IgVH Status Research Articles

Prognostic impact of genetic abnormalities in 536 first-line chronic lymphocytic leukaemia patients without 17p deletion treated with chemoimmunotherapy in two prospective trials: Focus on IGHV-mutated subgroups (a FILO study).

The potential prognostic influence of genetic aberrations on chronic lymphocytic leukaemia (CLL) can vary based on various factors, such as the immunoglobulin heavy variable (IGHV) status. We conducted an integrative analysis on genetic abnormalities identified through cytogenetics and targeted next-generation sequencing in 536 CLL patients receiving first-line chemo(immuno)therapies (CIT) as part of two prospective trials. We evaluated the prognostic implications of the main abnormalities, with specific attention to their relative impact according to IGHV status. In the entire cohort, unmutated (UM)-IGHV, complex karyotype, del(11q) and ATM mutations correlated significantly with shorter progression-free survival (PFS). Focusing on the subset of mutated IGHV (M-IGHV) patients, univariate analysis showed that complex karyotype, del(11q), SF3B1 and SAMHD1 mutations were associated with significant lower PFS. The prognostic influence varied based on the patient's IGHV status, as these abnormalities did not affect outcomes in the UM-IGHV subgroup. TP53 mutations had no significant impact on outcomes in the M-IGHV subgroup. Our findings highlight the diverse prognostic influence of genetic aberrations depending on the IGHV status in symptomatic CLL patients receiving first-line CIT. The prognosis of gene mutations and cytogenetic abnormalities needs to be investigated with a compartmentalized methodology, taking into account the IGVH status of patients receiving first-line BTK and/or BCL2 inhibitors.

Early Relapse after First Line Has a Significant Impact on Overall Survival in Patients with Mantle Cell Lymphoma (MCL)

Background:Mantle cell lymphoma (MCL) is still an incurable disease. While OS has significantly improved in recent years with more effective treatments, early relapse/refractoriness to induction treatment is associated with a dismal outcome (Visco, BJH 2019; Eskelund, HemaSphere 2021). Early identification of patients with MCL who will relapse early or not respond after induction could improve their management.Patients and methods:We retrospectively reviewed the characteristics of all consecutive patients diagnosed with MCL in our center from April 2000 to January 2021. The analysis included OS, PFS, DOR, age group (≤65 or >65 years old), morphology (classic vs pleomorphic/blastoid), Ki67 (<30% or ≥30%), MIPI, type of induction: HiDAC-based (high-dose citarabine-based) or non-HiDAC regimens, and early versus late relapse (defined as POD<24 or POD>24 when the relapse occurred ≤24 months or >24 months from diagnosis to the start date of the 1 st salvage treatment). Other factors currently known as relevant (SOX11, TP53 mutational status, karyotype, IGVH status, or other mutations) could not be evaluated due to the low number of patients with available data.Statistical analyses were performed with the R-4.1.0 software: Kaplan-Meier estimator for OS, PFS, DOR and median survival times, Cox regression for univariate and multivariate analyses and HR calculation, Log-rank test for comparison of OS, PFS, DOR between risk groups, and Fisher's exact test for the analysis of factors affecting POD24.Results:One hundred and twenty-five patients [Male:Female 91(72.8%):34(27.2%)] with MCL were analyzed. Age at diagnosis was ≤65 years in 60 patients (48%) and >65 years in 65 (52%), stage III-IV in 92.8%, MIPI was intermediate and high-risk in 22.6% and 53.6% respectively, Ki67 was ≥30% in 40 patients (64.5%) and MCL morphology was pleomorphic/blastoid in 40 patients (13.6%).Among 123 patients receiving at least 2 treatment lines (induction and 1 st salvage treatment), 36 (29.2%) received HiDAC and 87 (70.7%) a non-HiDAC induction. Among the 73 patients requiring salvage treatment, 45 (61.6%) were POD<24 and 28 (38.3%) were POD>24.At last follow-up (FU), 50 pts were alive (40%), 7 were lost to FU (5.6%) and 68 had died (54.4%), mainly due to lymphoma-related causes (69.1%). Median OS and PFS were 7.44 and 2.52 years respectively, and median DOR was 34.9 months. There were significant differences on outcome by age group, OS was 10 vs 4.89 years (p=.0021), PFS 4.11 vs 1.64 years (p=.00012) and DOR 57 vs 15.5 months (p=.014) for patients with ≤65 and >65 years, respectively. Regarding other prognostic factors, MIPI did not impact OS, PFS nor DOR. Ki67, blastoid variant and POD24 had impact on OS in the univariate analysis, and only POD<24 maintained its significant effect on OS in the multivariate analysis (p=0.05). Of note, POD24 affected OS in the full cohort (3.86 and 10 years in POD<24 and POD>24, respectively (p=.0001) (Figure 1)) and by age group (3.32 years in POD<24 and 10 years in POD>24 for ≤65 years (p=.00057) and 3.86 in POD<24 and 8.58 years in POD>24 for patients >65 years (p=.0037)).Finally, we analyzed factors potentially affecting POD24, and we found significant differences with the age group and type of induction.In patients >65 years, 75.6% relapsed early (POD<24), while in the younger group only 43.7% were POD<24 (p=.0076). Regarding the type of induction, in the HIDAC group only 33.3% were POD<24, while 68.9% were POD<24 in the non-HiDAC group (p=.017).Conclusions:In our cohort, POD<24 is a strong prognostic factor affecting outcome, with OS of 10 years in POD>24 vs 3.86 years in POD<24. Although younger patients receiving HiDAC-based induction were less likely to be POD<24 and had a significantly better outcome, a subgroup of them still relapse early or do not respond to induction, and for these higher-risk patients, newer and more effective treatment strategies are very much needed. [Display omitted] DisclosuresAbrisqueta: AstraZeneca: Consultancy, Honoraria; BMS: Consultancy, Honoraria; Roche: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Janssen: Consultancy, Honoraria. Bobillo: F. Hoffmann-La Roche Ltd: Consultancy, Speakers Bureau; Gilead: Speakers Bureau. Carpio: Regeneron, TAKEDA, Celgene, Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other. Iacoboni: BMS/Celgene, Gilead, Novartis, Janssen, Roche: Honoraria. Gironella Mesa: BMS, Janssen: Honoraria. Palacio: Bristol Myers Squibb (BMS): Consultancy. Bosch: Roche: Membership on an entity's Board of Directors or advisory committees, Other: Travel; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees, Other: Travel; Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel; TAKEDA: Membership on an entity's Board of Directors or advisory committees, Other: Travel. Marin-Niebla: Kiowa Kirin: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Janssen: Consultancy, Honoraria, Other: travel; Roche: Honoraria; Takeda: Consultancy, Honoraria, Other: travel.

