Splenic Marginal Zone Lymphoma Research Articles

Lymphomas with massive splenomegaly – a case series

Hairy cell leukaemia (HCL) is an uncommon chronic B-cell lymphoma. Classical HCL and its variant are separate entities. Splenic marginal zone lymphoma (SMZL) is rare accounting for <2%of lymphoid neoplasms2. A 62-year-old woman presented with constitutional symptoms and mild splenomegaly was having pancytopenia, monocytopenia and hairy cells in the peripheral blood. Bone marrow aspiration was a dry tap while trephine biopsy revealed diffuse lymphoid cell infiltration with characteristic fried egg appearance and fibrosis. Morphological and immunophenotypic diagnosis was HCL. She was successfully treated with IV-cladribine and G-CSF for chemotherapy induced severe neutropenia. She achieved complete remission in 6 months. A 56-year-old woman presented with abdominal distension and lower limb swelling was having marked splenomegaly, absolute lymphocytosis, mild neutropenia and anaemia. Blood film revealed lymphocytes showing moderate cytoplasm and cytoplasmic projections. With bone marrow biopsy and flow cytometry, splenic B-cell lymphoma/leukaemia with prominent nucleoli was diagnosed and was treated with rituximab monotherapy. A 76-year-old woman presented with progressive abdominal fullness, unintentional weight loss and early satiety was having splenomegaly, bicytopenia and absolute lymphocytosis consisting predominantly of small mature lymphocytes and occasional lymphocytes with polar villi. Morphological and immunopheno-typical results confirmed SMZL. She was treated with rituximab monotherapy.

Nodal Low-Grade B-Cell Lymphoma Co-Expressing CD5 and CD10 but Not CD23, IRTA1, or Cyclin D1: The Diagnostic Challenge of a Splenic Marginal Zone Lymphoma.

The diagnosis of lymphoma is based on histopathological and immunophenotypical features. CD5 and CD10 are traditionally considered a T-cell antigen and a germinal center B-cell antigen, respectively. It is very unusual for a low-grade B-cell lymphoma (BCL) to co-express CD5 and CD10. Although the biologic basis or clinical significance of such co-expression is unclear, this rare event may pose a significant diagnostic challenge. Here, we report a case of a 63-year-old male presenting with bilateral cervical lymphadenopathy and lymphocytosis. Histologically, the nodal tumor was largely diffuse with neoplastic small atypical lymphocytes co-expressing CD5, CD10, and CD20, but not CD23 or cyclin D1. The leukemic cells in the peripheral blood exhibited hairy projections. Taking together the marked splenomegaly, involvement of lymph nodes, bone marrow, and peripheral blood, a final diagnosis of splenic marginal zone lymphoma (SMZL) was reached. The patient was alive with partial response for 10 months after immunochemotherapy. The dual expression of CD5 and CD10 is extremely unusual for low-grade BCL and may lead to an erroneous diagnosis. Integrating the findings into peripheral blood smear tests, flow cytometry, histopathology, imaging, and clinical features is mandatory to exclude other lymphoma types and to reach a correct diagnosis, particularly for a case with nodal presentation.

Bendamustine and rituximab as first-line treatment for symptomatic splenic marginal zone lymphoma: long-term outcome and impact of early unmeasurable minimal residual disease attainment from the BRISMA/IELSG36 phase II study.

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Transformation risk and associated survival outcome of marginal zone lymphoma: A nationwide study.

Histological transformation into an aggressive B-cell lymphoma indicates a poor survival outcome for patients with indolent marginal zone lymphoma (MZL), which has been less studied. Large-scale data with long-term follow-up to investigate MZL transformation is limited. Here, by reporting a US-Nationwide cohort of 30,619 MZL patients diagnosed between 2000 and 2019, we found that transformation occurred in 2.08% (N = 624) of MZL cases, with the transformation incidence of 3.1 per 1,000 person-years. Advanced Ann Arbor stage, nodal MZL (NMZL) and splenic MZL (SMZL) were associated with an elevated risk of transformation. Certain subtype-specific characteristics, such as non-gastric extra-nodal MZL (vs. gastric, HR, 1.51, 95%CI 1.13-2.04; p = 0.006), and receiving splenectomy for SMZL (HR, 2.04, 95%CI 1.28-3.26; p = 0.003), also indicated a higher risk of transformation. Besides, transformation independently increased the overall mortality risk (HR, 1.38, 95%CI 1.24-1.53, p < 0.001), especially the higher lymphoma-caused mortality risk (HR, 3.21, 95%CI 2.81-3.67, p < 0.001). Transformation was also associated with a higher percentage of lymphoma-caused deaths. The post-transformation prognostic analyses demonstrated that female gender and age ≥ 65 years independently affected patients' mortalities. These findings, based on the largest cohort to date, contribute to a better understanding of transformed MZL, and provide valuable reference points for guidelines and patient counseling.

Optical genomic mapping is a helpful tool for detecting CCND1 rearrangements in CD5-negative small B-cell lymphoma: Two cases of leukemic non-nodal mantle cell lymphoma

Optical genome mapping (OGM) is a new DNA-based technology which provides comprehensive examination of the entire genome. We report two patients who presented with splenomegaly and leukocytosis with lymphocytosis including villous lymphocytes. Neither patient had lymphadenopathy. Bone marrow evaluation showed involvement by small B-cell lymphoma in a sinusoidal and interstitial distribution, and immunophenotypic analysis showed that the neoplastic cells were positive for B-cell markers and cyclin D1 but were negative for SOX11 and CD5. Initially, the clinicopathologic features in both patients were thought to be suspicious for hairy cell leukemia variant or splenic marginal zone lymphoma. However, OGM detected CCND1 rearrangement: t(2;11)/IGK::CCND1 in one case and t(11;14)/IGH::CCND1 in the other case. These cases illustrate the valuable role OGM can play in establishing the diagnosis of MCL. Case 1 also contributes to the paucity of literature on the rare occurrence of IGK::CCND1 in MCL.

CAD204520 Targets NOTCH1 PEST Domain Mutations in Lymphoproliferative Disorders.

NOTCH1 PEST domain mutations are often seen in hematopoietic malignancies, including T-cell acute lymphoblastic leukemia (T-ALL), chronic lymphocytic leukemia (CLL), splenic marginal zone lymphoma (SMZL), mantle cell lymphoma (MCL), and diffuse large B-cell lymphoma (DLBCL). These mutations play a key role in the development and progression of lymphoproliferative tumors by increasing the Notch signaling and, consequently, promoting cell proliferation, survival, migration, and suppressing apoptosis. There is currently no specific treatment available for cancers caused by NOTCH1 PEST domain mutations. However, several NOTCH1 inhibitors are in development. Among these, inhibition of the Sarco-endoplasmic Ca2+-ATPase (SERCA) showed a greater effect in NOTCH1-mutated tumors compared to the wild-type ones. One example is CAD204520, a benzimidazole derivative active in T-ALL cells harboring NOTCH1 mutations. In this study, we preclinically assessed the effect of CAD204520 in CLL and MCL models and showed that NOTCH1 PEST domain mutations sensitize cells to the anti-leukemic activity mediated by CAD204520. Additionally, we tested the potential of CAD204520 in combination with the current first-line treatment of CLL, venetoclax, and ibrutinib. CAD204520 enhanced the synergistic effect of this treatment regimen only in samples harboring the NOTCH1 PEST domain mutations, thus supporting a role for Notch inhibition in these tumors. In summary, our work provides strong support for the development of CAD204520 as a novel therapeutic approach also in chronic lymphoproliferative disorders carrying NOTCH1 PEST domain mutations, emerging as a promising molecule for combination treatment in this aggressive subset of patients.

