ABSTRACT:Natural killer (NK) large granular lymphocytic leukemia (LGLL) is a rare lymphoproliferative disorder lacking definitive clonality markers, complicating diagnosis and distinction from reactive NK cell expansions. We previously proposed an NK clonality score with high diagnostic accuracy, but a subset of patients remained unclassified. In this multicenter international study, we refined and validated updated diagnostic criteria using independent training (n = 78) and validation (n = 57) cohorts from 3 national registries (United States, Italy, and France). The revised framework integrates NK score parameters with CC motif chemokine ligand 22 (CCL22) mutations and bone marrow biopsy (BMB) findings. Four major criteria were defined: NK cell count of ≥1.0 × 109/L, killer-cell immunoglobulin-like receptor restriction, CD94/NKG2A overexpression, and somatic mutations in STAT3, TET2, or CCL22, the latter newly introduced. In the training cohort, 50 patients were classified as having NK-LGLL by an NK score of >4, 18 had intermediate scores (2-3), and 10 were diagnosed as reactive proliferations. CCL22 mutations were identified in 16 patients (20%), including 5 with intermediate scores who were reclassified as NK-LGLL; BMB supported the diagnosis in 2 additional cases, resulting in 57 NK-LGLL overall. These patients exhibited more cytopenias, higher treatment needs, and greater transfusion requirements than patients with alternative diagnoses. In the validation cohort (25 NK-LGLL and 32 reactive cases), CCL22 mutations were detected in 5 NK-LGLL (20%). Altogether, incorporation of CCL22 mutations reduced the fraction of unclassified patients, improved diagnostic sensitivity without compromising specificity, and may decrease reliance on invasive procedures. These revised international criteria represent a step toward standardized, molecularly guided NK-LGLL diagnosis.

Bone marrow biopsy (BMB) is a common procedure for the diagnosis, staging, and evaluation of several hematological diseases. BMB often causes anxiety and pain even when local anesthesia (LA), premedication, or an equimolar mixture of oxygen and nitrous oxide (EMONO) are used. Hypnosis has emerged as a promising non-pharmacological method to reduce anxiety and pain during invasive procedures. QUALHYBOM, a prospective, multicenter, randomized, open-label study was conducted to assess whether adding hypnosis to LA (LA + H) could improve patient comfort during BMB compared to LA alone. QUALHYBOM included 180 BMB-naive patients with suspected or confirmed hematological disorders, randomized (LA group n = 91, LA + H group n = 89) from June 2023 to November 2024 across four French hematological departments. The primary endpoint was patient comfort rated on a scale from 0 (no comfort) to 10 (maximum comfort), evaluated immediately after the procedure. Secondary outcomes included pain, anxiety (before and after BMB), practitioner comfort, and use of EMONO. Patient groups were well-matched at enrolment. Median comfort scores were 9 vs. 5, pain scores 2 vs.6, and post-procedure scores 1 vs.5, in the LA + H vs.LA groups, respectively, both with p < 10⁻4. Finally, the operator comfort was higher (10 vs. 7), and EMONO requirement lower (3

Abstract Background CAR T-cells have been approved for the treatment of relapsed/refractory (R/R) diffuse large B-cell lymphoma (DLBCL), mantle cell lymphoma (MCL), follicular lymphoma (FL), acute lymphoblastic leukemia (ALL), and multiple myeloma (MM). Although acute toxicities of CAR T-cells are well handled during hospitalization, delayed complications, including cytopenias, infections and relapses, are more challenging to manage after hospital discharge. To address this issue, a specific program (CARAMA) was initiated in Rennes University Hospital consisting in remote monitoring of CAR T-cell recipients by a specialized nurse. Methods Patients treated with CAR T-cells were followed remotely by a CARAMA nurse after hospital discharge. Adverse events and interventions were prospectively collected by the CARAMA nurse who called patients daily until day 21, twice a week until day 28, once a week until month 2, every other week until month 3, once a month until month 6, and once every 3 months until month 12 or until relapse/progression. Results Between September 2019 and December 2022, the program enrolled 150 patients with a median age of 64 years (range, 16-80). Patients received a median of 15 calls (range, 0-41). The CARAMA nurse recorded complications including cytopenias (111 patients, 73.5%) with transfusion (44 patients, 29.1%) and/or growth factor (110 patients, 72.8%) requirements, and infections in need of antibiotics (37 patients, 24.5%). Disease relapse was detected within CARAMA before a planned medical consultation in 24/65 patients (36.9%). The CARAMA nurse referred 75 patients (49.7%) to their general practitioner (N=71), hematologist (N=24) and/or other physician (N=29) and requested hospitalization for 41 patients (27.2%), due to infection (48.8%), profound cytopenias (14.6%), disease relapse (17.1%), and/or other causes (19.5%). Supportive measures were organized for 63 patients (41.7%). A survey at 3 months of the CARAMA program confirmed the high level of patient satisfaction. Conclusions Remote monitoring of patients treated with CAR T-cells, by a specialized dedicated nurse, allows early detection and management of delayed complications. It also saves medical time and resources. Finally, this close follow-up is reassuring for the patients and contributes to their well-being. These results support the development of CARAMA nurses in CAR T-cell centers.

