Teodora Kuzmanovic

Dynamics of clonal evolution in myelodysplastic syndromes

To elucidate differential roles of mutations in myelodysplastic syndromes (MDS), we investigated clonal dynamics using whole-exome and/or targeted sequencing of 699 patients, of whom 122 were analyzed longitudinally. Including the results from previous reports, we assessed a total of 2,250 patients for mutational enrichment patterns. During progression, the number of mutations, their diversity and clone sizes increased, with alterations frequently present in dominant clones with or without their sweeping previous clones. Enriched in secondary acute myeloid leukemia (sAML; in comparison to high-risk MDS), FLT3, PTPN11, WT1, IDH1, NPM1, IDH2 and NRAS mutations (type 1) tended to be newly acquired, and were associated with faster sAML progression and a shorter overall survival time. Significantly enriched in high-risk MDS (in comparison to low-risk MDS), TP53, GATA2, KRAS, RUNX1, STAG2, ASXL1, ZRSR2 and TET2 mutations (type 2) had a weaker impact on sAML progression and overall survival than type-1 mutations. The distinct roles of type-1 and type-2 mutations suggest their potential utility in disease monitoring.

Ubiquitination of the Transcription Factor IRF-3 Activates RIPA, the Apoptotic Pathway that Protects Mice from Viral Pathogenesis

The transcription factor IRF-3 mediates cellular antiviral response by inducing the expression of interferon and other antiviral proteins. In RNA-virus infected cells, IRF-3s transcriptional activation is triggered primarily by RIG-I-like receptors (RLR), which can also activate the RLR-induced IRF-3-mediated pathway of apoptosis (RIPA). Here, we have reported that the pathway of IRF-3 activation in RIPA was independent of and distinct from the known pathway of transcriptional activation of IRF-3. It required linear polyubiquitination of two specific lysine residues of IRF-3 by LUBAC, the linear polyubiquitinating enzyme complex, which bound IRF-3 in signal-dependent fashion. To evaluate the role of RIPA in viral pathogenesis, we engineered a genetically targeted mouse, which expressed a mutant IRF-3 that was RIPA-competent but transcriptionally inert; this single-action IRF-3 could protect mice from lethal viral infection. Our observations indicated that IRF-3-mediated apoptosis of virus-infected cells could be an effective antiviral mechanism, without expression of the interferon-stimulated genes.

Genomic determinants of chronic myelomonocytic leukemia

The biology, clinical phenotype and progression rate of chronic myelomonocytic leukemia (CMML) are highly variable due to diverse initiating and secondary clonal genetic events. To determine the effects of molecular features including clonal hierarchy in CMML, we studied whole-exome and targeted next-generation sequencing data from 150 patients with robust clinical and molecular annotation assessed cross-sectionally and at serial time points of disease evolution. To identify molecular lesions unique to CMML, we compared it to the related myeloid neoplasms (N=586), including juvenile myelomonocytic leukemia, myelodysplastic syndromes (MDS) and primary monocytic acute myeloid leukemia and discerned distinct molecular profiles despite similar pathomorphological features. Within CMML, mutations in certain pathways correlated with clinical classification, for example, proliferative vs dysplastic features. While most CMML patients (59%) had ancestral (dominant/co-dominant) mutations involving TET2, SRSF2 or ASXL1 genes, secondary subclonal hierarchy correlated with clinical phenotypes or outcomes. For example, progression was associated with acquisition of new expanding clones carrying biallelic TET2 mutations or RAS family, or spliceosomal gene mutations. In contrast, dysplastic features correlated with mutations usually encountered in MDS (for example, SF3B1 and U2AF1). Classification of CMML based on hierarchies of ancestral and subclonal mutational events may correlate strongly with clinical features and prognosis.

Origins of myelodysplastic syndromes after aplastic anemia

To the editor: The course of aplastic anemia (AA) is often complicated by the development of clonal disorders such as paroxysmal nocturnal hemoglobinuria (PNH) and secondary myelodysplastic syndromes (sMDS).[1][1][⇓][2][⇓][3][⇓][4]-[5][5] Identification of patients at risk for development of

A new mechanism of interferon’s antiviral action: Induction of autophagy, essential for paramyxovirus replication, is inhibited by the interferon stimulated gene, TDRD7

The interferon (IFN) system represents the first line of defense against a wide range of viruses. Virus infection rapidly triggers the transcriptional induction of IFN-β and IFN Stimulated Genes (ISGs), whose protein products act as viral restriction factors by interfering with specific stages of virus life cycle, such as entry, transcription, translation, genome replication, assembly and egress. Here, we report a new mode of action of an ISG, IFN-induced TDRD7 (tudor domain containing 7) inhibited paramyxovirus replication by inhibiting autophagy. TDRD7 was identified as an antiviral gene by a high throughput screen of an ISG shRNA library for blocking IFN’s protective effect against Sendai virus (SeV) replication. The antiviral activity of TDRD7 against SeV, human parainfluenza virus 3 and respiratory syncytial virus was confirmed by its genetic ablation or ectopic expression in several types of mouse and human cells. TDRD7’s antiviral action was mediated by its ability to inhibit autophagy, a cellular catabolic process which was robustly induced by SeV infection and required for its replication. Mechanistic investigation revealed that TDRD7 interfered with the activation of AMP-dependent kinase (AMPK), an enzyme required for initiating autophagy. AMPK activity was required for efficient replication of several paramyxoviruses, as demonstrated by its genetic ablation or inhibition of its activity by TDRD7 or chemical inhibitors. Therefore, our study has identified a new antiviral ISG with a new mode of action.