A Phase I Study Of The Oral Btk Inhibitor ONO-4059 In Patients With Relapsed/Refractory and High Risk Chronic Lymphocytic Leukaemia (CLL)

IntroductionThe B-cell receptor (BCR) pathway plays a central role in signal transduction pathways that regulate survival, activation, proliferation and differentiation of B-lineage lymphoid cells. Bruton's tyrosine kinase (Btk) is a critical kinase in BCR signal transduction and recent studies support that targeting Btk is effective in the treatment of B-cell malignancies. ONO-4059 is a highly potent and selective oral Btk inhibitor with an IC50 in the sub-nmol/L range that has demonstrated anti-tumour activity in several pre-clinical models (Yasuhiro T et al, AACR 2013). MethodsThis Phase I study was initiated to determine the safety, tolerability, dose-limiting toxicity (DLT), pharmacokinetics and pharmacodynamics of ONO-4059 given as monotherapy to patients with relapsed/refractory, high risk CLL for whom no therapy of higher priority was available. In this safety-driven, dose-escalating 3+3 design, ONO-4059 was administered as an oral, daily dose (flat dose) given continuously initially for up to 6 months, with the option of additional dosing up to 2 years. We present the safety and efficacy data on 16 evaluable patients, median age 67 yrs (range 40-77), with high risk cytogenetics (17p [38% (6/16)] or 11q [19% (3/16)], 12/16 displaying unmutated IgVH status, 8/16 displaying TP53 mutation), median baseline tumour burden 4,534mm2 [461-14,492 mm2]. Patients had received a median of 3 prior therapies [2-7], including fludarabine (16/16) and rituximab-containing therapy (13/16). Baseline median haematology parameters included hemoglobin 11.3 g/dL [8.5 -15.2], WBC 56.02 x 109/L [2.06-394.74], platelets 95.5 x 109/L [23-173], lymphocytes 50.6 x 109/L [0.5-390], neutrophils 2.46 x 109/L [0.5-11.1]. Patients were treated with ONO-4059 at doses ranging from 20mg-320mg (cohorts 1-5) and the study is currently ongoing with additional dose escalation cohorts to be completed. ResultsONO-4059 was found to be well tolerated, with no dose limiting toxicities (DLTs). A total of 15 ONO-4059-related Grade 1 (n=10) and 2 (n=5) adverse events were reported in 8 out of 16 patients, including one event of diarrhea (G2) and two events of rash (G1, G2). ONO-4059-related G3 or G4 haematological toxicities were infrequent and independent of dose; neutropenia (n=3 [G4 x1, G3 x 2]), febrile neutropenia (n=1 [G3]) occurring in 2 patients at the 20mg dose level only. Only one Grade 3 ONO-4059-related non-haematological event (pyrexia) was reported. Infection rate was low with only one event of bronchitis reported (G2, related). Three ONO-4059-related SAEs were reported in two patients (febrile neutropenia [G3] and pyrexia [G3] in one patient [20 mg]; rash [G2] in another [80 mg]). The pharmacokinetics of ONO-4059 reflects rapid absorption and elimination, a half-life of ∼6 hours, a dose dependent increase in exposure with no accumulation of ONO-4059 exposure and low inter- or intra-patient variability; with Btk occupancy in peripheral blood (as measured by phosphorylated Btk) being maintained for at least 24 hours across all dose levels.Responses have occurred across all dose levels. All patients experienced rapid reductions in lymphadenopathy observed early in treatment, between D1-D28 of the first cycle, accompanied by an increase in absolute lymphocyte count in some, but not all patients. Best overall response rate according to IWCLL criteria (including modified PR with lymphocytosis) was 70% [based on CT-scan and P/E for 7/10 evaluable patients]; with 2 PR, 5 PR with lymphocytosis (For 7 responding patients, median reduction of lymph nodes was 67% [Range 61%-79%]), 2 SD, 1 patient PD (possible Richter's). All patients showed an improvement in haematological parameters after ≥3 months on treatment. Patients (ONO-4059 20mg [1pt] and 40mg [1pt] respectively) who had grade 3 thrombocytopenia at study baseline had a significant improvement in platelet count (>100 x 109/L) with ONO-4059 treatment. To date, 15 of 16 patients remain on treatment with a median progression-free survival of 99 days [Range 8-268].In conclusion, ONO-4059 is a highly potent and selective oral Btk inhibitor with a favourable safety profile along with promising efficacy in this heavily pre-treated population, with responses observed in relapsed, refractory and 17p deleted patients. Disclosures:Salles:Janssen: Honoraria; Gilead: Honoraria; Celgene: Honoraria. Karlin:Janssen: Honoraria; Celgene: Honoraria. Morschhauser:ONO Pharma: Honoraria; Roche: Honoraria; Celgene: Honoraria; Mundipharma: Honoraria. Dyer:ONO Pharma: Honoraria, Research Funding. Hutchinson:ONO Pharma: Research Funding. Fegan:ONO Pharma: Honoraria. Cartron:ONO Pharma: Honoraria. Knurowski:ONO Pharma: Consultancy. Saunders:ONO Pharma: Consultancy; Pharmacyclics: Consultancy. Wright:ONO Pharma: Consultancy. Honda:ONO Pharma: Employment. Mazur:ONO Pharma: Consultancy. Yoshizawa:Ono Pharma: Employment. Kawabata:Ono Pharmaceutical Co., Ltd.: Employment. Birkett:Ono Pharma UK: Employment.

Neutrophil to T-Lymphocyte Ratio (NLR) and Neutrophil to Monocyte Ratio (NMR) Are Promising Biomarkers in Chronic Lymphocytic Leukemia Prognostic Assessment

