The use of immune checkpoint inhibitors (ICIs) has deeply improved the outcome of relapsed or refractory (R/R) primary mediastinal B-cell lymphoma (PMBCL) patients. However, real-world data are still limited, and several aspects, including the need for consolidation strategies, remain to be fully elucidated. We report the results of the multicenter Italian retrospective observational PRIMICI study on R/R PMBCL patients treated with ICIs in a real-life (off-label) setting. Seventy-four patients, 42 treated with pembrolizumab and 32 with nivolumab-brentuximab vedotin (nivo-BV), were enrolled. The median follow-up was 34 months. The best overall response and complete response (CR) rates were 64% (n = 50) and 50% (n = 37), respectively. Of the 37 patients in CR, 11 received a consolidation treatment and 26 patients continued ICIs; CR were stable, independently from the use of consolidation, with a 4-year disease-free survival of 100%. The estimated 5-year progression-free survival (PFS) and overall survival (OS) were 60.4% (95% confidence interval [C.I.] 48.3%-70.6%) and 77.5% (95% C.I. 65.8%-85.6%), respectively. Nivo-BV was associated with faster and higher response rates and better OS compared to pembrolizumab. OS significantly improved after 2020 due to the use of salvage treatment with CAR T-cells. In conclusion, this study supported the safety and effectiveness of pembrolizumab or nivo-BV as a salvage therapy in R/R PMBCL patients. Importantly, the chance to obtain long-lasting responses, regardless of the use of a consolidation treatment, indicates that ICIs may be used with a curative intent in a significant subset of patients.

Venetoclax (VEN) plus hypomethylating agents (HMAs) represents the standard of care for newly diagnosed acute myeloid leukaemia (AML) patients unfit for intensive chemotherapy, but prospective real-world observational data outside randomized clinical trials remain limited. The Gruppo Italiano Malattie Ematologiche dell'Adulto (GIMEMA) AML2320 is a prospective, multicentre, observational study (NCT04589728) including 193 newly diagnosed unfit AML patients receiving VEN plus azacitidine or decitabine between November 2020 and December 2021. Primary end-point was overall survival (OS); secondary end-points included composite complete remission (cCR), disease-free survival and safety. Median age was 74 years with 42% of patients being ≥75 years. After completing cycle 4, cCR was achieved in 73% of the patients, with 54% responding before cycle 2. After a median follow-up of 23 months, median OS was 13.0 months. cCR achievement within cycle 4 correlated with longer OS (19.1 vs. 9.1 months, p = 0.001). Patients receiving 400 mg VEN without azoles had improved OS compared with those on reduced doses with azoles (18.2 vs. 11.4 months, p = 0.015). An anchored matching-adjusted indirect comparison with the phase III VIALE-A trial (NCT02993523) showed comparable median OS (14.8 months vs. 14.9 months; p = 0.6). The GIMEMA AML2320 trial confirmed the efficacy of VEN/HMAs in a prospective, real-world unfit population. Early remission was associated with improved outcomes.

PURPOSE:Owing to prolonged treatment exposure, patients with relapsed/refractory multiple myeloma (RRMM), particularly those with low socioeconomic status (SES), may be especially vulnerable to financial toxicity (FT), which can negatively affect their health-related quality of life (HRQoL). We aimed to evaluate the association between SES and HRQoL in patients with RRMM and to explore the mediating role of FT in this relationship. METHODS:We conducted a cross-sectional analysis of baseline data from a multicenter, prospective, observational study of patients with RRMM in Italy and the United Kingdom. SES was assessed via a composite index including education, employment, and living arrangements. We evaluated HRQoL using the European Organisation for Research and Treatment of Cancer Quality of Life Questionnaire-Core 30 (EORTC QLQ-C30) and QLQ-MY20. Multivariable regression models adjusted for potential confounders were used to assess the association between SES and HRQoL. Mediation analysis was conducted to examine the role of FT in the relationship between SES and HRQoL. RESULTS:A total of 505 patients with RRMM were analyzed, and 15.6%, 49.3%, and 35.1% were classified as low, middle, and high SES, respectively. Patients with low versus high SES reported clinically meaningful worse scores in 11 EORTC QLQ-C30 scales and in the QLQ-MY20 disease symptoms scale. Compared with patients with high SES, those with low SES had significantly higher odds of reporting clinically important pain (odds ratio [OR], 3.55), financial difficulties (OR, 2.97), and impaired physical functioning (OR, 2.96). FT significantly and partially mediated the relationship between SES and HRQoL, accounting for 46.3% of the total effect. CONCLUSION:Patients with RRMM and low SES experienced worse HRQoL compared with those with higher SES, with FT partially mediating this association. Interventions addressing FT and broader social determinants of health in RRMM are warranted to reduce disparities and promote equity in cancer outcomes.

Background: Gaucher disease (GD) and Acid Sphingomyelinase Deficiency (ASMD) are rare, chronic, function progressive, and debilitating disorders caused by the altered lysosomal enzymes glucocerebrosidase (GCase) in GD and sphingomyelinase (ASM) in ASMD respectively. These pathologies share several clinical manifestations, and their real incidence is underestimated.

