Selecting eligible allogeneic haematopoietic stem cell transplantation (allo-HSCT) candidates with low-risk and intermediate-risk myelodysplastic syndrome (MDS) remains controversial. The International Working Group (IWG) for MDS prognosis identified a Revised International Prognostic Scoring System (IPSS-R) score >3.5 for benefits from early transplantation; the MDS-RIGHT group uses a broader set of poor risk features. We analysed 1145 lower risk MDS (lower risk and intermediate-risk IPSS-R) patients aged <75 years, using real-world European Myelodysplastic Syndromes registry data and identifying those meeting IWG or MDS-RIGHT criteria for allo-HSCT at baseline and 6-month follow-up. Fit patients were characterised by Karnofsky score ≥70 and Haematopoietic Cell Transplantation-specific Comorbidity Index <3. We evaluated clinical outcomes of transplant candidates, including survival, disease progression risk, new comorbidity risk and performance status decline. The IWG criterion, and not MDS-RIGHT criteria, identified patients with lower risk and intermediate-risk MDS with poorer baseline survival. Fit lower risk patients showed 2-year cumulative risks of incident comorbidity and performance status deterioration of 20% and 5% respectively. In summary, IWG and MDS-RIGHT features identify patients with lower or intermediate-risk MDS as candidates for early transplantation. Lower risk patients fit for transplantation have a cumulative incidence of adverse outcomes possibly jeopardising transplantation eligibility and should be carefully selected when planning delayed transplantation strategies.

Monoclonal gammopathy of undetermined significance (MGUS) is a common plasma cell disorder with well described risks of progression to myeloma and lymphoplasmacytic lymphoma. Using data from an established UK population-based cohort of hematological malignancies and premalignancies, we investigated patient and disease characteristics, subsequent hematological malignancy, and survival in 4,651 people diagnosed with MGUS between 2005 and 2019. The 5-year net (relative) survival (disease-specific estimate of the probability of survival) for MGUS patients was 87.8% (95% confidence interval [CI]: 85.9-89.7), with males (83.8%; 95% CI: 81.0-86.6) more affected than females (92.2%; 95% CI: 89.7-94.7). The proportion of subsequent hematological malignancies was also higher in males than females (8.8% vs. 5.3%; P<0.00001); the average annual rates of transition being 1.81% (95% CI: 1.44-2.18) and 0.99% (95% CI: 0.72-1.27), respectively. Furthermore, whilst annual rates of transformation to myeloma (1.04%) and lymphoplasmacytic lymphoma (0.11%) were as expected, both were higher in males (1.23% and 0.18%) than females (0.87% and 0.06%). With a median time to diagnosis of 40 months, the incidence of myeloid malignancy was also raised in males (relative risk=3.6; 95% CI: 2.5-4.9), but not females (relative risk=1.0; 95% CI: 0.3-1.9). No associations between MGUS and subsequent development of chronic lymphocytic leukemia were observed. Providing new data on the nature of MGUS progression, our analyses revealed previously undescribed sex disparities; including worse survival and increased rates of myeloid malignancy in males with non-IgM MGUS. These findings have implications for future research, as well as risk stratification and monitoring of patients with this highly prevalent plasma cell dyscrasia.

Abstract Dysregulated innate immunity contributes to clonal cytopenias and myeloid neoplasms, but its extent across disease stages and clinical relevance remain incompletely defined. We analyzed plasma ASC/NLRP3 double‐positive (DP) specks, ASC single‐positive (SP) specks, and 45 cytokines in 223 patients with idiopathic cytopenias of undetermined significance (ICUS)/clonal cytopenias of undetermined significance (CCUS), myelodysplastic syndromes (MDS), and chronic myelomonocytic leukemia (CMML) and 39 matched non‐inflammatory controls using adjusted regression, survival modeling, and paired longitudinal analyses. Inflammasome activation and cytokine perturbations were evident across the disease spectrum. DP‐ASC specks were elevated in MDS and CMML, whereas SP‐ASC specks were increased across all groups, indicating activation of ASC‐containing inflammasomes beyond NLRP3. Cytokines followed a graded ICUS → MDS → CMML pattern, with widespread upregulation of interleukins and chemokines (including IL‐7, IL‐8, IL‐11/CXCL11, and CCL7) alongside suppression of stem and progenitor support factors such as CSF3, FLT3LG, TRAIL, and TWEAK. At baseline, elevated IL‐15 and MMP1 predicted progression to acute myeloid Leukaemia, while higher IL‐10, CXCL8, and IL‐18 were associated with reduced survival; ASC specks were not independently prognostic. Longitudinal increases in selected cytokines distinguished progressors (area under the curve 0.82; 95% CI: 0.49–1.0). Cytokine patterns correlated with mutation categories, with the isolated SF3B1 mutation associated with higher DP‐ASC specks. These findings define early and progressive inflammasome engagement and nominate dynamic cytokine panels and the inflammasome–IL‐1 axis as actionable biomarkers and therapeutic targets.

When the World Health Organization (WHO) declared the global COVID-19 pandemic, it was recognised that patients with haematological cancers would be more susceptible to severe disease. Set within a UK population of ~4 million (https://hmrn.org/), all patients diagnosed with haematological neoplasms 2005-2019 who were alive on 1 January 2020 were followed up until March 2023. For comparative purposes, a similar age- and sex-matched general population cohort was also constructed. COVID-19 deaths were classified using ICD-10 codes and a multiple cause of death analysis was undertaken using a competing risk approach. Deaths of 486/18 883 haematology patients were attributed to COVID-19, yielding a cumulative incidence of 2.59% (95% confidence interval [CI]: 2.37-2.82) that was significantly higher than that of the general population (1.65; 95% CI: 1.58-1.72). In both cohorts, risks were higher in men, older people and those with co-morbidities. Within the patient cohort, excess mortality was largely concentrated in those suffering from more indolent conditions. Patients with the premalignancy MBL suffered from the highest excess mortality in the early phase of the pandemic when, unlike patients with malignancies, they were not advised to shield. Effects of shielding were evident and a clear vaccination benefit was demonstrated, with the exception of CLL and MCL; findings that warrant consideration in relation to other viruses.