Development of targeted drugs produced a significant changes in the management of patients with Chronic Lymphocytic Leukemia (CLL). The recent advances in molecular diagnostic techniques has provided a better prediction of disease progression but they are not easily accessible to all Institutions.CLL cells proliferation and survival are sustained by a complex microenvironmental network of cells interactions that are at least partially mediated through B cell receptor signaling.An increased number of studies describe the role of neutrophils and monocytes as markers of the inflammation and antitumor immunity that support tumorigenesis. Recently, authors published the increased NLR is associated to a prognosis in different cancer type.In our study we retrospectively evaluated the prognostic significance of peripheral blood neutrophils, monocytes and non-neoplastic lymphocytes in a unicentric, unselected, CLL cohort of 400 CLL patients at diagnosis (median age 64; stage Binet A 76%,B 18%,C 6%).Using blood count and the flow cytometric analysis reports, absolute neutrophil count (ANC), absolute monocyte count (AMC) and absolute T-lymphocyte count (ALC-CD3+) were evaluated and the neutrophil to T-lymphocyte ratio (NLR), the monocyte to T-lymphocyte ratio (MLR) and the neutrophil to monocyte ratio (NMR) were calculated at diagnosis, follow up and at relapse. Clinical and biological data from all patients were also retrieved to make a better comparison with the major clinical and prognostic markers commonly used.Compared to normal controls, CLL patients showed an increase AMC value (788±65 vs 469±51 cells/μL, p=0.005) and a slightly increased in the ALC-CD3+ value (2514±63 vs 2189±45 cells/μL, p=0.04). No difference in the ANC. The median NLR ratio was higher in CLL patients compared to healthy individuals (2.32 vs 1.7, p=0.02) and appeared more increased in early stage (2.65, 2.14, 1.95 mean value in Binet A, B, C respectively, p=0.0015). NMR median value was also significantly higher in early stage compared to advanced stage (8.0, 6.7, 5.1 in Binet A, B, C, p=0.0008).Conversely, MLR showed the opposite trend resulting more elevated in advanced stage at diagnosis, although with a low statistical significance.We did not found any correlation between these ratio and the IGVH status or genetic abnormalities.NLR and NMR higher level are associated with the absence of serum prognostic markers, such as CD38, and CD49d (p=0.002).160 out of 400 patients of our CLL cohort received a treatment during the course of disease. In our analysis, NLR and NMR are significantly higher in untreated patients than treated ones (median NLR 2.42 vs 1.95, p=0.0021; NMR 8.0 vs 7.0, p=0.0002). Among treated patients NLR and NMR significantly correlate with the time to treatment initiation (TTFT): lower NLR and NMR ratio correspond to a shorter TTFT (NLR p=0.03; NMR p=0.0035). For MLR the trend is opposite.A subgroup analysis conducted only on Rai 0/Binet A patients (n 219, 55%) showed that NLR and specially NMR are significantly associated with the risk of progression: lower NLR and NMR ratio correspond to an increased risk to symptomatic progression (p=0.0017), strengthening the hypothesis that high ratios are associated with a more indolent form of disease.In the 0/A population, analysis showed that a higher AMC and at a lesser extend a high ALC-CD3+ correlate with the progression rate and treatment initiation (Kaplan-Meier survival curve, p= 0.0001).Univariate analysis suggested that NLR ≥ 3.0, NMR ≥ 8.0 and AMC <900/mmc predicted a good prognosis in CLL patients. However, only NMR and AMC appeared as independent prognostic factors in multivariable analysis in Rai 0/Binet A patients.Our in vitro experiments supported these finding confirming: the non “immunosuppressive” role for neutrophils in CLL and the increased ability for monocytes to suppress T-lymphocytes activation and expand T-regulatory cell.In our study NLR ratio showed a positive prognostic role in CLL. A strongest protective meaning is displayed by NMR value. These combined value and AMC count provide more prognostic information at diagnosis and are associated with clinical outcome in CLL patients.These simple, cheap and reliable markers could be easily integrated into clinical practice reflecting both inflammatory status, immune response and microenviroment that contribute to disease progression. DisclosuresNo relevant conflicts of interest to declare.

Mantle Cell Lymphoma Mimicking Mucosa-Associated Lymphoid Tissue (MALT) Lymphomas: A Pathological Characterization (on behalf of the “Fondazione Italiana Linfomi (FIL) - Postgraduate Master Course”)

INTRODUCTION: Mantle Cell Lymphoma (MCL) is considered an aggressive B-cell lymphoma, but with a heterogeneous outcome and a median overall survival ranging from 3 to 5 years (yrs). Two different subsets are recognized, known as “classical” and “leukemic non-nodal” subtypes. Unfrequently MCL presents with isolated extranodal localization mimicking mucosa-associated lymphoid tissue (MALT) lymphomas; a large series of European Mantle Cell Network has been recently reported (Morello et al, ICML 2017). On the other side, pathological features of this peculiar presentation are not yet clarified.METHODS: We collected data on 34 patients (pts) with extranodal mantle cell lymphoma (EMCL) of MALT site, diagnosed between 2001 and 2016 in 10 Italian Centers of Fondazione Italiana Linfomi (FIL) network. The diagnosis of MCL was confirmed through detection of CyclinD1 overexpression and/or t(11;14). All pts were >18 yrs and HIV negative. Only minimal locoregional nodal involvement, with CT longest axis <2 cm, and bone marrow involvement with only extranodal disease were allowed. Pts with splenomegaly and leukemic disease were excluded. We performed a central pathological review of 21/34 cases (62%) in the Section of Anatomic Pathology of University of Pavia and compared their features with a cohort of nodal MCL.RESULTS: Clinical features and treatments are summarised in Table 1.Biopsied extranodal sites were as follows: 11 pts (32%) Waldeyer's ring, 11 (32%) gastrointestinal tract, 3 (9%) ocular adnexa, 3 (9%) oral cavity, 3 (9%) salivary glands, 2 (6%) thyroid and 1 (3%) liver. Detailed morphology was assessed in 28 pts: classical in 22 pts (78%), pleomorphic in 3 (11%) and blastoid in 3 (11%). CD5 expression was performed in 31 cases: 21 (68%) positive, 6 (19%) partially positive and 4 (13%) negative. Median Mib1/Ki67 was low (15%, range: 2-70%), <30% in 83% and ≥30% in 17% cases. CyclinD1 was available in 33 pts (88% positive, 3% negative, 9% partially positive) and t(11:14) in 13 pts (92% positive). SOX11 staining was performed in 21 cases: 17 (81%) resulted positive, 3 (14%) partially positive and only 1 (5%) negative. Overall, no statistically significant difference with nodal MCL series was observed, particularly regarding cytological features, CD5 and SOX11 expression and Mib1/Ki67. MIPI score was calculated in 33 pts: 15 (46%) low, 13 (39%) intermediate and 5 (15%) high; biologic-MIPI score was available in 29 pts: 7 (24%) low, 14 (48%) intermediate and 8 (28%) high.After a median follow-up of 5.1 yrs (0.5-15), overall survival (OS) was 93% at 5 yrs and 75% at 10 yrs with 4 deaths (2 for lymphoma, 2 for unknown cause). All the 12 pts treated with autologous stem cell transplantation was still alive at last follow-up. Progression-free survival (PFS) was 74% after 5 yrs and 64% after 10 yrs of follow-up. Univariate analysis results are overlapping with European series: pts with age <65 yrs (p=0.002), ECOG 0 (p=0.012), low MIPI score (p=0.005) and low biologic MIPI score (p=0.008) had a longer OS. In univariate analysis, a longer PFS was observed in pts with age <65 yrs (p=0.016), ECOG 0 (p=0.002), low MIPI score (p<0.001) and low biologic MIPI score (p=0.011). In multivariate analysis, age (p=0.022) confirmed its impact on PFS, after adjusting for biologic MIPI score.CONCLUSIONS: According to our preliminary data, we define EMCL as a subset with preferential involvement of Waldeyer's ring and gastrointestinal tract, mimicking the clinic-pathologic features of marginal zone lymphoma of extranodal sites, from which it should be differentiated via a thorough histologic characterization, and featuring the indolent course of the indolent variant of MCL. On the other hand, our cases harbored the morphologic and phenotypic features of the nodal form of MCL. On the basis, the true nature of this putative MCL subset should be deepened upon biologic characterization, including mutational and IGVH status analysis. [Display omitted] DisclosuresLoseto:gilead: Membership on an entity's Board of Directors or advisory committees; janssen: Consultancy; italfarmaco: Membership on an entity's Board of Directors or advisory committees. Arcaini:Pfizer: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sandoz: Consultancy; Gilead: Research Funding. Zaja:Novartis: Honoraria, Research Funding; Roche: Honoraria, Research Funding; Abbvie: Honoraria; Takeda: Honoraria; Gilead: Honoraria; Janssem: Honoraria; Celgene: Honoraria, Research Funding.

Outcome Prediction in CLL Patients Based on a Disease-Specific Oligonucleotide Microarray.

