myeloid Leukemia Diagnostics.

J Mol Diagn 2019 03 19;21(2):228-240. Epub 2018 Dec 19.

Molecular Biology Unit, Clinical Pathology Service, La Fe University and Polytechnic Hospital, Valencia, Spain; La Fe Health Research Institute, Hematology Research Group, Valencia, Spain; CIBERONC, Carlos III Health Research Institute, Madrid, Spain. Electronic address:

Next-generation sequencing (NGS) has redefined the genetic landscape of acute myeloid leukemia (AML), providing new molecular markers for diagnostic and prognostic classifications. However, its application in the clinical setting is still challenging. We hypothesized that a 19-gene AML-targeted NGS panel could be a valid approach to obtain clinically relevant information. Thus, we assessed the ability of this panel to classify AML patients according to diagnostic and prognostic indexes in a cohort of 162 patients. The assay yielded a median read depth >2000×, with 88% of on-target reads and a mean uniformity >93% without significant global strand bias. The method was sensitive and specific, with a valid performance at the clinical variant allele frequency cutoff of 3% for point mutations and 5% for insertions or deletions (INDELs). Three-hundred thirty-nine variants were found (36% INDELs and 64% single nucleotide variants). Concordance between NGS and other conventional techniques was 100%, but the NGS approach was able to identify more clinically relevant mutations. Finally, all patients could be classified into one of the 2016 World Health Organization diagnostic categories and virtually all into the recently proposed prognostic indexes (2017 European LeukemiaNet and Genomic classification). To sum up, we validate a reliable and reproducible method for AML diagnosis and demonstrate that small, well-designed NGS panels are sufficient to guide clinical decisions according to the current standards.
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http://dx.doi.org/10.1016/j.jmoldx.2018.09.009DOI Listing
March 2019

Clinical significance of complex karyotype at diagnosis in pediatric and adult patients with de novo acute promyelocytic leukemia treated with ATRA and chemotherapy.

Leuk Lymphoma 2019 05 11;60(5):1146-1155. Epub 2018 Dec 11.

g CIBERONC, Instituto Carlos III , Madrid , Spain.

Although additional cytogenetic abnormalities (ACA) do not affect the prognosis of patients with t(15;17) acute promyelocytic leukemia (APL), the role of a complex karyotype (CK) is yet to be clarified. We aimed to investigate the relationship of CK with relapse incidence in 1559 consecutive APL patients enrolled in three consecutive trials. Treatment consisted of AIDA induction followed by risk-adapted consolidation. A CK (CK) was defined as the presence of ≥2 ACA, and a very CK (CK+) as ≥3 ACA. Eighty-nine patients (8%) had a CK, of whom 41 (4%) had CK+. The 5-year cumulative incidence of relapse (CIR) in patients with CK was 18%, and 12% in those with <2 ACA (p=.09). Among patients with CK+, the 5-year CIR was 27% vs 12% (p=.003), retaining the statistical significance in multivariate analysis. This study shows an increased risk of relapse among APL patients with CK + treated with ATRA plus chemotherapy front-line regimens.
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http://dx.doi.org/10.1080/10428194.2018.1522438DOI Listing
May 2019

New mutations found by Next-Generation Sequencing screening of Spanish patients with Nemaline Myopathy.

PLoS One 2018 5;13(12):e0207296. Epub 2018 Dec 5.

Genomic Unit, Health Research Institute Hospital La Fe (IIS La Fe), Valencia, Spain.

