Chronic Lymphocytic Leukemia Group Research Articles

Ibrutinib followed by ofatumumab consolidation in previously untreated patients with chronic lymphocytic leukemia (CLL): GELLC-7 trial from the Spanish group of CLL (GELLC)

BTK inhibitors have been concurrently administered with anti-CD20 monoclonal antibodies (mAbs) in chronic lymphocytic leukemia (CLL). However, the optimal regimen for combining these two drugs remains pending. This multi-center phase 2 study aimed to analyze whether consolidation with ofatumumab improved the response in patients with CLL receiving front-line treatment with ibrutinib. Patients received 12 cycles of ibrutinib monotherapy. Those who achieved CR after this induction were maintained on ibrutinib. Conversely, those who did not attain CR continued with ibrutinib in addition to a consolidation, which involved 7 doses of ofatumumab. The primary objective was the complete response (CR) rate at cycle 20. This study is registered within the EU Clinical Trials Register (EudraCT 2016-004937-26). Between September 8, 2017, and May 21, 2018, 84 patients (median age, 69 years) were included. After completion of 12 cycles of ibrutinib (n=80), 4 patients (5%) were in CR, 67 (84%) in partial response (PR), and 6 patients (7%) had a PR with lymphocytosis (PRL). After consolidation with ofatumumab, 20 patients improved the response from PR to CR and 6 patients with PRL obtained a PR. Seventy-one patients (85%) completed 20 cycles of treatment, with a CR rate of 24/71 (34%). According to the intention-to-treat analysis at cycle 20, the ORR was 69/84 (82.2%), with a CRR of 24/84 (28.6%). Progression-free survival and overall survival at 48-months were 89.9% (CI: 82.4-95.5) and 92.2% (CI: 85.3-97.1), respectively. These findings underscore the potential for a consolidation strategy in CLL, wherein the addition of a mAb in patients with low tumor burden might enhance the quality of the response. The study was funded by Janssen that also supplied ibrutinib, whereas ofatumumab was supplied by Novartis.

The Utility of CD43 and CD200 in Differentiating Chronic Lymphocytic Leukemia from Other Mature B-Cell Neoplasms: A Cross Sectional Study.

To evaluate the utility of CD43 and CD200 in differentiating chronic lymphocytic leukemia (CLL) from other mature B-cell neoplasms. This was a cross-sectional study on patients diagnosed with B-cell neoplasms on flowcytometry. The median fluorescence intensity (MFI) of CD43, CD200 expressing neoplastic B-cells were compared between the CLL and non-CLL B-cell neoplasms followed by receiver operating characreristic curve (ROC) analysis. In addition, the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of CD43 and CD200 in diagnosing CLL were analysed. A total of 137 patients were included. The CLL group consisted 87 patients and non-CLL group consisted 50 patients. The Mann-Whitney U test showed significant CD43 expression (U = 997.5, Z= - 5.265, p < 0.001) and CD200 expression (U = 932.0, Z = - 5.5, p < 0.01) in CLL patients compared to non-CLL patients. The area under the curve were 0.771 and 0.786 for MFI of CD43 and CD200 in differentiating CLL from non-CLL group respectively. The optimal cut-off of MFI for CD43 and CD200 were 1323 and 1775 respectively. The sensitivity, specificity, PPV and NPV of CD43 in diagnosing CLL cases were 97.7%, 66%, 83.3% and 94.2% respectively. The sensitivity, specificity, PPV and NPV of CD200 in diagnosing CLL cases were 100%, 32%, 71.9% and 100% respectively. CD43 and CD200 are useful markers in differentiating CLL from other mature B-cell neoplasms with higher MFI expression of both markers found in CLL.

Abstract 18390: Ibrutinib Increases Ventricular Arrhythmia Burden Independently of Atrial Arrhythmia

Introduction: Use of Bruton Tyrosine Kinase inhibitors (BTKi) such as ibrutinib is frequently limited by atrial arrhythmia, possibly through off-target inhibition of C-Terminal-Src-Kinase (CSK). Whether BTKi can directly trigger ventricular arrhythmia is less clear. Case reports and retrospective analysis have suggested a link, but causal effect is not established. The association is further put into doubt by absent expression of CSK or BTK in ventricular cardiomyocytes, and absence of ventricular arrhythmia signals in meta-analyses of BTKi RCTs. Further study is critically needed. Hypothesis: Ibrutinib induces ventricular arrhythmia through a unique mechanism. Methods: We performed a retrospective crossover study of ibrutinib-treated patients with serial cardiac monitoring both on and off ibrutinib (pacemakers, loop recorders, or multiple holters) across a 10-year span in a large academic center to compare burden of atrial/ventricular ectopy on and off ibrutinib. A separate group of Chronic Lymphocytic Leukemia (CLL) patients with serial cardiac monitoring not on BTKi was used as additional controls. To determine biological correlation, we tested the ability of ibrutinib to induce arrhythmia in cultured human Induced Pluripotent Stem Cell derived ventricular cardiomyocytes (iPSC-vCMs). Results: 88% (15/17) of patients on ibrutinib with serial cardiac monitoring had increased burden of Premature Ventricular Contractions (PVCs), with an average increase of 155 PVCs/hour (p&lt;0.0001). An increase was not observed in the control cohort. Ventricular arrhythmia burden was not associated with atrial arrhythmia burden. Removing patient-related confounders, in vitro analysis of iPSC-vCMs demonstrated that ibrutinib induced arrhythmia following three days of treatment in a dose-dependent manner. Conclusions: In patients with continuous/serial cardiac monitoring, ibrutinib is associated with a significant increase in ventricular ectopy (average of 155 PVCs/hr). Lack of ventricular and atrial arrhythmia concordance suggests an alternative mechanism of arrhythmia induction. Increased arrhythmogenicity of human ventricular cardiomyocytes treated with ibrutinib indicates a direct myocardial mechanism.