BACKGROUND:Advancements in treating hematologic malignancies have improved survival, but health-related quality of life (HRQoL) remains a key concern due to the physical, emotional, and social impact of disease and treatment. AIMS:This study aimed to assess HRQoL in patients undergoing treatment for hematologic malignancies, identifying the most affected domains to guide supportive interventions. METHODS:HRQoL was evaluated using the 42-item Functional Assessment of Cancer Therapy-Lymphoma (FACT-Lym) in a prospective, multicenter observational study on rituximab safety across Italian hematology units (March 2018-June 2022). RESULTS:Of 772 patients with Non-Hodgkin Lymphoma (NHL) or Chronic Lymphocytic Leukemia (CLL), 412 (62.6%) completed the HRQoL assessment. Overall, patients reported acceptable quality of life. Key concerns included reduced enjoyment of life (45.1%), poor sleep (42.0%), and dissatisfaction with quality of life (41.0%). Functional impairments and emotional distress were also reported. CLL patients showed slightly better physical and lymphoma-specific well-being than NHL patients. Patients with other hematologic diseases had lower scores across several domains. CONCLUSIONS:While HRQoL is generally acceptable in NHL and CLL, functional well-being is most impacted. Routine integration of HRQoL assessments is essential to identify unmet needs and support patient-centered care.

In hematologic malignancies, neoplastic B cells induce T-cell exhaustion, impairing cytokine production and enabling tumor immune evasion. In the immunosuppressive tumor microenvironment, these cells disrupt immunological synapse (IS) formation, critical for T-cell activation. Soluble factors from CLL cells inhibit IS formation, pointing to an indirect mechanism of T-cell suppression. Identifying these molecules could enable therapies to restore T-cell activity. We analyzed the IS-suppressive effects of soluble factors from various B-cell malignancies, including CLL, hairy cell leukemia (HCL), Hodgkin lymphoma (HL), and non-Hodgkin lymphoma (NHL). Despite their distinct features, all these cancers impair T-cell responses through shared mechanisms. Conditioned supernatants from tumor cells isolated from patient samples of these malignancies were used to culture cytotoxic T cells (CTLs). We observed that all tumor cell types induced CTL exhaustion, as evidenced by elevated expression of PD-1, LAG-3, and CTLA-4, albeit with varying intensity. Additionally, tumor cell supernatants potently suppressed the capacity of CTLs to form IS. Multiplex ELISA assays revealed that each malignancy exhibits a complex and distinct panel of released cytokines. Uncovering the molecular signature of T-cell suppressive soluble factors in hematologic malignancies may potentially pave the way for the identification of novel therapeutic targets for these still-incurable diseases. Supported by PRIN-2022 PNRR P2022PSMX4_001. Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)

BACKGROUND:The authors previously demonstrated that in peripheral blood (PB) of chronic myeloid leukemia (CML), patients' leukemia stem cells (LSCs) CD26+ are detectable by flow cytometry at diagnosis, during tyrosine kinase inhibitor (TKI) therapy, and during treatment-free remission. METHODS:This study presents results of a prospective multicenter study including 242 newly diagnosed CML patients monitored for PB CD26+ leukemic stem cells (LSCs) quantification from diagnosis up to 24 months of TKI treatment. RESULTS:The bulk of CD26+ LSCs at diagnosis varied between patients with a median value of 7.14 cells/µL. During TKI treatment, it has been observed their consistent and rapid reduction without statistical differences according to type of first-line TKI. Instead, a significant correlation between a low amount of CD26+ LSCs at diagnosis and an optimal molecular response at 3, 12, and 24 months was documented (p = .03, p = .004, and p = .009, respectively). Three tertiles of CD26+ LSCs correlating to molecular response were identified: <3.21 cells/µL; between 3.21 and 19.21 cells/µL; and >19.21 cells/µL. The incidence of patients with optimal response was higher in the first CD26+ LSCs tertile respect to the third one (p = .027, p = .015, and p = .079, respectively) at all time points (3, 12 and 24 months). CONCLUSIONS:This study demonstrated a correlation between the amount of CD26+ LSCs at diagnosis and the molecular response, suggesting that the number of CD26+ LSCs at diagnosis could represent an additional tool for predicting TKI response.