BACKGROUND:In our previous study on erythropoiesis-stimulating agent (ESA) treatment in lower risk myelodysplastic syndromes from the European MDS (EUMDS) Registry, we showed that patients treated with ESAs had longer survival compared with patients who receive red blood cell transfusion (RBCT). In this study, with a longer follow up time and more patients included, we aimed to assess long-term effects on survival and health-related quality of life (HRQoL) of exposure to ESAs with or without RBCT in patients with lower risk myelodysplastic syndromes. METHODS:The EUMDS Registry is a non-interventional, longitudinal, real-world registry prospectively enrolling newly diagnosed patients older than 18 years with lower risk (International Prognostic Scoring System low or intermediate-1) myelodysplastic syndromes from 16 European countries and Israel. The analysis was restricted to patients with haemoglobin concentrations less than 100 g/L enrolled between Jan 1, 2008, and July 1, 2019, with last censoring of data on Dec 31, 2021. Patient management was recorded every 6 months, including treatment, transfusions, and HRQoL. ESA treatment followed local guidelines. The patients were separated into four groups at each study visit: no ESA or RBCT, ESA only, ESA plus RBCT, and RBCT only. The data were analysed longitudinally over time according to ESA and RBCT status during each 6-month interval, using propensity score matching. The main outcomes were median overall survival and leukaemia-free survival, and HRQoL. This study is registered with ClinicalTrials.gov, NCT00600860, as is ongoing. FINDINGS:2448 patients (the ESA-unexposed group [n=1265] and ESA-exposed group [n=1183]) were diagnosed before July 1, 2019; 1520 (62·1%) were male and 928 (37·9%) were female. Median follow-up time was 3·9 years (IQR 1·6-6·5). After applying eligibility criteria and propensity matching, there were 426 patients in the ESA-unexposed group and 744 patients in the ESA-exposed group. Median overall survival in the ESA exposed group was 44·9 months (95% CI 40·2-50·5) compared with 34·8 months (28·6-39·2) in the ESA unexposed group; the absolute difference was 10·1 months (95% CI 2·2-18·0; hazard ratio [HR] 0·70 [95% CI 0·59-0·83]; p<0·0001). Patients without RBCT in the presence or absence of ESA exposure maintained significantly better HRQoL than those with RBCT, irrespective of ESA exposure (linear mixed effect model of EQ-5d-3L index score, RBCT coefficient -0·04 [95% CI -0·06 to 0·03], p<0·0001; linear mixed effect model of VAS, -4·57 [-6·02 to -3·13], p<0·0001). INTERPRETATION:ESA treatment in patients with lower risk myelodysplastic syndromes significantly improves overall survival when started before or early after the onset of regular transfusion therapy. Avoiding RBCT is associated with significantly better HRQoL. FUNDING:H2020 European Research Council, Novartis Pharmacy B V Oncology Europe, Amgen, BMS/Celgene International, Janssen Pharmaceutica, Takeda Pharmaceuticals International, and Gilead Sciences.

Introduction The distinction between myelodysplastic neoplasms (MDS) and acute myeloid leukaemia (AML) is becoming increasingly blurred particularly at the arbitrary 20% blast cut-off at which AML is traditionally diagnosed. There is a move towards lowering this cut-off to 10% blasts allowing patients within the 10-20% range to access both AML and MDS therapies. There is however a clinical need for more precise methods to classify patients on the continuum between these diseases to inform treatment decisions. Using clustering techniques applied to targeted sequencing data derived from a large unselected population-based cohort we investigated whether molecular subtypes could be identified across AML and MDS which could better direct therapeutic interventions and improve outcomes. Methods Patients diagnosed with AML and MDS between 2004-2012 from a catchment population of ∼4 million (14 centres) were sequenced with a targeted 293-gene panel. A total of 1085 patients with complete genetic and cytogenetic data were available for analysis. Patients with previously well-defined good risk translocations were excluded from further analysis leaving a final cohort of 1007 patients (AML=445, MDS=561). A total of 69 genetic features (both mutational and structural) were analysed, which were either detected in at least 15 of the 1000 analysed patients or of established significance in myeloid neoplasm. To identify genetic subgroups, the data were modelled as a finite mixture of Bernoulli distributions, providing a data-driven probabilistic interpretation of group membership strength. The number of identifiable clusters was selected by using the Akaike Information Criterion (AIC) likelihood penalization method. Genetic subgroups were then correlated with overall survival, disease progression and treatment specific responses. All patients were followed up for mortality until 01 August 2023. Results A total of 970 patients (96%) harboured at least one of the 69 genetic features. Eight distinct clusters were identified and were named after the most discriminatory abnormality within each cluster. All clusters contained both AML and MDS cases however five clusters showed a predominance of MDS cases (EZH2, SF3B1, SRSF2, TP53, U2AF1_157) while AML cases predominated in the remaining 3 (BCOR, KMT2A-PTD, NPM1). The genomic profile within each cluster was strikingly similar across both AML and MDS. With respect to overall survival (OS) within the total cohort, the SF3B1 cluster showed the best OS while TP53 was by far the worst. These effects persisted when analysing the MDS and AML cohorts separately. The SF3B1 group had the best OS within MDS, but also in AML patients where the better outcome was equivalent to both the NPM1 (p=1.0) and U2AF1_157 groups (p=0.4). The TP53 group predicted the worst outcome across both diseases though the BCOR group was comparably poor in MDS, particularly after 2 years (TP53 vs BCOR OS; 1 year 15% (CI 9-26) vs 48% (CI 32-72), 2 years 5% (CI 2-14) vs 9% (CI 2-33). Within each cluster group, OS was not significantly different between MDS and AML patients for the BCOR (p=0.4), KMT2A_PTD (p=0.2), NPM1 (p=1.0) and U2AF1_157 (p=0.4) groups and approached significance for EZH2 (p=0.07). For other groups MDS patients had a significantly better OS and blast percentage was the main discriminating factor with respect to outcome. Those with <5% blasts had the best outcome while above this threshold OS was broadly comparable. The risk of progression to AML was highest for those MDS cases within the AML predominant clusters (BCOR HR 16.8; CI 6.3-44.5; NPM1 HR 10.6; CI 4.1-27.9) and TP53 cluster (HR 15.1; CI 6.5-35.1). A total of 269 patients received intensive chemotherapy as a first line treatment. Those patients within the NPM1, SF3B1 and U2AF1_157 had the best outcome following intensive therapy though these patients were significantly younger than other groups combined (p<0.0001). In contrast patients within the TP53 and SRSF2 groups had a very poor outcome with all treatment interventions. Conclusion By analysing MDS and AML cases on a continuum, distinct molecular subgroups can be identified with diverse outcomes and responses to treatment in many cases irrespective of blast percentage. While some of these groups are expected from previous studies, other groups are not currently recognised as distinct entities. Validation of the impact of these groups in the era of novel therapies is however needed.