Chronic Lymphocytic Leukemia (CLL) is characterized by a extreme variability in the clinical course; some patients survive only a few months whereas others have stable disease for more than 10 years. Individual outcome prediction in CLL is still a challenge, in spite of the existence of multiple newly described prognostic factors: cytogenetic alterations, mutational IgVH status, and expression of surrogate markers such as ZAP-70. The accuracy of outcome prediction in CLL may improve thanks to the inclusion of new additional biological variables identified by high-throughput molecular techniques. This more reliable risk-stratification of patients could allow to assign a risk-adapted individualized therapy. With this aim, we have analysed with oligonucleotide microarrays three different series of CLL patients in a three-step process.In a first-step, RNA extracted from tumour samples of a series of 26 CLL patients, was hybridised using commercial 22K oligonucleotide chips (Agilent Technologies). A total of 135 genes whose expression was associated with changes in survival probability were identified. An additional review of published data yielded a total number of 497 genes whose expression has been related with outcome in CLL. A CLL specific chip was then built in a second-step including, besides these 632 genes, control genes until a total of 1023 genes. This comprehensive CLL-chip was then hybridised with RNA extracted from peripheral-blood samples in a new series of consecutively diagnosed 100 CLL patients. Cox's univariate analysis was performed to look for molecular variables associated with Progression Free Survival (PFS) as endpoint. FDR significance levels were also used when appropriate, for adjusting the associated p-value. Expression of 94 genes was significantly associated with PFS. Profile patterns of the 94 resulting genes were clustered, and allow to identify 12 clusters. Based on biological function, we selected a representing gene of each cluster, to perform a multivariate Cox's analysis. This approach generated a model of 4 genes, implicated in BCR signalling, signal transduction and transcription regulation, that distinguishes four quartiles of CLL patients with PFS at 5 years of 95%, 70%, 44% and 8% respectively.These results have been validated in a new independent series of 60 consecutively diagnosed cases of CLL. If confirmed in additional studies, the availability of this CLL-specific chip could allow a more accurate prediction of the clinical course in CLL patients, this giving support to a stratification of patients into different risk-groups

Smudge cells percentage on blood smear is a reliable prognostic marker in chronic lymphocytic leukemia

ObjectiveWe evaluated the relevance of using the smudge cell percentage in the blood smear as a prognostic marker in CLL. MethodsIn this prospective study, 42 untreated Senegalese patients with CLL were enrolled. The diagnosis was established, based on the peripheral blood count and flow cytometry using the Matutes score. Cytogenetic aberrations, assessed by fluorescence in situ hybridization (FISH), were available for 30 patients, while the immunoglobulin heavy chain genes (IGVH) mutation status was performed by next-generation sequencing (NGS) in 24 patients. The SC percentage was determined in the blood smear, as previously described. Statistical analyses were executed using the GraphPad Prism 8. ResultsThe mean age was 63 years (48 - 85) and the male: female sex ratio was 4.66. A low SC (< 30%) percentage was correlated with Binet stage B/C (p = 0.0009), CD38 expression (p = 0.039), unmutated IGVH status (p = 0.0009) and presence of cytogenetic abnormalities (for del 13q, p = 0.0012, while for other cytogenetic aberrations, p = 0.016). An inverse correlation was found between the SC percentage and the absolute lymphocyte count (r = -0.51) and patients with higher percentage of SCs had a prolonged survival. However, there was no correlation between the SC percentage and age (p = 0.41) or gender (median, 19% for males vs. 20% for females; p = 0.76). ConclusionWhen less than 30%, the SC was associated with a poor prognosis in CLL. Easy and affordable, the percentage of SCs in a blood smear could be a reliable prognostic marker, accessible to all CLL patients, mainly those in developing countries.

Leucémie lymphoïde chronique : de la biologie à la thérapeutique

AbstractTreatment stategies for Chronic Lymphocytic Leukaemia drastically evolved since fifteen years. The increased knowledge of genetic and molecular anomalies resulted in a surge of new therapeutic targets. Chromosomal abnormalities (such deletion 17p with TP53 gene mutation) directs to poor pronostic and specific care. Molecular screening for IGVH status is now recommended before treatment. All influence clinical management and therapeutic guidelines balanced between conventional immunochemotherapy and targeted therapy. This review summarizes new insights for CLL care.

CLL-132: Mutational Status of IGVH in Correlation with Genetic Abnormalities of Patients with Chronic Lymphocytic Leukemia in Macedonia - A Single-Centre Experience

Context B-cell chronic lymphocytic leukemia (CLL) is a clinically heterogeneous disease. One of the most important molecular predictors, the immunoglobulin VH gene mutational status, divides CLL into two prognostic groups, where unmutated U-CLL is associated with remarkably worse prognosis than mutated M-CLL. In many studies, multiple recurring chromosomal aberrations appear to be effective prognostic markers in CLL. Objective The aim of the study was first-time evaluation of rearrangement of IG genes and genetic abnormalities in Macedonian CLL patients. Design Prospective study of Macedonian patients with CLL at early and advanced stage of disease in a period of time from January 2011 to January 2019. The median follow-up was 48 months (1-96 months). Setting Mutational status and configuration of IGHV-IGHD-IGHJ rearrangements in 90 treatment-naive CLL patients were analyzed using reverse transcriptase-polymerase chain reaction (RT-PCR) and sequencing methodology. We used multiplex ligation-dependent probe amplification (MLPA) for detection of most common genetic abnormalities. Interventions We introduced prognostic markers like cytogenetic aberrations, and mutational status of immunoglobulin heavy chain in prognostic stratification of Macedonian patients with CLL. Results Our evaluation has shown that 54.7% patients belonged to the U-CLL subset, whereas 45.3% belonged to the M-CLL subset. The most frequently expressed IGHV subgroup was IGHV3 (40%), followed by IGHV4 (29%), IGHV1(27%), and IGHV5 (3%). The most frequent subgroup with 8% were IGHV1-69*13 and IGHV4-34*0. In the IGHD and IGHJ sets, most frequently expressed genes were IGHD3 (59.7%) and IGHJ6(53.1%), respectively. We found that 80% of IGHV1, 65.6% of IGHD3, and 83.3% of IGHJ6 genes were U-CLL (p=0.0005). Frequency of genetic aberrations by MLPA were 13q14 deletion (20%) followed by 11q deletion (14%), then trisomy 12 (13%) and 17p deletion (6.6%), multiple mutations (6.7%), and NOTCH mutation (5.3%). Conclusions Our evaluation of mutational status on IGVH, IGDH, and IGJH genes in Macedonian CLL patients resulted with data that are consubstantial to those from the Mediterranean area. Novel mutations discovered in this study must be validated through large cohort studies and may offer clues to the mechanisms underlying the difference in CLL survival. In the future, therapeutic strategies targeting these genes should be evaluated and considered.