Nemaline Myopathy (NM) is a rare genetic disorder that encompasses a large spectrum of myopathies characterized by hypotonia and generalized muscle weakness. To date, mutations in thirteen different genes have been associated with NM. The most frequently responsible genes are NEB (50% of cases) and ACTA1 (15-25% of cases). In this report all known NM related genes were screened by Next Generation Sequencing in five Spanish patients in order to genetically confirm the clinical and histological diagnosis of NM. Four mutations in NEB (c.17779_17780delTA, c.11086A>C, c.21076C>T and c.2310+5G>A) and one mutation in ACTA1 (c.871A>T) were found in four patients. Three of the four mutations in NEB were novel. A cDNA sequencing assay of the novel variants c.17779_17780delTA, c.11086A>C and c.2310+5G>A revealed that the intronic variant c.2310+5G>A affected the splicing process. Mutations reported here could help clinicians and geneticists in NM diagnosis.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0207296PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6281284PMC
May 2019

Analytical performance assessment of a novel cartridge-based blood gas analyzer.

Clin Biochem 2019 Jan 24;63:154-155. Epub 2018 Oct 24.

Instrumentation Laboratory, 180 Hartwell Road, Bedford, MA 01730, United States.

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http://dx.doi.org/10.1016/j.clinbiochem.2018.10.013DOI Listing
January 2019

Time and Cost of Hospitalisation for Salvage Therapy in Adults with Philadelphia Chromosome-Negative B Cell Precursor Relapsed or Refractory Acute Lymphoblastic Leukaemia in Spain.

Pharmacoecon Open 2019 Jun;3(2):229-235

Haematology Department, Hospital Universitari i Politècnic La Fe, Avinguda de Fern ando Abril Martorell, 106, 46026, Valencia, Spain.

Background: Philadelphia chromosome-negative (Ph-) relapsed or refractory (R/R) B-cell precursor acute lymphoblastic leukaemia (ALL) is rare, and information on its impact on healthcare systems is scarce.

Objective: To quantify the time and reimbursement associated with hospitalisations of patients with R/R ALL in a Spanish hospital.

Methods: Retrospective review of medical charts identified patients aged ≥ 18 years with Ph- R/R ALL hospitalised between 1998 and 2014. Data were collected from the date of first diagnosis of R/R ALL (index) until death or loss to follow-up. The primary endpoint was the proportion of time hospitalised during chemotherapy. Reimbursement associated with hospitalisations (including associated chemotherapy) was also assessed.

Results: Thirty-two patients were eligible for inclusion. Their median age was 41 years, and 50% had a first remission duration of ≤ 1 year; 34% had undergone allogeneic haematological stem-cell transplantation (alloHSCT). Overall, 31 patients had received intensive salvage chemotherapy, during which there were 42 hospitalisations (mean 1.4/patient; mean duration 26 days). Patients spent a mean of 71% of the chemotherapy period in hospital. Total mean reimbursement was €26,417 per patient, almost all (€25,723) attributable to inpatient stays (€18,986/hospitalisation). From the index date to death or loss to follow-up (excluding alloHSCT-related hospitalisations), there were 80 hospitalisations (mean duration 24 days); mean reimbursement was €16,692 per hospitalisation and €41,730 per patient. AlloHSCT (n = 8) involved 18 hospitalisations (mean reimbursement €39,782/hospitalisation; €89,510/patient).

Conclusion: Data from this sample of patients suggest that hospitalisations in R/R ALL are lengthy and associated with high costs in Spain.
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http://dx.doi.org/10.1007/s41669-018-0098-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6533337PMC
June 2019

The modular network structure of the mutational landscape of Acute Myeloid Leukemia.

PLoS One 2018 10;13(10):e0202926. Epub 2018 Oct 10.

Hematology Service, Hospital Universitario y Politécnico La Fe, Valencia, Spain.

Acute myeloid leukemia (AML) is associated with the sequential accumulation of acquired genetic alterations. Although at diagnosis cytogenetic alterations are frequent in AML, roughly 50% of patients present an apparently normal karyotype (NK), leading to a highly heterogeneous prognosis. Due to this significant heterogeneity, it has been suggested that different molecular mechanisms may trigger the disease with diverse prognostic implications. We performed whole-exome sequencing (WES) of tumor-normal matched samples of de novo AML-NK patients lacking mutations in NPM1, CEBPA or FLT3-ITD to identify new gene mutations with potential prognostic and therapeutic relevance to patients with AML. Novel candidate-genes, together with others previously described, were targeted resequenced in an independent cohort of 100 de novo AML patients classified in the cytogenetic intermediate-risk (IR) category. A mean of 4.89 mutations per sample were detected in 73 genes, 35 of which were mutated in more than one patient. After a network enrichment analysis, we defined a single in silico model and established a set of seed-genes that may trigger leukemogenesis in patients with normal karyotype. The high heterogeneity of gene mutations observed in AML patients suggested that a specific alteration could not be as essential as the interaction of deregulated pathways.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0202926PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6179200PMC
March 2019

