ALBERT

Description: Peripheral T-cell lymphoma is a rare heterogeneous disease with generally poor outcome. Known risk factors include the International Prognostic Index (IPI) and b2 microglobulin. However, there is little information on molecular risk factors. In this single center analysis we have prospectively studied the prognostic value of a clonal T-cell receptor rearrangement (TCR) determined by conventional PCR for the γTCR (sensitivity 1 in 102) from DNA from peripheral blood MNC. Thirty nine consecutive patients diagnosed between 1987 and 2006 were assessed for clonality at diagnosis. Patient characteristics: Median age 53 years (range 28–89). M:F = 29 vs. 10. Histological diagnosis included: PTCL-NOS 7 (18%); Large cell 1 (2.5%); Large + medium-sized 5 (13%); Pleomorphic 7 (18%); Small cell 3 (8%); Small+medium mixed cell 1 (2.5%); Lennert’s type 4 (10%); Hepatosplenic 3 (8%); Intestinal 4 (10%); Kimura’s disease: 1 (2.5%); AILD: 3 (8%). Assessment of γTCR showed monoclonality in 24 (62%) and polyclonality in 15 (38%) patients. Patients with a monoclonal TCR had higher clinical stages, higher LDH levels, higher IPI scores, and higher ß2 microglobulin levels. Clinical stage: CS I–III (clonal 42%; polyclonal 58%) vs. CS IV (clonal 93%; polyclonal 7%). LDH above 240 U/L (clonal 78%; polyclonal 22%). IPI 0,1 (clonal 44%; polyclonal 56%), IPI 2 (clonal 75%; polyclonal 25%), IPI 3 (clonal 87%; polyclonal 13%), IPI 4,5 (clonal 60%; 40%). ß2 microglobulin above 2 mg/L (clonal 68%; polyclonal 32%). Patients with a clonal TCR tended to be older (57 vs. 49 yrs.). No major differences were found for age and sex. Induction treatment consisted of polychemotherapy (CHOP-like=34), prednisone (1), radiation (1). 3 patients received no therapy. Nine patients received subsequent involved field radiation and 6 pts. had an autologous stem cell transplant. Significant differences were observed in terms of disease outcome: The presence of a clonal γTCR (n=24) was associated with low clinical remission rates: CR 38%, PR 21%, SD 8%, PD 33%, relapse rate 38%. Remission rates in patients with polyclonal rearrangements (n=15) were: CR 66%, PR 20%, SD 7%, PD 1%, relapse rate 47%. The estimated 3-year overall survival for the monoclonal group was 26% vs. 69% for the polyclonal group (P=0.01). Importantly, patients with CS I–III and a monoclonal γTCR rearrangement (n=10) had a similarly poor survival when compared to the monoclonal group with CS IV (n=14) (18 months vs. 12 months; P=0.23). This indicates that γTCR positivity is of particular predictive value in patients with CS I–III. The data from this unique cohort suggest that γTCR-PCR is a useful prognostic tool in patients with peripheral T-cell lymphoma. Figure Figure

Description: Key Points Uridine diphospho glucuronosyltransferase 2B17 (UGT2B17) is overexpressed in poor prognostic chronic lymphocytic leukemia.

Description: Inhibition of Janus-kinase 1/2 (JAK1/2) is a mainstay to treat myeloproliferative neoplasms (MPN). Sporadic observations reported the co-incidence of B-cell non-Hodgkin lymphomas during treatment of MPN with JAK1/2 inhibitors. We assessed 626 patients with MPN, including 69 with myelofibrosis receiving JAK1/2 inhibitors for lymphoma development. B-cell lymphomas evolved in 4 (5.8%) of 69 patients receiving JAK1/2 inhibition compared with 2 (0.36%) of 557 with conventional treatment (16-fold increased risk). A similar 15-fold increase was observed in an independent cohort of 929 patients with MPN. Considering primary myelofibrosis only (N = 216), 3 lymphomas were observed in 31 inhibitor-treated patients (9.7%) vs 1 (0.54%) of 185 control patients. Lymphomas were of aggressive B-cell type, extranodal, or leukemic with high MYC expression in the absence of JAK2 V617F or other MPN-associated mutations. Median time from initiation of inhibitor therapy to lymphoma diagnosis was 25 months. Clonal immunoglobulin gene rearrangements were already detected in the bone marrow during myelofibrosis in 16.3% of patients. Lymphomas occurring during JAK1/2 inhibitor treatment were preceded by a preexisting B-cell clone in all 3 patients tested. Sequencing verified clonal identity in 2 patients. The effects of JAK1/2 inhibition were mirrored in Stat1−/− mice: 16 of 24 mice developed a spontaneous myeloid hyperplasia with the concomitant presence of aberrant B cells. Transplantations of bone marrow from diseased mice unmasked the outgrowth of a malignant B-cell clone evolving into aggressive B-cell leukemia-lymphoma. We conclude that JAK/STAT1 pathway inhibition in myelofibrosis is associated with an elevated frequency of aggressive B-cell lymphomas. Detection of a preexisting B-cell clone may identify individuals at risk.