Voruciclib plus venetoclax show high efficacy for CLL b cells on human stromal cells.

e20009 Background: Although treating B-Chronic Lymphocytic Leukemia (CLL) with small molecule inhibitors has shown promise, a lasting cure for this disease has yet to be found. Further treatments directed at key molecular targets in the CLL B cell need development for patients with adverse prognostic factors, and relapsed/refractory patients. We evaluated the impact of two novel drugs with non-overlapping mechanisms of action on CLL B cells: Voruciclib, a cyclin dependent kinase (CDK) inhibitor targeting CDKs 9, 6, 4, 1 and Venetoclax a BCL-2 inhibitor. To mimic the protective effect of the CLL micro-environment these experiments were done with CLL cells cocultured in the presence of bone marrow mesenchymal stromal cells (BMSC). Methods: Voruciclib, (MEI Pharma) and Venetoclax were tested for killing activity alone and in combination against CLL cells cocultured with healthy human bone marrow MSC. CLL B cells were from untreated patients stratified on risk of disease progression. Low risk had mutated and high risk patients had unmutated IVGH genetic status. CLL cells were cultured on BMSC’s with a series of escalating doses of individual drugs and drug combinations at fixed molar ratios then Annexin/PI stained for viability testing by flow cytometry. Killing curves generated for each drug/combination were analyzed by the combination index (C.I.) approach of Chou and Talalay with Calcusyn, characterizing interactions as synergistic, additive or inhibitory. C.I. value hierarchy classes synergy as moderate (0.7-0.9), synergistic (0.3-0.7), strong (0.1-0.3) and very strong (below 0.1). Results: CLL cells with unmutated IGVH status were more sensitive to Voruciclib than mutated IGVH both with and without BMSC’s. The combination Voruciclib +Venetoclax showed synergism for all patients regardless of risk; half strongly to very strongly (Table). Similar synergistic effects were seen with relapsed/refractory patients. Conclusions: Based on these preclinical data, Voruciclib + Venetoclax is a very promising combination to improve treatment of CLL patients. We speculate that CDK9 inhibition, which regulates transcription of Mcl-1, with Bcl-2 inhibition results in potent apoptosis induction in CLL B cells. [Table: see text]

Final Results from the Multicenter, Open-Label, Phase II GIBB Study of Obinutuzumab+Bendamustine in Previously Untreated Patients with Chronic Lymphocytic Leukemia

Final Results from the Multicenter, Open-Label, Phase II GIBB Study of Obinutuzumab+Bendamustine in Previously Untreated Patients with Chronic Lymphocytic Leukemia

Molecular Genetics and Prognosis in Younger Patients with Chronic Lymphocytic Leukemia (CLL)

Background: While CLL is commonly diagnosed in older patients, there are ~15% of patients diagnosed at ages younger than 50. Several past studies have investigated differences in clinical parameters and treatment outcomes in younger patients with CLL, including a shorter time to first treatment (TTFT) among younger patients (Parikh et al. 2014). However, few studies have reported on the genetic mutational differences between younger and older cohorts. To bridge this gap, we investigated the mutational landscape between younger and older patients and evaluated the clinical outcome TTFT, hypothesizing that our younger cohort of patients would associate with higher risk lesions and behave more aggressively. Methods: We conducted a single center retrospective database review of 557 patients diagnosed with CLL from 1980 to 2019 who underwent whole exome profiling between 2015 to 2019 with a lymphoid specific 75-gene next generation sequencing (NGS) panel (Genoptix Inc). A Pearson's chi-square test was used to compare categorical variables between groups and a Wilcoxon rank sum test was used to compare medians. The TTFT was estimated using Kaplan-Meier methods, and the difference between groups was compared using the log-rank test. Multivariate regression using a Cox proportional hazards model was used to compare TTFT between groups independent of well-accepted clinical risk factors for treatment initiation. Results: Of the 557 patients who underwent NGS testing, 92 (16.5%) were younger than 50 years old with a median age of 44.9 years old compared to a median age of 62.7 years old in the 465 (83.5%) patients older than 50 years old. There was no difference between the two groups with regards to previous treatment prior to NGS testing with 29.2% in the older patients and 26.1% in the younger patients having previously been treated (p=0.63). The median time from CLL diagnosis to initial NGS testing was 5.2 years in the younger cohort vs. 3.2 years in the older cohort (p=0.04). There were no differences in baseline prognostic factors between younger and older patients, including Rai stage, IGVH status, CD38 positivity, ZAP70 expression, and cytogenetic abnormalities. We found a lower frequency of TP53 mutations in younger compared to older patients (6.5% vs 15.7%, p=0.03) but otherwise found no differences in any other genetic mutations between the two groups, including NOTCH1, FAT1, ATM, and SF3B1 (Table 1). There was a longer TTFT in younger patients with a median TTFT of 7.61 years compared to 4.42 years in older patients (p=&lt;0.001) (Figure 1). The difference in TTFT was independent of TP53 mutation status in a multivariate analysis. Conclusions: We found no enrichment of specific genetic mutations in younger versus older patients except a lower prevalence of TP53 mutations in our younger patient population. We found younger patients have a longer TTFT, which is contrary to previous studies that have identified younger age as a negative prognostic marker. Future research is needed to determine why younger patients in our cohort appear to have a more indolent disease course in terms of TTFT despite similar baseline prognostic factors and molecular genetics to older patients. Disclosures Furman: Acerta Pharma: Consultancy; Genentech: Consultancy; Incyte: Consultancy; AstraZeneca: Consultancy; Abbvie: Consultancy; Beigene: Consultancy; TG Therapeutics: Consultancy; Sunesis: Consultancy; Oncotracker: Consultancy; Janssen: Consultancy; Pharmacyclics: Consultancy; Verastem: Consultancy. Allan:AbbVie, Inc: Consultancy, Membership on an entity's Board of Directors or advisory committees; Acerta Pharma: Consultancy; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics LLC, an AbbVie company: Consultancy; Bayer: Consultancy; Sunesis Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Honoraria; Verastem Oncology, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees.

Multicentre Genome Wide Association Study Identifies Risk Alleles for Progressive Chronic Lymphocytic Leukaemia