Splenic Marginal Zone Lymphoma complicated with cold autoimmune haemolytic anaemia

Splenic Marginal Zone Lymphoma (SMZL) is a distinct subtype of Marginal Zone Lymphoma (MZL) which is categorized under Non-Hodgkin Lymphoma. It is usually benign with the potential to transform into diffuse large B-cell lymphoma. Comorbid cold Autoimmune Haemolytic Anaemia(cAHA) is rare with SMZL. A case of a 53-year-old woman is presented here. She presented with fever and anaemic symptoms. Biochemical, radiological and histological investigations were done, and a diagnosis of SMZL complicated with cAHA was made.

Untangling hairy cell leukaemia (HCL) variant and other HCL-like disorders: Diagnosis and treatment.

The variant form of hairy cell leukaemia (HCL-V) is a rare disease very different from hairy cell leukaemia (HCL), which is a very well-defined entity. The 5th WHO edition (Leukemia, 36, 2022 and 1720) classification (WHO-HAEM5) introduced splenic lymphomas/leukaemias including four different entities: (1) HCL, (2) splenic marginal zone lymphoma (SMZL) with circulating villous cells in the peripheral blood, (3) splenic lymphoma with prominent nucleolus (SLPN), which replaced HCL-V and CD5 negative B-prolymphocytic leukaemia (B-PLL), and (4) splenic diffuse red pulp lymphoma (SDRPL). All these entities have to be distinguished because of a different clinical course and the need for a different treatment. The diagnosis can be challenging because of complex cases and overlap and/or grey zones between all the entities and needs integrating clinical, histologic, immunophenotypic, cytogenetic and molecular data. We review the diagnostic criteria including clinical, immunophenotypic and molecular characteristics of patients with HCL-V and other HCL-like disorders including HCL, SDRPL, SMZL, B-PLL and the Japanese form of HCL. We also discuss the different criteria allowing us to separate these different entities and we will update the recent therapeutic options that have emerged, in particular the advances with chemoimmunotherapy and/or targeted therapies.

Recognizing puzzling PD1 + infiltrates in marginal zone lymphoma by integrating clonal and mutational findings: pitfalls in both nodal and transformed splenic cases

BackgroundA marked increase in PD1-positive TFH cells in nodal MZL cases (NMZL) was reported previously and could prompt suspicion for a diagnosis of peripheral T-cell lymphoma (PTCL), especially angioimmunoblastic T-cell lymphoma (AITL).Case presentationTo demonstrate that the pitfall might exist not only in NMZL but also in transformed splenic MZL (tSMZL), two NMZL cases (70 y/o female with enlarged left cervical lymph node and 75 y/o male with generalized lymphadenopathy) and one case of tSMZL (47 y/o male with nodal and extranodal involvement) with obvious PD1-positive T-cell hyperplasia were described here. Although all their initial diagnoses were prompted to be AITL, they were comprehensively characterized by clinical features, morphologic, immunophenotypic, clonality, and targeted exosome sequencing (TES) findings. Case 1 and Case 2 were NMZL with increased PD1 + T cells in the “peripheral pattern” or “mixed peripheral and central pattern”, and Case 3 was SMZL with abundant PD1-positive T cells in the “nodular pattern” that transformed to tSMZL (DLBCL) with PD1-positive T cells distributed in the “diffuse pattern.” In addition to the monoclonal IG rearrangement and polyclonal TCR rearrangement results, TES demonstrated enriched and recurrent mutations in MZLs and failed to find aberrations described in AITL- or TFH-derived lymphomas.ConclusionsIt is important to realize that this pitfall can also occur in more diagnostically difficult tSMZL cases; the integration of histopathology with clonality and mutation studies is also highlighted.

Risk of Transformation By Frontline Management in Follicular Lymphoma and Marginal Zone Lymphoma: A US Population-Based Analysis

INTRODUCTION Follicular lymphoma (FL) and marginal zone lymphoma (MZL) are commonly characterized by an indolent nature and favorable overall survival (OS). However, patients (pts) may experience high-grade transformation (HGT) into DLBCL, with an estimated annual risk of 1%-3% and cumulative incidence of HGT are 8% for FL and 10% for MZL at 10 and 12 years, respectively. The impact of treatment and surveillance strategies on the incidence of HGT remains uncertain, with some studies indicating reduced risk in treated pts without significant OS difference post-HGT. This study uses population-based data to assess the cumulative incidence and risk factors associated with HGT, post-HGT OS and lymphoma-specific survival (LSS) across lymphoma subtypes, with either treatment or surveillance as initial strategies. METHODS We extracted data for pts diagnosed with grades 1-2 FL, splenic MZL (SMZL), nodal MZL (NMZL), and extranodal MZL (MALT) from SEER database (period 2000-2015 with follow-up through 2018). We only included pts with histologic confirmed lymphoma as first malignancy, complete survival and treatment data, and a minimum of 3 months from FL and MZL diagnosis to HGT. Competing risk methods were used to estimate HGT cumulative incidence rates, with time to first event (HGT or death) defined from lymphoma diagnosis to date of first event; event-free pts were censored at date of last contact. Gray's test and Fine-Gray (FG) proportional subdistribution hazard modelling was employed to identify variables associated with risk of HGT. Post-HGT OS and LSS analyses utilized Cox and FG regression models, respectively. RESULTS 26827 pts met inclusion criteria. FL comprised 14229 pts (53%) while MZL types were SMZL 1182 (4.4%), NMZL 3916 (14.6 %) and MALT 7500 (28%). Treatment was given at diagnoses in 60.2%, 33.7%, 53.8% and 49.8% of pts with FL, SMZL, NMZL, MALT, respectively. 864 (3.2%) pts developed HGT to DLBCL, 553 (3.9%) in FL and 311 (2.5%) in MZL pts. After a median follow-up of 84 months (range 1-227), the 10-year cumulative incidence of HGT were for FL 4.24% (95%CI 3.88%-4.62%), SMZL 6.46% (95%CI 4.98%-8.20%), NMZL 3.28% (95%CI 2.70%-3.95%) and MALT 1.73% (95%CI 1.42%-2.09%). In FL, the 10-year cumulative incidence of HGT was 3.83% (95%CI 3.40-4.31%) in treated pts and 4.88% (95%CI 4.26%-5.54%) for those under surveillance (SHR=0.80, 95%CI 0.68-0.94; p=0.009) (Figure 1). This finding was confirmed in a multivariable analysis (p=0.001) that conversely identified female gender (p&amp;lt;0.001) and advanced stage at diagnosis (p&amp;lt;0.001) with higher risk of HGT (Figure 1). In MZL, the 10-year cumulative incidence rate of HGT in treated pts versus those managed with surveillance depicted by subtypes were as follows: SMZL 6.93% (95%CI 4.40%-10.2%) vs 6.29% (95%CI 4.53%-8.43%); NMZL 3.71% (95%CI 2.87%-4.70%) vs 2.78% (95%CI 2.03%-3.71%); MALT 2.01% (95%CI 1.54%-2.57%) vs 1.45% (95%CI 1.07%-1.93%). No significant difference between treatment versus surveillance were detected in univariable and multivariable analyses for SMZL (p=0.72 and 0.54) and NMZL (p=0.07 and 0.07), whereas surveillance pts had lower risk of HGT in MALT (p=0.02 and 0.01) (Figure 2). There were 429 (49.7%) deaths, including 337 (39%) lymphoma-specific deaths recorded in 864 pts with HGT in the entire cohort; and the median OS after HGT was 3.67 years (95%CI 2.5-6.25). Frontline therapy for FL was associated with higher risk of overall and lymphoma-specific deaths post-HGT (HR=1.32, 95%CI 1.03-1.69, p=0.02; and SHR=1.33, 95%CI 1.02-1.75, p=0.03); however, there was no significant difference in OS between frontline therapy and surveillance approach in SMZL (p=0.11), NMZL (p=0.73) and MALT (p=0.70). Interestingly, in SMZL worse LSS post-HGT was observed in those managed with frontline therapy (SHR=2.68, 95%CI 1.32-5.45, p=0.006). CONCLUSIONS Frontline treatment was associated to reduced HGT risk in FL, with advanced stage and female gender identified as risk factors for HGT. Contrarily, for SMZL, NMZL, and MALT, initial treatment neither diminished HGT risk nor improved overall survival post-HGT. Interestingly, in MALT initial treatment exhibits a higher HGT risk, likely due to selection bias, even though HGT cumulative incidence in MALT was lower compared to other subtypes. SMZL displayed a particularly high cumulative incidence of HGT and poorer LSS.