Lower risk (LR) myelodysplastic syndromes (MDS) are heterogeneous hematopoietic stem and progenitor disorders caused by the accumulation of somatic mutations in various genes including epigenetic regulators that may produce convergent DNA methylation patterns driving specific gene expression profiles. The integration of genomic, epigenomic, and transcriptomic profiling has the potential to spotlight distinct LR-MDS categories on the basis of pathophysiological mechanisms. We performed a comprehensive study of somatic mutations and DNA methylation in a large and clinically well-annotated cohort of treatment-naive patients with LR-MDS at diagnosis from the EUMDS registry (ClinicalTrials.gov.NCT00600860). Unsupervised clustering analyses identified six clusters based on genetic profiling that concentrate into four clusters on the basis of genome-wide methylation profiling with significant overlap between the two clustering modes. The four methylation clusters showed distinct clinical and genetic features and distinct methylation landscape. All clusters shared hypermethylated enhancers enriched in binding motifs for ETS and bZIP (C/EBP) transcription factor families, involved in the regulation of myeloid cell differentiation. By contrast, one cluster gathering patients with early leukemic evolution exhibited a specific pattern of hypermethylated promoters and, distinctly from other clusters, the upregulation of AP-1 complex members FOS/FOSL2 together with the absence of hypermethylation of their binding motif at target gene enhancers, which is of relevance for leukemic initiation. Among MDS patients with lower-risk IPSS-M, this cluster displayed a significantly inferior overall survival (p < 0.0001). Our study showed that genetic and DNA methylation features of LR-MDS at early stages may refine risk stratification, therefore offering the frame for a precocious therapeutic intervention.

Introduction The long-term outcome of patients with clonal cytopenia of undetermined significance (CCUS) remains ill defined. Selected datasets show a very high prevalence of disease progression to a confirmed myeloid malignancy though these are predominantly retrospective datasets from tertiary referral centres. There is also increasing interest in the non-haematological effects of clonal haematopoiesis with regards to general health outcomes and there is a requirement to have long term outcome data on prospective unselected CCUS cohorts. We have previously reported 5-year follow-up data on such a cohort of 400 CCUS patients identified through a large haematopathology service. This study represents 10-year follow-up of this cohort including progression and survival dynamics over time, predictive models of both progression and survival and cause of death. Methods All patient samples (n=2083) referred for investigation of unexplained cytopenia or suspected MDS to the Haematological Malignancy Diagnostic Service (HMDS) in the UK over a 2-year period (July 2014-2016) were subjected to targeted myeloid panel sequencing of 27 genes. This was alongside the established diagnostic pathway of morphology, flow cytometry and cytogenetics. A total of 400 CCUS patients were identified during this analysis. Outcome data was captured on these patients including subsequent diagnoses, overall survival and cause of death (the latter data capture is ongoing). Both pathologists and clinicians were blinded to the results of this analysis removing diagnostic and treatment bias. Results Median follow-up of this cohort was 9.6 years. The cohort included 256 males and 144 females, and the majority had only 1 somatic mutation (205/400; 51%) with TET2, SRSF2 and DNMT3A mutations predominating (47%, 22%, 19% respectively). At the point of analysis 317 patients were deceased (79%) while 79 patients had progressed to a confirmed myeloid malignancy (20%). The majority of progressions were to MDS (36/79; 46%) or myelodysplastic/myeloproliferative (MDS/MPN) overlap syndromes (25/79; 32%). The cumulative hazard for overall survival was near linear meaning hazard was constant over time. In contrast, most progressions occurred within the first 5 years following diagnosis (66/79; 84%) with approximately piecewise constant hazard estimates reducing from 0.061 (CI 0.047, 0.076) to 0.026 (CI 0.012, 0.041) after 5.14 years. Importantly there was an absence of reported progressions during the COVID-19 pandemic lockdown period. Confirming, that the majority of patients were dying as a consequence of CCUS, 5-year net survival was 63.3% (95%CI: 56.4-70.2) compared to 47.0% (95%CI: 42.0-51.8) 5-year overall survival. Time to death analysis confirmed that the number of mutated genes had a significant impact on survival when incorporated into age and sex adjusted models while number of mutations was not significant. A predictive model including age, sex (M), ASXL1, BCOR, TP53, maximum VAF (variant allele fraction) and >1 mutated gene had a concordance index of 0.67. Importantly maximum VAF had a protective effect on survival becoming significant only when number of mutated genes was also included, consistent with a dominant clone predicting a better outcome versus multiple lower-level clones. Time to progression analysis, in contrast, showed an overwhelming effect of the number of mutated genes in predictive models. A model including number of mutated genes and sex had a concordance index of 0.79 with no improvement in predictive power when other variables, such as maximum VAF were included. Progression free survival analysis confirmed the strongest effects identified for both survival and progression, producing a model containing age, gender, maximum VAF and number of mutated genes with a concordance index of 0.65. Conclusion Long term outcome analysis of this unselected CCUS cohort has provided further clarity on the natural history of this condition. At 10 years the progression rate is much lower than previously reported, though this is likely impacted by the very high mortality rate. Progression events most commonly happen early and are driven by mutation number while in those with multiple mutated genes, maximum VAF had a protective effect on survival. Net survival estimates confirmed the majority of CCUS patients were dying as a consequence of their disease; specific causes will be further explored.