The increased incidence of chronic lymphocytic leukemia (CLL) in first-degree relatives of affected patients indicates an element of genetic susceptibility to this malignancy, borne out in large scale genome-wide association studies (GWAS), which have identified over 40 constitutional risk alleles. Given the important genetic contribution to CLL susceptibility we hypothesized that constitutional genetic variants also affect disease progression. We employed GWAS methods in a large United Kingdom multi-center cohort study of well-characterized predominantly early-stage CLL cases to identify risk alleles for progressive CLL. We conducted six GWAS for single nucleotide polymorphisms (SNPs) associating with progressive CLL incorporating a total of 774 cases of European ancestry recruited to 6 clinical centers across the United Kingdom. CLL cases were genotyped on the Illumina OmniExpress platform and genotypes were determined using Illumina GenomeStudio software. After imputation, we combined the association test statistic for 5,199,911 autosomal SNPs common to all 6 GWAS after exclusion of those with an imputation quality score of &lt;0.9 and a minor allele frequency (MAF) of &lt; 2.5%, and conducted a meta-analysis under a fixed-effect model. The primary outcome assessed was time to first treatment (TTFT), defined as the interval between CLL diagnosis and first treatment or last follow-up. Pooling data from the 6 GWAS identified 5 SNPs at two genomic locations that surpassed genome-wide significance (P ≤ 5 x 10-8) for association with TTFT. The strongest statistical evidence for an association with progressive disease was for rs736456 (hazard ratio (HR) = 1.76, 95% confidence interval (CI) = 1.45-2.14; P = 1.26 x 10-8), which maps to chromosome 10q26.13. The second strongest association with progressive disease was for rs3778076 (HR = 2.03, 95% CI = 1.58-2.62; P = 3.89 x 10-8) which maps to chromosome 6p. Both markers showed consistent direction and magnitude of effect sizes across all six GWAS with no evidence of heterogeneity, and retained prognostic significance in multivariate models for disease progression, particularly in models restricted to Binet A patients. Whilst not as powerful as IGVH status, rs736456 and rs3778076 had prognostic utility equivalent to CD38 status, and are particularly powerful when considered together, identifying 5% of CLL patients carrying 2 or more risk alleles at high risk of progressive disease (Figure). To identify cis-regulated genes at each locus associated with progressive disease we interrogated gene expression data derived from a meta-analysis of 31,624 blood samples collated by the eQTLGen consortium. Of the thirteen genes annotated to within 1Mb of the chromosome 10 association signal rs736456 is eQTL for the PLEKHA1 gene (TAPP-1)(Benjamini-Hochberg corrected P-value [PBH] = 1.29 x 10-15). Of the 27 genes annotated to within 1Mb of the chromosome 6 signal, rs3778076 is eQTL for 5 genes including UHRF1BP1 (PBH = 6.73 x 10-139) and C6ORF106 (PBH = 3.54 x 10-64). Annotated genes at both loci have been implicated in modulating B-cell receptor or innate immune responses, key pathways in CLL pathogenesis. Data on post-treatment survival were available on 390 CLL cases, with 231 deaths and 159 censored at last follow-up, and neither the lead SNP at chromosome 10 or chromosome 6 were significantly associated with post-treatment survival in patients primarily treated with regimens that included chlorambucil, fludarabine or cyclophosphamide. It will be important to determine whether these markers predict overall survival in patients treated with novel targeted therapies. Taken together, these data identify rs736456 and rs3778076 as prognostic in early stage CLL patients demonstrating that progression of CLL from asymptomatic to symptomatic disease is determined by constitutional genetic variation as well as the more established somatic drivers. Constitutional genetic markers have the advantage of being easy to perform, highly reproducible and inexpensive making them ideal for incorporation into multivariate prognostication models for early stage CLL. Figure Disclosures Fegan: Roche: Honoraria; Janssen: Honoraria; Gilead: Honoraria; Abbvie: Consultancy, Other: Conference attendance sponsorship. Forconi:Gilead Sciences: Research Funding; Janssen-Cilag: Consultancy, Honoraria, Other: Travel, Accommodations, Expenses, Speakers Bureau; Novartis: Honoraria; Menarini: Consultancy; Abbvie: Consultancy, Honoraria, Other: Travel, Accommodations, Expenses, Speakers Bureau; Roche: Honoraria. Schuh:AbbVie: Consultancy, Speakers Bureau; Genentech: Consultancy, Speakers Bureau; Pharmacyclics: Consultancy, Speakers Bureau; Verastem: Speakers Bureau; Gilead: Speakers Bureau; Seattle Genetics: Speakers Bureau; Jazz Pharmaceuticals: Speakers Bureau; Bristol-Myers Squibb: Research Funding; Janssen: Speakers Bureau; Kite: Speakers Bureau. Hillmen:Pharmacyclics LLC, an AbbVie Company: Consultancy, Honoraria, Research Funding; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Expenses, Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Expenses, Research Funding; Roche: Research Funding; Gilead: Research Funding; Apellis: Research Funding; Acerta: Membership on an entity's Board of Directors or advisory committees. Pratt:Binding Site, Amgen, Takeda, Janssen, Gilead: Consultancy, Honoraria, Other: Travel support.

Next-Generation Optical Mapping Reveals Numerous Previously Unrecognizable Structural Variants in Chronic Lymphocytic Leukemia

Introduction: Chronic lymphocytic leukemia (CLL) is the most common form of leukemia in adults in western countries. Although a number of genetic aberrations (complex karyotype, del(17p), del(11q), unmutated IGVH status, TP53 mutations) having predictive and prognostic significance in CLL was reported, there is still a lack of information about large structural variants, repetitive regions, copy number variations, and inversions in the CLL genome due to limitations of current methods (NGS, cytogenetics). Aim: Using novel next-generation optical mapping technology, we analyzed leukemic cells from peripheral blood of CLL patients at different stages of the disease. Methods and Patients: High-molecular weight DNA was isolated from peripheral blood (&gt;90% CD5+/CD19+/CD20+/CD23+ B-cells) from two CLL patients (P1: 70-y-old male, diagnosis of CLL in 2018, Binet Stage B, unmutated IGVH, untreated; P2: 59-y-old male, diagnosis of CLL in 2005, Binet Stage A, unmutated IGVH, on ibrutinib (2nd line) since 4/2018). For optical mapping, DNA was labeled by DLS chemistry and visualized on Saphyr system (BioNano Genomics). The optical genome maps (480 Gbp, coverage 124X) were aligned to a human reference genome (GRCh38) and the detected structural variants (SVs) were compared against the database of healthy controls; only rare de novo variants were analyzed. In both patients, cytogenetic analysis (karyotype, FISH, and arrayCGH) and mutational analysis by NGS (TP53, ATM, BIRC3, NOTCH1, SF3B1, POT1 genes) were available. Results: For both CLL patients, the optical mapping method identified 97% of chromosomal abnormalities previously reported by standard cytogenetic methods such as deletions on 6q, 13q, 11q, 17p and trisomy 12. In both patients, same large deletions of 17.5 kbp in 9q21 and 7.1 kbp in 14q21 chromosome region were detected, undetectable by standard methods. Both patients also carried intra-chromosomal translocation t(13;13) located in 13q14 region. Additionally, P1 genome carried 2 kbp deletion in oncogene EGFR and 500 bp insertion affecting tumor suppressor TCF4, both genes previously associated with CLL. Moreover, numerous additional copy number variations and large somatic SVs, including on average tens of deletions, insertions, inversions, duplications, and translocations were detected in both patients. Conclusion: Our study revealed numerous novel genomic rearrangements in CLL genome, unrecognizable by standard methods. These data highlights a high potential of optical maps for refinement of genomic variability in CLL, mainly for detection of large structural variations and copy number variants. The study on larger patient cohort and longer follow-up of patients may identify genetic aberrations associated with the clinical course and therapy response. Support: MZ ČR VES16-32339A, MZ ČR - RVO (FNOl, 00098892), IGA UP_2019_014, IGA UP_2019_01 Disclosures Minarik: Celgene: Consultancy, Honoraria, Research Funding; Amgen, BMS, Janssen-Cilag, Takeda: Consultancy, Honoraria.