Background Thrombocytopenia is a frequent hematologic abnormality in patients with myelodysplastic syndromes (MDS), occurring in approximately 40–60% of cases. It has been consistently associated with adverse clinical outcomes and is incorporated into the Revised International Prognostic Scoring System (IPSS-R), where platelet (PLT) counts <100,000/mm³ indicate a higher-risk disease category. In patients with lower-risk MDS (LR-MDS), the prognostic significance of thrombocytopenia is less well defined but has been incorporated in a prognostic model by the MD Anderson Cancer Center together with anemia, age, marrow blast percentage, and cytogenetic risk (Garcia-Manero G et al. Leukemia 2008). The objective of this study was to evaluate the prognostic impact of thrombocytopenia at diagnosis on overall survival (OS) and leukemia-free survival (LFS) in a large cohort of patients with LR-MDS enrolled in the registry of the Italian Foundation for the Study of Myelodysplastic Syndromes (FISiM). Methods We retrospectively analyzed clinical and hematologic data from patients diagnosed with LR-MDS between January 1, 2007, and October 29, 2024, and included in the nationwide FISiM registry. All patients were classified according to the 2016 WHO diagnostic criteria. Categorical variables were compared using the chi-square test. OS and LFS were estimated using the Kaplan-Meier method and compared by log-rank test. Cox proportional hazards regression models were used for multivariate analyses to identify variables independently associated with outcomes. A two-tailed p-value <0.05 was considered statistically significant. Results A total of 2,213 patients with LR-MDS were included in the analysis. Of these, 610 patients (27.6%) presented with PLT counts <100,000/mm³ at diagnosis, while 1,603 (72.4%) had counts ≥100,000/mm³. The cohort comprised 1,314 males and 899 females, with a median age of 75 years (range: 19–98). The median follow-up was 32.1 months. The overall median OS was 80.0 months, with 1-, 2-, and 3-year OS rates of 96%,87% and78%, respectively. According to WHO classification, MDS with multilineage dysplasia (MDS-MLD) was the most common subtype in the thrombocytopenic group (61% vs. 38%), followed by MDS with single-lineage dysplasia (15% vs. 28%). In univariate analysis, thrombocytopenia was significantly associated with inferior OS and LFS (p<0.001 for both comparisons). This finding was also correlated with low megakaryocyte cellularity and megakaryocytic dysplasia. Interestingly, thrombocytopenia was more frequently observed in patients with hemoglobin levels >10 g/dL, suggesting it may define a distinct biological subgroup. In multivariate analysis, PLT <100,000/mm³ remained an independent predictor of worse OS (hazard ratio [HR] 1.25; 95% confidence interval [CI]: 1.15–1.37; p<0.001), along with older age, female sex, hemoglobin <10 g/dL (all p<0.001), and marrow blasts ≥5% (p=0.003). Leukemic transformation occurred in 65 patients (10.7%) in the thrombocytopenic group and in 131 patients (8.2%) with PLT ≥100,000/mm³ (p=0.06). The median time to progression was significantly shorter in thrombocytopenic patients: 139.5 months (95% CI: 129.3–149.7) vs. 155.7 months (95% CI: 149.6–161.8); p=0.005. Overall, 723 patients (32.7%) died, with a higher mortality observed in the thrombocytopenic group (36.4% vs. 31.3%). In the multivariate analysis for LFS, PLT <100,000/mm³ remained an independent adverse factor (HR: 1.20; 95% CI: 1.02–1.40; p=0.03), together with female sex (p=0.003) and blast count ≥5% (p<0.001). Conclusions Our study confirms that thrombocytopenia at diagnosis is an independent predictor of worse overall and leukemia-free survival in patients with lower-risk MDS. These findings underscore the importance of incorporating platelet count into risk stratification strategies even within the LR-MDS population. Closer monitoring and potentially earlier therapeutic intervention should be considered in this subgroup to improve clinical outcomes.

Background Clinical decisions for patients with relapsed/refractory multiple myeloma (RRMM) are challenging and assessment of frailty is critical to inform treatment strategies. The International Myeloma Working Group Frailty Index (IMWG-FI) is widely used in patients with newly diagnosed multiple myeloma (NDMM), given its ability to identify patients with distinct survival outcomes. However, very limited evidence exists on its prognostic value in the setting of RRMM and on the potential role of patient-reported outcomes (PROs) in the definition of frailty this setting. Objective The primary objective of this analysis was to investigate the prognostic value for overall survival (OS) of the IMWG-FI in patients with RRMM. A secondary objective was to examine whether PROs data could provide prognostic information for OS beyond the IMWG-FI and other key sociodemographic and clinical factors. Methods Adult patients with RRMM were enrolled in an international prospective cohort observational study by the GIMEMA. Eligibility criteria also included the availability of all the individual components of the geriatric assessment needed to calculate the IMWG-FI and a PRO assessment including the EORTC QLQ-C30 and the QLQ-MY20. OS was defined as the time from study entry to death from any cause. The prognostic value of the IMWG-FI was evaluated using Kaplan-Meier estimates and log-rank tests to compare OS across IMWG-FI groups. Additionally, we investigated the prognostic value of EORTC QLQ-C30 and QLQ-MY20 scales using univariate and multivariate Cox regression models. The initial univariate model included the following key variables: sex, time since initial MM diagnosis, living arrangements, education level, number of previous therapy lines, transplantation, best response to previous therapies, and IMWG frailty group. Likelihood ratio tests and C-indexes were used to evaluate potential improvement in prognostic information when adding PROs to the IMWG-FI. A bootstrap resampling procedure (1000 iterations) was applied to evaluate prognostic importance of each variable through inclusion frequencies across bootstrap generated samples. Results Between November 2017 and September 2022, 511 patients were enrolled from 31 centers. Patients had a median age at study entry of 69.8 years, and 287 (56%) were male. Median time since initial diagnosis of MM was 5 years (IQR 3- 7), 428 (84%) patients had received ≥2 lines of therapy before study entry, and 201 (39%) had at least one comorbidity. According to the IMWG-FI, 270 (52.8%) patients were classified as fit, 116 (22.7%) as intermediate, and 125 (24.5%) as frail. The 3 original IMWG-FI groups had distinct OS (p=0.001). However, similar OS were observed for the fit and the intermediate group, leading to the development of a two-tier classification by grouping fit/intermediate vs. frail. Indeed, well distinct median OS were observed for these two newly derived groups, being 38 and 33 months for fit/intermediate and frail patients (p<0.001), respectively. OS probabilities at 1, 2, and 3 years were clearly differentiated between the two IMWG-FI groups. For example, the two-year OS probabilities were 75.2%. and 62.1%, for fit/intermediate and frail patients, respectively. Prognostic analysis of PROs data indicated that physical functioning (PF) had the highest inclusion frequency (90%) across the 1000 bootstrap generated samples, thereby pointing to the prognostic importance of this variable. Multivariate analysis adjusting for key potential confounders, including IMWG-FI, showed that PF remained independently associated with OS (HR 0.933, 95% CI 0.875–0.995; p=0.034), which translates into a 7% increase in the hazard of death for every 10-point decrease (worsening) in the EORTC QLQ-C30 PF scale. Given these results, we investigated whether adding PF to a survival model based only on IMWG-FI (fit/intermediate vs. frail) could enhance its predictive accuracy. This resulted in a statistically significant increase in the likelihood ratio test (p=0.006) with the addition of PF, along with an improvement in the C-index (from 0.56 to 0.61). Conclusions In the setting of RRMM, the IMWG-FI is able to well discriminate two patients' groups with distinct OS. Our findings also indicate that patient-reported PF provides prognostic information for OS beyond the IMWG-FI, thereby laying the groundwork for the future development of a simplified patient-centric frailty index which would include PROs.