Description: Introduction Lymphoma patients are at increased risk of thromboembolic events (TE), however, thromboprophylaxis in these patients is largely under utilized. Actual guidelines recommend different models for thromboembolic risk estimation in cancer patients. Proposed models are of limited use in lymphoma patients as their development is not based on specific characteristics for this patient population. Previously, we developed and internally validated a simple model, based on individual clinical and laboratory patient characteristics that would classify lymphoma patients at risk for a TE. The variables independently associated with the risk for thromboembolism were: previous venous and/or arterial events, mediastinal involvement, BMI〉30 kg/m2, reduced mobility, extranodal localization, development of neutropenia and hemoglobin level 〈 100g/L. For patients classified at risk in derivation cohort (n=1236), the model revealed positive predictive value of 25.1%, negative predictive value of 98.5%, sensitivity of 75.4%, and specificity of 87.5%. The diagnostic performance measures retained similar values in the internal validation cohort (n=584). The aim of this study was to perform external validation of the previously developed thrombosis lymphoma (Throly) score. Methods The study population included patients with a confirmed diagnosis of non-Hodgkin lymphoma (NHL), Hodgkin lymphoma (HL), and chronic lymphocytic leukemia (CLL)/ small lymphocytic lymphoma (SLL) from 8 lymphoma centers from USA, France, Spain, Croatia, Austria, Switzerland, Macedonia, and Jordan. During 2015 to 2016, data were prospectively collected for venous TE events from time of diagnosis to 3 months after the last cycle of therapy for newly diagnosed and relapsed patients who had completed a minimum of one chemotherapy cycle. The score development and validation were done according to TRIPOD suggested guidelines. Sensitivity analyses were carried out to test the model robustness to possible different settings, according to in/out patient settings and according to different countries included. Results External validation cohort included 1723 patients, similar to the developed group and consisted of 467 indolent NHL, 647 aggressive NHL, 235 CLL/SLL and 366 HL patients, out of which 121 (7%) patients developed venous thromboembolic events. For patients classified at risk in external validation cohort, the model resulted in positive and negative predictive values of 17% and 93%, respectively. Based on new available information from this large prospective cohort study this model was revised to include the following variables: diagnosis/clinical stage, previous VTE, reduced mobility, hemoglobin level 〈 100g/L and presence of vascular devices. In the new score we divided patients in two groups: low risk patients, score value ≤ 2; and high risk patients, score value 〉 2. For patients classified at risk by the revised model, the model produced positive predictive value of 22%, negative predictive value of 96%, sensitivity of 51%, and specificity of 72%. In sensitivity analysis, the final model proved its robustness in different settings of major importance for lymphoma patients. The final model presented good discrimination and calibration performance. Concordance C statistics was 0.794 (95% CI 0.750-0.837). Conclusions Revised Thrombosis Lymphoma - ThroLy score is more specific for lymphoma patients than any other available score targeting thrombosis risk in solid cancer patients. We included biological characteristic of lymphoma, indolent vs aggressive, as well as data about dissemination of disease, localized vs advanced stage, reflecting specificity of lymphomas comparing to other types of cancer. Also, we pointed out significance of central vascular devices as risk factor having considered the role of vascular damage during insertion as a potential trigger for activation of the clotting cascade. This score is user friendly for daily clinical practice and provides a very good predictive power to identify patients who are candidates for pharmacological thromboprophylaxis. Disclosures Cheson: AbbVie, Roche/Genentech, Pharmacyclics, Acerta, TG Therapeutics: Consultancy. Ghielmini:Roche: Consultancy, Honoraria, Research Funding, Speakers Bureau. Jaeger:Gilead: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; AOP Orphan: Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; MSD: Research Funding; Bioverativ: Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Honoraria; Mundipharma: Membership on an entity's Board of Directors or advisory committees; Takeda-Millenium: Membership on an entity's Board of Directors or advisory committees; Takeda-Millenium: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; GSK: Membership on an entity's Board of Directors or advisory committees; Infinity: Membership on an entity's Board of Directors or advisory committees.