Rapid Dose Escalation of Venetoclax in Patients with Relapsed/Refractory Chronic Lymphocytic Leukemia Previously Treated with B-Cell Receptor Inhibitor Therapy

Rapid Dose Escalation of Venetoclax in Patients with Relapsed/Refractory Chronic Lymphocytic Leukemia Previously Treated with B-Cell Receptor Inhibitor Therapy

PS1152 THE USE OF THE BCL‐2 INHIBITOR IN CLL PATIENTS WHO PROGRESSED AFTER B‐CELL‐RECEPTOR INHIBITORS: A RETROSPECTIVE MULTICENTER ITALIAN EXPERIENCE

Background:B‐cell receptor signaling inhibitors (BCRi), such as Ibrutinib (Ibr) and Idelalisib (Idela), have changed the treatment paradigm for chronic lymphocytic leukemia (CLL), and the BCL‐2 inhibitor Venetoclax (Ven) has shown promising efficacy across clinical studies in relapsed/refractory (R/R) CLL. To date, the exact timing on the use of new pathways inhibitors in the real life practice is not well established.Aims:In this multicenter retrospective real life study we evaluated the overall response rate (ORR), progression‐free survival (PFS), overall survival (OS) of CLL patients (pts) treated with Ven after one or two BCRi. Also the reasons for BCRi discontinuation ‐ adverse events (AE) vs progressive disease (PD) ‐ were evaluated.Methods:We report a multi‐center retrospective analysis that included 76 CLL pts from 18 different Italian centers treated with Ibr and/or Idela prior to Ven for progressive naïve CLL or with relapsed or refractory disease. Twenty‐four pts were treated with one BCRi, while 52 pts with both BCRi before Ven. Median age was 63 years (26–87); 78% pts had an unmutated IgVH status, 33% pts had 17p deletion and 32% pts had TP53 mutation. The sequence of the BCRi prior to Ven was the following: Ibr, Idela, Ven in 16 pts; Idela, Ibr, Ven in 8 pts; Ibr, Ven in 37 pts; Idela, Ven in 15 pts.Results:No statistically significant difference was found in terms of ORR to Ven in patients who had been exposed to one or two BCRi (p = 0.114). At the same time, we observed that pts who received only one BCRi prior to Ven showed a significantly better PFS and OS at 12 months: 76% vs. 35% (p = 0.011) and 88% vs. 57% (p = 0.015), respectively. BCRi treatment was stopped in 70% of pts because of PD and in 30% due to AE. When we considered the impact of the reasons of the first BCRi discontinuation ‐ AE vs PD ‐ on Ven response, we found that the ORR was significantly better in pts who stopped due to AE: 91% vs 49% (p = 0.03). PFS and OS were also significantly better in pts who discontinued treatment due to AE compared to PD, with a PFS of 84% vs 45% (p = 0.003) and an OS of 93% vs 62% (p = 0.028). When we considered the impact of discontinuation of both BCRi on Ven response, the PFS was similar in CLL patients who stopped BCRi due to AE (p = 0.31), while pts who experienced PD to both BCRi showed a worse PFS during Ven (p = 0.06).Summary/Conclusion:The number of BCRi received before BCL2i does not affect ORR to Ven. Single BCRi is better than double BCRi in terms of PFS and OS following Ven treatment, regardless of the BCRi used. Discontinuation of the first BCRi because of AE is associated with a better PFS and OS to Ven compared to pts who experienced PD. The discontinuation of double BCRi for AE did not impact on PFS to Ven. On the other hand, pts who stopped both BCRi for PD showed a tendency to a worse PFS on Ven, without reaching significance. Finally, these preliminary data seem to suggest the use of double BCRi before Ven for pts who discontinued due to AE; on the contrary, we suggest the use of Ven after single BCRi when the reason of discontinuation is disease progression.image

PB1901 IBRUTINIB, SINGLE AGENT BTK INHIBITOR, FOR RELAPSED/REFRACTORY (R/R) CHRONIC LYMPHOCYTIC LEUKEMIA: A REAL‐WORLD EXPERIENCE FROM RETE EMATOLOGICA PUGLIESE (REP).

Background:Ibrutinib has showed to improve Progression Free Survival (PFS) and Overall Survival (OS) in patients with Chronic Lymphocytic Leukemia (CLL) maintaining a good tolerability profile. Moreover a good tolerability profile was showed in previously published data.Aims:Within the Rete Ematologica Pugliese (REP) seven hematologic centers verified the safety and the efficacy of Ibrutinib as a single agent treatment. Response rate, PFS and OS have been evaluated in all treated patients and in poor cytogenetic risk groups.Methods:One hundred twenty‐two patients from seven hematologic centers of REP were enrolled in the study. Ibrutinib was administred as salvage therapy across different lines as reported: 2nd line n = 75 (61%) (BR n = 48, FCR n = 24, other n = 3); 3rd line n = 35 (28,7%); 4th line n = 12 (8,2%). All patients received Ibrutinib 420 mg once daily until progression or unacceptable toxicity. PFS, OS and Overall Response Rate (ORR) were calculated. Kaplan Meier method was used to estimate PFS/OS and differences in survival times between groups were assessed with a log‐rank test.Results:Patients have a median age of 65 years (range: 28‐84), 50% of them had more than 65 yrs. Cytogenetic FISH was performed in 66% of patients (n = 80). No abnormalities were found in 31% of patients (n = 25): while del17p, del13q, del11q and trisomy12 were found in n = 33, n = 13, n = 3 and n = 6 of patients, respectively. Unmutated and mutated IgVH status was performed in 73 patients (60%); of these, 46(63%) patients had unmutated and 27 (37%) had mutated IgVH.Overall, ORR was achieved in 94 patients (77%) (complete response [CR] was reached in 10%) of patients). In the subgroup having a del17p ORR was achieved in 21 (61%) patients. Median of follow up time is 17 months (range 1‐45). The estimated PFS at 30 months was 66,7% from the entire study population. Patients with del17p displayed a median PFS of 39 months while it was not reached in patients having no history of cytogenetic abnormalities (Figure 2). Median OS was not reached from all treated patients and the estimated OS at 30 months was 72.7%, while no difference in terms of OS was found when patients with and without del17p were compared (81.7% vs 90.2%), (overall logrank p = 0.807) (Figure 4).Out of all treated patients, 47% of them showed the following adverse events (AEs) with grade ≥3: neutropenia (16%), hypertension (12%), atrial fibrillation (3%), anemia (3%), diarrhea (2%).Summary/Conclusion:In REP real‐world experience, treatment for R/R CLL patients with Ibrutinib as a single agent demonstred to be effective across negative and positive cytogenetic groups and to be well tolerated. AES did not result in treatment interruptions in the most of cases. In fact, over a median follow up of 17 months, 80% of patients remain on Ibrutinib‐treatment. Clearly, these results will require longer follow up time to be further confirmed.image

Limited value of routine follow-up visits in chronic lymphocytic leukemia managed initially by watch and wait: A North Denmark population-based study.

IntroductionThe majority of newly diagnosed chronic lymphocytic leukemia (CLL) patients are followed initially by watch and wait (WAW). Clinical practice varies and the value of frequent follow-up visits remains unclear. Thus, in this study we investigated the clinical value of follow-up visits for patients with CLL.MethodsWe collected data from diagnosis and follow-up visits for patients diagnosed with CLL and managed by WAW in the North Denmark Region between 2007–2014. High- and low-risk group patients were determined by Binet stage, IgVH status, and cytogenetics at diagnosis. The effect of risk group allocation on the probability of receiving CLL-directed treatment within two years was included in a multivariable logistic regression model adjusted for age and blood test results.Results273 patients were included in the study with a median follow-up of 3 years (IQR: 1.6–5.4). Overall, the median interval between follow-up visits was 98 days (95% CI: 96–100) (high-risk patients: 91 days [95% CI: 86–95] vs. low-risk patients: 105 days [95% CI: 100–110]). Among 2,312 follow-up visits, only 387 (17%) were associated with interventions. At the following time points: 6 months, 1 year, and 1.5 years, patients with low-risk CLL had significantly lower odds of initiating treatment compared to patients with high-risk CLL.ConclusionWAW plays an important role in managing CLL. Interventions at follow-up visits were infrequent and low-risk patients had significantly lower risk of treatment initiation. We question the value of routine follow-up in CLL in the absence of changes in symptoms and/or blood test results.