AbstractChimeric antigen receptor T cells (CAR T cells) can induce prolonged remission in a substantial subset of patients with relapse/refractory lymphoma. However, little is known about patients' life after CAR T‐cell therapy. We prospectively assessed the multidimensional recovery of lymphoma patients in remission, before leukapheresis, before CAR T‐cell infusion, and 3, 6, and 12 months thereafter. Validated tools were used to measure lymphoma‐related and global health‐related quality of life (HRQoL; Functional Assessment of Cancer Therapy‐Lymphoma [FACT‐Lym] and EQ‐5D‐5L), cognitive complaint (FACT‐Cognition), fatigue (FACIT‐Fatigue subscale), psychological status (Hospital Anxiety and Depression Scale, Post‐Traumatic Check List Scale), and sexuality (Relationship and Sexuality Scale). Beyond 12 months of remission, we also surveyed physical, professional, sexual, and general life status. At 3, 6, and 12 months, 53, 35, and 23 patients were evaluable, respectively. Improvement in lymphoma‐related HRQoL was clinically relevant at 3, 6, and 12 months with a mean change from baseline of 10.9 (95% confidence interval [CI]: 5.8; 16.1), 12.2 (95% CI: 4.2; 20.1), and 11.72 (95% CI: 2.06; 21.38), respectively. Improvement in global HRQoL, fatigue, and anxiety was clinically relevant, but 20%–40% of patients experienced persistent fatigue, psychological distress, and cognitive complaints over time. Beyond 12 months after CAR T cells, 81.8% of 22 evaluable patients were satisfied with their daily life. Physical activity, professional, sexual, and global well‐being had returned to prediagnosis levels in nearly half of the patients. We found an improvement in HRQoL after CAR T‐cell therapy including anxiety, depression, sexual satisfaction, and general well‐being. However, not all patients recover a “normal life.” Further research is needed to determine which patients are at risk of quality‐of‐life impairment to improve recovery after CAR T‐cell infusion.

Chimeric antigen receptor (CAR) T-cell therapy has transformed the care of patients with relapsed/refractory B-cell-derived hematologic malignancies. To date, six CAR T-cell therapies, targeting either CD19 or B-cell maturation antigen, have received regulatory approval. Along with the promising survival benefit, CAR T-cell therapy is associated with potentially life-threatening adverse events, including cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome. While clinical trials evaluating CAR T-cell therapy consistently report the incidence of these adverse events, most trials do not collect health-related quality of life (HRQoL) data. As such, the impact of the CAR T-cell therapy process and related adverse events on the physical and psychological well-being of patients remains uncertain. HRQoL and other patient-reported outcome (PRO) assessments in patients with relapsed or refractory hematologic malignancies are of utmost importance, as individuals may have unmet needs and a high demand for tolerable therapy if a cure is not obtained. In addition, it is important to standardize methods of data collection to better assess the impact of CAR T-cell therapy on quality of life, optimize patients' care and costs, and enable comparisons between different studies. We conducted a literature search up to June 2023 to identify the HRQoL tools used in clinical trials and in real-world studies investigating CAR T-cell therapy in patients with lymphomas or leukemias. In the present comprehensive review, we summarize the most commonly used CAR T-cell specific and non-specific HRQoL tools and discuss how the use of HRQoL and other PRO tools may be optimized.

Large gran u lar lym pho cyte (LGL) leu ke mia is a rare lymphoproliferative dis or der char ac ter ized by an expan sion of clonal T or nat u ral killer lym pho cytes. Neutropenia-related infec tions and ane mia rep re sent the main man i fes ta tions. LGL leu kemia is frequently associated with autoimmune disorders such as rheumatoid arthritis, Sj & ouml;gren's syndrome, auto immune endocrinopathies, vasculitis, or autoimmune cytopenia. Recent advances in the phenotypic and molecular characteriza tion of LGL clones have underscored the piv otal role of a chronic anti genic stim u la tion and a dysregulation of the Jak/STAT signaling pathway in the pathophysiology linking leukemic-cell expansion and autoimmunity. In more than half of patients, there is a somatic STAT3 muta tion. The dis ease is char ac ter ized by an indo lent course, but approx i ma tely half of all patients will even tu ally require ther apy. The first-line treat ment for LGL leu ke mia is his tor i cally based on immunosuppressive agents (methotrexate, cyclophosphamide, or cyclosporine). However, cytokines blocking molecules or Jak/STAT inhib i tors rep re sent a new con cep tual ther a peu tic approach for LGL leu ke mia. In this review, we pres ent an over view of the spec trum of LGL pro lif er a tions, poten tial links between LGL expan sion and auto im mu nity, and ther a peutic approaches.

Large granular lymphocytic (LGL) leukaemia is a rare chronic lymphoproliferative disorder characterized by an expansion of cytotoxic T or NK cells. Despite a usually indolent evolution, most patients will require a treatment over the course of the disease because of cytopenia or symptomatic associated autoimmune disorders. First-line treatment is based on immunosuppressive agents, namely cyclophosphamide, methotrexate and ciclosporin. However, relapses are frequent, and there is no consensus on the management of relapsed/refractory patients. The implication of the JAK/STAT pathway in the pathogenesis of this disease has prompted our group to propose treatment with ruxolitinib. A series of 21 patients who received this regimen is reported here. Ten patients (47.6%) were refractory to the three main immunosuppressive drugs at the time of ruxolitinib initiation. Ruxolitinib yielded an overall response rate of 86% (n = 18/21), including 3 complete responses and 15 partial responses. With a median follow-up of 9 months, the median response duration was 4 months. One-year event-free survival and 1-year overall survival were 57% and 83% respectively. Mild side effects were observed. Biological parameters, notably neutropenia and anaemia, improved significantly, and complete molecular responses were evidenced. This study supports ruxolitinib as a valid option for the treatment of relapsed/refractory LGL leukaemia.

