Overview on WHO-HAEM5 and the diagnostic relevance of genetic alterations for the classification.

The landscape of haematological malignancies is constantly evolving, driven by advances in our understanding of their genetic basis. This has cumulated within the 5th Edition of the World Health Organization (WHO) Classification of Haematolymphoid Tumours published in short form in 2022 [1, 2] and being available in full length both as "Blue Book" (in print expected early 2024) as well as web-based classification (see: https://tumourclassification.iarc.who.int/welcome/). Similarly, the importance of genetic alterations for the classification is highlighted in other classification systems related to haematologic neoplasms [3-5]. In this special issue of the Medizinische Genetik, we present a comprehensive overview of the genetic alterations contributing to the classification of haematolymphoid neoplasms in the 5th Edition of the WHO classification (WHO-HAEM5) and its diagnostic relevance in the context of various haematological malignancies.

Genetic studies in clonal haematopoiesis, myelodysplastic neoplasms and acute myeloid leukaemia - a practical guide to WHO-HAEM5.

In recent years, technology developments and increase in knowledge have led to profound changes in the diagnostics of haematologic neoplasms, particularly myeloid neoplasms. Therefore an updated, fifth edition of the World Health Organization (WHO) classification of haematolymphoid neoplasms (WHO-HAEM5) will be issued in 2024. In this context, we present a practical guide for analysing the genetic aspects of clonal haematopoiesis of indeterminate potential (CHIP), clonal cytopenia of undetermined significance (CCUS), myelodysplastic neoplasms (MDS), and acute myeloid leukaemia (AML) based on WHO-HAEM5. This guide navigates through the genetic abnormalities underlying myeloid neoplasms which are required to be detected for classification according to WHO-HAEM5 and provides diagnostic algorithms.

Personalized Timing for Allogeneic Stem-Cell Transplantation in Hematologic Neoplasms: A Target Trial Emulation Approach Using Multistate Modeling and Microsimulation.

PURPOSE: Decision about the optimal timing of a treatment procedure in patients with hematologic neoplasms is critical, especially for cellular therapies (most including allogeneic hematopoietic stem-cell transplantation [HSCT]). In the absence of evidence from randomized trials, real-world observational data become beneficial to study the effect of the treatment timing. In this study, a framework to estimate the expected outcome after an intervention in a time-to-event scenario is developed, with the aim of optimizing the timing in a personalized manner. METHODS: Retrospective real-world data are leveraged to emulate a target trial for treatment timing using multistate modeling and microsimulation. This case study focuses on myelodysplastic syndromes, serving as a prototype for rare cancers characterized by a heterogeneous clinical course and complex genomic background. A cohort of 7,118 patients treated according to conventional available treatments/evidence across Europe and United States is analyzed. The primary clinical objective is to determine the ideal timing for HSCT, the only curative option for these patients. RESULTS: This analysis enabled us to identify the most appropriate time frames for HSCT on the basis of each patient's unique profile, defined by a combination relevant patients' characteristics. CONCLUSION: The developed methodology offers a structured framework to address a relevant clinical issue in the field of hematology. It makes several valuable contributions: (1) novel insights into how to develop decision models to identify the most favorable HSCT timing, (2) evidence to inform clinical decisions in a real-world context, and (3) the incorporation of complex information into decision making. This framework can be applied to provide medical insights for clinical issues that cannot be adequately addressed through randomized clinical trials.

Clinical and Genomic-Based Decision Support System to Define the Optimal Timing of Allogeneic Hematopoietic Stem-Cell Transplantation in Patients With Myelodysplastic Syndromes.