Outcomes of Conservative Indication Criteria for First-Line Treatment in Chronic Lymphocytic Leukemia: Insights from the Brazilian Registry of CLL

Introduction: Chronic lymphocytic leukemia (CLL) exhibits diverse treatment requirements, with some patients receiving immediate treatment while others remain under observation. In 2008, the International Workshop on Chronic Lymphocytic Leukemia (IWCLL) established criteria for treatment indications, widely adopted in clinical practice and trials. The Brazilian Group of CLL (BGCLL) has been evaluating a more conservative approach to treatment indication performed in multiple centers, which avoids predefined cutoff levels for cytopenias and refrains from considering progressive lymphocytosis, extranodal involvement, or disease-related symptoms as isolated criteria. However, the safety and potential different outcomes of this conservative approach compared to the strict IWCLL criteria remain uncertain. Objective: This study aims to compare the outcomes of CLL patients without indication for treatment according to the BGCLL criteria, who were either treated or not based on the center's decision. Methods: We conducted a retrospective analysis of CLL patients enrolled in the Brazilian Registry of CLL, observed between January 2009 and March 2023, meeting the minimum data availability criteria for analysis and following IWCLL inclusion guidelines. Results: A total of 2396 patients from 41 centers were included. Among them, 1313 patients (55%) met the IWCLL criteria for treatment indication, while only 558 patients (23%) fulfilled the more conservative BGCLL indication. Of the 1240 untreated patients, 247 (20%) had a treatment indication based on IWCLL guidelines. Common indications for treatment initiation included cytopenias in 159 patients (64%), disease-related constitutional symptoms in 41 patients (17%), and symptomatic lymphadenopathy in 22 patients (9%). Non-treatment despite having an indication was attributed to BGCLL's more conservative criteria in 67% of cases, severe comorbidities in 6%, and death during follow-up in 27%. Among the 1156 treated patients, 90 patients (8%) received treatment despite lacking a treatment indication according to the IWCLL criteria. According to BGCLL's conservative criteria, 600 (48%) treated patients did not have a treatment indication. After a median follow-up of 58 months (range: 3-506), the overall survival (OS) was 70% at 8 years. Among patients without an indication for treatment according to BGCLL's conservative criteria, OS was significantly worse in treated patients (79%) compared to untreated patients (64%, P&amp;lt;0.0001, figure 1). Conclusion: These real-world data demonstrate that a conservative approach in indicating first-line treatment for CLL is safe and associated with improved survival, possibly by mitigating toxicities and treatment-related complications such as clonal selection. Furthermore, this strategy may conserve resources, enhancing drug accessibility for a larger number of patients in clear need of treatment, particularly in low-income countries.

Outcomes of a digital health coaching program for individuals with acute myeloid and chronic lymphocytic leukemia.

496 Background: Chronic lymphocytic leukemia (CLL) accounts for ~19,000 and acute myeloid leukemia (AML) accounts for ~20,000 new cases a year, according to SEER data. Individual management and overall survival rates vary by cancer type, each with unique and potentially long-term care needs. Digital interventions continue to expand, connecting with individuals beyond the clinical setting to support health self-efficacy and address overall wellness needs. However, data related to their application for hematologic malignancies is limited. Methods: The aim of this randomized, longitudinal cohort study was to evaluate the effect of a 3-month digital health coaching program on health self-efficacy, measured by the Cancer Behavior Inventory (CBI), and other patient reported outcomes (PROs) among individuals with AML or CLL. We aimed to enroll 500 participants, 250 each with CLL and AML, randomized to either the digital intervention or usual care. PROs were collected at baseline and up to once per month for 3 months via REDCAp. Clinical outcomes were tracked via electronic health record. The change in CBI scores from baseline to 3 months was calculated with 95% confidence intervals. A 2-sample t-test was used to compare change between intervention and control arms. PRO trends over time were analyzed using linear mixed models (LMMs). Results: The study enrolled 147 individuals from July 2020 through December 2022. Thirty-seven were enrolled in the AML group (19 control), and 110 in the CLL group (55 control). The mean change in CBI from baseline to 3 months in the AML control arm was -3.57 (95% CI: -17.30 – 10.15) and in the AML intervention arm was 7.03 (95% CI: -2.67 – 16.74), with higher scores indicating greater efficacy for coping. While not significant, mean change in CBI in the CLL control arm was 1.43 (95% CI: -3.45 – 6.31; p=0.556) and in the CLL intervention arm was 3.46 (95% CI: -1.44 – 8.37; p=0.219). In the CLL group, self-efficacy was associated with improved quality of life (p &lt;0.001); For every unit increase in FACT-LEU-TOI, CBI increased on average by 0.81 units (95% CI: 0.62 – 1.00). Conclusions: Due to limited study enrollment during the COVID-19 pandemic, the study was not powered to detect a difference between groups. Data from the engaged cohort suggest a non-statistically significant trend over time in improvement in health self-efficacy for the participants engaged in digital health coaching. A recent meta-analysis suggests self-efficacy has a strong negative correlation with distress and positive correlation with quality of life and improved self-efficacy may mitigate negative effects of a cancer diagnosis and treatment. Data related to trends in PRO over time within and between AML and CLL cohorts provide further insights into physiologic and psychosocial outcomes for these populations. Clinical trial information: NCT04774744 .