Topic: 35. Quality of life and palliative care Background: CD19 CAR-T cells have been approved for the treatment of relapse/refractory (R/R) lymphoma. Although CAR T-cells are able to induce prolonged remission (and potentially cure) in a significant number of patients, little is known about patients’ life after CAR-T cells. Here, we conducted a prospective study to evaluate health-related quality of life (HRQol), as well as physical, social and professional outcomes after CAR-T cell therapy. Aims: Primary objective was to describe HRQol changes during the first year after CAR-T cell treatment. Secondary objectives were to describe psychological factors and to evaluate satisfaction about normal life recovery. Methods: This prospective study was conducted at the University Hospitals of Rennes and Toulouse (France). All adult patients with R/R lymphoma treated with CAR T-cells, including Axicabtagene ciloleucel (axi-cel), Tisagenlecleucel (tisa-cel) and Brexucabtagene autoleucel (brexu-cel), were eligible. HRQol changes were assessed using the FACT-Lym questionnaire. Other questionnaires evaluated cognitive state (FACT-Cog), fatigue (FACT-Fatigue), anxiety and depression (HAD scale), post-traumatic stress disorder (PTSD), and sexuality (VICAN). Questionnaires were collected at baseline (before leucapheresis), immediately before CAR-T infusion, 3, and 6 months after CAR T-cell infusion. Patients were censored in case of relapse or death. Among patients in remission after 12 months, professional, physical, social, sexual and global life information were collected. Results: From March 2020 to August 2022, 59 patients were included in the study, including 46 LBCL (78.0%), 5 MCL (8.5%) and 8 FL (13.5%). Median age was 63 years (range, 19-78). Patients were treated with axi-cel (n=37, 62.7%), tisa-cel (n=18, 30.5%) or brexu-cel (n=3, 5.1%). The median follow-up after CAR T-cell infusion was 11 months. At 3 months and 6 months respectively, there were 53 and 38 patients still in remission. The FACT-Lym score showed a clinically relevant improvement of HRQoL at 3 and 6 months after CAR T-cell infusion, with a mean change from baseline of 10.94 points (95%CI 5.83;16.05) and 12.16 (95%CI 4.19;20.12), respectively. The fatigue subscale showed a clinically relevant improvement at 3 and 6 months, with a mean change from baseline of 5.87 points (95%CI 2.93;8.8) and 4.70 (95%CI 0.2;9.2), respectively. The FACT-Cog score did not show any clinically relevant change for perceived cognitive impairments. The HAD score showed that 43% and 24% of patients presented anxiety and depression at baseline, respectively, versus 26% and 7% after 6 months. Overall, 26% of patients presented a significant PTSD (i.e. >44) at 6 months. Sexual life satisfaction improved from 30% at baseline to 50% after 6 months. After 12 months with a median follow-up of 19 months, 22 patients were in remission. Among the working age population (n=10), 50% patients returned to work in a median time of 9 months (range, 3-13). All patients practiced a physical activity again, although less intensively. Overall, 54% of patients felt less fit than before initial diagnosis. Three patients out of 8 (37.5%) return to their social activity. Most patients were satisfied with their sexual life (68.2%) and, in comparison with before initial diagnosis 59.1% considered that it was back to normal. Overall, 54.5% of the patients considered that their life was back to normal, i.e. similar to what it was before lymphoma diagnosis. These results are shown in the figure below. Summary/Conclusion: Our study shows an clinically significant improvement in HRQol after CAR-T cells therapy with an improvement in anxiety, depression, sexual satisfaction.. Nevertheless, half of the working age population returned to work and 54% considered their life back to normal.Keywords: Lymphoma, Quality of life, CAR-T