Splenic B Cell Lymphoma Innate Immune Cells: Detection of Red Versus White Pulp Lymphoma Differences in NK Cell Populations By High-Parameter Flow Cytometry

Introduction Splenic B-cell lymphomas, (SLs, total incidence ~1:100,000/year) comprise four distinct indolent B-cell non-Hodgkin lymphomas. Three predominantly involve the red pulp of the spleen (classical hairy cell leukemia (HCL), hairy cell leukemia variant (HCLv) and splenic diffuse red pulp small B-cell lymphoma (SDRPL)) and the fourth predominantly involves the white pulp (splenic marginal zone lymphoma (SMZL)). HCL is usually diagnosed by detection of an activation mutation in BRAF and responds well to treatment. We focused our study on the remaining three diseases, which have overlapping clinical presentations with only subtle differences in their morphology and phenotypes, but have considerable differences in prognosis, response to treatment and outcome. The white pulp lymphoma SMZL generally has good treatment responses and longer survival, whereas the red pulp SLs, HCLv and SDRPL are less responsive to treatments with shorter survival. Cells of the tumor microenvironment (TME) may have distinct characteristics between these three diseases. We hypothesized that TME innate immune cells from red pulp SLs (HCLv, and SDRPL) would be similar to each other and distinct from the white pulp SL, SMZL.In this study, we report that natural killer (NK) cells, which are important innate immune cells critical for immune surveillance, vary in proportions among red and white pulp SLs. Methods We studied splenectomy specimens from patients with SMZL (n=8), HCLv (n=8), SDRPL (n=7), and controls (healthy, post-trauma, n=8). Cryo-preserved bulk splenic suspension cell isolates from fresh (&amp;lt;4 hour) surgical specimens and intact formalin-fixed paraffin embedded (FFPE) tissue sections were used. Human specimen collection and usage was conducted with written informed consent after approval of the Institutional Research Subjects Review Board. FFPE tissue sections were studied by immunohistochemical (IHC) staining and splenic suspension cells were analyzed by high-parameter fluorescence flow cytometry using a Cytek Aurora spectral flow cytometer with a 25-color myeloid and a 28-color NK/B-cell panel. Results IHC characterization showed an extensive network of innate immune cells as reflected by CD163 + macrophages. Using high-parameter flow cytometry, we focused on innate immune cells in the non-B- non-T-cell (CD3 -CD19 -) population. The average proportions of this population were: NK cells 25-36%, monocyte/macrophages 20-34%, dendritic cells 5-11%, and granulocytes 2%. Among the patient groups, the median percentage of NK cells was greater for controls (34%) and SMZL (35%) as compared to red pulp lymphomas (HCLv (28%), SDRPL (24%)). Next, we examined NK subtypes: 1) cytotoxic NK cells (CD56 +CD16 +), 2) cytokine-secreting weakly cytotoxic NK cells (CD56 +CD16 -), and 3) undefined NK cells (CD56 -CD16 +) as a median percentage of total NK cells. The major subtype for control and SMZL was CD56 +CD16 - with values of 58 and 44%, respectively. This subtype comprises only 38%-40% of red pulp SLs. Proportionally the CD56 +CD16 + subtype was the largest for red pulp SLs at 52-57% as compared to 30% for control and 38% for SMZL. The remaining CD56 -CD16 + NK cell subtype was generally higher among red pulp SLs (HCLv (9%), SDRPL (12%)) as compared to control (6%) and SMZL (4%) patients. The increase in cytotoxic CD56 +CD16 + subtype among red pulp SLs was confirmed by measuring the percentage of granzymeB +perforin + cells among mature cytotoxic NK cells (CD56 +CD16 +/ CD57 +NKG2a -). SLs had higher proportions of this NK phenotype (78-86% compared to control 69%) and red pulp SLs had the most granzymeB +perforin + proportions with 82 and 86% for HCLv and SDRPL, respectively. Discussion In summary, spleens from patients with SMZL had similar percentages of total NK cells to controls, with lower percentages in red pulp SLs. All SLs had a higher percentage of cytotoxic NK cells than control, with the red pulp SLs having the most. These data suggest that monoclonal antibody treatments targeting SL antigen, such as anti-CD20, may be effective by activating cytotoxic NK cell to target lymphoma cells by antibody-dependent cellular cytotoxicity. Thus, uncovering the characteristics of the TME may aid the development of non-invasive diagnostic procedures and/or precision therapies.