Background Somatic mutations are frequently reported in individuals with cytopenia but without a confirmed haematological diagnosis (clonal cytopenia of undetermined significance; CCUS). These patients have an increased risk of progression to a myeloid malignancy and worse overall survival than those with no such mutations. To date, studies have been limited by retrospective analysis or small patient numbers. We aimed to establish the natural history of CCUS by prospectively investigating outcome in a large, well defined patient cohort.Methods This prospective cohort study was conducted at the Haematological Malignancy Diagnostic Service, a diagnostic laboratory in Leeds, UK. Patients aged at least 18 years who were referred for investigation of cytopenia were eligible for inclusion; those with a history of myeloid malignancy were not eligible. Targeted sequencing was conducted alongside routine clinical testing. Baseline mutation analysis was then correlated with the main study outcomes: longitudinal blood counts, disease progression to a myeloid malignancy, and overall survival with a median follow-up of 454 years (IQR 403-504). Data were collected manually from hospital records or extracted from laboratory or clinical outcome databases.Findings Bone marrow samples from 2348 patients were received at the Haematological Malignancy Diagnostic Service between July 1, 2014, and July 31, 2016. Of these, 2083 patients (median age 72 years [IQR 63-80, range 18-99]; 854 [410%] female and 1229 [590%] male) met the inclusion criteria and had samples of sufficient quality for further analysis. 598 (287%) patients received a diagnosis on the basis of their biopsy sample, whereas 1485 (713%) samples were classified as non-diagnostic; of these, CCUS was confirmed in 400 (269%) patients (256 [640%] male and 144 [360%] female). TET2, SRSF2, and DNMT3A were the most frequently mutated genes in patients with CCUS, with 320 (80%) of 400 patients harbouring a mutation in at least one of these genes. Age (p<00001), sex (p=00027), and mutations in ASXL1 (p=00009), BCOR (p=00056), and TP53 (p=00055) correlated with a worse overall survival; however, the number of mutations was the strongest predictor for progression to a myeloid malignancy (two mutations, p=00024; three or more mutations, p=00004). Extended sequencing of samples from a subgroup of patients with sequential samples and no mutations in the initial myeloid gene panel showed recurrent mutations in both DDX41 and UBA1, suggesting that these genes should be included in clinical test panels.Interpretation Mutation analysis is advised in patients who have undergone bone marrow examination and have an otherwise-unexplained cytopenia. High-risk genetic mutations and increased numbers of mutations are predictive of both survival and progression within 5 years of presentation, warranting clinical surveillance and, when necessary, intervention.

The precise link between inflammation and pathogenesis of myelodysplastic syndrome (MDS) is yet to be fully established. We developed a novel method to measure ASC/NLRP3 protein specks which are specific for the NLRP3 inflammasome only. We combined this with cytokine profiling to characterise various inflammatory markers in a large cohort of patients with lower risk MDS in comparison to healthy controls and patients with defined autoinflammatory disorders (AIDs). The ASC/NLRP3 specks were significantly elevated in MDS patients compared to healthy controls (p < 0.001) and these levels were comparable to those found in patients with AIDs. The distribution of protein specks positive only for ASC was different to ASC/NLRP3 ones suggesting that other ASC-containing inflammasome complexes might be important in the pathogenesis of MDS. Patients with MDS-SLD had the lowest levels of interleukin (IL)-1β, tumour necrosis factor (TNF), IL-23, IL-33, interferon (IFN) γ and IFN-α2, compared to other diagnostic categories. We also found that inflammatory cytokine TNF was positively associated with MDS progression to a more aggressive form of disease and IL-6 and IL-1β with time to first red blood cell transfusion. Our study shows that there is value in analysing inflammatory biomarkers in MDS, but their diagnostic and prognostic utility is yet to be fully validated.