PURPOSE: Allogeneic hematopoietic stem-cell transplantation (HSCT) is the only potentially curative treatment for patients with myelodysplastic syndromes (MDS). Several issues must be considered when evaluating the benefits and risks of HSCT for patients with MDS, with the timing of transplantation being a crucial question. Here, we aimed to develop and validate a decision support system to define the optimal timing of HSCT for patients with MDS on the basis of clinical and genomic information as provided by the Molecular International Prognostic Scoring System (IPSS-M). PATIENTS AND METHODS: We studied a retrospective population of 7,118 patients, stratified into training and validation cohorts. A decision strategy was built to estimate the average survival over an 8-year time horizon (restricted mean survival time [RMST]) for each combination of clinical and genomic covariates and to determine the optimal transplantation policy by comparing different strategies. RESULTS: Under an IPSS-M based policy, patients with either low and moderate-low risk benefited from a delayed transplantation policy, whereas in those belonging to moderately high-, high- and very high-risk categories, immediate transplantation was associated with a prolonged life expectancy (RMST). Modeling decision analysis on IPSS-M versus conventional Revised IPSS (IPSS-R) changed the transplantation policy in a significant proportion of patients (15% of patient candidate to be immediately transplanted under an IPSS-R-based policy would benefit from a delayed strategy by IPSS-M, whereas 19% of candidates to delayed transplantation by IPSS-R would benefit from immediate HSCT by IPSS-M), resulting in a significant gain-in-life expectancy under an IPSS-M-based policy (P = .001). CONCLUSION: These results provide evidence for the clinical relevance of including genomic features into the transplantation decision making process, allowing personalizing the hazards and effectiveness of HSCT in patients with MDS.

A case of familial donor-derived acute myeloid leukemia with underlying pre-leukemic mutations.

Not available.

Analysis of 3760 hematologic malignancies reveals rare transcriptomic aberrations of driver genes.

BACKGROUND: Rare oncogenic driver events, particularly affecting the expression or splicing of driver genes, are suspected to substantially contribute to the large heterogeneity of hematologic malignancies. However, their identification remains challenging. METHODS: To address this issue, we generated the largest dataset to date of matched whole genome sequencing and total RNA sequencing of hematologic malignancies from 3760 patients spanning 24 disease entities. Taking advantage of our dataset size, we focused on discovering rare regulatory aberrations. Therefore, we called expression and splicing outliers using an extension of the workflow DROP (Detection of RNA Outliers Pipeline) and AbSplice, a variant effect predictor that identifies genetic variants causing aberrant splicing. We next trained a machine learning model integrating these results to prioritize new candidate disease-specific driver genes. RESULTS: We found a median of seven expression outlier genes, two splicing outlier genes, and two rare splice-affecting variants per sample. Each category showed significant enrichment for already well-characterized driver genes, with odds ratios exceeding three among genes called in more than five samples. On held-out data, our integrative modeling significantly outperformed modeling based solely on genomic data and revealed promising novel candidate driver genes. Remarkably, we found a truncated form of the low density lipoprotein receptor LRP1B transcript to be aberrantly overexpressed in about half of hairy cell leukemia variant (HCL-V) samples and, to a lesser extent, in closely related B-cell neoplasms. This observation, which was confirmed in an independent cohort, suggests LRP1B as a novel marker for a HCL-V subclass and a yet unreported functional role of LRP1B within these rare entities. CONCLUSIONS: Altogether, our census of expression and splicing outliers for 24 hematologic malignancy entities and the companion computational workflow constitute unique resources to deepen our understanding of rare oncogenic events in hematologic cancers.

The E592K variant of SF3B1 creates unique RNA missplicing and associates with high-risk MDS without ring sideroblasts.

Among the most common genetic alterations in the myelodysplastic syndromes (MDS) are mutations in the spliceosome gene SF3B1. Such mutations induce specific RNA missplicing events, directly promote ring sideroblast (RS) formation, and generally associate with more favorable prognosis. However, not all SF3B1 mutations are the same, and little is known about how distinct hotspots influence disease. Here we report that the E592K variant of SF3B1 associates with high-risk disease features in MDS, including a lack of RS, increased myeloblasts, a distinct co-mutation pattern, and a lack of the favorable survival seen with other SF3B1 mutations. Moreover, compared to other hotspot SF3B1 mutations, E592K induces a unique RNA missplicing pattern, retains an interaction with the splicing factor SUGP1, and preserves normal RNA splicing of the sideroblastic anemia genes TMEM14C and ABCB7. These data have implications for our understanding of the functional diversity of spliceosome mutations, as well as the pathobiology, classification, prognosis, and management of SF3B1-mutant MDS.

Hyperactive STAT5 hijacks T cell receptor signaling and drives immature T cell acute lymphoblastic leukemia.