Chronic leukemias and COVID-19: An analysis of mortality and morbidity outcomes in the United States.

e18783 Background: COVID-19 pandemic has resulted in a global health crisis with widespread mortality and morbidity. Patients with chronic leukemias (CL) are at an increased risk for severe outcomes due to their compromised immune system. Despite the potential impact of COVID-19 on this population, there is a scarcity of information on the outcomes for CL patients. Our study aims to evaluate the mortality and morbidity of COVID-19 in CL patients in the United States. Methods: In this retrospective cohort study, we employed the National Inpatient Sample database of 2020, the largest inpatient database in the United States, to include adult patients (age ≥ 18 years) with chronic leukemias (CL) who were hospitalized for COVID-19, as identified by ICD-10-CM codes. The primary outcome was all-cause in-hospital mortality, while the secondary outcomes were acute respiratory failure (ARF) and the requirement for mechanical ventilation (MV). To identify the predictors of mortality, we conducted multivariable logistic regression analyses, with a p-value of less than 0.05 being considered statistically significant. Additionally, we conducted sensitivity analyses based on the type of CL (chronic lymphocytic leukemia (CLL) or chronic myeloid leukemia (CML)) and the remission status (in remission, not achieved remission, and relapse). The statistical analysis was performed using STATA software. Results: Among 91,375 patients with CL, 5.2% were hospitalized for COVID-19, with 87% of these patients having CLL. The majority of the CL patients (88.6%) had not achieved remission. The mean age of the patients was 72.7 years, with 62.4% being male and 78.9% being white. The mortality rate among CL-COVID-19 patients was 17%, with no significant difference in mortality rates between patients with and without CL (odds ratio [OR] 0.96, 95% confidence interval [CI] 0.81-1.14, p-value 0.65). About 61.5% of CL-COVID-19 patients had ARF and only 10.5% required MV. Patients with CL showed a 16% increase in the odds of ARF (OR 1.16, 95% CI 1.01-1.32, p-value 0.03) while no significant difference was found in the odds of MV. Sensitivity analysis showed no significant differences in mortality, ARF, or MV rates between CLL and CML groups. Another analysis by remission status revealed that patients who had not achieved remission had higher odds of ARF (OR 1.64, 95% CI 1.09-2.46, p-value 0.018) compared to those in remission or relapse. Conversely, patients in remission had a significantly lower likelihood of ARF (OR 0.63, 95% CI 0.41-0.97, p-value 0.036). No significant differences in mortality and MV odds were found according to remission status. Conclusions: Our findings emphasize the importance of vigilant monitoring and management of CL patients during the COVID-19 pandemic, particularly for those who have not achieved remission, to reduce their risk of severe respiratory outcomes.

Differential diagnosis of lymphocytosis in routine laboratory practice: Contribution of lymphocyte parameters using the Sysmex-XN9000 haematology analyzer.

This study investigates the potential contribution of structural and dispersion parameters, as well as alarms provided by Sysmex XN9000 haematology analyzer. The objective was to assess the need for a microscopic examination in the context of lymphocytosis. It also aims to contribute in differentiating rapidly lymphoproliferative disorders such as chronic lymphocytic leukaemia (CLL), non-chronic lymphocytic leukaemia (NON-CLL) and non-infectious reactive lymphocytosis (REAC). We prospectively assessed lymphocyte parameters (Ly-X, Ly-Y, Ly-Z, Ly-WX, Ly-WY, Ly-WZ) provided by the Sysmex XN9000 analyzer; they were measured in the white blood cell differential (WDF) channel which also provides alarms via the precursor/pathological cellular channel (WPC). Blood samples from 71 subjects with CLL, NON-CLL lymphoproliferative and REAC non-infectious reactive lymphocytosis, as well as a control (NORM) group of 12 subjects without any abnormalities were analyzed. The most discriminating parameters to distinguish the different groups were Ly-X, Ly-Z and Ly-WZ. The lymphoid structural parameters Ly-X and Ly-Z significantly discriminated the CLL group from the other groups (p < 0.001), and the CLL group from the REAC group (p < 0.01), respectively. The Ly-WZ parameter discriminated the CLL group from the NON-CLL, REAC (p < 0.001) and NORM (p < 0.01) groups. Alarms were higher in all study groups compared to the NORM group. An algorithm integrating these structural and alarm parameters is proposed. This study demonstrated that Ly-X, Ly-Z, and Ly-WZ lymphocyte parameters are useful for detecting morphological changes in lymphocytes; they provide useful information for the differential diagnosis of lymphocytosis, and this before the examination of the blood smear. An algorithm combining the WDF (parameters) and the WPC (alarms) makes it possible to decide whether or not to use a microscopic examination or flow cytometry immunophenotyping.

Additional malignancies shorten overall survival in chronic lymphocytic leukemia irrespective of chromosomal aberrations: A retrospective cohort study.

The aim of this study was to determine the incidence of other malignancies (OMs) in patients with chronic lymphocytic leukemia (CLL) and to identify parameters associated with the occurrence of OMs in addition to CLL. This retrospective cohort study was conducted by examining the records of CLL patients who applied to a tertiary hospital between January 2013 and December 2021. The cases were divided into 2 groups, CLL (n = 107) and CLL + OM (n = 25), according to the presence of additional malignancy. Lymphocyte count (P = .014), white blood cell count (P = .006), and hemoglobin (P = .034) were significantly higher in the CLL group. Rai stage IV percentage (P = .015), Binet stage B percentage (P = .043), progression, and sepsis percentages (P = .008) were significantly higher in the CLL + OM group. Overall survival time was significantly lower in the CLL + OM group (P = .032). Most OMs had been diagnosed before CLL (63.64%) in the no-treatment group, while the majority of OMs were diagnosed after CLL (78.57%) in the treatment group (P = .032). CLL patients with OM had a more advanced CLL stage, and survival was significantly shorter in these patients. In addition, CLL-associated OM appears to occur more frequently in the post-treatment period.