Description: Background. Anaplastic lymphoma kinase positive systemic anaplastic large cell lymphoma (ALK+ ALCL) represents an aggressive T-cell malignancy that primarily affects children and younger adults. Upon treatment with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) or CHOP-like regimens its prognosis is relatively favourable compared to other aggressive T-cell lymphomas, however, the rare event of disease reoccurrence remains particularly challenging. ALCL is CD30 positive, and mono treatment with brentuximab vedotin (BrV) can achieve long term remissions in only few relapsed patients not receiving consolidating stem cell transplantation (Pro B. et al. Blood 2017). A subset of ALCL patients express platelet derived growth factor receptor (PDGFR) alpha or beta in their tumour cells which is induced by ALK via AP-1 transcription factor activation. We previously demonstrated that PDGFR blockade by the Abl/c-Kit/PDGFR kinase inhibitor imatinib is an effective treatment concept for ALK+ ALCL in experimental mouse models (Laimer D. et al. Nature Med. 2012). Here we aimed to assess the prognostic value of PDGFR expression and to probe the clinical value of its targeting by imatinib in ALK+ ALCL patients. Methods. The impact of PDGFR expression to predict clinical outcome was analyzed in samples of 98 ALK+ ALCL patients included in the studies NHL-BFM 90, 95 and ALCL99 between 1992 and 2006. Six patients (age 18-45) with chemo-refractory ALK+ ALCL were prospectively treated with imatinib plus (4) or minus (2) BrV. 33% (2/6) had previously received salvage treatment with stem cell transplantation (SCT). Three of the patients had been enrolled in a prospective single-arm study combining imatinib and BrV, a trial that was terminated due to poor patient recruitment (AGMT ALCL1 trial, EudraCT No.: 2013-003505-26, supported by Takeda®). PDGFR alpha and beta expression analyses were performed on tumour samples of treated patients. Results. ALK+ ALCL patients with high PDGFR expression on tumor cells (n=11) had a significantly lower event-free survival rate (EFS) at 5 years compared to patients with no or low expression (n=87) (27±13% versus 70±5%, respectively, p

Description: Abstract 2685 Genetic alterations such as C-MYC, BCL2, BCL6 translocations and TP53 alterations have a prognostic impact in patients with aggressive B-cell lymphomas (Johnson et al JCO 2012; Sietse et al Blood 2011; Young et al Blood 2009). Here we provide novel information about the influence on survival of sole C-MYC translocation compared with additional BCL2 and/or TP53 alterations. The clinical outcome of 28 patients with C-MYC positive diffuse large B-cell lymphoma (DLBCL) (N=4) or B-cell lymphoma, unclassifiable, with features intermediate between DLBCL and Burkitt lymphoma (B-UNC/DLBCL/BL) (N=24) was retrospectively evaluated according to their genetic alterations (GA) and therapy (R-CHOP N=10 (36%) patients vs. B-ALL-GMALL containing HD-MTX, N=18 (64%) patients). C-MYC, BCL2 and TP53 alterations were screened by FISH. TP53 mutations were investigated by immunohistochemistry, positive cases were further sequenced. Cases were grouped in 4 categories: C-MYC only (32%), C-MYC & BCL2 (32%), C-MYC & TP53 (21%) and triple hit (14%). Statistical analyses were performed using R for Windows 2.12.1 (R Development Core Team) software. P-values 〈 0.05 were