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive immature T cell cancer. Mutations in IL7R have been analyzed genetically, but downstream effector functions such as STAT5A and STAT5B hyperactivation are poorly understood. Here, we studied the most frequent and clinically challenging STAT5BN642H driver in T cell development and immature T cell cancer onset and compared it with STAT5A hyperactive variants in transgenic mice. Enhanced STAT5 activity caused disrupted T cell development and promoted an early T cell progenitor-ALL phenotype, with upregulation of genes involved in T cell receptor (TCR) signaling, even in absence of surface TCR. Importantly, TCR pathway genes were overexpressed in human T-ALL and mature T cell cancers and activation of TCR pathway kinases was STAT5 dependent. We confirmed STAT5 binding to these genes using ChIP-Seq analysis in human T-ALL cells, which were sensitive to pharmacologic inhibition by dual STAT3/5 degraders or ZAP70 tyrosine kinase blockers in vitro and in vivo. We provide genetic and biochemical proof that STAT5A and STAT5B hyperactivation can initiate T-ALL through TCR pathway hijacking and suggest similar mechanisms for other T cell cancers. Thus, STAT5 or TCR component blockade are targeted therapy options, particularly in patients with chemoresistant clones carrying STAT5BN642H.

Classical meets malignant hematology: a case of acquired εγδß-thalassemia in clonal hematopoiesis.

Hemoglobinopathies including thalassemias are among the most frequent genetic disorders worldwide. Primarily, these entities result from germline variants in the globin gene clusters and their cis-acting regulatory elements, and thus the WHO classifies thalassemias as inherited diseases. Non-inherited disorders of globin chain synthesis mimicking the phenotype of thalassemias have also been described and are referred to as acquired thalassemias. These forms mainly affect the alpha-globin genes and are observed at much lower frequencies...

Molecular and clinical analyses of PHF6 mutant myeloid neoplasia provide their pathogenesis and therapeutic targeting.

PHF6 mutations (PHF6MT) are identified in various myeloid neoplasms (MN). However, little is known about the precise function and consequences of PHF6 in MN. Here we show three main findings in our comprehensive genomic and proteomic study. Firstly, we show a different pattern of genes correlating with PHF6MT in male and female cases. When analyzing male and female cases separately, in only male cases, RUNX1 and U2AF1 are co-mutated with PHF6. In contrast, female cases reveal co-occurrence of ASXL1 mutations and X-chromosome deletions with PHF6MT. Next, proteomics analysis reveals a direct interaction between PHF6 and RUNX1. Both proteins co-localize in active enhancer regions that define the context of lineage differentiation. Finally, we demonstrate a negative prognostic role of PHF6MT, especially in association with RUNX1. The negative effects on survival are additive as PHF6MT cases with RUNX1 mutations have worse outcomes when compared to cases carrying single mutation or wild-type.

Prognostic impact of CEBPA mutational subgroups in adult AML.

Despite recent refinements in the diagnostic and prognostic assessment of CEBPA mutations in AML, several questions remain open, i.e. implications of different types of basic region leucin zipper (bZIP) mutations, the role of co-mutations and the allelic state. Using pooled primary data analysis on 1010 CEBPA-mutant adult AML patients, a comparison was performed taking into account the type of mutation (bZIP: either typical in-frame insertion/deletion (InDel) mutations (bZIPInDel), frameshift InDel or nonsense mutations inducing translational stop (bZIPSTOP) or single base-pair missense alterations (bZIPms), and transcription activation domain (TAD) mutations) and the allelic state (single (smCEBPA) vs. double mutant (dmCEBPA)). Only bZIPInDel patients had significantly higher rates of complete remission and longer relapse free and overall survival (OS) compared with all other CEBPA-mutant subgroups. Moreover, co-mutations in bZIPInDel patients (e.g. GATA2, FLT3, WT1 as well as ELN2022 adverse risk aberrations) had no independent impact on OS, whereas in non-bZIPInDel patients, grouping according to ELN2022 recommendations added significant prognostic information. In conclusion, these results demonstrate bZIPInDel mutations to be the major independent determinant of outcome in CEBPA-mutant AML, thereby refining current classifications according to WHO (including all dmCEBPA and smCEBPA bZIP) as well as ELN2022 and ICC recommendations (including CEBPA bZIPms).

Comparing malignant monocytosis across the updated WHO and ICC classifications of 2022.