Background Arising in blood and lymph-forming tissues, haematological malignancies (leukaemias, lymphomas and myelomas) are the fifth most common group of cancers. Around 60% are currently incurable and follow a chronic, remitting–relapsing pathway often initially managed by ‘watch & wait’. This involves hospital-based monitoring, followed by treatment if the cancer progresses (which not all do) and then further observation, in a process that may continually repeat. New treatments are constantly emerging, survival is improving and prevalence is rising, but population-based data documenting entire care pathway are sparse. Hence, empirically-based incidence and prevalence estimates about various treatment states (watch and wait, first-line treatment, observation, second-line treatment, etc.) and patterns of healthcare activity are lacking. Likewise, despite complex trajectories, anxiety-provoking watch and wait, and therapies that impede quality of life and incur marked healthcare costs, evidence about patient preferences for information sharing and treatment decisions is scant. Objectives Primary – to generate high-quality, evidence-based information about the care pathways of the general population of patients with chronic haematological malignancies. Secondary – to produce information resources suitable for testing in routine National Health Service practice. Design Population-based cohort of ≈ 8000 patients with chronic haematological malignancies, incorporating five nested work packages, each with its own individual design: (1) exploration of patient experiences: information and treatment decisions; (2) population-based analyses; (3) health economics; (4) development of information resources to support decision-making; and (5) patient well-being and decision-making survey. Setting This programme is predicated on the infrastructure of the United Kingdom’s Haematological Malignancy Research Network (www.hmrn.org); which provides ‘real-world’, robust, generalisable data to inform research and clinical practice, nationally and internationally. Set in Yorkshire and Humberside, the Haematological Malignancy Research Network’s catchment population of ≈ 4 million has a comparable sex, age, urban/rural, and area-based deprivation (Index of Multiple Deprivation, income domain) distribution to the United Kingdom as a whole; and in terms of ethnic diversity the region is centrally ranked, with around 80% of residents identifying as White British, 9% as Asian and 2% as black. Within the Haematological Malignancy Research Network, clinical practice adheres to national guidelines, and all patients with blood cancers are centrally diagnosed (≈ 2500 each year), tracked through their treatment pathways and linked to national databases (deaths, cancer registrations and Hospital Episode Statistics). Linked to the same national databases, the Haematological Malignancy Research Network also contains an age- and sex-matched general-population cohort. Participants Patients aged ≥ 18 years, resident in the study region, and diagnosed with chronic lymphocytic leukaemia, follicular lymphoma or myeloma. Methods Core Haematological Malignancy Research Network data were used to compare the hospital activity of patients with chronic lymphocytic leukaemia, follicular lymphoma and myeloma with that of the general population. Following additional linkages to genetic and clinical data, follicular lymphoma prognostic factors were examined. Two self-administered questionnaires addressing (1) quality of life and well-being and (2) decision-making were iteratively developed, piloted and deployed. Linkage to quality of life, clinical information and Hospital Episode Statistics enabled economic (myeloma) model development. In-depth interviews were conducted with 35 patients (10 alongside relatives). Results Trajectories of ≈ 8000 patients were mapped, and patient-pathway visualisations summarising individual and aggregate information were developed. As expected, patients with chronic blood cancers experienced higher levels of hospital activity than their general population counterparts, the largest effects being for myeloma. Following survey deployment, 3153 patients were recruited across 14 hospitals, 1282 with chronic lymphocytic leukaemia, follicular lymphoma or myeloma. Over half of the questionnaires were completed by patients on watch and wait; the remainder were completed during treatment or post-chemotherapy monitoring. Information gathered, coupled with in-depth interviews, demonstrated patients’ marked anxiety and fluctuating preferences for information sharing and decision-making, contingent on complex, inter-related factors. In turn, prognostic and microsimulation economic models were used to predict individual-level trajectories across multiple treatment lines, examining associated overall survival, costs and quality-adjusted life-years. Limitations Survey mapping to individual care pathways could not be completed because the COVID-19 pandemic delayed clinical data collection. Patients who attended clinics and participated in the survey were more likely than non-attenders to have had first-line chemotherapy, be slightly younger and live in more affluent areas. Conclusions This programme collated high-quality, population-based evidence. Previously lacking, this, coupled with new findings on preferences for information sharing and treatment decisions, provides the foundation for future research. Future work The translation of information accrued into resources suitable for testing in routine NHS practice is key. In this regard, COVID-19 has changed the communication landscape. The visualisations developed by this programme require further refinement/testing using participatory co-design with stakeholder groups. Underpinned by a suitable protocol applied within a single multidisciplinary team setting, prior to further evaluation within/outside the region, such outputs require testing in a cluster-randomised trial. Funding This award was funded by the National Institute for Health and Care Research (NIHR) Programme Grants for Applied Research programme (NIHR award ref: RP-PG-0613-20002) and is published in full in Programme Grants for Applied Research ; Vol. 12, No. 5. See the NIHR Funding and Awards website for further award information.