considered as significant. Using Kaplan Meier survival analysis no significant difference was found between therapeutic regiments indicating no advantage for aggressive chemotherapy (Fig. 1A). Based on the GA overall survival (OS) was significantly different between the four groups (p=0.048). OS at five years was 75% for C-MYC, 71% for triple hit, 35% for C-MYC & BCL2 and 17% for C-MYC & TP53 (Fig. 1B). These data confirm previous results in MYC-induced murine lymphomas (Schuster et al Blood 2011). In the log-rank test only low International Prognostic Index (IPI) scores (0–1) predicted survival correctly. Due to this fact, we aimed to define a new prognostic index containing genetic parameters. Univariate and multivariate analyses were performed to identify predictive parameters. Patient characteristics were tested for potential influence on OS using the Cox proportional hazards model in combination with all-possible-subset regression. Covariates were: age, LDH (U/ml), BM infiltration (%) and factor variables: sex, LDH high vs. low, presence or absence of BM infiltration, localization (extranodal vs. nodal), clinical stage, IPI-score, therapy and GA. Model selection was performed using the all-possible subset approach, calculating all models with j=1 to 11 covariates. A model was considered valid if all predictors exhibit p-values below 0.05. A factor variable was considered significant if at least one level differs significantly from a predefined base level (base level for GA: sole C-MYC; therapy: GMALL-ALL protocol; stage: CS I; IPI: 0 points). The final model was checked for possible evidence of non-proportional hazards and for influential observations. Table 1. shows the characteristics of the multivariate Cox regression model including the covariates BM and GA (explained variation R2=0.48). Table 1. Model characteristics for the two-variable model containing genetic alterations and BM infiltration as significant covariates. Covariate/factor level Regression coefficient βj Relative Risk exp(βj) p-value 95 % Confidence Interval C-MYC & BCL2 2.8 16.7 0.008 [2.1; 135] C-MYC & TP53 4.3 71.4 〈 0.001 [5.6; 904] C-MYC & BCL2 & TP53 0.2 0.8 0.87 [0.07; 9.1] BM 3.2 25.0 0.001 [3.5; 178] It became obvious that the presence of C-MYC & BCL2 or C-MYC & TP53 increased the relative risk for lymphoma related death whereas C-MYC only and interestingly also triple hit cases showed favorable prognosis in this rare entity. These finding motivated the definition of a new prognostic index: assigning one point each to the risk factors 1) BM infiltration, 2) C-MYC & BCL2; two points to 3) C-MYC & TP53 and no point to 4) presence of sole C-MYC or 5) triple hit aberrations, respectively, yielding scores between 0 and 3. Based on these scores two prognostic groups “good” (scores 0–1) vs. “poor” (score 2–3) were classified. OS stratified by the new risk groups was highly significant (p-value 〈 0.001) (Fig. 1C). Our data indicate that 1) C-MYC and TP53 simultaneous alterations constitute the highest risk group. This risk is considerably ameliorated when an additional BCL2 aberration is present. 2) GMALL-ALL protocol is non-superior to R-CHOP. 3) Our prognostic index integrating clinical and genetic features is able to predict outcome and should be evaluated in larger studies. Disclosures: Jaeger: Emergent Product Development: Consultancy, Research Funding.