Chronic Lymphocytic Leukemia With Two B-Cell Populations of Discordant Light Chain Restrictions in Individual Patients: Parallel Development of Biclonal B-Cell Neoplasms or Clonal Evolution With Isotype Switch?

To evaluate clinicopathologic characteristics of biclonal chronic lymphocytic leukemia (CLL). Retrospectively analyze clinical data and pathologic features. Ten cases were identified in which flow cytometry demonstrated an abnormal B-cell population with a CLL-like immunophenotype but showed no definitive light chain restriction. All had cytogenetic abnormalities detected, including seven with two CLL-related abnormalities. Four of these showed features suggestive of clonal evolution, all having del(13q) as a "stem-line" abnormality and three showing del(11q) as a "side-line" abnormality. Five (50%) cases demonstrated deleterious NOTCH1 mutations, in contrast to 11.8% in a control group of monoclonal CLL (P < .05). Of the 10 patients, 5 received treatment, with good/partial response in three cases and therapeutic resistance in one case. The median treatment-free survival was estimated at 68 months. Despite a polytypic pattern of light chain expression, the neoplastic nature of biclonal CLL is suggested by a characteristic CLL phenotype and can be confirmed by cytogenetic and genomic analyses. The two clones with discordant light chain isotypes may share a "stem-line" cytogenetic abnormality, suggesting possible clonal evolution. Biclonal CLL is associated with NOTCH1 mutations, which may occur in a small subclone and gradually evolve in clonal size. Genomic analysis on light chain-sorted and/or chronologically collected samples may provide insight into clonal evolution in CLL.

Application of modified Matutes score containing CD200 in diagnosis of chronic lymphocytic leukemia.

Immunophenotyping technique is a powerful tool for the diagnosis and differential diagnosis of chronic lymphocytic leukemia (CLL) and other B-cell chronic lymphoproliferative diseases (B-CLPD). CD200 is strongly expressed in CLL. This study aims to analyze the clinical value of modified Matutes score (MMS) containing CD200 in the diagnosis of CLL. We retrospectively analyzed 103 B-CLPD patients diagnosed from January 2020 to July 2021, including 64 CLL patients, 11 follicular lymphoma (FL) patients, 14 mantle cell lymphoma (MCL) patients, 6 marginal zone lymphoma (MZL) patients, 1 hairy cell leukemia (HCL) patient, and 7 lymphoplasmic lymphoma/Waldenstrom macroglobulinemia (LPL/WM) patients. The expression of CD markers between the CLL group and the non-CLL group was compared, and the sensitivity, specificity, and clinical consistency of MMS and Royal Marsden Hospital (RMH) immunophenotyping score system were analyzed. There were significant differences in the expressions of CD5, CD23, FMC7, CD22, CD79b, CD200, and sIg between the CLL group and the non-CLL group (χ2 values were 37.42, 54.98, 30.71, 11.67, 55.26, 68.48, and 17.88, respectively, all P<0.01). When the RMH immunophenotyping score≥4, the sensitivity was 79.7%, and the specificity was 100%. When the MMS≥3, the sensitivity was 95.3%, and the specificity was 100%. The Kappa coefficient of RMH immunophenotyping system was 0.677, and the Kappa coefficient of MMS system was 0.860. The MMS system containing CD200 has better sensitivity and same specificity compared with RMH immunophenotyping system, and MMS system may be more useful in the diagnosis of CLL.

Low plateletcrit is associated with reduced progression: Free and overall survival in chronic lymphocytic leukemia.

Alterations of plateletcrit and mean platelet volume (MPV) and pathogenesis of chronic lymphocytic leukaemia (CLL) have been linked to various inflammatory disorders. The prognostic impact of plateletcrit and MPV were evaluated. MPV and plateletcrit levels of both CLL and control group were compared and then in CLL patients, additional diseases, leukocyte count, platelet count, lactate dehydrogenase, Rai stage, progression-free and overall survival, mutations, if any, and chemotherapy, if any, were recorded. Then, the relationship between MPV and plateletcrit values and these parameters were evaluated in CLL patients. Platelet and plateletcrit values were found to be significantly lower in CLL patients than the control group (p<0.001) for both. Plateletcrit and MPV values of patients who did not receive chemotherapy were higher than those who received chemotherapy (p=0.03, p=0.02, respectively). Being over 75 years old, plateletcrit value less than 0.1565 %, platelet level below 175 x 109/L, and leukocyte count greater than 53.5 x 109/L was found to significantly reduce overall survival. Male gender, each stage increase, plateletcrit less than 0.1565 % and leukocyte count greater than 53.5 x 109/L was related to reduce treatment-free survival in CLL patients. Plateletcrit can be a viable prognostic marker for defining both treatment free and overall survival.