BACKGROUND:Improved outcome has been reported in chronic myeloid leukemia (CML) patients treated with tyrosine kinase inhibitors (TKIs) in sponsored trials. METHODS:This is a multicenter prospective cohort study of consecutive patients with newly diagnosed chronic phase CML from 19 regions in Italy. Baseline treatments and prognostic factors on time to first optimal molecular response (≥ molecular response 3, MR3), time to disease progression, time to death from CML, and overall survival (OS) were analyzed using multivariable Fine and Gray models. RESULTS:The authors included 1433 CML patients: 49% (median age, 70 years) treated with frontline imatinib (IMA), and 51% treated with second-generation TKIs (2G-TKIs; median age, 52 years). EUTOS long-term survival (ELTS) was low in 68.1% of 2G-TKIs patients, compared to 50.4% of IMA patients. Faster molecular responses were observed with 2G-TKIs within the first 6 months and maintained thereafter (subhazard ratio [sHR], 1.31; 95% confidence interval [CI], 1.15-1.50). Female gender and low ELTS risk had faster time of response. Achieving major molecular response (MMR or MR3) was associated with reduced risk of progression at 6 and 12 months. Overall, 41 patients progressed without differences between IMA and 2G-TKIs. Intermediate and high risk ELTS showed higher risk of progression and death from CML. Twenty-two CML-related deaths (16.5%) occurred mostly in the first 2 years from diagnosis, higher in 2G-TKIs patients (sHR, 1.75; 95% CI, 0.52-5.87). OS at 5 years was 88% with no clear differences between IMA and 2G-TKIs treatment after adjustment for potential confounders. CONCLUSIONS:The study confirms faster responses with 2G-TKIs compared to IMA but similar clinical outcomes and a strong prognostic effect of ELTS.

Background/Objectives: The aim of this study was to assess the unmet needs of myelodysplastic neoplasm (MDS) patients and their caregivers, focusing on how these needs impact quality of life (QoL) and daily functioning. MDS predominantly affects older adults. It is often complicated by severe red blood cell transfusion-dependent anemia and may require frequent hospital visits, conferring a substantial burden on patients and caregivers. Methods: A national survey was conducted between June 2022 and May 2023 in 46 hematology centers across Italy, involving 259 patients and 105 caregivers. The survey included validated QoL tools (QOL-E and HM-PRO) to measure the impact of disease and treatments on health-related QoL and symptoms. Results: Of the 259 patients surveyed, 42% were transfusion-dependent, with 45% reporting distress related to hospital travel, which was significantly associated with lower QoL scores (QOL-E physical score 50.0 vs. 62.5, p < 0.001). Transfusion dependency led to worse outcomes across physical, emotional, and social domains (HM-PRO Part A score 59.8 vs. 23.7, p < 0.001). Anxiety affected 66% of patients, while 56% reported feeling emotionally distressed. Forty-eight percent of patients required a caregiver, and among caregivers, 29% reported significant disruption to their work, including changing their job or reduced hours. Patients requiring frequent hospital visits showed notably worse QoL scores (HM-PRO emotional score 56.8 vs. 31.8, p < 0.001). Conclusions: This study identified substantial unmet needs for MDS patients, particularly in addressing the heavy burden of transfusions and hospital visits. Both patients and caregivers experienced significant impact on daily life and on QoL, highlighting the urgent need for treatments that reduce hospital dependency, improve patient outcomes, and alleviate the caregiver burden.