Favorable Outcomes of Splenic Marginal Zone Lymphoma in an International Study of 934 Patients with Long Follow-up

Introduction: Splenic marginal zone lymphoma (SMZL) is a rare, indolent disease. There is no universally accepted standard therapy and long-term outcome data are limited. Rituximab monotherapy is increasingly used over splenectomy in the first-line setting despite a paucity of comparative studies. In the absence of randomized trials, large-scale international real-world data studies are critical to evaluate outcomes of different treatment interventions and to understand the natural history of SMZL in the rituximab-era. Methods: Adult patients with SMZL diagnosed between Jan 1 st 2000 and Dec 31 st 2018 were identified using regional/nationwide population-based registries or hospital registries in Europe, the UK, North America, and Australia. Medical records and pathology reports from eligible patients were reviewed locally by clinicians with specialty experience in hemato-oncology. SMZL cases were required to meet the diagnostic criteria by Matutes et al. Detailed data on clinico-pathological features, comorbidities, treatments (including treatment intention), treatment responses and survival outcomes were entered in a central study database. Overall survival (OS) was defined as time from diagnosis until death from any cause or censoring at last follow-up unless stated otherwise. Event-free survival (EFS) was defined as the time from diagnosis until histological transformation, initiation of new chemotherapy or death from any cause. Survival probabilities were estimated using the Kaplan-Meier estimator. Cumulative risk of transformation was estimated with death as a competing risk. Primary endpoints were 10-year OS and 5-year EFS respectivly. Results: A total of 934 patients from 25 sites were included. Median age at diagnosis was 68 years (IQR 60-76) and 54% were female. Bone marrow involvement was present in 96% (564/590) of those assessed, anemia (Hb &amp;lt;12g/dL for women or &amp;lt;13g/dL for men) in 58%, LDH &amp;gt;UNL in 46%, lymphocytosis (&amp;gt;5 x10 9/L) in 34%, and B-symptoms in 34%. Treatment data were available for 97% of patients. Median follow-up was 9.1 years (range 0-23). Watch and wait was the initial strategy in 28% and 88% of patients received at least one line of treatment during follow-up with the most common being splenectomy (45%), immuno-chemotherapy (19%) or rituximab monotherapy (8%). Fifteen percent received “other” treatments (i.e., combinations of listed therapies and/or radiation). The overall response rate to first-line treatment was 88% (95%CI: 85-90%) with 61% (95%CI: 57-64%) achieving CR. Second line (2L) therapy was needed in 34% during follow-up, 3L in 20% and 4L+ in 9%. The 5-year EFS and OS for all patients were 61% (95%CI: 57-65%) and 77% (95%CI: 74-80%) respectively. The 10-year OS and lymphoma-specific mortality (with non-SMZL deaths as competing risk) estimates were 60% (95%CI: 57-64%) and 11% (95%CI: 9-14%) respectively. The 5-year OS after 2L and 3L were 61% (95%CI: 55-68%) and 51% (95%CI: 42-61%) respectively (figure 1). The 10-year cumulative risk of histological transformation was 17% (95%CI: 13-21%), with 5-year post-transformation OS of 42% (32-54%). At the time of transformation, 60% were chemo-naive and 52% had never been treated for SMZL. The 5-year OS for chemo-naive patients with transformation was 54% (95%CI: 41-71%) versus 24% (95% CI: 13-44%) for transformations after (immuno)chemotherapy (p = 0.01). When comparing patients intended for treatment in first line with splenectomy (n=312) versus rituximab (n=70), patients treated with splenectomy more frequently presented with extra-hilar lymphadenopathy (35% vs 18%), splenomegaly (92% vs 87%) and anemia (64% vs 48%). OS was similar for the two groups (Figure 2, p = 0.53). The 5-year EFS were 63% (95%CI: 57-68%) and 64% (95%CI: 54-77%), respectively (p = 0.31). Conclusions: To date, this is the largest real-world study of SMZL with detailed treatment and outcome data. The study shows that histological transformation was not uncommon with 10-year risk of 17%. However, outcomes for the entire cohort were excellent irrespective of initial treatment and both OS and EFS were similar after first line rituximab versus splenectomy. Long-term survival remained relatively high even following multiple lines of therapy.

DNA Methylation-Based Classification of Hairy Cell Leukemia and Splenic B Cell Lymphoma

Improved classification of rare lymphoid neoplasms would be aided by a deeper understanding of their underlying molecular features and is important for diagnosis, prognosis and therapy. Tumor entities classified within the WHO category of splenic B cell lymphomas and leukemias often exhibit heterogenous, transecting features, and include hairy cell leukemia (HCL), splenic diffuse red pulp lymphoma (SDRPL), splenic marginal zone lymphoma (SMZL), and the newly described WHO entity, splenic B cell lymphoma/leukemia with prominent nucleoli (SBLPN); the latter including patients formerly classified as HCL-variant (HCL-V). Genome-wide epigenetic information provides a tumor cell fingerprint combining cell-of-origin and tumor-specific events. Here we used DNA methylation to perform an unbiased molecular subclassification and to explore novel biological aspects of these patients. Samples from patients with a pathological diagnosis of HCL, HCL-V, SDRPL and SMZL (made prior to the 5 th WHO revision and ICC classifications) were obtained from 19 institutions across 9 countries, totaling 367 patients. Cells were FACS-purified where necessary and DNA was analyzed by 450/850K Illumina DNA methylation arrays. Genetic mutations were assessed by whole-exome or targeted sequencing, IGHV-D-J sequences by Sanger sequencing, and copy number alterations (CNAs) by Illumina arrays. The 1000 most variable CpG methylation sites were used for k-means clustering. Recursive feature elimination/random forest algorithms were used to develop a classifier for DNA methylation-based subgroups with 98% accuracy. Unsupervised clustering of 197 patients diagnosed with HCL, HCL-V or SDRPL revealed 5 distinct DNA methylation (M) subgroups ( Figure 1). Subgroup assignment was stable throughout longitudinal sampling (including pre/post-treatment) and consistent between splenic, bone marrow and PBMC derived cells. A subgroup with universally clonal BRAF-V600E mutations and majority diagnosed as HCL was termed the M-HCL subgroup ( Table 1). Four other groups termed M-SBLPN1-4 contained all HCL-V and SDRPL diagnosed samples and were devoid of BRAF-V600E mutations. M-SBLPN1 comprised MAP2K1 mutations (91%) and was enriched for CREBBP, ARIDIA and TERT-promoter mutations. These patients displayed an HCL-like immunophenotype (64.3% CD25+) with 1/3 diagnosed as HCL. M-SBLPN2 exhibited the highest prevalence of TP53 mutations and concomitant genomic instability. Patients in M-SBLPN1,2 were enriched in unmutated IGHV4-34 rearrangements. M-SBLPN3,4 subgroups displayed an immunophenotype more dissimilar to HCL, mutated IGHV genes, and enrichment of IGLL5, SYK and BIRC3 mutations. M-SBLPN4 contained the most SDRPL samples, suggesting it may represent the SDRPL entity retained by the WHO. We next uncovered that 29/170 SMZL patients displayed DNA methylation patterns mapping to M-SBLPN2-4. These patients were phenotypically and molecularly similar to SBLPN (70% displaying villous morphology and depleted in IGHV1-2*04, NOTCH2, KLF2 mutations), likely representing SMZL patients suggested for reassignment to SBLPN in the updated WHO classification. To elucidate molecular pathways governing the biology of M-SBLPN subgroups, transcription factor motif enrichment analysis in hypomethylated genomic regions revealed selective activation of AP-1 in M-HCL along with ETS in M-SBLPN1,2. Both transcription factors are downstream of MAPK signaling, consistent with activating BRAF and MAP2K1 mutations in these subgroups. However, we observed strong ETS enrichment in the absence of MAP2K1 in M-SBLPN along with mutual exclusivity of MAP2K1 and TP53 mutations, suggesting TP53 mutations are driving ETS activation. Although lymphoid neoplasms rarely exhibit TERT promoter mutations, 83% of M-SBLPN1 patients showed the c.-124C&amp;gt;T mutation commonly observed in other cancers producing an ETS binding site and ectopic TERT activation. ETS activation and gain of an ETS site by mutation implies oncogenesis involves aberrant TERT activation in this subgroup. In summary, we have developed a DNA methylation-based classifier that resolves 4 SBLPN subgroups with distinct molecular features, and reclassifies a subset of SMZL and HCL patients, adding further information to the updated WHO/ICC entities. We reveal distinct biological pathways operating in M-SBLPN subgroups that may aid targeted therapy approaches.