Abstract A39: Targeting tumor-microenvironmental dependencies as a therapeutic approach in Chronic Lymphocytic Leukemia (CLL)

Abstract Interaction between CLL and its microenvironment is a major factor in drug resistance. Bone marrow derived stroma cells provide a protective and supportive niche to allow CLL cells to survive drug-induced apoptosis. Therefore, a current challenge is to develop new strategies to target the microenvironment. To understand the role of tumor microenvironment in CLL survival, we studied conditioned media (CM) derived from NKTert cells. Human NKTert stroma cells provide a survival advantage to primary CLL cells by protecting them from spontaneous apoptosis in ex-vivo culture. We identified three cytokines (IL6, IL8 and CCL2) from CM as candidates for mediating survival. We exposed CLL cells to neutralizing antibodies against each cytokine and found significant decrease in CLL viability in presence of anti-CCL2 suggesting its role in CLL survival. BH3 profiling revealed an increase in apoptotic priming with anti-CCL2 treatment as well. Using dynamic BH3 profiling (DBP), we found that anti-CCL2 increased dependence on BCL-2 as well as BCL-xL in primary CLL cells suggesting a potential role of CCL2 on CLL survival. To our surprise, we also identified a CLL group (CM independent, CMI) that were resilient to cell death in the absence of stroma support in ex-vivo culture. In addition to this, anti-CCL2 antibody showed no effect on their survival. Next, we exposed CMI CLL cells to venetoclax and found that these CLL cells were less sensitive to apoptosis compared to the CLL samples that are stroma dependent (CM dependent, CMD). BH3 profiling showed that CMI CLL cells are less primed for apoptosis. Consistent with our findings, DBP with anti-CCL2, venetoclax or the combination didn’t increase any drug induced apoptotic priming in CMI CLL samples. In our study, 80% of CMI harbor unmutated IgHV. To provide additional insight to the molecular mechanism, we performed western blot analyses for BCL-2 family proteins and found decreased BAX ad BCL-2 expression in CMI samples. This result suggests altered BCL-2 family proteins expression might be a mechanism for differential priming. Bulk RNA seq analysis revealed upregulation of focal adhesion genes and Ras signaling pathway in CMI samples. In summary, our study suggest that in the CMD subset of CLL, anti-CCL2 therapy enhances apoptotic priming and venetoclax efficacy. Furthermore, we have also functionally defined two biologically distinct CLL subgroups based on stroma dependence. Citation Format: Salma Parvin, Aditi Aryal, Shanye Yin, Matthew S. Davids, Geoffrey G. Fell, Catherine J. Wu, Anthony Letai. Targeting tumor-microenvironmental dependencies as a therapeutic approach in Chronic Lymphocytic Leukemia (CLL) [abstract]. In: Proceedings of the Third AACR International Meeting: Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2022 Jun 23-26; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2022;3(5_Suppl):Abstract nr A39.

Increased serum level of alpha-2 macroglobulin and its production by B-lymphocytes in chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL), the most common adult’s leukemia in the western world, is caused in 95% of the cases by uncontrolled proliferation of monoclonal B-lymphocytes. The complement system in CLL is chronically activated at a low level via the classical pathway (CP). This chronic activation is induced by IgG-hexamers, which are formed after binding to alpha-2-macroglobulin (A2M). The study investigated for the first time the serum levels of A2M in CLL patients, their association with the disease severity, and A2M production by the malignant B-lymphocytes. Blood samples were collected from 65 CLL patients and 30 normal controls (NC) subjects, and used for quantifications of the A2M levels, the complement activation marker (sC5b-9), the complement components C2, C3 and C4, and clinical biochemistry and hematology parameters. The production of A2M was studied in B-lymphocytes isolated from blood samples as well as in CLL and non-CLL cell lines.The serum A2M levels were significantly higher in CLL patients vs NCs, showing values of 3.62 ± 0.22 and 1.97 ± 0.10 mg/ml, respectively. Within the CLL group, A2M levels correlated significantly with the disease stage, with sC5b-9, and with clinical indicators of the disease severity. Increased A2M production was showed in three out of four CLL B-lymphocytic lines that were studied, as compared to non-CLL lines, to a non-lymphocytic line, and to blood-derived primary B-lymphocytes. A2M production was further increased both in primary cells and in the CLL cell-line after incubation with CLL sera, compared to NC sera. This study shows for the first time that serum A2M levels in CLL are significantly increased, likely due to A2M production by the malignant B-lymphocytes, and are correlated with the disease severity and with chronic complement activation. The moderate change in A2M production after incubation with NC sera in-vitro supports the hypothesis that inhibition of excess A2M production can be achieved, and that this may potentially down-regulate the IgG-hexamerization and the resulting chronic CP activation. This may also help restore complement system activity, and eventually improve complement activity and immunotherapy outcomes in CLL.

IGH Translocations in Chinese Patients With Chronic Lymphocytic Leukemia: Clinicopathologic Characteristics and Genetic Profile.

Immunoglobulin heavy chain translocations (IGH-t) have occasionally been reported in Chinese patients with chronic lymphocytic leukemia (CLL). The objective of the present study was to identify the clinicopathologic features of patients with IGH-t CLL and compare them with those of patients with non-IGH-t CLL. We performed fluorescence in situ hybridization (FISH) based on a routine CLL prognostic FISH panel using IGH, IGH-BCL2, BCL3, IGH-CMYC, and BCL6 FISH probes. Furthermore, we retrospectively evaluated the clinical features of 138 newly diagnosed CLL patients via chromosome banding analysis (CBA), FISH, and targeted next-generation sequencing. IGH-t was identified in 25 patients (18.1%). Patients with IGH-t CLL had lower flow scores than those with non-IGH-t CLL. The most frequent translocation was t(14;18) (10 patients), followed by t(14;19) (3 patients), and t(2;14)(p13;q32), t(7;14)(q21.2;q12), t(9;14)(p13;q32) (3 patients). The remaining nine patients included three with abnormal karyotypes without translocation involving 14q32, four with a normal karyotype, and two who failed CBA. The most frequently concomitant FISH-detected aberrations were 13q deletion, followed by +12 and TP53 deletion, while one case involved ATM deletion. Complex karyotypes were detected in five patients with IGH-t CLL, in whom all partner genes were non-BCL2. Available mutational information indicated that KMT2D mutation was the most frequent mutation among tested 70 patients, while TP53 mutation was the most frequent mutation in the IGH-t group. Moreover, the IGH-t group had higher FBXW7 (P=0.014) and ATM (P=0.004) mutations than the non-IGH-t group, and this difference was statistically significant. Our study demonstrates that IGH-t is not uncommon among Chinese CLL patients, and that its partner genes are multiple. The gene mutational profile of the IGH-t group was distinct from that of the non-IGH-t group, and the concomitant chromosomal abnormalities within the IGH-t CLL group differed. Thus, identification of IGH-t and its partner genes in CLL patients may help further refine risk stratification and strengthen the accurate management in CLL patients.