Health-related quality of life (HRQoL) of patients with myeloproliferative neoplasms (MPNs) may be impaired across several domains. In this multicenter observational study, we evaluated HRQoL and symptoms in a cohort of MPN patients with validated measures, including the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire-Core 30 (EORTC QLQ-C30), the Myeloproliferative Neoplasm Symptom Assessment Form Total Symptom Score (MPN-SAF TSS), and the Functional Assessment of Chronic Illness Therapy-Fatigue Scale (FACIT-Fatigue) questionnaire. The primary objective was to compare the HRQoL profile of patients, by disease subtype, with that of the general population according to the EORTC QLQ-C30. A total of 572 patients with essential thrombocythemia (ET, n = 228), polycythemia vera (PV, n = 207), and myelofibrosis (MF, n = 137) were assessed. Worse statistically and clinically significant differences were observed for role functioning (ET: ∆ = 8.9, P < 0.001; PV: ∆ = 11, P < 0.001; MF: ∆ = 16.7, P < 0.001) and fatigue (ET: ∆ = 5, P < 0.001; PV: ∆ = 8.3, P < 0.001; MF: ∆ = 11.5, P < 0.001) in all three diagnostic groups. However, patients with MF also reported impairments in other important health domains. Fatigue was the most frequently reported and burdensome symptom, with greater severity correlating with a broader and more complex array of associated symptoms. Our analysis also revealed a substantial underestimation of symptoms by treating hematologists in paired physician-patient reports. Current findings may help to disentangle specific HRQoL limitations and symptomatology experienced by patients with MPNs, and underscore the importance of incorporating patient-reported outcomes into routine practice to better reflect the patient's perspective of the disease and treatment-related burden.

INTRODUCTION:The International Myeloma Working Group Frailty Index (IMWG FI) is one of the most used frailty assessment tools in patients with multiple myeloma (MM). A patient-centered frailty tool based on patient-reported outcomes (PROs) has been recently proposed for patients with relapsed/refractory MM (RRMM): the Patient-Reported Frailty Phenotype (PRFP). This cross-sectional analysis aimed to replicate the PRFP within a real-world setting and to describe health-related quality of life (HRQoL) profiles based on this new patient-centered frailty classification. MATERIALS AND METHODS:This analysis was based on baseline data from a multi-center prospective observational study that included adult patients with RRMM. Frailty was assessed using the IMWG FI, and HRQoL was evaluated with the EORTC QLQ-C30 and its myeloma module (QLQ-MY20). For this analysis, the PRFP was also calculated, and Cohen's kappa was computed to examine the agreement between the two frailty assessment approaches. Descriptive statistics were used to investigate the HRQoL profiles of patients classified as frail, pre-frail, and fit. RESULTS:Five hundred eleven patients were enrolled. The median age was 69.8 years, and 44 % were female. According to IMWG FI and PRFP, 24.5 % and 25.1 % of patients were classified as frail, with a weighted Cohen's kappa of 0.27, indicating fair agreement. Frail patients, as classified by the PRFP, reported higher treatment side effects, disease symptoms, and worse future perspectives and body image compared to pre-frail and fit patients. DISCUSSION:Current findings suggest that the PRFP may be a valuable tool to assess frailty in patients with RRMM. However, further prospective studies are needed to better understand the value of PROs in a more accurate assessment of frailty in the RRMM setting. TRIAL REGISTRATION:ClinicalTrials.gov: NCT03190525.

Haematological malignancies represent a heterogenous group of diseases, encompassing lymphomas, leukaemia, and multiple myeloma. Among these, the involvement of human endogenous retroviruses (HERVs) has been most consistently reported in lymphoma, while their role in leukaemia and multiple myeloma remains limited. This study investigated the humoral response to the envelope proteins of HERV-K and HERV-H in the peripheral blood of patients with multiple myeloma and non-Hodgkin lymphoma and assessed HERV-K envelope gene expression through an approach combining indirect ELISA and quantitative PCR. The study revealed an increased humoral response against the HERV-K envelope epitope in patients with non-Hodgkin lymphoma compared to matched healthy controls. However, no differences were observed in patients with multiple myeloma. Although limited to the humoral level, these findings support the relevance of HERV-K specific immune responses in non-Hodgkin lymphoma and provide a rationale for further investigation.

