Actas de las V Jornadas ScienCity 2022. Fomento de la Cultura Científica, Tecnológica y de Innovación en Ciudades Inteligentes

ScienCity es una actividad que viene siendo continuada desde 2018 con el objetivo de dar a conocer los conocimientos y tecnologías emergentes siendo investigados en las universidades, informar de experiencias, servicios e iniciativas puestas ya en marcha por instituciones y empresas, llegar hasta decisores políticos que podrían crear sinergias, incentivar la creación de ideas y posibilidades de desarrollo conjuntas, implicar y provocar la participación ciudadana, así como gestar una red internacional multidisciplinar de investigadores que garantice la continuación de futuras ediciones. En 2022 se recibieron un total de 48 trabajos repartidos en 25 ponencias y 24 pósteres pertenecientes a 98 autores de 14 instituciones distintas de España, Portugal, Polonia y Países Bajos.Fundación Española para la Ciencia y la Tecnología-Ministerio de Ciencia, Innovación y Universidades; Consejería de la Presidencia, Administración Pública e Interior de la Junta de Andalucía; Estrategia de Política de Investigación y Transferencia de la Universidad de Huelva; Cátedra de Innovación Social de Aguas de Huelva; Cátedra de la Provincia; Grupo de investigación TEP-192 de Control y Robótica; Centro de Investigación en Tecnología, Energía y Sostenibilidad (CITES

Conditional Expression of E2A-HLF Induces B-Cell Precursor Death and Myeloproliferative-Like Disease in Knock-In Mice

<div><p>Chromosomal translocations are driver mutations of human cancers, particularly leukemias. They define disease subtypes and are used as prognostic markers, for minimal residual disease monitoring and therapeutic targets. Due to their low incidence, several translocations and their biological consequences remain poorly characterized. To address this, we engineered mouse strains that conditionally express <i>E2A-HLF</i>, a fusion oncogene from the translocation t(17;19) associated with 1% of pediatric B-cell precursor ALL. Conditional oncogene activation and expression were directed to the B-cell compartment by the Cre driver promoters <i>CD19</i> or <i>Mb1</i> (Igα, CD79a), or to the hematopoietic stem cell compartment by the <i>Mx1</i> promoter. <i>E2A-HLF</i> expression in B-cell progenitors induced hyposplenia and lymphopenia, whereas expression in hematopoietic stem/progenitor cells was embryonic lethal. Increased cell death was detected in <i>E2A-HLF</i> expressing cells, suggesting the need for cooperating genetic events that suppress cell death for B-cell oncogenic transformation. <i>E2A-HLF/Mb1</i>.<i>Cre</i> aged mice developed a fatal myeloproliferative-like disorder with low frequency characterized by leukocytosis, anemia, hepatosplenomegaly and organ-infiltration by mature myelocytes. In conclusion, we have developed conditional <i>E2A-HLF</i> knock-in mice, which provide an experimental platform to study cooperating genetic events and further elucidate translational biology in cross-species comparative studies.</p></div

Conditional E2A-HLF transgenic mice develop B cell lymphopenia and hyposplenia.

<p><b>(A)</b> Total bone marrow cells from 2-month-old wild-type (WT, n = 12) and E2A-HLF/Mb1.Cre (n = 11) transgenic mice were enumerated by trypan blue exclusion assay. Horizontal bars denote the mean. Statistical analysis was done by Mann-Whitney U test. <b>(B)</b> Dot plots from flow cytometry analysis show mature B cell (B220+CD19+) subpopulations in peripheral blood (PB, upper panel) and progenitor B cell subpopulations (Lin-CD19-CD43+, Lin-CD19+CD43+, Lin-CD19+CD43-) in bone marrow (BM, lower panel) of representative wild-type and E2A-HLF/Mb1.Cre transgenic mice. Lin, lineage markers (CD3, CD4, CD8, NK1-1, Mac1, Gr1, Ter119). <b>(C)</b> Graph summarizes relative frequencies of mature B cells in peripheral blood (upper panel) and progenitor B cells in bone marrow (lower panel) of wild-type (WT, n = 9), E2A-HLF/CD19.Cre 2-month-old mice (n = 4), E2A-HLF/Mb1.Cre 2-month-old (n = 6) and 6-month-old (n = 3) mice. Columns denote mean and bars denote standard error of the mean. <b>(D)</b> Spleens are shown for representative WT (n = 3) and transgenic E2A-HLF/Mb1.Cre mice (n = 3). <b>(E)</b> Graph shows spleen weights from WT (n = 14), E2A-HLF/CD19.Cre (n = 4) and E2A-HLF/Mb1.Cre (n = 11) mice, horizontal bars denote the mean. Statistical analysis was done by Mann-Whitney U test; n.s., not significant.</p

Conditional E2A-HLF transgenic mice.