The Proliferative History Index, Epicmit, Derived from Genome-Wide Epigenomic Profiling, Is a Key Driver of Clinical Survival in Splenic Marginal Zone Lymphoma

The epiCMIT (epigenetically-determined Cumulative MIToses) mitotic clock (Durran-Ferrer, 2020) tracks DNA hypo- and hyper-methylation changes in heterochromatin and H3K27me3-containing chromatin, respectively, providing a comprehensive record of B cell mitotic history (pre- and post-malignant transformation). Higher epiCMIT levels correlate with poor survival in CLL and MCL, suggesting that a greater mitotic history predicts future proliferative capacity. However, no studies have investigated epiCMIT in splenic marginal zone lymphoma (SMZL). We studied 142 SMZL cases, a rare lymphoid malignancy with specific genetic characteristics [del(7q), KLF2and NOTCH2 mutations, biased use of IGHV1-2*04]. After calculating epiCMIT scores from 450/850K Illumina arrays we compared these data with gene mutations from targeted resequencing and WGS, copy number variations, telomere length, RNA-Seq, miRNA-Seq data, and clinical outcome data. By defining epiCMIT-hyper [mean: 0.6, range: 0.29-0.91] and -hypo [mean: 0.71, range: 0.39-0.90] values in our cohort, we demonstrate a strong positive correlation between hyper- and hypo- scores, as previously observed in MCL and CLL (Durran-Ferrer, 2020). The highest score obtained for epiCMIT-hyper or -hypo per sample was then selected as the epiCMIT score per patient [mean: 0.72, range: 0.39-0.91]. We identified significant correlations between epiCMIT and key clinico-biological characteristics, including a significant negative correlation between epiCMIT and telomere length (R 2=0.67 p&amp;lt;0.001) supporting cell proliferation and concomitant telomere attrition. High epiCMIT scores were associated with female gender (p=0.02), IGHV1-2*04 alleles (p&amp;lt;0.001) with intermediate levels of IGHV somatic hypermutation (SHM) load (97-99.9%, p=0.04), driver somatic gene mutations, including those in KLF2 (p&amp;lt;0.001), NOTCH2 (p&amp;lt;0.01), TP53 (p=0.01) and KMT2D (p&amp;lt;0.001) and deletion of 7q (p=0.01) ( Fig1). Using previously defined classification systems (Arribas, 2015 and Bonfiglio, 2022), we show enrichment of high-risk subgroups [Arribas- high global methylation subgroup (High-M, p&amp;lt;0.001) and Bonfiglio- NFKB, NOTCH and KLF2 genetic subgroup (NNK, p=0.01)] in cases with high epiCMIT. In a sub-cohort of 42 SMZL cases, with a spectrum of epiCMIT values and WGS/ (mi)RNA-Seq data available, we demonstrated 1) a significantly higher genome-wide somatic mutational burden in patients with high epiCMIT-total scores (25 v 17mut/Mb, p=0.001), and 2), correlations between epiCMIT and specific gene/miRNA expression associated with cellular survival and proliferation ( CARD11 [R 2=0.6, p&amp;lt;0.01], MAP2K1 [R 2=0.61, p&amp;lt;0.01], miRNA-155 [R 2=0.58, p=0.018]), apoptosis ( BCOR [R 2=0.61, p&amp;lt;0.01]), and epigenetic regulation ( EZH2 [R 2=0.58, p&amp;lt;0.01]). We then conducted univariate Cox Proportional Hazards analysis to investigate the prognostic significance of clinical and molecular features on time to first treatment (TTFT; most prevalent treatments: splenectomy, n=51; rituximab, n=20) and overall survival (OS). The epiCMIT score and epiCMIT-hyper values showed the highest hazard ratios (HR) for TTFT (HR=26.6, p=0.001) and OS (HR=9.45, p=0.043) respectively ( Fig2). Other factors associated with shorter TTFT included TP53 mutation, 3q gain, IGHV1-2*04 usage, High-M, short telomeres and female gender. Shorter OS was significantly (p&amp;lt;0.05) linked to mutations in key genes (including TP53 and NOTCH2), genomic complexity and del(7q). To avoid redundancy and collinearity, only the most significant epiCMIT metric was included in a multivariate Cox Proportional Hazards model (backwards selection, 94 patients/63 events), and epiCMIT-total emerged as an independent marker for shorter TTFT (HR=44.4, p=0.004), together with gain of chromosome 3q (HR=2.6, p=0.002). Kaplan-Meier analysis further confirmed the importance of epiCMIT-total, with higher scores (&amp;gt;median: 0.73) associated with significantly shorter TTFT (median 23 vs 3.5 months) and higher mortality (p=0.004). To conclude, we demonstrate the potential clinical utility of epiCMIT score in SMZL patients, identifying that SMZL patients with high epiCMIT harbour specific genetic characteristics, and exhibit reduced treatment-free survival. EpiCMIT could be valuable in clinical practice, helping to identify those patients destined to progress and require closer monitoring.

Epidemiology and Survival of Patients with Marginal Zone Lymphoma Treated at the National Cancer Institute of Peru. a Retrospective Analysis of 187 Cases

Background Marginal zone lymphoma (MZL) represents a heterogeneous subgroup, classified as extranodal, splenic and nodal lymphoma. Extranodal cases, also termed mucosa-associated lymphoid tissue (MALT), account for 70% of MZLs. Splenic marginal zone B-cell lymphoma (SMZL) accounts for &amp;lt;2% of all NHL. There is no data about marginal zone lymphoma in Latin American patients. Methods We evaluated retrospectively all patients diagnosed with marginal zone lymphoma at the National Cancer Institute from 2010 to 2019. Baseline characteristics, treatments and survival were retrieved from medical records. Kaplan Meier method was used to analyze the overall survival. The estimate of the survival curves was performed by the Kaplan-Meier method, and the differences were computed by the log-rank test. The Cox proportional hazard models were used to identify predictors of survival of MZL. A level of p &amp;lt;0.05 was considered for a statistical significance. Results During the study period 2010-2019, 4437 patients with B-cell non-Hodgkin lymphoma were evaluated at the INEN, 187 of them were marginal zone lymphomas, representing 4.2% of all cases. The most frequent marginal zone lymphoma was MALT, followed by nodal and splenic lymphoma with 89.8%, 5.3% and 4.8% of cases, respectively. The median age for the entire cohort was 59 years (range 17-89). The median age for the MALT, nodal and splenic MZL cohort were 58.5 years (range 17-89), 75 years (range 52-88), 68 years (range 43-87), respectively. 37.4% were male for the entire cohort, and 37%, 40%, 44% for the MALT, nodal and splenic MZL group. Elevated LDH was seen in 13% of the entire cohort and in 11% and 24% of the MALT and splenic MZL group. Transformed lymphoma was reported in 6.4% of the entire cohort, but in the 20% of nodal MZL and no one case of the splenic MZL. More data you can see in the table 1. The most frequent primary site locations for MALT lymphoma were gastric, ocular adnexal, parotid gland, cutaneous, intestinal and CNS with 36%, 30%, 7%, 5%, 5% and 5%, respectively. The median overall survival was 114 months (IQR 76-133) for the entire cohort and 133 months, 59 and 59 months for the MALT, nodal MZL, and splenic MZL, respectively (p=0.0011), see the figure 1. The 5-year OS was 81.4% (95% CI, 71.6-88.1) for the entire cohort and 85.7% (95% CI, 76.5-91.5) for the MALT lymphoma group. No one patients were alive at 5 years in both nodal and splenic lymphoma groups. Conclusion Marginal zone lymphoma is a relatively rare type of B-cell lymphoma. The most frequent type is MALT lymphoma, followed by nodal MZL. The median age for the entire group was 59 years, however patients with nodal MZL were older with a median age of 75 years. Transformed lymphoma was present in 6% of the entire cohort and in the 20% of the nodal MZL. Gastric, ocular adnexal and parotid gland were the most common primary site of MALT lymphoma. MALT lymphoma had long survival, but nodal and splenic MZL had shorter survival with no one surviving 5 years from diagnosis.