Evaluation of Serum Level of CXCL12 in Correlation with Several Clinical and Hematological Parameters in Patients with Chronic Lymphocytic leukemia

Background: Chronic lymphocytic leukemia (CLL) is the commonest leukemia in western countries. The disease typically occurs in elderly patients and has a highly variable clinical course, transformation is initiated by specific genomic changes that impair apoptosis of clonal B‐cells. C-X-C Motif chemokine Ligand 12(CXCL12) is a chemotactic factor for T cells and monocytes, and in B cell lymphopoiesis and bone marrow myelopoiesis. CXCL12 is a 68-amino acid small (8 kDa) cytokine that belongs to the CXC chemokine family. It had been reported as key mediator for angiogenesis and inflammation, also it expressed by bone marrow stromal cells in CLL patients and it is responsible for CLL cells chemotaxis in to the tissue microenvironments.Aims of the study: To assess CXCL12 expression in patients with chronic lymphocytic leukemia and to correlate their expression with different clinicopathological and hematological parameters . Materials and Methods: the study included 80 subjects divided in to two groups: Patient group included 40 adult patients with CLL and control group included 40 healthy age and gender-matched controls. All hematological parameters were done by automated hematological analyzer.Serum CXCL12 was estimated in both patients and control subjects by ELISA technique. Analysis of data was carried out using the available statistical package of SPSS-25 (Statistical Packages for Social Sciences-version 25). Results: Serum CXCL12 expression was significantly higher in CLL patients compared to controls .There was significant inverse correlation between CXCL12 expression and hemoglobin level and significant direct correlation between its expression and total W.B.C. Count and absolute lymphocyte count , significant direct correlation had been also found between CXCL12 expression and advanced Ria stage .

Incidence of Non-Melanoma Skin Cancers in Patients with Chronic Lymphocytic Leukaemia: A Retrospective Study in a Bay of Plenty, New Zealand Population

Background/Aims: Chronic lymphocytic leukaemia (CLL) incurs an increased risk of developing second primary malignancies (SPMs). The pathogenesis underpinning this increased SPM risk is attributable to synergism between genetic aberrations, chronic immune suppression / dysfunction and CLL treatments. Specifically, CLL associated skin cancer is a well-documented phenomenon, with common malignancies such as squamous (SCC) and basal cell (BCC) carcinomas occurring at higher frequencies in CLL patients and are often of a more aggressive phenotype. The incidence of non-melanoma skin cancers (NMSCs) among New Zealand CLL patients is unknown, as NMSCs are not required to be reported to the New Zealand Cancer Registry. Accordingly, this study aimed to analyse the incidence of NMSC in CLL patients from the Bay of Plenty region of New Zealand.Methods: We conducted a retrospective analysis of 213 CLL patients, who were referred to Bay of Plenty District Health Board (BOPDHB) between 2015 and 2020, for the occurrence of SPMs. Clinical notes were reviewed for the demographics of sex, age at time of CLL diagnosis, CLL treatments received and occurrence and type of NMSCs. A control group of 76 patients referred to BOPDHB haematology, for anaemia or thrombocytopenia, of which the cause subsequently remained unknown and was not associated with lymphoproliferative disease, was reviewed for primary malignancy (PM), occurrence and type. NMSC incidence was also compared to a local data, on overall NMSC incidence in the general Bay of Plenty population, in 2015.Results: The predominant SPM types occurring in this CLL patient group were NMSCs. A total of 249 and 170 histological diagnoses of SCC and BCC were made in our group of CLL patients, respectively. Of the 213 CLL patients, 48.8% had a diagnosis of at least one NMSC, with median time between CLL and NMSC diagnosis of 5 years (range, -6 to 19 years). CLL patients that were diagnosed with NMSC developed a mean number of 3.93 ± 2.94 separate lesions, compared to 2.6 ± 1.7 lesions in our control population. Note: comparison of NMSC incidence data between CLL group and Bay of Plenty populationConclusion: Patients with CLL are at increased risk of developing multiple NMSC lesions. Therefore, we believe CLL patients should be assessed by dermatologists or general practitioners for whole-body skin examination, 6 monthly following their CLL diagnosis. This strategy promotes early diagnosis, and patient education on importance of sun protection strategies to reduce NMSC risk. This is essential to the ongoing care of CLL patients in a NZ context, given the increased incidence of NMSC associated with CLL, in combination with the high levels of UV exposure and skin damage among the NZ general population. This study also highlights how NMSC disease burden is likely to be significantly underestimated due to a lack of data collection on these malignancies by the New Zealand Cancer Registry. DisclosuresNo relevant conflicts of interest to declare.