Background: Extranodal NK/T cell lymphoma (ENKTCL) nasal type is a rare subset of peripheral T lymphoma characterized by a constant association to Epstein Barr Virus (EBV). Asparaginase containing regimen is the current standard of care in this pathology. Despite recent improvements in the therapeutic strategy, patient’s prognosis remains poor, especially for those who experienced relapsed or refractory (R/R) disease. Recent studies highlighted that immune escape mechanisms are involved in ENKTCL pathogenesis. Particularly, the frequent PD-L1 upregulation led to assess the efficacy of anti-PD1 (aPD1) therapy in small cohorts of R/R ENKTCL patients, with encouraging results. Thus, immune checkpoint blockade represents a promising approach for R/R ENKTCL to date. Aims: Our study aims to evaluate the efficacy of aPD1 therapy alone or in combination in 37 patients with R/R ENKTCL. We also performed a comparative analysis with a historic cohort of 38 patients treated for R/R ENKTCL before immunotherapy era. Methods: 37 patients from 24 French centers, with R/R ENKTCL treated with at least one cycle of aPD1 as salvage therapy between 2017 and 2022 were analyzed in this study. Among them, 12 were included in the prospective ACSE Unicancer study and received aPD1 as monotherapy. The historic cohort used for the comparative analysis was established from the ENKTCL national observatory (DRC Limoges) and composed of 38 R/R ENKTCL patients who received at least one cycle of first salvage therapy without aPD1 between 2006 and 2019. All patients were treated with Asparaginase-containing regimen as frontline therapy. Results: Patient characteristics were as follows: median age of 52 years (19-79), sex ratio M/F of 2/1, disseminated stage in 57% of the cases, high PINK score in 41% of the cases. The overall response rate (ORR) at the last follow-up was 40.5% (15 patients) in the aPD1 group. With a median follow-up time of 6.5 months for the whole cohort and 23.4 months for survivors, at 2 years progression free survival (PFS) was 22.4% and overall survival (OS) was 50.2%. Median PFS was 6.9 months and median OS was unreached. Among the 22 patients who experienced progression or relapse after the initiation of aPD1 therapy,14 patients (64%) received salvage therapy, mostly containing gemcitabine in association with immunotherapy continuation. Regarding prognostic factors at relapse, we identified that patients with performance status (PS) ≥2, B symptoms and ≥2 extranodal sites involved at relapse had significantly worse OS in univariate analysis. Only patients with ≥2 extranodal sites involved at relapse had significantly decreased OS in multivariate analysis. This cohort was then compared to the historic cohort after matching on a propensity score using the greedy nearest neighbor 1:1. 23 patients in each group were included in this analysis. Both groups had a comparable PFS (p=0.16) but OS was significantly improved in patients treated with aPD1 as salvage therapy alone or in combination (48.4% versus 23.1%, p=0,011). Summary/Conclusion: We report here the largest cohort of R/R ENKTCL treated with aPD1. Our study confirms the efficacy of aPD1 therapy in R/R ENKTCL and highlights its superiority as compared to other types of salvage therapy used in this setting before immunotherapy era. Our results will prompt us to now prospectively evaluate the benefit of aPD1 therapy as first line therapy in patients with high risk disseminated stage of ENKTCL, in combination with chemotherapy agents able to enhance anti-tumor immune response. To improve response to immunotherapy, it is also crucial to decipher mechanisms underlying immune escape mechanisms in ENKTCL and to identify biomarkers predictive of response or resistance to immune checkpoint blockade.Keywords: Lymphoma therapy, Immunotherapy, Lymphoma, NK-T cells

Large granular lymphocyte leukemia (LGLL) is a chronic lymphoproliferative disorder characterized by the proliferation of T or NK cytotoxic cells in the peripheral blood, the spleen and the bone marrow. Neutropenia leading to recurrent infections represents the main manifestation of LGLL. One specificity of LGLL is its frequent association with auto-immune disorders, among them first and foremost rheumatoid arthritis, and other hematologic diseases, including pure red cell aplasia and bone marrow failure. The large spectrum of manifestations and the classical indolent course contribute to the diagnosis difficulties and the frequency of underdiagnosed cases. Of importance, the dysimmune manifestations disappear with the treatment of LGLL as the blood cell counts normalize, giving a strong argument for a pathological link between the two entities. The therapeutic challenge results from the high rate of relapses following the first line of immunosuppressive drugs. New targeted agents, some of which are currently approved in autoimmune diseases, appear to be relevant therapeutic strategies to treat LGLL, by targeting key activated pathways involved in the pathogenesis of the disease, including JAK-STAT signaling.

Extranodal NK/T cell lymphoma (ENKTCL) is a rare subset of peripheral T lymphoma of which Asparaginase containing regimens are the standard of care. Despite recent improvements in the therapeutic strategy, patient’s prognosis remains poor, especially for those who experienced relapsed or refractory (R/R) disease. Recent studies highlighted that immune escape mechanisms are involved in ENKTCL pathogenesis. Particularly, the frequent PD-L1 upregulation led to assess the efficacy of anti-PD1 (aPD1) therapy in small cohorts of R/R ENKTCL patients, with encouraging results. Our study aims to evaluate the efficacy of aPD1 therapy alone or in combination in 37 patients with R/R ENKTCL. We also performed a comparative analysis with a historic cohort of 38 patients treated for R/R ENKTCL before immunotherapy era. 37 patients from 24 French centers, with R/R ENKTCL treated with at least one cycle of aPD1 as salvage therapy between 2017 and 2022 were analyzed in this study. Among them, 12 were included in the prospective ACSE Unicancer study and received aPD1 as monotherapy. The historic cohort used for the comparative analysis was established from the ENKTCL national observatory and composed of 38 R/R ENKTCL patients who received at least one cycle of first salvage therapy without aPD1 between 2006 and 2019. All patients were treated with Asparaginase-containing regimen as frontline therapy. Patient characteristics were as follows: median age of 52 years (19–79), sex ratio M/F of 2/1, disseminated stage and high PINK score respectively in 57% and 41% of the cases. The overall response rate at the last follow-up was 40.5% in the aPD1 group. With a median follow-up time of 6.5 months for the whole cohort and 23.4 months for survivors, progression free survival and overall survival (OS) at 2 years were 22.4% and 50.2%. Among the 22 patients who experienced progression or relapse after aPD1 initiation, 14 patients (64%) received salvage therapy, mostly containing gemcitabine in association with immunotherapy continuation. This cohort was then compared to the historic cohort after matching on a propensity score. 23 patients in each group were included in this analysis. OS was significantly improved in patients treated with aPD1 as salvage therapy alone or in combination (48.4% versus 23.1%, p = 0011). We report here the largest cohort of R/R ENKTCL treated with aPD1. Our study confirms the efficacy of aPD1 therapy in R/R ENKTCL and highlights its superiority as compared to other types of salvage therapy used in this setting before immunotherapy era. Our results will prompt us to now prospectively evaluate the benefit of aPD1 therapy as first line therapy in patients with high risk disseminated stage of ENKTCL, in combination with chemotherapy agents able to enhance anti-tumor immunity. Encore Abstract - previously submitted to regional or national meetings (up to <1’000 attendees), EHA 2023 Keywords: Extranodal non-Hodgkin lymphoma, Immunotherapy No conflicts of interests pertinent to the abstract.