Importance of FDG-PET (Fluorodeoxyglucose Positron Emission Tomography) in Staging and Response Assessment of Hairy Cell Leukemia (HCL)

Alessandro Mancini and Luca De Carolis are Co-first authors. Nicodemo Baffa, Brunangelo Falini and Enrico Tiacci are Co-last authors. INTRODUCTION HCL is an indolent BRAF-mutated leukemia usually presenting with cytopenias, splenomegaly, little or no lymphadenopathy, heavy bone marrow (BM) infiltration and few circulating leukemic cells. Imaging exams for staging include abdominal echography and computed tomography (CT), while the role of PET is largely unknown. METHODS 84 patients (pts) with relapsed/refractory HCL participating to our clinical trials of BRAF inhibitor-based therapies prospectively underwent PET/CT pre- and post-treatment, in addition to usual examinations for response assessment. Since FDG uptake is normally higher in liver than spleen, BM and lymph nodes, uptake in the latter tissues was considered abnormal if equal to or higher than the liver. Splenomegaly was defined by a longest spleen diameter &amp;gt;13 cm on PET/CT. The standard definition of complete remission (CR) in HCL just requires no palpable splenomegaly on physical examination, in addition to resolution of cytopenias as well as no hairy cells visible in the blood smear and the BM biopsy by non-immunological stains. RESULTS Pre-therapy, 58/77 (75%) non-splenectomized pts showed abnormal splenic FDG uptake (maximal standardized uptake value/SUVmax: median 4.8, vs 3.9 for the liver), which was always diffuse without focal lesions. Conversely, focal splenic uptake is not infrequent in splenic marginal zone lymphoma (18%, 7/39 pts - Abdom Radiol 2018;43:2721; vs of 0/58 HCL pts, p=0.0003), which may aid in the differential diagnosis with this HCL-mimicker. Moreover, 14/58 (24%) HCL pts. with a PET+ spleen did not have splenomegaly, and splenic uptake returned to normal in 10/11 evaluable cases (91%) achieving at least a partial response post-therapy, which suggests leukemic spleen involvement even without splenomegaly. Abnormal diffuse BM uptake was observed in 41/82 evaluable pts (50%; median SUVmax 5, vs 3.4 for the liver). In these cases, leukemic involvement in the BM biopsy was greater (median 80%) than in the other 41 pts (median 65%; p-value &amp;lt;0.01), pointing to HCL infiltration as the cause of abnormal BM uptake rather than to reactive hyperplasia of non-involved BM. Post-therapy, pathologic spleen uptake normalized in 46/51 evaluable pts (90%), and their CR rate was higher (32/46 pts, 70%) than in the other 5 pts remaining PET+ (1/5, 20%; p=0.047). Among the 46 PET- cases, 31 (67%) were not splenomegalic while 15 (33%) had residual splenomegaly (up to 17 cm; Fig. 1A); interestingly, within PET- cases relapse-free survival (RFS) was similar not only in conventionally defined CR cases with (n=8) vs without (n=24) residual non-palpable splenomegaly (p=0.84 after a median follow-up of 19 months), but also when the 6 cases meeting all conventional CR criteria except for still palpable splenomegaly were added to the 8 CR pts with residual non-palpable splenomegaly (p=0.7 after a median follow-up of 20.5 months - Fig. 1B). These findings suggest that metabolic status reflects splenic HCL involvement more reliably than spleen size or palpability, a concept that could improve the current definition of CR in HCL. Abnormal BM uptake resolved after treatment in 34/37 evaluable pts (92%), and their CR rate was again higher (23/34, 68%) than in the other 3 pts remaining PET+ (0/3; p=0.047). Leukemic infiltration of the BM biopsy was lower in the 34 PET- vs the 3 PET+ pts (median 5% vs 70%-80%, respectively; p=0.0004), indicating that BM abnormal uptake reflects HCL infiltration load rather than reactive BM hyperplasia post-therapy. Finally, abnormal lymph node uptake was observed pre-therapy in 9/84 (11%) pts (median SUVmax 7.1, vs 3.4 for the liver) and normalized after treatment in 7/8 evaluable pts (88%), including one case with residual lymph node enlargement (2.2 cm); all these 7 pts had achieved a CR while the remaining pt had no response to therapy. CONCLUSIONS PET is potentially useful in the differential diagnosis with splenic marginal zone lymphoma and may aid in the clinical management of HCL pts by detecting metabolic involvement of the main disease sites even in the absence of organomegaly. Importantly, PET signal tracks with response to therapy even in case of persistent organomegaly, which may lead to refine the definition of CR in HCL.

Pirtobrutinib, a Highly Selective, Non-Covalent (Reversible) BTK Inhibitor in Relapsed / Refractory Marginal Zone Lymphoma: Results from Phase 1/2 BRUIN Study