Myelodysplastic syndromes (MDS) are a group of heterogeneous disorders caused by the accumulation of somatic mutations in the hematopoietic stem and progenitor compartment. Besides del(5q), SF3B1 or TP53 mutations, referred to as defining genetic abnormalities, mutation patterning hardly structured the classification of MDS. Mutations in epigenetic factors occur early in the development of clonal hematopoiesis leading to precocious alterations of DNA methylation implicated in oncogenesis. Convergent DNA methylation patterns related to both mutations and microenvironment imprinting may induce specific gene expression profiles and contribute to phenotypic variations. To refine a pathophysiological classification of lower-risk MDS, we combined genetic profiling to DNA methylation and transcriptome data. Methods: We enrolled 543 lower-risk treatment-naïve MDS patients at diagnosis and performed DNA-sequencing of 24 genes. DNA methylation and RNA-sequencing data were generated in a representative subset of 175 cases with available material (75 MDS-SLD/MLD, 40 MDS-EB1, 53 MDS-RS-SLD/MLD, 5 MDS del(5q), 2 MDS-U) and 7 age-matched healthy controls. IPSS-R was very low, low or intermediate in 87.4%. During a median follow-up of 2 years, 16% of MDS cases progressed to AML (MDSp). Infinium EPIC 850K array allowed after filtering the examination of methylation of 723,612 CpG sites with enrichment at enhancers and promoters. Differentially methylated regions with differentially methylated CpG sites (DMR-CpG) between cases and controls were defined with a minimum of 2 probes, a Benjamini-Hochberg-adjusted P-value cut-off of 0.05, a false discovery rate of 0.001 and a mean Δβ-value >20% at CpG sites. Consensus motifs for transcription factors were identified using Homer. Results: Unsupervised clustering on genetic profiling of 543 patients identified distinct subsets of lower-risk MDS with different clinical features: one cluster (group A) was enriched in del(5q), three clusters were characterized by SF3B1 mutations, either isolated (group B) or associated with DNMT3A (group E) or TET2 (group F), while the two remaining clusters were enriched in either complex pattern of mutations (group C) or SRSF2 mutations (group D). Genetic clusters showed significantly different clinical outcomes, clusters C, D and E having lower overall survival and higher risk of progression to high risk MDS or AML compared to other clusters. In the subset of 175 cases, a total of 2,953 unique DMR-CpGs allowed a robust repartition of patient samples in 4 groups using the k-means method. The 4 methylation groups were clinically distinct in terms of age, hemoglobin, neutrophils, monocytes, platelets, BM blasts, WHO classification, IPSS-R, and mutation pattern. A significant correlation was found between genetic and methylation groups ( P<0.001). Hypomethylated CpGs defined group 4 which was enriched in MDS-RSwith low blast count and few co-mutations, while groups 1 and 3 demonstrated hypermethylated profiles driven by TET2/ IDH1-2 mutations and TET2/ SRSF2 mutations, respectively. TET2-mutated cases exhibited an increased proportion of hypermethylated DMR-CpGs at enhancers (DME) significantly enriched in C/EBP and ETS family transcription factor motifs. In MDS without evidence of progression, we identified 1,418 DME and their gene targets of which those significantly deregulated in RNA-seq were involved in the control of translation and response to TGF-β. By contrast, in MDS with rapid progression to AML, DMR-CpGs were significantly enriched at CpG islands and promoters ( P<0.0001) and a set of 10-20 genes downstream of these promoters were specifically deregulated. Conclusion: Somatic mutations and DNA methylation at enhancers and promoters identify distinct subsets of lower-risk MDS with different clinical behavior. Hypermethylation at enhancers with C/EBP or ETS family motifs is a hallmark of MDS and may account for changes in transcription factor recruitment and cell fate. Hypermethylation of promoters with downstream effects on gene expression may indicate a propensity to rapid evolution to AML.

Background: Allogeneic hematopoietic stem cell transplantation is the only curative treatment for myelodysplastic syndrome (MDS). How to select lower-risk (LR) patients (pts) for transplantation is a matter of debate. The International Working Group for Prognosis in MDS (IWG) defined intermediate IPSS-R risk pts as a higher-risk if IPSS-R score is above 3.5. MDS-RIGHT criteria recommend transplantation in LR-MDS if: very/poor risk cytogenetics, >50% blasts increase from baseline or >15% BM blasts, neutrophils <0.3x109/l, platelets <30x109/l, high transfusion intensity ≥2 units/month for 6 months, progression to higher risk IPSS-R group or drop of platelets >25% in 6 months (MDS Europe. https://www.mds-europe.org). Aims: The primary aim of the study was to compare the survival of LR-MDS pts candidate for transplant by IWG and MDS-RIGHT criteria with non-candidate pts using registry data. The secondary aim was to compare the survival of IWG and MDS-RIGHT registry based cohorts with survival in published transplant cohort. Methods: 2284 pts from the EUMDS registry with LR-MDS (very/low and intermediate risk IPSS-R) were included in the registry based cohort. Survival analysis of pts meeting IWG and MDS-RIGHT criteria at baseline, 6 and 12 months was done in all candidates and those fit for transplantation, defined by Karnofsky ≥70, HCT-CI <3 according to MDS-RIGHT. Systematic review of transplantation studies in MDS was conducted. Published survival from transplant cohort were compared to the registry based cohorts. Results: Number of all and fit [Nall (Nfit, %)] candidates for transplant at baseline, 6 and 12 months was: IWG: 280 (146, 52%), 15 (5, 33%) and 22 (0, 0%); MDS-RIGHT: 182 (97, 53%), 428 (112, 26%) and 125 (39, 31%), respectively. Median OS, PFS and LFS survival in candidate and non-candidate patients at baseline were significantly different (p<0.01). Median OS was: IWG: 2.5 vs 4.6 years; MDS-RIGHT: 3.3 vs 4.6 years (Fig.1A and B). MDS-RIGHT pts with IPSS-R >3.5 (28% of IWG group) shown unfavorable median OS comparing with other pts (1.5 vs 4.8 years, p <0.001, Fig.1C). Published OS of transplanted intermediate IPSS-R risk pts (Scheid et al., BMT 2017) at 12, 24 and 36 months was: 59%, 48%, and 43%. Survival in IWG and MDS-RIGHT pts was better at all time points. MDS-RIGHT pts with IPSS-R >3.5 had higher survival at 1 year (68%) but worse survival at 2 and 3 years (42% and 33%) (p=0.07, Fig.1D). Summary/Conclusion: Both transplant selection criteria identify LR-MDS patients with unfavorable survival. MDS-RIGHT pts with IPSS-R above 3.5 have particularly unfavorable prognosis. There is no clear evidence for upfront transplantation in any of prognostic group.Keywords: Myelodysplastic syndrome, Prognosis, Allogeneic hematopoietic stem cell transplant

