A clinical-molecular update on azanucleoside-based therapy for the treatment of hematologic cancers

The azanucleosides azacitidine and decitabine are currently used for the treatment of acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) in patients not only eligible for intensive chemotherapy but are also being explored in other hematologic and solid cancers. Based on their capacity to interfere with the DNA methylation machinery, these drugs are also referred to as hypomethylating agents (HMAs). As DNA methylation contributes to epigenetic regulation, azanucleosides are further considered to be among the first true "epigenetic drugs" that have reached clinical application. However, intriguing new evidence suggests that DNA hypomethylation is not the only mechanism of action for these drugs. This review summarizes the experience from more than 10 years of clinical practice with azanucleosides and discusses their molecular actions, including several not related to DNA methylation. A particular focus is placed on possible causes of primary and acquired resistances to azanucleoside treatment. We highlight current limitations for the success and durability of azanucleoside-based therapy and illustrate that a better understanding of the molecular determinants of drug response holds great potential to overcome resistance

A clinical-molecular update on azanucleoside-based therapy for the treatment of hematologic cancers

The azanucleosides azacitidine and decitabine are currently used for the treatment of acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) in patients not only eligible for intensive chemotherapy but are also being explored in other hematologic and solid cancers. Based on their capacity to interfere with the DNA methylation machinery, these drugs are also referred to as hypomethylating agents (HMAs). As DNA methylation contributes to epigenetic regulation, azanucleosides are further considered to be among the first true "epigenetic drugs" that have reached clinical application. However, intriguing new evidence suggests that DNA hypomethylation is not the only mechanism of action for these drugs. This review summarizes the experience from more than 10 years of clinical practice with azanucleosides and discusses their molecular actions, including several not related to DNA methylation. A particular focus is placed on possible causes of primary and acquired resistances to azanucleoside treatment. We highlight current limitations for the success and durability of azanucleoside-based therapy and illustrate that a better understanding of the molecular determinants of drug response holds great potential to overcome resistance

Polycomb protein RING1A limits hematopoietic differentiation in myelodysplastic syndromes

Altres ajuts: This project was supported by grants from Deutsche José Carreras Leukaemie Stiftung DJCLS R 14/16 (KSG and MB), Radiumhemmets forskningsfonder, the Swedish Cancer foundation and the Swedish Research council (AL), German Cancer Consortium DKTK (AKG), the German Research Council DFG FOR2033 Go 713/2-1 and SFB 1243 A09 (KSG). Research in the Buschbeck lab is further supported by AFM-Téléthon (AFM-18738), the Marie Skłodowska Curie Training network 'ChroMe' (H2020-MSCAITN-2015-675610), and AGAUR (2014-SGR-35). Research at the IJC is supported by the 'La Caixa' Foundation, the Fundació Internacional Josep Carreras, Celgene Spain and the CERCA Programme / Generalitat de Catalunya.Genetic lesions affecting epigenetic regulators are frequent in myelodysplastic syndromes (MDS). Polycomb proteins are key epigenetic regulators of differentiation and stemness that act as two multimeric complexes termed polycomb repressive complexes 1 and 2, PRC1 and PRC2, respectively. While components and regulators of PRC2 such as ASXL1 and EZH2 are frequently mutated in MDS and AML, little is known about the role of PRC1. To analyze the role of PRC1, we have taken a functional approach testing PRC1 components in loss- and gain-of-function experiments that we found overexpressed in advanced MDS patients or dynamically expressed during normal hematopoiesis. This approach allowed us to identify the enzymatically active component RING1A as the key PRC1 component in hematopoietic stem cells and MDS. Specifically, we found that RING1A is expressed in CD34 + bone marrow progenitor cells and further overexpressed in high-risk MDS patients. Knockdown of RING1A in an MDS-derived AML cell line facilitated spontaneous and retinoic acid-induced differentiation. Similarly, inactivation of RING1A in primary CD34 + cells augmented erythroid differentiation. Treatment with a small compound RING1 inhibitor reduced the colony forming capacity of CD34 + cells from MDS patients and healthy controls. In MDS patients higher RING1A expression associated with an increased number of dysplastic lineages and blasts. Our data suggests that RING1A is deregulated in MDS and plays a role in the erythroid development defect