Background: Marginal zone lymphoma (MZL) is an indolent lymphoma that is typically treated with chemoimmunotherapy, but often requires sequential therapy. Covalent (c) Bruton tyrosine kinase inhibitors (BTKi) are increasingly being used in the relapsed or refractory (R/R) setting, but options for pts with MZL after cBTKi therapy may be limited. Pirtobrutinib, a highly selective, non-covalent (reversible) BTKi has favorable once daily oral pharmacology that enables continuous BTK inhibition throughout the dosing interval, regardless of intrinsic rate of BTK turnover. Pirtobrutinib has demonstrated promising efficacy and tolerability in pts with poor-prognosis B-cell malignancies following prior therapy, including prior cBTKi. Pirtobrutinib is approved in the USA to treat R/R mantle cell lymphoma after at least two lines of systemic therapy including prior BTKi treatment. Here we report the safety and efficacy of pirtobrutinib in pts with MZL from the BRUIN study (NCT03740529). Methods: Pts with previously treated B-cell malignancies, including MZL, were eligible for pirtobrutinib monotherapy treatment in either the dose escalation or expansion portion of the multicenter, phase 1/2 BRUIN study. MZL diagnosis required local pathologic review of an adequate biopsy. Endpoints include overall response rate (ORR) assessed by investigator per Lugano 2014 criteria, duration of response (DOR), progression-free survival (PFS), overall survival (OS), and safety. A 05 May 2023 data cut was used. Results: Among the 36 pts with MZL, 34 (94%) received the recommended phase 2 dose of pirtobrutinib, 200mg once daily.Pts had a median age of 68 (range 22-83), a majority were female (56%, n=20), and median prior lines of systemic therapy were 3 (range 1-10). Pts had nodal (47%, n=17), splenic (36%, n=13), and extranodal (17%, n=6) MZL subtypes. Most pts had Ann Arbor stage III/IV disease (81%, n=29). Most pts were in high risk MALT-IPI group (53%, n=19), 10 (28%) were intermediate, 1 (3%) was low, and missing in 6 (17%) pts. Baseline LDH was elevated in 15 (42%) pts and hemoglobin was &amp;lt;12 g/dL in 29 (81%) pts. All 36 MZL pts were previously treated with an anti-CD20 antibody, and 31 (86%) pts had received prior chemotherapy. Of the 26 pts (72%) with MZL who received a prior cBTKi, 20 (77%) pts discontinued due to progressive disease (PD) and 6 (23%) discontinued due to toxicity/other. Median time on treatment was 9 months (range 1.1-40.8), with 15 (47.1%) pts still receiving pirtobrutinib. The ORR of the MZL cohort was 50% (95% confidence interval [CI], 32.9- 67.1), including 1 (2.8%) complete response, 17 (47.2%) partial responses ( Figure). There were 15 (41.7%) pts with stable disease. The ORR in pts with nodal, splenic, and extranodal MZL were 58.8% (95%CI, 32.9-81.6), 30.8% (95%CI, 9.1-61.4), and 66.7% (95%CI, 22.3-95.7), respectively. The ORR for MZL pts with prior cBTKi therapy was 46.2% (95%CI, 26.6-66.6). With a median follow-up time of 17.6 months, median DOR was 12.7 months (95%CI, 5.6-non-estimable [NE]). Median PFS was 16.5 months (95%CI, 7.43-22.1) with a median follow up of 15.8 months. With a median follow up of 21.5 months, the median OS was NE (95%CI, 27.6-NE) and the 24-mo OS rate was 77.5% (95%CI, 56.3-89.3). In the MZL cohort, the most frequent treatment-emergent adverse events (TEAE) of any grade regardless of attribution were diarrhea (36.1%, n=13), fatigue (33.3%, n=12), and contusion (30.6%, n=11). Grade ≥3 hemorrhage/hematoma was not observed, and any grade hypertension (5.6%, n=2) was infrequent. No atrial fibrillation/flutter was observed. The most frequent grade ≥3 TEAE was neutropenia/neutrophil count decreased (27.8%, n=10) and anemia (13.9%, n=5). Treatment-related AE led to dose reductions in 4 pts (11.1%) and pirtobrutinib discontinuation in 2 (5.6%) pts (diarrhea and platelet count decreased/neutropenia). No fatal TEAEs were observed. Conclusions: In this cohort of heavily pre-treated R/R MZL pts, pirtobrutinib showed preliminary efficacy and was well tolerated with low rates of discontinuation.Pirtobrutinib appears to be a promising therapeutic option after cBTKi treatment in R/R MZL pts.

Marsun, a Phase III, Multicenter, Open Label, Randomized, Controlled Study Investigating Mosunetuzumab-Lenalidomide Versus Investigator Choices in Patients with Relapsed or Refractory Marginal Zone Lymphoma (R/R MZL)

Background MZLs are the second most common indolent NHL (iNHL) and comprise three distinct subtypes: (i) extranodal MZL (EMZL), mostly represented by MALT lymphoma; (ii) nodal MZL (NMZL); and (iii) splenic MZL (SMZL). For patients with high tumor burden based on GELF criteria, and patients with symptoms related to disease, first-line treatments rely on immuno-chemotherapy to reach high response rates at the cost of substantial toxicities notably in elderly patients. To date, R/R MZL remain a therapeutic challenge without clear standard of care treatment between immuno-chemotherapy, targeted therapies (i.e. BTK/PI3K inhibitors or the immunomodulatory agent lenalidomide (Len)), mainly because of an underrepresentation of MZL in the “indolent NHL” trials. Mosunetuzumab is a CD20xCD3 T-cell-engaging bispecific antibody designed to redirect T-cell cytotoxic activity against B-cells with promising efficacy results as monotherapy in various B-cell NHL. Lenalidomide was approved in the USA for R/R MZL based on data from MAGNIFY (MZL=45 patients) and AUGMENT (MZL=63 patients) and previous results of a phase IB study showed that mosunetuzumab in combination with Lenalidomide has a manageable safety profile and encouraging activity in patients with R/R follicular lymphomas (Morschhauser F. et al. ASH 2021) Here we present details of a trial in progress investigating the efficacy and the safety of mosunetuzumab-lenalidomide, a chemo-free regimen, compared to investigator choice, in patients with R/R MZL in need of systemic treatment and not eligible for local therapy such as radiotherapy or surgery. Methods This is an open label, multi-center, international, randomized phase III trial to compare the efficacy of Mosunetuzumab-Lenalidomide with investigator choices exclusively in R/R MZL patients. Patients with a proven diagnosis of EMZL, SMZL or NMZL and previously treated with at least one and no more than three prior systemic treatments are eligible. Previous treatment lines must include at least one systemic line with a drug targeting CD20 (monoclonal antibody at least 2 cycles) with or without chemotherapy (CHOP, bendamustine, CVP, chlorambucil) or targeted treatment such as ibrutinib. Patients will be stratified according to MZL subtype and time to progression of disease after first-line within 2 years. Mosunetuzumab will be administered subcutaneously (21 days C1, then 28 days C2-12) and lenalidomide will be given PO 20 mg/day from Day 1 to Day 21 from cycles C2 to C6. For each patient, investigator choice must be decided before randomization between R-Len (12 cycles) and R-chemotherapy (R-Bendamustine or R-CHOP followed by a 6 months maintenance). Both arms will have the same one-year fixed duration. The primary efficacy endpoint for comparison is progression-free survival (PFS) as determined by investigator (Lugano criteria 2014). Secondary objectives include CR24 (CR at 24 months) as determined by investigator and by central review according to Lugano criteria 2014 based on PET results, overall response rate and CR other than CR24 as determined by investigator, or by central review based on PET result according to Lugano criteria 2014. 260 patients are planned to be enrolled in France, Belgium, Germany, Italy and Portugal with FPI expected in Q3 2023 and approximately 4-year accrual period. The study will comprise a first stage of 10 patients in a safety cohort with mosunetuzumab-lenalidomide followed by a stage 2 with patients randomized 1:1 to receive M-len or investigator choice (125 patients per arm). The sponsor of the trial is the Lymphoma Academic Research Organisation (LYSARC) in collaboration with GLA and FIL with the support of F.Hoffmann-La Roche Ltd.