Follicular lymphoma (FL) is morphologically and clinically diverse, with mutations in epigenetic regulators alongside t(14;18) identified as disease-initiating events. Identification of additional mutational entities confirms this cancer’s heterogeneity, but whether mutational data can be resolved into mechanistically distinct subsets remains an open question. Targeted sequencing was applied to an unselected population-based FL cohort (n = 548) with full clinical follow-up (n = 538), which included 96 diffuse large B-cell lymphoma (DLBCL) transformations. We investigated whether molecular subclusters of FL can be identified and whether mutational data provide predictive information relating to transformation. DNA extracted from FL samples was sequenced with a 293-gene panel representing genes frequently mutated in DLBCL and FL. Three clusters were resolved using mutational data alone, independent of translocation status: FL_aSHM, with high burden of aberrant somatic hypermutation (aSHM) targets; FL_STAT6, with high STAT6 & CREBBP mutation and low aSHM; and FL_Com, with the absence of features of other subtypes and enriched KMT2D mutation. Analysis of mutation signatures demonstrated differential enrichment of predicted mutation signatures between subgroups and a dominant preference in the FL_aSHM subgroup for G(C>T)T and G(C>T)C transitions consistent with previously defined aSHM-like patterns. Of transformed cases with paired samples, 17 of 26 had evidence of branching evolution. Poorer overall survival (OS) in the aSHM group (P = .04) was associated with older age; however, overall tumor genetics provided limited information to predict individual patient risk. Our approach identifies 3 molecular subclusters of FL linked to differences in underlying mechanistic pathways. These clusters, which may be further resolved by the inclusion of translocation status and wider mutation profiles, have implications for understanding pathogenesis as well as improving treatment strategies in the future.

Information on causes of death (CoDs) and the impact of myelodysplastic syndromes (MDS) on survival in patients with lower-risk MDS (LR-MDS) is limited. A better understanding of the relationship between disease characteristics, clinical interventions and CoDs may improve outcomes of patients with LR-MDS. We prospectively collected data on patients with LR-MDS in the European MDS registry from 2008 to 2019. Clinical, laboratory and CoDs data were obtained. To examine MDS-specific survival, relative survival (RS) was estimated using national life tables. Of 2396 evaluated subjects, 900 died (median overall survival [OS]: 4.7 years; median follow-up: 3.5 years). The most common CoDs were acute myeloid leukaemia/MDS (20.1%), infection (17.8%) and cardiovascular disease (CVD; 9.8%). Patients with isolated del(5q) and with red cell transfusion needed during the disease course, had a higher risk of fatal CVD. The 5-year OS was 47.3% and the 5-year RS was 59.6%, indicating that most patients died due to their underlying MDS. Older patients (aged >80 years) and the lowest-risk patients were more likely to die from competing causes. This study shows that MDS and its related complications play crucial role in the outcome of patients with LR-MDS.

Cohort Profile Update: The Haematological Malignancy Research Network (HMRN) UK population-based cohorts Eve Roman ,* Eleanor Kane, Debra Howell, Maxine Lamb, Timothy Bagguley, Simon Crouch, Daniel Painter, Russell Patmore, and Alexandra Smith Epidemiology and Cancer Statistics Group, Department of Health Sciences, University of York, YO10 5DD, UK and Queens Centre for Oncology, Castle Hill Hospital, UK

Diffuse large B-cell lymphoma (DLBCL) is characterised by pronounced genetic and biological heterogeneity. Several partially overlapping classification systems exist – developed from mutation, rearrangement or gene expression data. We apply a customised network analysis to nearly five thousand DLBCL cases to identify and quantify modules indicative of tumour biology. We demonstrate that network-level patterns of gene co-expression can enhance the separation of DLBCL cases. This allows the resolution of communities of related cases which correlate with genetic mutation and rearrangement status, supporting and extending existing concepts of disease biology and delivering insight into relationships between differentiation state, genetic subtypes, rearrangement status and response to therapeutic intervention. We demonstrate how the resulting fine-grained resolution of expression states is critical to accurately identify potential responses to treatment. Significance statement We demonstrate how exploiting data integration and network analysis of gene expression can enhance the segregation of diffuse large B-cell lymphoma, resolving pattens of disease biology and demonstrating how the resolution of heterogeneity can enhance the understanding of treatment response. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This work was supported by Cancer Research UK program grant (C7845/A17723 and C7845/A29212) (M.C, G.D., D.W, and R.T). HMRN is supported by Cancer Research UK program grant A29685 (D.P., S.C., A.S., E.R.). D.J.H. was supported by a fellowship from Cancer Research UK (CRUK) (RCCFEL∖100072) and received core funding from Wellcome (203151/Z/16/Z) to the Wellcome-MRC Cambridge Stem Cell Institute and from the CRUK Cambridge Centre (A25117). D.J.H is supported by the National Institute for Health and Care Research (NIHR) Cambridge Biomedical Research Centre (BRC-1215-20014). R.T., G.D., E.R., A.S., D.P., D.W. are supported by the National Institute for Health and Care Research Leeds Biomedical Research Centre. The views expressed are those of the authors and not necessarily those of the NIHR or the Department of Health and Social Care. For the purpose of Open Access, the authors have applied a CC BY public copyright licence to any Author Accepted Manuscript version arising from this submission. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: GSE4475, GSE4732, GSE10846,GSE12195,GSE19246_FF,GSE22470,GSE31312,GSE32918,GSE34171,GSE53786,GSE87371,GSE9858,GSE181063,GSE117566 PubMedID15550490 PudMedID29641966(NCICCR-DLBCL) EGAS00001002606) I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes All data produced are available at