Distinct clinical and biological implications of various DNMT3A mutations in myeloid neoplasms

Distinct clinical and biological implications of various DNMT3A mutations in myeloid neoplasms

Deletion of Ptpn1 induces myeloproliferative neoplasm

Abstract Leukemia accepted article preview online, 23 January 2017. doi:10.1038/leu.2017.31.

Distinct Genomic Associations to Predict Acute Myeloid Leukemia (AML) Progression from Myelodysplastic Syndromes (MDS)

Approximately 25% of MDS patients (pts) progress to AML; 15% of those with lower-risk, and up to 40-50% of those with higher-risk disease. Predicting those who are likely to progress to AML early in their disease course could directly impact treatment decisions: lower-risk pts with higher risk of AML transformation may be offered transplant, while higher-risk pts may be considered for AML-like therapy. In this study, we developed a genomic model that evaluates mutational patterns and their association with AML progression. Development of this model mimics Netflix or Amazon9s recommender system in which customers who bought products A and B, are likely to buy C: pts who have a mutation in genes A and B, are then likely to progress to AML. Clinical and genomic data ofMDS pts diagnosed between 1/1996 and 9/2016 were analyzed. A panel of 60 gene mutations obtained by next generation targeted deep sequencing was included. Association rules using Apriori algorithm was used to study the relationship between multiple genes/cytogenetic abnormalities and AML progression in an unbiased approach. Association rules are a machine learning method that uncovers relationships between variables in a given dataset. Rules with highest confidence (>90% that an association exists) and highest lift (strength of the association) were chosen. Univariate and multivariate analyses were used to evaluate the impact of mutations on AML progression, adjusting for covariates (age, IPSS-R). A total of 527 pts with lower-/higher-risk MDS were analyzed, of those 105 (20%) progressed to AML, with a median time to progression (TTP) of 13 months (interquartile range, 6.3-29.8). Median age at diagnosis was 67 years (range, 19-99) and 38% were female. Risk categories by IPSS-R included: 78 (15%) very low, 200 (38%) low, 95 (18%) intermediate, 98 (18.5%) high, and 56 (10.5%) very high; IPSS-R cytogenetic risk groups were 15 (3%) very good, 331 (63%) good, 87 (16%) intermediate, 37 (7%) poor, and 57 (11%) very poor. The most commonly mutated genes were SF3B1 (14%), ASXL1 (13%), TET2 (12%), SRSF2 (11%), DNMT3A (10%), STAG2 (9%), and TP53 (8%). Per IPSS-R, 14% of pts with very low/low, 21% with intermediate and 30% with high/very high categories progressed to AML. Univariate analyses identified the following cytogenetic and mutational abnormalities to be associated with AML progression: complex karyotype (CK, P =.019), ASXL1 ( P =.023), DNMT3A ( P =.011), FLT3 ( P =.031), PHF6 ( P =.018), PTPN11 ( P =.037) , RUNX1 ( P =.001) , STAG2 ( P =.023) , TET2 ( P =.047) , U2AF1 ( P =.038), and ≥3 gene mutations( P =.002), whereas SF3B1 was associated with lower risk of AML progression ( P =.025). In multivariate Cox regression analyses, including significant mutations, age ( P =.5) and IPSS-R scores ( P RUNX1 (HR2.8 ; 95% CI 1.6-5.3; P= .001), and ≥3 gene mutations (HR=1.65; 95% CI 1.1-2.5; P= .021) were independent factors that associated with AML progression, while SF3B1 (HR=.417; 95% CI .205-.851, P =.016) was associated with lower risk of progression. Association rules identified the following combinations of cytogenetic/genomic abnormalities for AML progression: (ASXL1, RUNX1, STAG2/BCOR), (ASXL1, STAG2, ZRSR2/SRSF2), (ASXL1, TET2, Trisomy8 ), (RUNX1, STAG2, Trisomy 8 ), ( CK , TP53, TET2) . These combinations were associated with a 5 month reduction in time to AML progression.[Figure 1] Using genomic data to reliably identify patients at highest risk for progression to AML early in their disease course can dramatically alter treatment recommendations. Independent factors such as RUNX1 and ≥3 gene mutations predicted for AML progression, while SF3B1 was associated with a reduced risk. Certain cytogenetic/genomic abnormalities were associated with shorter time to AML progression. This data suggests that MDS pts with ≥3 genomic abnormalities or certain genomic combinations could be offered more aggressive therapy early in their disease course including those with lower risk disease. Disclosures Sekeres: Celgene: Membership on an entity9s Board of Directors or advisory committees. Gerds: Incyte: Consultancy; CTI BioPharma: Consultancy. Advani: Takeda/ Millenium: Research Funding; Pfizer: Consultancy. Maciejewski: Alexion Pharmaceuticals, Inc.: Consultancy, Membership on an entity9s Board of Directors or advisory committees, Other: Speaker Fees; Apellis Pharmaceuticals: Consultancy; Ra Pharma: Consultancy.

Germline loss of function SAMD9 and SAMD9L alterations in adult myelodysplastic syndromes

TO THE EDITOR: Familial bone marrow failure (BMF) syndromes present typically in children and younger adults.[1][1][⇓][2]-[3][3] A number of germline (GL) mutations in genes such as DDX41 ,[4][4] RUNX1 ,[5][5] ETV6 ,[6][6] GATA2 ,[7][7] and ANKRD26 [8][8] have been implicated in the pathogenesis