Hairy Cell Leukemia with Severe Leukocytosis

Introduction Hairy cell leukemia (HCL) is a rare indolent mature B-cells neoplasm that typically manifests with pancytopenia and splenomegaly. Nevertheless, significant splenomegaly, high hairy cells count in the blood, elevated β 2 microglobulin, and increased LDH can predict resistance to purine analogs. The absence of BRAF V600E mutation in HCL is rare and is associated with poorer outcomes. Case Presentation A 68-year-old female patient presented with symptomatic massive splenomegaly, leukocytosis (89,000 /microliter), monocytopenia, and thrombocytopenia. β 2microgobulin level was 4.4 mcg/L. Lactate dehydrogenase was 521 U/L. Peripheral blood and bone marrow examinations revealed typical hairy cells morphology, and the cells were positive for CD19, CD20, CD103, CD25, CD11c, CD200 (dim), and surface-Kappa. CD38 was negative. BRAF V600E result was wild-type. IGHV was mutated (IGHV3-49). She received first-line therapy with a 5-day course of Cladribine (0.15 mg/kg/day) with initial resolution of leukocytosis, normalization of platelets, and a 50% reduction in spleen size. This was followed by consolidation with an 8-weekly course of Rituximab (375 mg/m 2). Soon after the 2 nd dose of Rituximab, leukocytosis recurred, and she was initiated on second-line therapy with Bendamustine (90 mg/m 2 on days 1 and 2) along with Rituximab (BR). She has not achieved any durable response following two cycles of BR. Unfortunately, she suffered a septic shock requiring admission to our intensive care unit. Upon routine mental status evaluation, brain MRI revealed innumerable supratentorial and infratentorial bilateral acute/subacute and old microhemorrhages (Figure 1). This prompted further evaluation of unexplained recurrent episodes of pulmonary hemorrhages while on mechanical ventilation that were found to be secondary to pulmonary end-arterial cavitating lesions suggestive of hairy cells infiltration (Figure 2 arrowheads). As she was clinically unstable, we attempted to control leukocytosis with corticosteroids, leukapheresis aggressively and started the patient on Ibrutinib, but she succumbed to her disease. Discussion Recently, the 5 th edition of the World Health Organization classification of hematolymphoid tumors highlighted the splenic B-cell lymphoma and leukemia family that encompasses hairy cell leukemia (HCL), splenic B-cell lymphoma/leukemia with prominent nucleoli (SBLPN), formerly known as hairy-cell leukemia variant, splenic diffuse red pulp small B-cell lymphoma (SDRPL) and splenic marginal zone lymphoma (SMZL). HCL is typically an indolent mature B-cell neoplasm with characteristic pathologic features that preferentially insinuate the bone marrow and the spleen causing impaired hematopoiesis and splenomegaly. Unlike other disorders within this group, HCL commonly harbors BRAF V600E somatic mutation in almost all cases. Rarely, patients with HCL can have extramedullary involvement by the malignant cells. Leukocytosis in HCL appears to increase the risk of developing extramedullary lesions that can present unique clinical challenges necessitating early and effective therapy. According to the literature, brain lesions in HCL were described in three other cases, and leukocytosis was a common denominator. Our patient with significant leukocytosis had brainand pulmonary lesions, and the latter was the cause of her demise due tounusually aggressive and refractory disease. Therefore, we hypothesize that leukocytosis in HCL can be “the tip of the iceberg”. The higher the leukocytosis, the higher the burden of extramedullary disease, as in our case, who had simultaneous lesions in the brain and lungs. Patients with HCL and leukocytosis may warrant initial diagnostic evaluation to ensure the absence of potentially sinister lesions, which can inform subsequent monitoring and management plans. Additionally, more novel therapies could be considered sooner in the treatment course to control the disease and these lesions. Finally, our patient did not harbor BRAF V600E mutation, which likely resulted in further worse outcomes and limited the usage of targeted therapies like BRAF inhibitors.

Effectiveness and Safety of Orelabrutinib Combined with Rituximab As First-Line Treatment in Marginal Zone Lymphoma

Background: Marginal zone lymphoma (MZL) collectively is the third most common type of B-cell non-Hodgkin's lymphoma (NHL) worldwide, with an increased trend in incidence during the last two decades (Rossi D E et al., N Engl J Med. 2022). The lack of predominant first-line systemic therapy and toxicities associated with current rituximab-based therapies have inspired the exploration of more optimal treatment strategies (Becnel MR et al., Br J Haematol, 2019). Given the crucial role of bruton tyrosine kinase (BTK) in the B-cell receptor signaling pathway, BTK inhibitor has been proven to yield robust outcomes with improved tolerability in the development of NHL (Dhillon S et al., Drugs, 2021; Xu W et al., Blood, 2019; Song Y et al., Blood, 2019). Orelabrutinib is a novel BTK inhibitor with high selectivity and has been approved in China for relapsed/refractory MZL recently (Dhillon S et al., Drugs, 2021). Meanwhile, several clinical studies reported that orelabrutinib in combination with rituximab in NHL could preserve natural killer cell-mediated antibody-dependent cellular cytotoxicity induced by rituximab and enhance the clinical efficacy (Yu H et al., Mol Ther Oncolytics, 2021; Qu C et al., Blood, 2023). However, orelabrutinib plus rituximab as the first-line treatment for MZL remains largely understudied, thus, we conducted a retrospective study to evaluate its effectiveness and safety. Methods: Between July 27, 2021 and March 18, 2023, 10 patients with MZL who received orelabrutinib (150 mg, qd, po) with rituximab (375 mg/m 2, iv, day 1 of each 28-day cycle) as first-line therapy were included in this retrospective study. After 6 cycles of treatment, patients underwent orelabrutinib monotherapy (150 mg, qd, po) as maintenance for a year. Dynamic dose adjustment was performed according to the patient's clinical status. Effectiveness was assessed by positron emission tomography/computed tomography-based 2014 Lugano criteria. The primary endpoint was the overall response rate (ORR); secondary endpoints were progression-free survival (PFS), overall survival (OS), and safety. Results: Ten patients (6 males; median age 63.5 years, range 45-76) were included in this study, 4 with extranodal MZL of mucosa-associated lymphoid tissue (40.0%), 2 with nodal MZL (20.0%), 1 with splenic MZL (10.0%), and 3 with unknown subtype (30%). Among a total of 10 patients, 3 (30%) achieved complete response (CR) and 6 (60%) attained partial response (PR) as their best response for an ORR of 90% ( Figure 1). After a median follow-up of 13.0 months (range 7.8-24.7), the median PFS was not reached and a 6-month PFS rate was 100% ( Table 1). OS could not be assessed due to there were no deaths occurred. As of the cut-off date (May 6, 2023), 8 patients received orelabrutinib maintenance treatment and the median duration of maintenance treatment was 9.6 months (range 3.0-17.8). The ORR was 75% (6/8) during maintenance treatment, while 1 patient had stable disease (SD) and 1 developed progressive disease (PD). In addition, abnormal hematology due to disease gradually recovered with the therapy within 4 months. No serious adverse events were observed. The off-target related AEs such as atrial fibrillation, diarrhea, and major hemorrhage were not reported. Conclusion: This retrospective data suggests that orelabrutinib with rituximab has an encouraging anti-tumor activity in MZL, with a favorable safety profile. These results provide a potential first-line treatment strategy in MZL. Further verification in prospective clinical trials of clinical outcomes of orelabrutinib in first-line MZL is warranted in the future.