We present a noninvasive Web-based app to help exclude or diagnose myelodysplastic syndrome (MDS), a bone marrow (BM) disorder with cytopenias and leukemic risk, diagnosed by BM examination. A sample of 502 MDS patients from the European MDS (EUMDS) registry (n > 2600) was combined with 502 controls (all BM proven). Gradient-boosted models (GBMs) were used to predict/exclude MDS using demographic, clinical, and laboratory variables. Area under the receiver operating characteristic curve (AUC), sensitivity, and specificity were used to evaluate the models, and performance was validated using 100 times fivefold cross-validation. Model stability was assessed by repeating its fit using different randomly chosen groups of 502 EUMDS cases. AUC was 0.96 (95% confidence interval, 0.95-0.97). MDS is predicted/excluded accurately in 86% of patients with unexplained anemia. A GBM score (range, 0-1) of less than 0.68 (GBM < 0.68) resulted in a negative predictive value of 0.94, that is, MDS was excluded. GBM ≥ 0.82 provided a positive predictive value of 0.88, that is, MDS. The diagnosis of the remaining patients (0.68 ≤ GBM < 0.82) is indeterminate. The discriminating variables: age, sex, hemoglobin, white blood cells, platelets, mean corpuscular volume, neutrophils, monocytes, glucose, and creatinine. A Web-based app was developed; physicians could use it to exclude or predict MDS noninvasively in most patients without a BM examination. Future work will add peripheral blood cytogenetics/genetics, EUMDS-based prospective validation, and prognostication.

Available evidence suggests that in most patients with LR-MDS the risk of death is not related to disease progression but is mainly attributable to non-leukemic death. 2,17 In addition, a proportion of these patients have prolonged survival that precludes the design of clinical trials adopting OS as a primary endpoint. These challenges have resulted in potentially biased assessment of the effectiveness and appropriate use of the available interventions in this patient population. The EUMDS Registry has identified novel meaningful outcome indicators and clinical endpoints, and reliable measures of response to HCI (Figure 4). The results of our analysis indicate that RBCT density is strongly associated with a decreased OS, even at relatively low dose densities. In addition, we observed that an early decrease in platelet count is an independent adverse prognostic indicator in LR-MDS, and combining relative platelet drop and transfusion dependency allows early identification of patients at risk of rapid progression, and may guide early therapeutic interventions, including allogeneic hematopoietic stem cell transplantation or experimental interventions. Taken together, these results indicate that regular RBCT requirement, early platelet count kinetics, and restriction in HRQoL are early independent and meaningful outcome indicators, and reliable measures of effectiveness of therapeutic interventions, evaluated in this set of studies. These findings support the integration of RBCT requirement and HRQoL in the general core outcome sets and in response criteria in patients with LR-MDS, and have important implications for clinical practice and the design of clinical endpoints. Our results strongly support the adoption of freedom from transfusion as a meaningful clinical endpoint in patients with LR-MDS. Anemia is the main determinant of therapeutic intervention in patients with LR-MDS, and ESA are recommended as first-line treatment for patients with symptomatic anemia. 10 The observational studies within the EUMDS Registry showed that the response rate, as well as the capacity of these agents to delay the onset of a regular RBCT need, is most pronounced in RBCT-naive patients. These results identified early initiation of treatment with ESA as a major treatment response indicator, and indicate that ESA should be recommended in LR-MDS patients with symptomatic anemia before starting regular RBCT. After the onset of RBCT dependency, patients with LR-MDS are prone to long-term accumulation of iron. 1,43 The EUMDS Registry studies provided evidence that elevated LPI levels are associated with reduced survival in RBCT dependent patients, whereas iron chelation therapy normalizes LPI levels. These findings suggest that NTBI and LPI may serve as early indicators of iron toxicity and a means to measure the effectiveness of iron chelation therapy in patients with LR-MDS. However, qualified NTBI and LPI are only currently available in specialized laboratories. 44 Large observational cohorts with detailed clinical and laboratory data, like the EUMDS cohort, are the ideal framework in which to identify well defined MDS subtypes that may benefit from novel targeted treatments. An example of such a subtype is MDS with loss of parts of chromosome 5, namely del5q; these patients have a relatively favorable outcome on lenalidomide treatment. In order to identify homogeneous subsets of patients within MDS, preliminary evidence has suggested that recently identified mutations in splicing factors may recognize distinct disease entities within myeloid neoplasms. 45 Splicing modulators are now in pre-clinical testing, and are very likely to lead to the introduction of effective drugs for specific groups of MDS patients. Luspatercept, a specific inhibitor of growth and differentiation factor-11, a member of the transforming growth factor β superfamily, induced substantial improvement of anemia, especially in patients with ring sideroblasts. 46 Characterization of individual cases by new genetic markers (one of the main objectives of the MDS-RIGHT project) will allow refined classification of patients into biological subgroups that are expected to respond differently to therapeutic interventions to guide discontinuation of those interventions that are less effective or less cost-effective. The main question is whether RCT data and retrospective cohort data in selected tertiary care centers are representative of the 'real world' data of the older patients with LR-MDS in the general population. A careful comparison of the 'real world' data and the RCT data will be needed in order to provide a clear answer to these questions. Meanwhile, the current analyses of data collected over 10 years in the EUMDS Registry provides relevant and important information which could help assess prognosis and response to standard interventions in this older patient group.