Outcomes of Patients with Hematologic Malignancies and COVID-19: Perspective of a Large Hematology Center in Greece

The emergence of SARS-CoV-2, as of July 2021 has affected 469,042 individuals and accounted for 12,851 deaths nationally in Greece, according to WHO database. Mortality rate is higher in elderly patients (pts) and in pts with comorbidities, including malignancies. However, there is a growing interest on COVID-19 outcomes in pts with hematologic diseases. The aim of this study was to perform a systematic registration and analysis of the outcomes of pts with hematologic disease and COVID-19 in our center.The study is a single-center, retrospective study, conducted at a Hematology Department and HCT unit of a tertiary Hospital after approval from local Ethics Committee. We included pts with a hematologic disease and RT-PCR confirmed COVID-19 infection between October 2020 and July 2021. We reviewed hematological medical records to extract demographic and clinical data of COVID-19 infections. Most of the data have already been intergraded in ASH Research Collaborative Data Hub.Hematologic diseases were categorized to: Acute Myeloblastic Leukemia (AML), Acute Lymphoblastic Leukemia (ALL), Non-Hodgkin Lymphomas (NHL), Chronic Lymphocytic Leukemia (CLL), Hodgkin Lymphoma (HL), Multiple Myeloma (MM), Myelodysplastic Syndromes (MDS), Chronic Myeloid Leukemia (CML), Myeloproliferative Neoplasms (MPN, including all Philadelphia-negative MPN) and other hematologic conditions.We evaluated a total of 89 pts, 54% were male and 46% female, with a median age of 64.5 (20-86) and 59.5 (21-85) years respectively. 83% of pts were ≥40 years and 27% ≥70 years old. Most of them (92%) acquired infection outside a hospital setting. 13% of pts were asymptomatic and diagnosis was confirmed only with positive RT-PCR test.The most common represented malignancies were NHL 26%, CLL 20% and acute leukemias 13.5%, while 15% of pts underwent transplantation (HCT).Pts presented with moderate/severe COVID-19 were 55%, while 43% of hospitalized pts required Intensive Care Unit (ICU) admission. Overall, the death rate was 24%, while remarkably almost all pts required ICU support did not survive (mortality 94%). Higher mortality observed in patients with MDS (50%), MM (43%), CLL (39%), ALL (33%) and NHL (30%).Further analysis showed a positive correlation between mortality and male gender with 16 deaths out of 21 (p = .0245), as well as mortality and ICU admission (p < .001). A chi-square test of independence was performed to examine the relation between age and COVID-19 severity, without any statistically significant result [x 2 (2, N = 87) = 3.475, p = .176]. Whereas the only significant correlation between age and mortality was among age groups 18-39 and >70 years (p = .0146).Regarding treatment, pts were divided into two subgroups, 78% of them received anticancer therapy at least once in their lives, while 22% of them have never been on treatment, mainly pts with CLL and indolent NHL. 62% of the first subgroup manifested moderate/severe COVID-19 infection requiring hospitalization with 28% death rate, while the same rates in the 2 nd subgroup were 30% and 10% respectively. Although there was a significant correlation between the treatment status and the severity of COVID-19 infection (p = .020), the above was not translated in statistically higher death rate in the first subgroup (p = .14).There was also a correlation between HCT and COVID-19 severity in general (p = .005), with autologous HCT having statistically higher mortality than the allogeneic subgroup (p = .032). Α similar analysis in CLL and NHL groups showed no relation among treatment status, COVID-19 severity, and mortality (p values .638 and .115/ .34 and .62 respectively).As anticipated in hematological pts, the immunocompromised nature of the underlying disease makes them extremely vulnerable to COVID-19 infection regardless of their treatment status, a fact that is also reflected in mortality despite ICU admission and support. In general, the severity of infection is correlated to anticancer therapy, while mortality to male sex, ICU admission and autologous HCT.Larger number of pts are necessary for further studies to better understand the parameters that impact the outcome of COVID-19 in hematological pts. Hematology departments should remain COVID-19 free zones, dedicated only to hematologic treatment and pts should strictly comply with social distancing. It remains to see if vaccines can play a key role to protect this special group of pts. [Display omitted] DisclosuresAnagnostopoulos: Abbvie: Other: clinical trials; Sanofi: Other: clinical trials ; Ocopeptides: Other: clinical trials ; GSK: Other: clinical trials; Incyte: Other: clinical trials ; Takeda: Other: clinical trials ; Amgen: Other: clinical trials ; Janssen: Other: clinical trials; novartis: Other: clinical trials; Celgene: Other: clinical trials; Roche: Other: clinical trials; Astellas: Other: clinical trials .

Epidemiological Characterization and Determination of TP53 and IGHV Mutational Status of a Large Series of Previously-Untreated Chronic Lymphocytic Leukemia (CLL) Patients in Spain: The Epicll Study

Epidemiological Characterization and Determination of TP53 and IGHV Mutational Status of a Large Series of Previously-Untreated Chronic Lymphocytic Leukemia (CLL) Patients in Spain: The Epicll Study

Specific T Cell Receptor Gene Repertoire Profiles in Subgroups of CLL Patients with Distinct Genomic Aberrations