The regulatory capacity of B cells directs the aggressiveness of CLL

ABSTRACTChronic lymphocytic leukemia (CLL) is associated with abnormal T-cell responses responsible for defective anti-tumor activities. Intriguingly, CLL B cells share phenotypical characteristics with regulatory B (Breg) cells suggesting that they might negatively control the T-cell activation and immune responses. We elaborated an in vitro co-culture system with T cells to evaluate the Breg capacities of CLL B cells following innate Toll-like receptor 9 (TLR9) engagement. We demonstrated that B cells from half of the patients exhibited regulatory capacities, whilst B cells from the remaining patients were unable to develop a Breg function. The T cell sensitivities of all patients were normal suggesting that defective Breg activities were due to intrinsic CLL B cell deficiencies. Thus, TLR-dedicated gene assays highlighted differential signature of the TLR9 negative regulation pathway between the two groups of patients. Furthermore, correlations of the doubling time of lymphocytosis, the time to first treatment, the mutational status of IgVH and the Breg functions indicate that patients with efficient Breg activities have more aggressive CLL than patients with defective Breg cells. Our in vitro observations may open new approaches for adjusting therapeutic strategies targeting the Breg along with the evolution of the disease.

Real Life Dates of Chronic Lymphocytic Leukemia (CLL) in Germany. Interim Analysis of 234 Consecutively New Diagnosed Patients in a German Epidemiological-Clinical Register

Background:Therapy studies yield important insights into clinical features and therapeutic options of CLL. However studies in Germany represent less then 20% of all CLL patients (pts); pts >75 years (yrs) are included in 3% only. Furthermore initial phases are not reflected in multicenter studies and data representing real life situation are scarce.Methods:Out of 1.443 patients consulting our center because of leucocytosis (>10 Gpt/l) between 2003/07/01 and 2018/06/30 we diagnosed CLL consecutively in 234 pts (16,2%): 143 male (m), 91 female (f). Median age at diagnosis is 71,4 yrs. Median leucocyte count is 18,0 Gpt/l. BINET - Stages are: A in 82%, B in 11% and C in 7% of pts. Diagnoses are strictly based on WHO definition: Pts with small lymphocytic lymphomas (sLL, n=11) are included but those with monoclonal lymphocytosis of unknown significance (MLUS) or monoclonal B - lymphocytoses with uncertainly flow cytometry (FC) results are excluded. Histopathology, FC and genetics have been performed by external hematological reference laboratories. All investigations followed the rules of best clinical and laboratory practice. Dates of death are given by the record sections of the involved communities (deadline 2018/06/30).To address different questions we defined 3 groups of pts: A: “collective group”, (n=234), i.e. all pts.diagnosed from 2003/07/01 to 2018/06/30. B: “epidemiological group” (n=129), i.e. pts from 3 communities (113.000 inhabitants - inc - in 2009/12/31) in close proximity referred to our center. C: “genetic features group” (n=99), i.e. pts diagnosed continuously between 2012/07/01 and 2018/06/30 with systematically performed genetics, (i.e. at least 80% of pts in this group. Genetics are: Cytogenetic (banding) n=88; FISH (del 6q21/6q, del 11q22.3, +12/+12q, del 13q14/ 13q34, del 17p13.1, 14q32) n=85. PCR (IgVH status - mutated vs. unmutated; TP53, NOTCH1, SF3B1 mutation) n=82.Results:“Collective group” (A): 7,5 years OS is 72% and 15 years OS is 33% respectively (KAPLAN-MEIER). Age of pts acts as a predominant factor for long term OS: 77% in pts < 60 yrs vs. 40% in pts ≥60 to <75 yrs vs. 16% in pts ≥75 yrs. 15 yrs OS of treated pts is 32% (n=78, median age at diagnosis: 73,3 yrs, m : f = 1,50); 15 yrs OS of untreated patients is 33% (n=146, median age at diagnosis: 71,0 yrs, m : f = 1,56). Mean time to 1st treatment was 16,0 month (0,1 to 132,6 months).“Epidemiological group” (B): Raw incidence is 7,56/105 inh. (m: 9,13, f: 6,06), age adjusted incidence is 10,3/105 inh (m: 12,5, f: 8,3), with highest rate of age between 75 to < 80 yrs. Standard incidence is calculated as 6,27/105 (BRD 1987), 4,78/105 (Europe) and 5,06/105 (USA 2000). Prevalence could be measured directly (2017-12-31): 79,1/105 inh.“Genetic features group” (C): IgVH mutation status is hypermutated in 48 pts. vs. unmutated in 39 pts. Results of FISH analyses (n = 85) are: del 6: 5,5%, del 11: 17,6%, +12: 3%, del 13q: 70,3%, del 17p: 8,8%; normal status: 13,2%. PCR revealed mutations of NOTCH1 in 7%, of SF3B1 in 10,5% and of TP53 in 8,6% of 82 pts, respectively. Combined del17p by FISH and molecular TP53 mutations at diagnosis are found in 3 of 81 pts (3,7%).Cytogenetic findings different to results of FISH and/or PCR were found in 36 of 82 pts (43,9%).Calculated 5 yrs freedom of therapy is 75% in IgVH hypermutated pts vs. 45% in unmutated pts (p = 0,0005; log rank test); calculated 5 yrs OS is 92% vs. 59% (p=0,023).Special situations in CLL: 13 of 234 pts (5,5%) have one parent or sibling with low grade NHL, mostly CLL. In contrast there is non spouse suffering of CLL / low grade NHL. Deficiency in Immunoglobuline G (Ig G < 4,0 g/l) was found in 17 of 199 pts at diagnose (8,5%), 29 pts (12,4%) were substituted with Ig G's in the Course of CLL. In 14 (6,0%) pts. we diagnosed a 2nd hematologic malignancy (3x cMPN, 1x CML, 3x MDS, 3x AML, 1x FL, 1x MCL, 1x MZL, 1x DLCBL-independent of CLL; monoclonal gammopathies are not considered). RICHTER syndrome was found in 1 case, but progression to plasmocytic/plasmoblastic disease was seen in 3 cases. Li FRAUMENI Syndrome was revealed in 1 male patient, he has been in stable disease for 13 yrs.Conclusion:CLL seems to be more frequent than yet considered. IgVH mutation status seems to be very important as a single prognostic factor concerning time of 1st therapy as well as OS. Clinical features of CLL are very different and impressing. DisclosuresBöttcher:Genentech: Research Funding; Janssen: Honoraria; AbbVie: Honoraria, Research Funding; Celgene: Research Funding; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.