<p><b>(A)</b> Schematic representation of wild type, targeted, and recombined E2A alleles. After Cre recombinase expression, 3’ E2A exons (13, E12, E47 and 16) and the PGKneo cassette are deleted and the human HLF cDNA linked with EGFP by an IRES element is fused in frame to E2A. The expression of Cre-recombinase was driven by the B-cell specific promoters CD19 or Mb1 (CD79a, Igα), or in hematopoietic stem cells by the Mx1 promoter. The phenotypes arising depending on the Cre-recombinase driven promoter are indicated on the right. MPD-like, myeloproliferative disease like. <b>(B)</b> Flow cytometry analysis shows GFP expression from fetal liver (FL) cells of an E2A-HLF/Mx1.Cre embryo and from total bone marrow (BM) of E2A-HLF/Mb1.Cre and E2A-HLF/CD19.Cre mice. <b>(C)</b> Representative western blots show E2A and E2A-HLF protein levels in FACS-sorted progenitor B cells from wild-type (WT, lin-CD19+CD43+) and 2-month-old transgenic (lin-CD19+CD43+GFP+) E2A-HLF/Mb1.Cre and E2A-HLF/CD19.Cre mice. For comparison, E2A and E2A-HLF expression is shown from the non-E2A-HLF cell line REH and from the E2A-HLF+ cell line HAL-01. GAPDH was used as loading control. The E2A/E2A-HLF ratio (shown below) was determined by densitometry.</p

The epigenetics of breast cancer – Opportunities for diagnostics, risk stratification and therapy

The stage and molecular pathology-dependent prognosis of breast cancer, the limited treatment options for triple-negative carcinomas, as well as the development of resistance to therapies illustrate the need for improved early diagnosis and the development of new therapeutic approaches. Increasing data suggests that some answers to these challenges could be found in the area of epigenetics. In this study, we focus on the current research of the epigenetics of breast cancer, especially on the potential of epigenetics for clinical application in diagnostics, risk stratification and therapy. The differential DNA methylation status of specific gene regions has been used in the past to differentiate breast cancer cells from normal tissue. New technologies as detection of circulating nucleic acids including microRNAs to early detect breast cancer are emerging. Pattern of DNA methylation and expression of histone-modifying enzymes have been successfully used for risk stratification. However, all these epigenetic biomarkers should be validated in larger clinical studies. Recent preclinical and clinical studies show a therapeutic benefit of epigenetically active drugs for breast cancer entities that are still difficult to treat (triple negative, UICC stage IV). Remarkably, epigenetic therapies combined with chemotherapies or hormone-based therapies represent the most promising strategy. At the current stage, the integration of epigenetic substances into established breast cancer therapy protocols seems to hold the greatest potential for a clinical application of epigenetic research

Monitoring of Measurable Residual Disease Using Circulating DNA after Allogeneic Hematopoietic Cell Transplantation

Relapse of the underlying disease is a frequent complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT). In this study, we describe the clinical utility of measurable residual disease (MRD) and mixed chimerism (MC) assessment in circulating cell-free DNA (cfDNA) analysis to detect earlier relapse in patients with hematological malignancies after allo-HSCT. A total of 326 plasma and peripheral blood mononuclear cell (PBMCs) samples obtained from 62 patients with myeloid malignancies were analyzed by droplet-digital PCR (median follow-up: 827 days). Comparison of MC in patients at relapse and in complete remission identified an optimal discriminating threshold of 18% of recipient-derived cfDNA. After performing a targeted next-generation sequencing (NGS) panel, 136 mutations in 58 patients were detected. In a total of 119 paired samples, the putative mutations were detected in both cfDNA and PBMCs in 73 samples (61.3%). In 45 samples (37.8%) they were detected only in cfDNA, and in only one patient (0.9%) were they detected solely in DNA from PBMCs. Hence, in 6 out of 23 patients (26%) with relapse after allo-HSCT, MRD positivity was detected earlier in cfDNA (mean 397 days) than in DNA derived from PBMCs (mean 451 days). In summary, monitoring of MRD and MC in cfDNA might be useful for earlier relapse detection in patients with myeloid malignancies after allo-HSCT

Epigenetic priming of AML blasts for all-trans retinoic acid-induced differentiation by the HDAC class-I selective inhibitor entinostat.