Introduction. IMIDs occur in about 15% of MDS and CMML (Mekinian et al, Rheumatology (2016;55:291-300), but the precise relationship between those disorders remains uncertain. The impact of IMIDs on OS in those patients is also still unclear. Published data may include methodological biases, especially the consideration of IMIDs as a time-dependent event, as it can be diagnosed before, concomitantly, or after MDS/CMML diagnosis. We studied the impact of IMIDs on OS in MDS/CMML, considering this time-dependent relationship. Methods. We constructed a multicenter European cohort of non-VEXAS patients, to which 5 centers participated (University Hospitals of Bordeaux, Vall d'Hebron (Barcelona), Sassari (Italy), Paris Saint-Louis, and Poitiers (France)). A first analysis compared MDS patients who, at MDS/CMML diagnosis, were already diagnosed with IMID (irrespective of diagnostic interval between the 2 disorders) versus those without IMID; a second analysis was performed in the patient subgroup without IMIDs at MDS/CMML diagnosis to estimate the impact of developing IMIDs as a time-dependent event. Cox multivariate regressions included R-IPSS variables, age, and gender. An updated Cox regression was used for the second analysis. Models were stratified on the center. Multiple imputation was performed to manage missing values. HR and confidence intervals (CI) were calculated by bootstrapping the imputed datasets. Results. Data from 1132 patients (810 MDS and 322 CMML) diagnosed between 2000 and 2022 were collected. Median age was 74 (range 21-95), with 61% males; R-IPSS was very high in 6.6%, high in 12%, intermediate in 29%, low in 35%, and very low risk in 17%. Molecular biology was unavailable in close to 50% of the patients, diagnosed before NGS techniques were available, so that the WHO 2016 classification was used. An IMID was identified in 196 patients (17.3%), including autoimmune cytopenias (20%), connective diseases (6.6%), inflammatory arthritis (32%), neutrophilic dermatosis (7.7%), vasculitis (21%), and other less frequent IMIDs (13%). In some male patients diagnosed before 2020 and in whom UBA1mutation analysis was not performed subsequently, VEXAS could however not be excluded. No difference was found in aseline characteristics between those with and without IMIDs, except for neutrophils, which were higher in the IMIDs group (median 3.0 versus 2.3 G/L, p=0.007). When comparing patients with or without IMID at MDS/CMML diagnosis (first analysis, n=1003) presence of an IMID had no impact on OS in the multivariate analysis (HR 1.02; 95% CI 0.77-1.35), while prognostic factors for OS were, as expected, lower Hb level (HR 1,11), higher log(neutrophils) (HR 1.14), platelets < 50 G/L (HR 1.66,), marrow blasts 5-9% (HR 1.68), and 10-19% (HR 1.66) high-risk (HR 2.41) and very-high risk karyotype (HR 3.88). In the second analysis (n=966), however, the occurrence of IMID after MDS/CMML diagnosis had a negative impact on OS (HR 1.84; 95% CI 1.05-3.20) in the multivariate time-dependent analysis. Other prognostic factors remained generally similar including older age (HR 1.03), lower Hb level (HR 1.12), higher log(neutrophils) (HR 1.26), platelets < 50 G/L (HR 1.69), marrow blasts 10-19% (HR 1.68), high-risk (HR 2.11; 95% CI 1.35-3.42) and very-high risk karyotype (HR 3.83; 95% CI 2.39-6.26). No specific type of IMID was associated with worse survival.Conclusion. Development of an IMID during the course of MDS/CMML negatively impacts OS. We are currently reviewing in detail possible worsening of MDS/CMML characteristics (regarding WHO classification, IPSS-R, karyotype, and, in more recent patients, somatic mutations) just before or after diagnosis of IMID, potentially explaining poorer outcome. This would possibly also contribute to better knowledge of the physiopathological link between IMIDs and MDS/CMML, and in particular whether occurrence of an inflammatory disease can lead to MDS/CMML progression or if clonal evolution of MDS/CMML can trigger or worsen an IMID, as suggested by our group (Zhao et al. Leukemia. 2023; 37:1186-1190). These two explanations may not be exclusive.