Chronic lymphocytic leukemia (CLL) B cells engage in multifaceted bi-directional interactions with bystander cells, including T cells. Immunogenetic studies in CLL revealed clonal expansions of T cells and shared T cell clonotypes between different patients, strongly implying clonal selection by antigens. Although the exact nature of these antigens remains largely elusive, evidence exists that the clonotypic B cell receptor immunoglobulin (BcR IG) may serve as a source of antigenic epitopes for T cells. That said, recurrent genomic aberrations associated with distinct abnormal expression profiles could represent an alternative, non mutually exclusive, source of potent immunogenic onco-antigens that might shape the T cell repertoire in CLL. On these grounds, here we interrogated the T cell receptor (TR) gene repertoire of CLL patients with different genomic aberration profiles with the aim to identify unique signatures that would allude to distinct antigen selection pressures. The study group included 46 patients with CLL, sampled before treatment initiation, who were categorized in 5 subgroups defined by a unique genomic aberration, as follows: +12, n=18; del(11q), n=10; del(13q), n=7; del(17p)/TP53mut, n=6; NOTCH1mut, n=5. Confounding effects of multiple aberrations have been minimized, as we previously established through comprehensive characterization (including FISH, SNP arrays and gene panels) that the analyzed patients carried only one of the above aberrations. Starting material was RNA extracted from blood mononuclear cells. TRBV-TRBD-TRBJ gene rearrangements were RT-PCR amplified and subjected to paired-end next generation sequencing (NGS). Raw NGS reads (n=13,213,563| median: 294,757/sample) were processed through a purpose-built bioinformatics pipeline. Only productive rearrangements (n=9,249,546 | median=199,184/sample) were taken into consideration for the computation of clonotypes i.e. TRB rearrangements with identical TRBV gene usage and amino acid complementarity-determining region 3 (CDR3) sequence. Overall, 513,984 distinct clonotypes (median=10,304 clonotypes/sample) were assessed. The main measure of clonality employed in this study was the median cumulative frequency of the 10 most expanded T cell clonotypes/sample (MCF-10). For comparisons of the clonality profiles, a group of 17 aged-matched healthy individuals were used as controls. All patients displayed oligoclonal T cell expansions with the following MCF-10 values: del(11q): 21.6%, +12: 25%, del(13q): 20.6%, NOTCH1mut: 9.1%, del(17p)/TP53mut: 12.9%; the difference between the del(11q) and +12 groups versus the NOTCH1mut group was statistically significant (p<0.05). The MCF-10 value of the control group was estimated at 17.5%, supporting the notion of age-related decrease in TR repertoire diversity. However, the del(11q), +12 and del(13q) CLL groups displayed elevated clonality, reaching statistical significance (p<0.002) in the case of +12. TRBV gene repertoire analysis revealed that the TRBV12-3 gene predominated in most groups, except for the del(17p)/TP53mut, where the predominant gene was TRBV10-3. Clonotype comparisons disclosed the presence of shared TR clonotypes both within a particular group but also between groups. Overall, 446/513,984 clonotypes were found to be shared by at least two patients across all groups; the vast majority (392/446, 88%) of shared clonotypes appeared to be CLL-biased since they did not match entries in public databases of TR clonotypes from various contexts. Subgroup-specific clonotypes were identified for all aberrations examined; these emerged as unique to the particular subgroups, as revealed by extensive comparisons against both public databases but also a large TR clonotype database from CLL available to us from our previous studies. In conclusion, recurrent genomic aberrations, especially large chromosomal abnormalities, display an oligoclonal TR gene repertoire. The distinct immunogenetic profile of each group examined here and, most importantly, the existence of subgroup-specific clonotypes, suggest that abnormal protein expression and gene dosage effects likely represent a relevant source of CLL-specific selecting antigens. DisclosuresScarfo: Janssen: Honoraria, Other: Travel grants; Astra Zeneca: Honoraria; Abbvie: Honoraria. Anagnostopoulos: Abbvie: Other: clinical trials; Sanofi: Other: clinical trials ; Ocopeptides: Other: clinical trials ; GSK: Other: clinical trials; Incyte: Other: clinical trials ; Takeda: Other: clinical trials ; Amgen: Other: clinical trials ; Janssen: Other: clinical trials; novartis: Other: clinical trials; Celgene: Other: clinical trials; Roche: Other: clinical trials; Astellas: Other: clinical trials . Ghia: AbbVie: Consultancy, Honoraria, Research Funding; Acerta/AstraZeneca: Consultancy, Honoraria, Research Funding; AstraZeneca: Consultancy, Honoraria, Research Funding; ArQule/MSD: Consultancy, Honoraria; BeiGene: Consultancy, Honoraria; Celgene/Juno/BMS: Consultancy, Honoraria; Gilead: Consultancy, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria; Sunesis: Research Funding. Haferlach: MLL Munich Leukemia Laboratory: Other: Part ownership. Rosenquist: Roche: Honoraria; Janssen: Honoraria; Illumina: Honoraria; AstraZeneca: Honoraria; Abbvie: Honoraria. Stamatopoulos: Gilead: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding; AstraZeneca: Honoraria, Research Funding; Janssen: Honoraria, Research Funding. Baliakas: Janssen: Honoraria; Gilead: Honoraria, Research Funding; Abbvie: Honoraria. Chatzidimitriou: Abbvie: Honoraria, Research Funding; Janssen: Honoraria, Research Funding.

Safety and Efficacy of Idelalisib Treatment of Chronic Lymphocytic Leukemia (CLL) or Lymphoma Relapsing after Allogeneic Hematopoietic Cell Transplantation (alloHCT): A Survey By the EBMT Chronic Malignancies and Lymphoma Working Parties

Safety and Efficacy of Idelalisib Treatment of Chronic Lymphocytic Leukemia (CLL) or Lymphoma Relapsing after Allogeneic Hematopoietic Cell Transplantation (alloHCT): A Survey By the EBMT Chronic Malignancies and Lymphoma Working Parties