Introduction: CD19 CAR T-cells can induce prolonged remission in a significant number of patients with relapse/refractory (R/R) lymphoma. However, little is known about patients’ life after CAR T-cell therapy. Here, we conducted a prospective study to evaluate health-related quality of life (HRQol), as well as physical, social and professional outcomes after CAR T-cell therapy. Methods: This prospective study was conducted at the University Hospitals of Rennes and Toulouse (France). All adult patients with R/R lymphoma treated with CAR T-cells were eligible. HRQol changes were assessed using the FACT-Lym questionnaire. Other questionnaires evaluated cognitive state (FACT-Cog), fatigue (FACT-Fatigue), anxiety and depression (HAD scale), post-traumatic stress disorder (PTSD), and sexuality (Vican). Questionnaires were collected at baseline (before leukapheresis), immediately before CAR-T infusion, 3 and 6 months after CAR T-cell infusion. Patients were censored in case of relapse or death. Among patients in remission after 12 months, professional, physical, sexual and global life information were collected. Results: From March 2020 to August 2022, 59 patients were included in the study (46 LBCL, 5 MCL, 8 FL). Median age was 63 years (range, 19–78). Patients were treated with axi-cel (N = 37), tisa-cel (N = 18) or brexu-cel (N = 3). The median follow-up after CAR T-cell infusion was 11 months. There were 53 and 38 still in remission at 3 and 6 months after CAR T-cell infusion, respectively. The FACT-Lym score showed a clinically relevant improvement of HRQol, with a mean change from baseline of 10.9 points (95%CI 5.8; 16.1) and 12.2 (95%CI 4.2; 20.1), respectively. The fatigue subscale showed a clinically relevant improvement at 3 and 6 months, with a mean change from baseline of 5.9 points (95%CI 2.9; 8.8) and 4.7 (95%CI 0.2; 9.2), respectively. The FACT-Cog score did not show any clinically relevant change. The HAD score showed that 43% and 24% of patients presented anxiety and depression at baseline, respectively, versus 26% and 7% after 6 months. Overall, 26% of patients presented a significant PTSD (i.e., >44) at 6 months. 54.5% of the patients considered that their life was back to normal (before lymphoma diagnosis); 68.2% patients were satisfied with their sexual life, 59.1% considered that it was back to normal; 54% felt less fit than before lymphoma; 21/21 patients practicing a physical activity before lymphoma resumed their activity; 3/8 resumed their social activity; Among the working age population (n = 10), 50% patients had returned to work. Encore Abstract - previously submitted to EHA 2023 Keywords: Aggressive B-cell non-Hodgkin lymphoma, Cellular therapies, Other Conflicts of interests pertinent to the abstract F. Colin Consultant or advisory role Gilead, Novartis A. Anota Consultant or advisory role AMGEN, IPSEN, ASTRAZENECA, KITE-GILEAD Educational grants: KITE GILEAD L. Ysebaert Consultant or advisory role AstraZeneca, Beigene, BMS/Celgene, Janssen, Gilead/Kite R. Houot Consultant or advisory role Kite/Gilead Honoraria: Bristol Myers Squibb/Celgene, MSD, Kite/Gilead, Roche, Novartis, Janssen A. Moignet Consultant or advisory role Kite, Novartis

Large granular lymphocytic leukemia is a rare lymphoproliferative disorder characterized by a clonal expansion of T-lineage lymphocyte or natural killer (NK) cells in 85 and 15% of cases respectively. T and NK large granular leukemia share common pathophysiology, clinical and biological presentation. The disease is characterized by cytopenia and a frequent association with autoimmune manifestations. Despite an indolent course allowing a watch and wait attitude in the majority of patients at diagnosis, two third of the patient will eventually need a treatment during the course of the disease. Unlike T lymphocyte, NK cells do not express T cell receptor making the proof of clonality difficult. Indeed, the distinction between clonal and reactive NK-cell expansion observed in several situations such as autoimmune diseases and viral infections is challenging. Advances in our understanding of the pathogenesis with the recent identification of recurrent mutations provide new tools to prove the clonality. In this review, we will discuss the pathophysiology of NK large granular leukemia, the recent advances in the diagnosis and therapeutic strategies.

Large granular lymphocyte leukaemias (LGL) are rare lymphoproliferative syndromes characterised by clonal expansion of T or NK lymphocytes in 85 and 15% of cases respectively. Interestingly, T and NK LGL leukaemias share a common pathophysiology and similar clinical and biological presentations. This lymphoproliferative syndrome is characterised by cytopenias and a frequent association with autoimmune diseases or manifestations. It is an indolent disease that in most cases allows for an abstinence-only strategy at diagnosis. However, the majority of patients will require initiation of treatment during follow-up. As NK cells lack a TCR, obtaining evidence of clonality in NK-ALL leukaemias is difficult. This is crucial in view of possible reactive expansions in the context of viral infections or dysimmune diseases. The diagnostic approach has been facilitated by the progress made in recent years in the understanding of the pathophysiology and the recent identification of recurrent mutations. In this review, we will discuss the pathophysiology of NK LGL leukaemias, present recent advances in diagnostic strategies before discussing therapeutic management.