Different Gene Sets Are Associated With Azacitidine Response In Vitro Versus in Myelodysplastic Syndrome Patients

Myelodysplastic syndromes (MDS) are a heterogeneous group of hematopoietic disorders characterized by dysplasia, ineffective hematopoiesis, and predisposition to secondary acute myeloid leukemias (sAML). Azacitidine (AZA) is the standard care for high-risk MDS patients not eligible for allogenic bone marrow transplantation. However, only half of the patients respond to AZA and eventually all patients relapse. Response-predicting biomarkers and combinatorial drugs targets enhancing therapy response and its duration are needed. Here, we have taken a dual approach. First, we have evaluated genes encoding chromatin regulators for their capacity to modulate AZA response. We were able to validate several genes, whose genetic inhibition affected the cellular AZA response, including 4 genes encoding components of Imitation SWItch chromatin remodeling complex pointing toward a specific function and co-vulnerability. Second, we have used a classical cohort analysis approach measuring the expression of a gene panel in bone marrow samples from 36 MDS patients subsequently receiving AZA. The gene panel included the identified AZA modulators, genes known to be involved in AZA metabolism and previously identified candidate modulators. In addition to confirming a number of previously made observations, we were able to identify several new associations, such as NSUN3 that correlated with increased overall survival. Taken together, we have identified a number of genes associated with AZA response in vitro and in patients. These groups of genes are largely nonoverlapping suggesting that different gene sets need to be exploited for the development of combinatorial drug targets and response-predicting biomarkers

Diagnostic performance of four SARS-CoV-2 antibody assays in patients with COVID-19 or with bacterial and non-SARS-CoV-2 viral respiratory infections

SARS-CoV-2 antibody assays are used for epidemiological studies and for the assessment of vaccine responses in highly vulnerable patients. So far, data on cross-reactivity of SARS-CoV-2 antibody assays is limited. Here, we compared four enzyme-linked immunosorbent assays (ELISAs; Vircell SARS-CoV-2 IgM/IgA and IgG, Euroimmun SARS-CoV-2 IgA and IgG) for detection of anti-SARS-CoV-2 antibodies in 207 patients with COVID-19, 178 patients with serological evidence of different bacterial infections, 107 patients with confirmed viral respiratory disease, and 80 controls from the pre-COVID-19 era. In COVID-19 patients, the assays showed highest sensitivity in week 3 (Vircell-IgM/A and Euroimmun-IgA: 78.9% each) and after week 7 (Vircell-IgG: 97.9%; Euroimmun-IgG: 92.1%). The antibody indices were higher in patients with fatal disease. In general, IgM/IgA assays had only limited or no benefit over IgG assays. In patients with non-SARS-CoV-2 respiratory infections, IgG assays were more specific than IgM/IgA assays, and bacterial infections were associated with more false-positive results than viral infections. The specificities in bacterial and viral infections were 68.0 and 81.3% (Vircell-IgM/IgA), 84.8 and 96.3% (Euroimmun-IgA), 97.8 and 86.0% (Vircell-IgG), and 97.8 and 99.1% (Euroimmun-IgG), respectively. Sera from patients positive for antibodies against Mycoplasma pneumoniae, Chlamydia psittaci, and Legionella pneumophila yielded particularly high rates of unspecific false-positive results in the IgM/IgA assays, which was revealed by applying a highly specific flow-cytometric assay using HEK 293 T cells expressing the SARS-CoV-2 spike protein. Positive results obtained with anti-SARS-CoV-2 IgM/IgA ELISAs require careful interpretation, especially if there is evidence for prior bacterial respiratory infections

Inhibition of CBP synergizes with the RNA-dependent mechanisms of Azacitidine by limiting protein synthesis