All-trans retinoic acid (ATRA) has only limited single agent activity in AML without the PML-RARα fusion (non-M3 AML). In search of a sensitizing strategy to overcome this relative ATRA resistance, we investigated the potency of the HDAC class-I selective inhibitor entinostat in AML cell lines Kasumi-1 and HL-60 and primary AML blasts. Entinostat alone induced robust differentiation of both cell lines, which was enhanced by the combination with ATRA. This "priming" effect on ATRA-induced differentiation was at least equivalent to that achieved with the DNA hypomethylating agent decitabine, and could overall be recapitulated in primary AML blasts treated ex vivo. Moreover, entinostat treatment established the activating chromatin marks acH3, acH3K9, acH4 and H3K4me3 at the promoter of the RARβ2 gene, an essential mediator of retinoic acid (RA) signaling in different solid tumor models. Similarly, RARβ2 promoter hypermethylation (which in primary blasts from 90 AML/MDS patients was surprisingly infrequent) could be partially reversed by decitabine in the two cell lines. Re-induction of the epigenetically silenced RARβ2 gene was achieved only when entinostat or decitabine were given prior to ATRA treatment. Thus in this model, reactivation of RARβ2 was not necessarily required for the differentiation effect, and pharmacological RARβ2 promoter demethylation may be a bystander phenomenon rather than an essential prerequisite for the cellular effects of decitabine when combined with ATRA. In conclusion, as a "priming" agent for non-M3 AML blasts to the differentiation-inducing effects of ATRA, entinostat is at least as active as decitabine, and both act in part independently from RARβ2. Further investigation of this treatment combination in non-M3 AML patients is therefore warranted, independently of RARβ2 gene silencing by DNA methylation

Long-term follow-up of patients with acute myeloid leukemia undergoing allogeneic hematopoietic stem cell transplantation after primary induction failure

Abstract Primary induction failure (PIF) in acute myeloid leukemia (AML) patients is associated with poor outcome, with allogeneic hematopoietic stem cell transplantation (HCT) being the sole curative therapeutic option. Here, we retrospectively evaluated long-term outcomes of 220 AML patients undergoing allogeneic HCT after PIF who never achieved remission, and identified clinical and molecular risk factors associated with treatment response and ultimate prognosis. In this high-risk population, disease-free survival was 25.2% after 5 years and 18.7% after 10 years, while overall survival rates were 29.8% and 21.6% after 5 and 10 years of HCT, respectively. 10-year non-relapse mortality was 32.5%, and 48.8% of patients showed disease relapse within 10 years after allogeneic HCT. Adverse molecular risk features determined at initial diagnosis, poor performance status at the time of allogeneic HCT, and long diagnosis-to-HCT intervals were associated with unfavorable prognosis. Collectively, our data suggests that immediate allogeneic HCT after PIF offers long-term survival and cure in a substantial subset of cases and that high-risk AML patients who never achieved complete response during induction might benefit from early donor search

SETDB2 Links E2A-PBX1 to Cell-Cycle Dysregulation in Acute Leukemia through CDKN2C Repression

Summary: Acute lymphoblastic leukemia (ALL) is associated with significant morbidity and mortality, necessitating further improvements in diagnosis and therapy. Targeted therapies directed against chromatin regulators are emerging as promising approaches in preclinical studies and early clinical trials. Here, we demonstrate an oncogenic role for the protein lysine methyltransferase SETDB2 in leukemia pathogenesis. It is overexpressed in pre-BCR+ ALL and required for their maintenance in vitro and in vivo. SETDB2 expression is maintained as a direct target gene of the chimeric transcription factor E2A-PBX1 in a subset of ALL and suppresses expression of the cell-cycle inhibitor CDKN2C through histone H3K9 tri-methylation, thus establishing an oncogenic pathway subordinate to E2A-PBX1 that silences a major tumor suppressor in ALL. In contrast, SETDB2 was relatively dispensable for normal hematopoietic stem and progenitor cell proliferation. SETDB2 knockdown enhances sensitivity to kinase and chromatin inhibitors, providing a mechanistic rationale for targeting SETDB2 therapeutically in ALL. : Lin et al. report that the protein lysine methyltransferase SETDB2 is a direct target of chimeric transcription factor E2A-PBX1 and required for pathogenesis in B cell precursor leukemia. SETDB2 suppresses expression of the cell-cycle inhibitor CDKN2C, establishing an oncogenic pathway that silences a major tumor suppressor in acute leukemia. Keywords: SETDB2, E2A-PBX1, CDKN2C, preB-ALL, acute leukemia, cell-cycle inhibitio