Myelodysplastic syndromes (MDS) are associated with an increased risk of progression to acute myeloid leukemia (AML). Disease evolution involves the sequential acquisition of somatic mutations and clonal selection over time. The Molecular International Prognostic Scoring System (IPSS-M) represents the state-of-the-art for risk stratification at diagnosis but its applicability in a longitudinal context has not been validated. The FISIM-NGS-MDS study was designed to prospectively collect longitudinal clinical and molecular data (from peripheral blood, PB) to investigate clonal evolution and identify patterns predictive of progression (NCT04212390). Methods: Adult patients with a diagnosis of MDS according to the 2016 WHO Classification were prospectively enrolled at diagnosis at 28 Italian hospitals. PB samples were collected at diagnosis, annually during follow-up, before/after treatment, and at disease progression or AML evolution. Targeted NGS was performed at Humanitas Research Hospital. To validate mutation detection accuracy, a subset of PB samples was analyzed in parallel with paired bone marrow (BM) samples. Dynamic validation of the IPSS-M was performed using time-dependent Cox regression models, with model performance assessed by concordance index (c-index). Clonal evolutionary trajectories were reconstructed using cancer cell fractions (CCF), derived from copy number–adjusted variant allele frequencies (VAF), focusing on mutations occurring in at least 1% of the study population. Patient-level directed acyclic graphs were generated through pairwise CCF comparisons and aggregated into cohort-level temporal graphs. A minimum-agony ranking algorithm was applied to infer the most likely order of mutation acquisition. Baseline findings were validated using the original IPSS-M development cohort. Longitudinal validation, comparing inferred versus observed evolutionary directionality, was performed using serial sequencing data from the FISIM cohort. Finally, a time-dependent Cox model with 100-iteration bootstrap validation was fitted to identify CCF dynamics consistently associated with AML evolution. The study included 1,002 patients with a median age at diagnosis of 74 years; 315 patients (31%) were classified as IPSS-M Moderate High or higher at baseline. Analysis of paired PB/BM samples from 115 patients revealed 97.1% concordant variants. The few discordant variants had VAF <3%. The rate of discordant events remained low (3.8%) for variants with VAF <5%. VAF for BM was slightly higher than paired PB (median difference 2%, p<0.01). IPSS-M risk classification changed over time in 217 patients (28.6%). Compared with baseline assessment, dynamic IPSS-M showed improved predictive performance across all clinically relevant outcomes, with c-index for overall survival of 0.80 vs 0.74 for baseline IPSS-M, and for leukemia-free survival 0.81 vs 0.77, respectively. Evolutionary modelling using CCF at diagnosis identified 46 recurrent mutation trajectories (present in >10 patients), defined as pairs of co-occurring mutations with a consistent temporal relationship—i.e., the presence of an earlier mutation increased the likelihood of acquiring a subsequent one. Longitudinal validation confirmed consistent directionality for 29 out of 46 trajectories. External baseline validation in the original IPSS-M cohort (n=2,957) was concordant, with only 5 trajectories showing divergent directionality. Time-dependent Cox regression, adjusted for IPSS-M and IPSS-R, showed that CCF for TP53, RUNX1, TET2, PHF6, U2AF1, STAG2, and PTPN11 independently predicted AML evolution. Each gene was retained in over 30% of bootstrap iterations. All associations had a positive hazard direction, suggesting that progressive clonal expansion and/or acquisition of new mutations within these evolutionary trajectories correlates with worse clinical outcomes. Conclusions. Dynamic IPSS-M validation showed superior prognostic performance vs conventional assessment, supporting its use for re-evaluating patient risk over time. CCF-based evolutionary modeling reconstructed mutation sequences and identified genes whose clonal expansion independently predicts AML evolution. Longitudinal clonal monitoring with PB samples was reliable, enabling early, non-invasive identification of high-risk trajectories and improved patient management. Overall, these findings support the concept that novel MDS prognostic tools should be based on longitudinal data.

Background: In a cross-sectional study we previously demonstrated that in peripheral blood (PB) of chronic myeloid leukemia (CML) patients leukemia stem cells (LSCs) CD26+ are detectable by flow-cytometry at diagnosis, during TKI therapy and during treatment free remission (TFR). No prospective data are available regarding the behavior of PB CD26+LSCs from diagnosis and the correlation, if any, between the bulk of this staminal compartment at diagnosis with the attainment of molecular response. Methods: We here present final results of a prospective Italian multicenter study including newly diagnosed chronic phase (CP) CML patients centrally monitored by flow-cytometry for PB CD26+LSCs quantification from diagnosis up to 24 months of TKI treatment. Results: 242 consecutive CP-CML patients were enrolled (132 imatinib, 72 nilotinib and 38 dasatinib). The bulk of CD26+LSCs at diagnosis varied between patients with a median value of 7,1454 cells/µl (range 0,0126-698,746 cells/µl; IQR 2,18-33,26 cells/µl). During TKI treatment, we observed a consistent and rapid reduction of them achieving median values of 0,0132 cells/µl (IQR 0-0,034 cells/µl), 0,011 cells/µl (IQR 0-0,031 cells/µl) and 0,0071 cells/µl (IQR 0-0,0259 cells/µl) at 3, 12 and 24 months, respectively. No statistically significant differences in terms of CD26+LSCs log-reduction were noted according to the type of TKI treatment at any time points evaluated. However, a significant correlation between a low amount of CD26+LSCs at diagnosis and an optimal molecular response at 3, 12 and 24 months (BCR::ABL1<10% and BCR::ABL1<0.1%, respectively) was documented. Indeed, CML patients with optimal molecular response at 3 months had a median CD26+LSCs of 6,21 cells/µl (IQR 1,79-31,50 cells/µl) while suboptimal responders patients showed a median of 19,87 cells/µl (IQR 5,37-39,81 cells/µl) (p=0.03); moreover, patients with BCR::ABL1<0.1% at 12 and 24 months, revealed median CD26+LSCs at diagnosis of 5,50 cells/µl (IQR 1,81-22,64 cells/µl) and 6,05 cells/µl (IQR 1,79-29,90 cells/µl) respectively, compared to suboptimal responders showing 16,87 cells/µl (IQR 2,82 -71,77) and 20,52 cells/µl (IQR 4,24-106,91) (p=0.004, p=0.009). Additionally, evaluating the cohort of CML patients who switched TKI treatment after failure with respect to patients who did not change TKI, we observed that the former had a significantly higher median CD26+LSCs at diagnosis (14,59 cells/µl; IQR: 3,76-46,00 cells/µl) compared to the no-switch group (median of 5,82 cells/µl; IQR: 2,35-26,70 cells/µl) (p=0.034). Three ranges of CD26+LSCs correlating to molecular response were identified: <3,21 cells/µl (1° tertile); between 3,21-19,21 cells/µl (2° tertile); >19,21 cells/µl (3° tertile). In particular, considering the molecular response at 3 months the incidence of CML patients with BCR::ABL1<10% was 93.5% in the first CD26+LSCs tertile, while 78.8% in the third tertile (p=0.027). At 12 months the incidence of optimal response in the first tertile was 78.5% and 62.8% in the third one (p=0.015). At 24 months the two incidences were 90.8% and 77.9%, respectively (p=0.079). Conclusions: This prospective study demonstrated a rapid rate of reduction of CD26+LSCs during TKI treatment, however confirming their long-lasting persistence even if at very low levels. For the first time, a correlation between the amount of CD26+LSCs at diagnosis and the response to TKI treatment, was documented. Given these results, the bulk of CD26+ LSCs at diagnosis could represent an easily and rapidly measurable, new prognostic tool for predicting TKI response.