Introduction: CD19 CAR‐T cells have been approved for the treatment of relapse/refractory (R/R) diffuse large B‐cell lymphoma (DLBCL). With these adoptive immunotherapies, about 30‐40% of patients achieve prolonged remissions, most of whom are likely to be cured. However, little is known about life after CAR‐T cells. Thus, we initiated a prospective study to evaluate the personal, social and professional outcomes after CAR‐T cell therapy in lymphoma patients. Methods: This is a prospective, observational, multicenter study, conducted at the University Hospitals of Rennes and Toulouse (France). All adult patients treated with CAR‐T cells (Yescarta or Kymriah) for R/R DLBCL are eligible. Primary objective is to describe health‐related quality of life (HRQoL) changes during the first year after CAR‐T cell therapy using the EORTC QLQ‐C30 and FACT‐Lym questionnaires. Secondary objectives are to describe the dynamic changes of other personal, social, and professional parameters including anxiety and depression (HAD scale), cognitive state (FACT‐Cog), fatigue (FACT‐Fatigue), sexuality (SqoL), post‐ traumatic stress disorder (PTSD), social and professional reintegration, supportive care measures. These outcomes are correlated with baseline characteristics of the patients and his disease, as well as with response to therapy, adverse events, and biological parameters. Patients fill the questionnaires before leukapheresis, immediately before CAR‐T infusion, and then 3 months, 6 months and one year after CAR‐T infusion. Adverse events are prospectively collected by dedicated nurses who call patients daily until day 21, twice a week until day 28, once a week until month 2, every other week until month 3, once a month until month 6, and once every 3 months until month 12. Results: This first patient was enrolled in November 2020. To date, 30 patients were included in the study: 19 male and 11 female, median age is 65 years (range, 19‐75) and median number of prior lines is 2 (range, 2‐5). Median follow up is 3.2 months (range, 0‐12) for the entire cohort. Six patients have completed their 6 months follow‐up. Among the 5 patients with available HRQoL at 6 months, 3 patients (60%) presented a clinically significant improvement of their global HRQoL level. Conclusions: This prospective study will provide a better understanding of the personal, social, and professional outcomes of patients who have been treated with CAR‐T cells. These data will be of importance to better inform patients who undergo CAR‐T cell therapy, identify needs for improvement in quality of life after CAR‐T cell therapy, and better appreciate the social and economic impact of these treatments.

Distinguishing chronic lymphoproliferative disorders of NK cells (CLPD-NK) from reactive NK-cell expansion is challenging. We assessed the value of killer immunoglobulin-like receptor(KIR) phenotyping and targeted high-throughput sequencing in a cohort of 114 consecutive patients with NK cell proliferation, retrospectively assigned to a CLPD-NK group (n = 46) and a reactive NK group (n = 68). We then developed an NK-cell clonality score combining flow cytometry and molecular profiling with a positive predictive value of 93%. STAT3 and TET2 mutations were respectively identified in 27% and 34% of the patients with CLPD-NK, constituting a new diagnostic hallmark for this disease. TET2-mutated CLPD-NK preferentially exhibited a CD16low phenotype, more frequently displayed a lower platelet count, and was associated with other hematologic malignancies such as myelodysplasia. To explore the mutational clonal hierarchy of CLPD-NK, we performed whole-exome sequencing of sorted, myeloid, T, and NK cells and found that TET2 mutations were shared by myeloid and NK cells in 3 of 4 cases. Thus, we hypothesized that TET2 alterations occur in early hematopoietic progenitors which could explain a potential link between CLPD-NK and myeloid malignancies. Finally, we analyzed the transcriptome by RNA sequencing of 7 CLPD-NK and evidenced 2 groups of patients. The first group displayed STAT3 mutations or SOCS3 methylation and overexpressed STAT3 target genes. The second group, including 2 TET2-mutated cases, significantly underexpressed genes known to be downregulated in angioimmunoblastic T-cell lymphoma. Our results provide new insights into the pathogenesis of NK-cell proliferative disorders and, potentially, new therapeutic opportunities.