ABSTRACT: The World Health Organization (WHO) classification of hematolymphoid tumors and the International Consensus Classification (ICC) of 2022 introduced major changes to the definition of chronic myelomonocytic leukemia (CMML). To assess its qualitative and quantitative implications for patient care, we started with 3311 established CMML cases (according to WHO 2017 criteria) and included 2130 oligomonocytosis cases fulfilling the new CMML diagnostic criteria. Applying both 2022 classification systems, 356 and 241 of oligomonocytosis cases were newly classified as myelodysplastic (MD)-CMML (WHO and ICC 2022, respectively), most of which were diagnosed as myelodysplastic syndrome (MDS) according to the WHO 2017 classification. Importantly, 1.5 times more oligomonocytosis cases were classified as CMML according to WHO 2022 than based on ICC, because of different diagnostic criteria. Genetic analyses of the newly classified CMML cases showed a distinct mutational profile with strong enrichment of MDS-typical alterations, resulting in a transcriptional subgroup separated from established MD and myeloproliferative CMML. Despite a different cytogenetic, molecular, immunophenotypic, and transcriptional landscape, no differences in overall survival were found between newly classified and established MD-CMML cases. To the best of our knowledge, this study represents the most comprehensive analysis of routine CMML cases to date, both in terms of clinical characterization and transcriptomic analysis, placing newly classified CMML cases on a disease continuum between MDS and previously established CMML.

Collaborative effect of Csnk1a1 haploinsufficiency and mutant p53 in Myc induction can promote leukemic transformation.

ABSTRACT: It is still not fully understood how genetic haploinsufficiency in del(5q) myelodysplastic syndrome (MDS) contributes to malignant transformation of hematopoietic stem cells. We asked how compound haploinsufficiency for Csnk1a1 and Egr1 in the common deleted region on chromosome 5 affects hematopoietic stem cells. Additionally, Trp53 was disrupted as the most frequently comutated gene in del(5q) MDS using CRISPR/Cas9 editing in hematopoietic progenitors of wild-type (WT), Csnk1a1-/+, Egr1-/+, Csnk1a1/Egr1-/+ mice. A transplantable acute leukemia only developed in the Csnk1a1-/+Trp53-edited recipient. Isolated blasts were indefinitely cultured ex vivo and gave rise to leukemia after transplantation, providing a tool to study disease mechanisms or perform drug screenings. In a small-scale drug screening, the collaborative effect of Csnk1a1 haploinsufficiency and Trp53 sensitized blasts to the CSNK1 inhibitor A51 relative to WT or Csnk1a1 haploinsufficient cells. In vivo, A51 treatment significantly reduced blast counts in Csnk1a1 haploinsufficient/Trp53 acute leukemias and restored hematopoiesis in the bone marrow. Transcriptomics on blasts and their normal counterparts showed that the derived leukemia was driven by MAPK and Myc upregulation downstream of Csnk1a1 haploinsufficiency cooperating with a downregulated p53 axis. A collaborative effect of Csnk1a1 haploinsufficiency and p53 loss on MAPK and Myc upregulation was confirmed on the protein level. Downregulation of Myc protein expression correlated with efficient elimination of blasts in A51 treatment. The "Myc signature" closely resembled the transcriptional profile of patients with del(5q) MDS with TP53 mutation.

Tumor necrosis factor α promotes clonal dominance of KIT D816V+ cells in mastocytosis: role of survivin and impact on prognosis.

ABSTRACT: Systemic mastocytosis (SM) is defined by the expansion and accumulation of neoplastic mast cells (MCs) in the bone marrow (BM) and extracutaneous organs. Most patients harbor a somatic KIT D816V mutation, which leads to growth factor-independent KIT activation and accumulation of MC. Tumor necrosis factor α (TNF) is a proapoptotic and inflammatory cytokine that has been implicated in the clonal selection of neoplastic cells. We found that KIT D816V increases the expression and secretion of TNF. TNF expression in neoplastic MCs is reduced by KIT-targeting drugs. Similarly, knockdown of KIT or targeting the downstream signaling cascade of MAPK and NF-κB signaling reduced TNF expression levels. TNF reduces colony formation in human BM cells, whereas KIT D816V+ cells are less susceptible to the cytokine, potentially contributing to clonal selection. In line, knockout of TNF in neoplastic MC prolonged survival and reduced myelosuppression in a murine xenotransplantation model. Mechanistic studies revealed that the relative resistance of KIT D816V+ cells to TNF is mediated by the apoptosis-regulator BIRC5 (survivin). Expression of BIRC5 in neoplastic MC was confirmed by immunohistochemistry of samples from patients with SM. TNF serum levels are significantly elevated in patients with SM and high TNF levels were identified as a biomarker associated with inferior survival. We here characterized TNF as a KIT D816V-dependent cytokine that promotes clonal dominance. We propose TNF and apoptosis-associated proteins as potential therapeutic targets in SM.