The nucleotide analogue azacitidine (AZA) is currently the best treatment option for patients with high-risk myelodysplastic syndromes (MDS). However, only half of treated patients respond and of these almost all eventually relapse. New treatment options are urgently needed to improve the clinical management of these patients. Here, we perform a loss-of-function shRNA screen and identify the histone acetyl transferase and transcriptional co-activator, CREB binding protein (CBP), as a major regulator of AZA sensitivity. Compounds inhibiting the activity of CBP and the closely related p300 synergistically reduce viability of MDS-derived AML cell lines when combined with AZA. Importantly, this effect is specific for the RNA-dependent functions of AZA and not observed with the related compound decitabine that is only incorporated into DNA. The identification of immediate target genes leads us to the unexpected finding that the effect of CBP/p300 inhibition is mediated by globally down regulating protein synthesis. © 2021. The Author(s)

Inhibition of CBP synergizes with the RNA-dependent mechanisms of Azacitidine by limiting protein synthesis

This project was supported by the FEDER/Ministerio de Ciencia e Innovación - Agencia Estatal de Investigación through the grant PIE16/00011 RESPONSE (to M.B.), the European Research Council ERC-StG-336860 (to J.Z.), the grant Juan de la Cierva- Formación FJCI-2014-22983 (to J.D.), the grant Sara Borrell CD17/00084 (to J.D.), the Marie Skłodowska Curie Training network "ChroMe" H2020-MSCA-ITN-2015-675610 (to M.M.), the FPI predoctoral fellowship BES-2016-077251 (to M.M.L.P.), the SFB 1243 DFG (to K.S.G.), the Austrian Science Fund SFB-F4710 (to J.Z.) and the Deutsche José Carreras Leukämie Stiftung DJCLS 14R/2018 (to K.G. and M.B.). Research in the Buschbeck lab is further supported by the following grants: MINECO grant RTI2018-094005-B-I00 (to M.B.), the Marie Skłodowska Curie Training network 'INTERCEPT-MDS' H2020-MSCA-ITN-2015-953407 (to M.B. and K.S.G.); AGAUR 2017-SGR-305 (to M.B.) and Fundació La Marató de TV3 257/C/2019 (to M.B.). Research at IMP is supported by Boehringer Ingelheim, the Austrian Research Promotion Agency (Headquarter grant FFG-852936) and the Austrian Academy of Sciences. Research at the IJC is generously supported by the 'La Caixa' Foundation, the Fundació Internacional Josep Carreras, Celgene Spain and the CERCA Programme/Generalitat de Catalunya. A.G. received funds by "Agencia Estatal de Investigación" (AEI) through the Plan Nacional "Excelencia" grant number SAF2017-84301-P, by the "Associación Española Contra el Cancer" (AECC) grant number LABAE20040GENT and by the Agency for Management of University and Research Grants (AGAUR) of the Catalan Government grant 2017SGR01743. Proteomic analyses were performed in the IJC Proteomic Unit, which are part of Proteored PRB3 and are supported by grant PT17/0019 from the PE I + D + i 2013-2016, funded by ISCIII and ERDF. We thank Kaoru Tohyama for providing MDS-L cells, members of the Buschbeck lab, the RESPONSE network (PIE16/00011), Blanca Xicoy and Francesc Solé for valuable discussions. We thank the IJC Biobanking unit for sample preparation and storage, Bernat Cucurull from the IJC Proteomics unit for the ClickIT sample preparation, Marco Fernandez for advice and training in flow cytometry and all other staff of IJC and IGTP core facilities for excellent support.The nucleotide analogue azacitidine (AZA) is currently the best treatment option for patients with high-risk myelodysplastic syndromes (MDS). However, only half of treated patients respond and of these almost all eventually relapse. New treatment options are urgently needed to improve the clinical management of these patients. Here, we perform a loss-of-function shRNA screen and identify the histone acetyl transferase and transcriptional co-activator, CREB binding protein (CBP), as a major regulator of AZA sensitivity. Compounds inhibiting the activity of CBP and the closely related p300 synergistically reduce viability of MDS-derived AML cell lines when combined with AZA. Importantly, this effect is specific for the RNA-dependent functions of AZA and not observed with the related compound decitabine that is only incorporated into DNA. The identification of immediate target genes leads us to the unexpected finding that the effect of CBP/p300 inhibition is mediated by globally down regulating protein synthesis