American Journal of HematologyVolume 96, Issue 10 p. E368-E370 CORRESPONDENCEFree Access Ruxolitinib for refractory large granular lymphocyte leukemia Aline Moignet, Centre Hospitalier Universitaire de Rennes, Service d'Hématologie Clinique, Rennes, FranceSearch for more papers by this authorCédric Pastoret, orcid.org/0000-0002-5675-3154 Centre Hospitalier Universitaire de Rennes, Laboratoire d'Hématologie, Rennes, France Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche U1236, Rennes, FranceSearch for more papers by this authorGuillaume Cartron, Department of Hematology, CHU Montpellier, UMR 5535, Université de Montpellier, Montpellier, FranceSearch for more papers by this authorPaul Coppo, Service d'Hématologie, Centre de Référence des Microangiopathies Thrombotiques (CNR-MAT), Hôpital Saint-Antoine, AP-HP – Sorbonne – Université, Paris, France INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, FranceSearch for more papers by this authorThierry Lamy, Corresponding Author thierry.lamy@univ-rennes1.fr orcid.org/0000-0003-2960-4855 Centre Hospitalier Universitaire de Rennes, Service d'Hématologie Clinique, Rennes, France Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche U1236, Rennes, France Correspondence Thierry Lamy, Hematology Department, CHU Pontchaillou, 2 rue Henri Le Guilloux, F-35033 Rennes cedex 9, France. Email: thierry.lamy@univ-rennes1.frSearch for more papers by this author Aline Moignet, Centre Hospitalier Universitaire de Rennes, Service d'Hématologie Clinique, Rennes, FranceSearch for more papers by this authorCédric Pastoret, orcid.org/0000-0002-5675-3154 Centre Hospitalier Universitaire de Rennes, Laboratoire d'Hématologie, Rennes, France Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche U1236, Rennes, FranceSearch for more papers by this authorGuillaume Cartron, Department of Hematology, CHU Montpellier, UMR 5535, Université de Montpellier, Montpellier, FranceSearch for more papers by this authorPaul Coppo, Service d'Hématologie, Centre de Référence des Microangiopathies Thrombotiques (CNR-MAT), Hôpital Saint-Antoine, AP-HP – Sorbonne – Université, Paris, France INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, FranceSearch for more papers by this authorThierry Lamy, Corresponding Author thierry.lamy@univ-rennes1.fr orcid.org/0000-0003-2960-4855 Centre Hospitalier Universitaire de Rennes, Service d'Hématologie Clinique, Rennes, France Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche U1236, Rennes, France Correspondence Thierry Lamy, Hematology Department, CHU Pontchaillou, 2 rue Henri Le Guilloux, F-35033 Rennes cedex 9, France. Email: thierry.lamy@univ-rennes1.frSearch for more papers by this author First published: 16 June 2021 https://doi.org/10.1002/ajh.26275AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinked InRedditWechat To the Editor: Large granular lymphocyte (LGL) leukemia is a T or NK cell clonal disorder characterized by tissue invasion of marrow, spleen, and liver. Clinical presentation is dominated by recurrent infections associated with neutropenia, anemia, splenomegaly and auto-immune diseases, particularly rheumatoid arthritis (RA). It accounts for 2%–5% of chronic lymphoproliferative disorders. Indications for starting therapy are based on occurence of infections related neutropenia, symptomatic or transfusion-dependent anemia or associated autoimmune diseases requiring therapy.1 There is no standard treatment for patients with LGL leukemia. Immunosuppressive therapy remains the foundation of first line treatment including single agents, that is methotrexate, oral cyclophosphamide or cyclosporine (cyclosporin, INN). Usually, refractory patients may be treated with purine analogs, bendamustine or alemtuzumab while polychemotherapy does not provide any good results. There is a clear unmet medical need for such patients whose prognosis is poor. Before moving to targeted therapy, it is important to better understand the pathogenesis of LGL leukemia. Constitutive activation of STAT3 pathway in all LGL leukemia patients has been demonstrated in 2001, while common somatic gain-of-function STAT3 mutations was reported in 28% to 60% of T-LGL leukemia and 30% to 48% of NK LGL lymphocytosis in 2012. Whether or not STAT3 mutation acts as an initial trigger or a late event in LGL leukemia proliferation is still debated. Dysregulations of several signaling pathways including MAPK, PI3K-AKT, NF-ĸB, and JAK/STATs constitute a hallmark of the pathogenesis of LGL leukemia. However, the landscape of mutational events includes also epigenetic and DNA methylation mutated genes.1 We and others have recently reported TET2 mutations in 30% of NK LGL leukemia.2 All these findings open a new window to explore targeted therapy in LGL leukemia. The JAK inhibitors are indeed of potential interest in the treatment of the disease. The JAK3 specific inhibitor tofacitinib has shown clinical efficacy in a series of nine RA patients presenting with refractory T-LGL leukemia.3 Hematologic response was observed in six patients with neutropenia improvement in five patients. One patient experienced an increase of hemoglobin level. The JAK2 inhibitor ruxolitinib is commonly used in myeloproliferative disorders. Interestingly, its effectiveness is not dependent on JAK2V617F mutation status. We therefore evaluated the use of ruxolitinib in two cases of refractory T–LGL leukemia. The initial diagnosis was based on a chronic LGL-T cell expansion (>1 × 109/L) with a typical phenotype (CD3+/CD5dim/CD8+/CD57+) and TCRγ clonal rearrangement. Both patients had a long term medical history of transfusion-dependent anemia and neutropenia, failing or partially responding to immunosuppressive or chemotherapeutic agents. Informed consent was obtained in both cases according to IRB protocol. Case 1: A 36-year-old female was diagnosed with pure red cell aplasia associated to STAT3 mutated T-LGL leukemia. From 2009 to 2017, she successively received high dose immunoglobulin, prednisone, methotrexate, ciclosporin (cyclosporine) and