Introduction: Pomalidomide represents the backbone for the treatment of relapsed/refractory Multiple Myeloma in combination with several agents as monoclonal antibodies and proteosome inhibitors. Since October 2020 three associations of Pomalidomide with monoclonal antibodies (Isatuximab, Daratumumab and Elotuzumab) have been available in Italy for the treatment of relapsed or refractory Multiple Myeloma (RRMM). The triplets Isatuximab -Pomalidomide-Dexamethasone (IsaPd) and Elotuzumab-Pomalidomide-Dexamethasone (EloPd) are licensed after two lines of therapy with lenalidomide and a proteosome inhibitor while Daratumumab association is available from second line of therapy. Here, we report preliminary efficacy and safety data for the combinations of monoclonal antibodies with Pomalidomide in a Sardinian study of RRMM patients as salvage therapy outside of controlled clinical trials. The primary objective of this study was to evaluate the toxicity profile and quality of respons e to EloPd, IsaPd and DaraPd administered as salvage therapy in a real-world setting. Methods: The 3 cohorts included 54 patients with RRMM from 4 Sardinian centers who received at least one cycle of IsaPd (20 patients), EloPd (15 patients) or DaraPd (19 patients) as salvage therapy between February 2021 and June 2024. Results: The median age of the 20 IsaPd-treated patients was 67.5 years (range 49-84 years), 74 years (range 59-82 years) for the EloPd cohort and 69 (36-81years) for DaraPd population; 50%, 80 and 53% were male, respectively (Table 1). The median number of prior therapies was 2 (range 2-4) for the IsaPd, 3 (range 2-7) for the EloPd and 2 (2-5) cohort; 13 (65%), 7 (46%) and 12 (63%) received prior autologous stem cell transplantation (ASCT), respectively. In the EloPd cohort 12 patients (80%) are more than 70 years and 66% shows a creatinine clearance less than 60 ml/min. Overall 4 patients out of 54 have a clearance creatinine ≤ 30 ml/min. Only 15 patients (28%) of the three cohorts started the treatments because of refractoriness. The median number of courses of IsaPd administered to date was 5 (range, 1-29). The overall response rate (ORR) was 53%, with 3 stringent complete remissions (sCR) (16%) and 2 complete response (CR) (10%). The median time to first response was 1 month, while the median time to best response was 2 months. In the EloPd population, it was documented ORR of 60%, with 1 sCR (7%), 3 CR (20%), and 3 very good partial remissions (VGPRs) (20%). A median time to first response of 1 month was also recorded in this population, whereas the median time to best response was 2 months. The ORR in the DaraPd populations was of 58%, with 2 sCR (11%), 4 CR (21%), and 5 VGPRs (26%). Renal insufficiency and age > 70 years had no significant impact on the likelihood of achieving a response in either population. A median time to first response of 1 month was also recorded in this population, whereas the median time to best response was 2 months. In the IsaPd cohort, after a median follow-up of 8 months (range 1-26 months), 16 patients discontinued treatment, 14 due to disease progression, 1 due to toxicity, and 1 due to patient choice). Eleven patients died (9 patients from progressive disease, 1 from secondary acute myeloid leukemia and 1 for Hodgkin Lymphoma). Common grade 3 or 4 adverse events were thrombocytopenia (20%), neutropenia (45%), anemia (15%), and infection (15%). Infusion reactions occurred in only 2 patients (10%) and were grade 1 or 2 without treatment discontinuation. In the EloPd population, the median follow-up time was 8 months (range 3-25 months). Eight patients discontinued treatment, 7 for progression and 1 for toxicity: 3 died due to disease progression. Grade 3 or 4 adverse events were fatigue (18%), thrombocytopenia (18%), neutropenia (18%), anemia (36%), and infection (63%). The median follow-up in DaraPd population was of 9 months (range 1-16 months), 3 patients discontinued treatment, 1 due to disease progression, 1 due to toxicity, and 1 due to refractoriness). Only 1 patient died from progressive disease. Common grade 3 or 4 adverse events were neutropenia (21%), anemia (15%), and infection (15%), fatigue (53%). Conclusions: Despite the limited number of cases, our preliminary real-world data confirm that the associations of Pomalidomide with monoclonal antibodies represent effective and safe therapies for RRMM patients, broadly consistent with results from controlled clinical trials.