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

Int J Lab Hematol 2019 Feb 5;41(1):109-117. Epub 2018 Oct 5.

Hematology, Hospital Universitario de Salamanca-IBSAL, Salamanca, Spain.

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. In step 1, we used an automated informatics system to screen 137 453 hemograms for cases of anemia and/or macrocytosis (n = 2702). In step 2, we excluded all patients whose anemia appeared to be due to a known cause. This left 290 patients had anemia of uncertain etiology. In step 3, we conducted further investigations, including a peripheral blood smear, and analysis of iron, vitamin B12, folate, and thyroid hormone levels.

Results: A differential diagnosis was obtained in 139 patients (48%). The primary causes of anemia were iron deficiency (n = 59) and megaloblastic anemia (n = 39). In total, 25 hematologic disorders were diagnosed, including 14 patients with MDS (56%). The median age of MDS patients was 80 years, 12 had anemia as an isolated cytopenia, and most (n = 10) had lower-risk disease (IPSS-R ≤ 3.5). SF3B1 mutations were most frequent (n = 6) and correlated with the presence of ring sideroblasts (100%) and associated with better prognosis (P = 0.001).

Conclusions: Our prospective, four-step approach is an efficient and logical strategy to facilitate the diagnosis of MDS on the basis of unexplained anemia and/or macrocytosis, and may allow the early diagnosis of the most serious causes of anemia. Molecular analysis of genes related to MDS could be a promising diagnostic and prognostic approach.
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http://dx.doi.org/10.1111/ijlh.12933DOI Listing
February 2019

Clonal architecture in patients with myelodysplastic syndromes and double or minor complex abnormalities: Detailed analysis of clonal composition, involved abnormalities, and prognostic significance.

Genes Chromosomes Cancer 2018 11 24;57(11):547-556. Epub 2018 Sep 24.

Department of Hematology and Medical Oncology, University of Göttingen, Göttingen, Germany.

The study analyzes the clonal architecture and the abnormalities involved in a series of 191 patients with myelodysplastic syndromes (MDS) and 2-3 clonal abnormalities. All patients were extracted from an international database. The patients were classified into six clonal subtypes (2A-3C) based on the number of abnormalities and the presentation of unrelated clones (UC) and/or a clonal evolution. UC were detected in 23/191 patients (12%). The composition of UC showed great variability. The only recurrent combination of abnormalities was del(5q) and + 8 in 8 of 23 patients (35%). In patients with clonal evolution, the clone size of the primary and secondary clone varied: Patients with -7 and + 8 in the primary clone showed a larger primary and a smaller secondary clone (-7: median 74% vs 10%; +8 73% vs 18%) while patients with del(5q) in the primary clone showed a smaller primary and a larger secondary clone (33% vs 61%). Univariate and multivariate analyses showed no significant differences regarding overall or AML-free survival between the clonal subtypes. Only the subtype 3C (3 abnormalities and clonal evolution) was an independent risk factor for developing AML (Hazard Ratio 5.5 as compared to subtype 2A, P < .05). Finally, our study confirms that the number of abnormalities clearly defines a significant risk factor for overall- as well as AML-free survival. Importantly, in patients with more than one clone, the calculation of the number of abnormalities in the entire sample instead of the number of abnormalities per clone allows a higher prognostic accuracy.
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http://dx.doi.org/10.1002/gcc.22667DOI Listing
November 2018