cyclophosphamide leading to transitory and partial remission. In 2018, one course of 2CdA (cladribine) was administered due to recurrence of severe anemia and neutropenia without any hematological improvement. The patient was transfusion dependent and experienced recurrent infections. In July 2019, sixth line therapy with ruxolinitib was then introduced at an oral dose of 15 mg twice per day and resulted in rapid improvement of anemia and neutropenia with complete hematological response. At month 20, patient had hematologic complete response with normalization of blood cell count: Hb 11.3 g/dl, neutrophils 1.78 × 109/L, platelets 335 × 109/L (Figure 1(A)). FIGURE 1Open in figure viewerPowerPoint (A) Course of treatment in a patient with a refractory T–LGL leukemia treated successfully with ruxolitinib. (B) Bone marrow histology and flow cytometry quantification of LGL in blood samples during the course of ruxolutinib. Bone marrow biopsy performed before ruxolitinib showed a left shift of erythroid precursors and CD8+/granzyme B+ intrasinusoidal lymphocytes, with several linear arrays of cells, and clusters. After 12 months of ruxolitinib, erythroid precursors were normally distributed and immunohistochemical analysis revealed a significant decrease of CD8+/granzymeB+ intrasinusoidal lymphocytes (MGG: May-Grünwald-Giemsa staining, original magnification; CD8 and Granzyme B immunohistochemistry, original magnification x200). T-cell LGL defined as CD3pos, CD8pos, CD5low, CD57pos cells expressing Vb7.1 accounted for 77% of lymphocytes before ruxolutinib initiation and significantly decreased to 16% after 12 months of treatment. (C) After 12 months of ruxolitinib, absolute LGL count (defined by the total number of peripheral lymphocytes × percent of CD8+/Vβ7.1 cells) decreased from 1.2 × 109/L to 0.08 × 109/L correlating with a significant reduction of STAT3 variance allelic frequency from 25% to 4% After 12 months of therapy, bone marrow biopsy evidenced a slight persistence of LGL infiltration (Figure 1(B)). Flow cytometric and molecular analyses showed a drastic and stable reduction of clonal LGL expansion in serial blood samples (Figure 1(C)). At month 20, patient showed a persistent hematologic complete response with normalization of blood cell count: Hb 11.3 g/dl, neutrophils 1.78 × 109/L, platelets 335 × 109/L. Case 2: The second case was related to 70-year-old female with severe pancytopenia and splenomegaly related to STAT3 un-mutated T-LGL leukemia diagnosed in 1990. She failed methotrexate, ciclosporin, cyclophosphamide and 2CdA (cladribine). The patient was transfusion dependent despite 4 months EPO delivery before starting ruxolitinib. In March 2020, ruxolitinib was administered at a dose of 10 mg taken orally twice per day and induced a very good partial remission: the patient became transfusion independent by the third month of therapy. In February 2021, complete blood cell count showed: hemoglobin 10.4 g/dl (EPO not stopped), neutrophils 1.5 × 109/L (G-CSF once a week), Platelets 110 × 109/L. The large spleen was no longer palpable. Both patients did not present any adverse event secondary to ruxolitinib. The JAK inhibitors may be considered as relevant targeting agents for the treatment of cytopenic LGL leukemia patients.4, 5 Different compounds targeting JAK/STAT3 signaling pathway have demonstrated in vitro efficacy on leukemic samples from T-LGL leukemic and aggressive NK cell leukemia patients, depending on the basal activity of mutant STAT3. It has been shown that IL6/IL15 promoted T/NK LGL leukemic expansion through STAT/JAK pathway activation. The ability of STAT3 to promote IL6 gene expression then results in a feed forward autocrine loop. Therefore, we hypothesize that JAK inhibition could induce LGL clone apoptosis through a blockade of a feedback IL6 activation. Clinical efficacy of ruxolitinib was recently reported in a case of refractory severe enterocolitis harboring a STAT3 gene mutation.6 In the two present cases, ruxolitinib was able to induce durable complete and very good partial response (lasting more than 1 year) in two refractory LGL patients. These results make ruxolitinib a potential and promising alternative therapeutic agent in LGL leukemia. This option needs to be further explored in prospective clinical trials. CONFLICT OF INTEREST The authors declare no conflict of interest. Open Research DATA AVAILABILITY STATEMENT Data available on request from the authors: The data that support the findings of this study are available from the corresponding author upon reasonable request. REFERENCES 1Lamy T, Moignet A, Loughran TP Jr. LGL leukemia: from pathogenesis to treatment. Blood. 2017; 129(9): 1082- 1094. CrossrefCASPubMedWeb of Science®Google Scholar 2Pastoret C, Desmots-Loyer F, Drillet G, et al. Linking the KIR phenotype with STAT3 and TET2 mutations to identify chronic lymphoproliferative disorders of NK cells. Blood. 2021; 137: 3237- 3250. CrossrefCASPubMedWeb of Science®Google Scholar 3Bilori B, Thota S, Clemente MJ, et al. Tofacitinib as a novel salvage therapy for refractory T-cell large granular lymphocytic leukemia. Leukemia. 2015; 29(12): 2427- 2429. CrossrefCASPubMedWeb of Science®Google Scholar 4Kuusanmäki H, Dufva O, Parri E, et al. Drug sensitivity profiling identifies potential therapies for lymphoproliferative disorders with overactive JAK/STAT3 signaling. Oncotarget. 2017; 8(57): 97516- 97 527. CrossrefPubMedGoogle Scholar 5Dufva O, Kankainen M, Kelkka T, et al. Aggressive natural killer-cell leukemia mutational landscape and drug profiling highlight JAK-STAT signaling as therapeutic target. Nat Commun. 2018; 9(1): 1567. CrossrefPubMedWeb of Science®Google Scholar 6Parlato M, Charbit-Henrion F, Nader EA, et al. Efficacy of ruxolitinib therapy in a patient with severe enterocolitis associated with a STAT3 gain-of-function mutation. Gastroenterology. 2019; 156(4): 1206- 1210. CrossrefPubMedWeb of Science®Google Scholar Volume96, Issue101 October 2021Pages E368-E370 FiguresReferencesRelatedInformation