Description: Fludarabine and cyclophosphamide are the backbone of therapy for patients with CLL. The addition of rituximab leads to further improvement of response rates and progression free survival (results from the randomized CLL8 study of the GCLLSG). However, a considerable number of patients have insufficient or short responses to FC or RFC and there is a need to identify factors influencing response or resistance to therapy. The aims of this study are to identify gene expression associated with response or resistance before the start of therapy, to investigate changes in the expression of specific genes or pathways associated with response or resistance during the first cycle of FC and RFC, to provide a rationale for the additional use of novel drugs to improve remission and overcome resistance. We investigated peripheral blood samples from 20 patients receiving FC (n=10) or RFC (n=10) by gene expression profiling, flow cytometry, RT-PCR and western blotting before and during therapy. Sixteen patients received FC or RFC as first line (8 within the CLL8 study) and 4 as second line treatment. All patients were in stage Binet B or C. Gene expression was analyzed and correlated to good (CR or PR) or poor clinical response (SD or PD) at the end of therapy based on NCI-WG/IWCLL criteria. CD19+ cells were harvested by cell sorting before therapy, 24 hours after FC (FC arm), 24 hours after rituximab, and 24 hours after FC (RFC arm). Microarray analysis was performed using Affymetrix U133A gene chips. Genes with a consistent pattern of expression (high or low) in the majority of samples in the good or poor response group were further analyzed. Overall, 9 patients responded adequately to therapy (3 CR, 7 PR), while 11 did not (7 SD, 3PD). Unmutated IgVH status and poor risk cytogenetics were more frequent in poor responders. Gene expression signature before treatment showed that overexpression of 39 genes strongly correlated with response, while overexpression of 20 genes (including HSPA1B, IFI6, APP, CEACAM1, CD9, GAB1, INPP5F) was associated with resistance. Changes in expression after initiation of treatment was also analyzed. Seven genes (including CENTD1, HBA2, COL9A2 and APRIN) were significantly upregulated after rituximab in non-responders. Upregulation of 13 genes (including PMAIP1, SFRS11, CLK1, EFHC1, MRPL39, TUG1, TBRG1, CD49d, PTPRC) after R-FC and 7 genes (including ITPKB, LOC641298, CD44, TAF5) after FC was associated with poor response (resistance) to RFC and FC respectively. Many of these genes are involved in regulation of apoptosis, cell cycle, integrin and PI3-K signaling Therapeutic antibodies or inhibitors against some of these targets are already available. RT-PCR analysis demonstrated a significant downregulation of Akt1 mRNA 24 hours after rituximab infusion in RFC group but no significant changes were observed in patients receiving FC alone. In vitro exposure to rituximab confirmed its in vivo effect and resulted in a significant downregulation of Akt1 and PI3-K-p85 mRNA expression. FACS analysis demonstrated a decrease in the percentage and mean fluorescence intensity (MFI) of surface CD20 after rituximab infusion. This effect was associated with a significant change in total amount and phosphorylation state of CD20 in the RFC group. There was also a decrease in the MFI of CD44 and CD23 after rituximab in the majority of patients in the RFC group but this effect was not consistent in the FC group. In conclusion, we have identified a set of markers associated with good or poor response to FC or RFC before therapy and during the first cycle of treatment. The data provide a rationale for targeted drug combinations to overcome resistance and improve response to therapy in CLL.

Description: Background. Inhibition of Bruton´s tyrosine kinase (BTK) with the small molecule ibrutinib has significantly improved the survival of patients with chronic lymphocytic leukemia (CLL). BTK is also expressed in platelets. Collagen- and von Willebrand Factor (vWF)-dependent (ristocetin-induced) impairment of platelet function has recently been described (Levade M et al., Blood 2014, 124:3991-5;Kamel S et al., Leukemia 2015, 29:783-787) . Bleeding events were observed in 61% of patients in a recently published 3 year follow-up (Byrd JC et al., Blood 2015, 125:2497-2506). Bleeding under ibrutinib is generally mild (CTC grade 1-2 corresponding to spontaneous bruising or petechiae), but grade 3 or 4 bleeding can be observed, particularly after trauma. We hypothesized that quantitative assessment of platelet aggregation in ibrutinib CLL patients could help (1) to predict bleeding tendency, and (2) to guide patients through invasive procedures. Patientsand Methods. Twenty-four adult patients with previously treated CLL (16 male/8 female, median age 67 years, range 55-84) received ibrutinib orally at a planned dose of 420mg/day and were regularly monitored and thoroughly investigated for bleeding tendency. The median time on ibrutinib was 7.5 months, (range 1-27). Bleeding events (any CTC grade) occurred in 13 (54%) and dose-reductions to 280 (N=12) or 140mg (N=3) (for bleeding, infections, or neutropenia) were made in 15 (63%) of patients during a median observation period of 5 months (range 1-12). Bleeding was observed in 4 of 6 patients with concomitant anticoagulation. Of note, only 1 of the 24 patients had a CTC grade 3 bleeding event, and no grade 4 or 5 events were observed. Ristocetin-induced platelet aggregation (RIPA, herein referred to as RCoF) was quantitatively measured in fresh hirudin-blood by whole blood aggregometry with a Multiplate® Analyzer (Roche Diagnostics). Platelet aggregation was expressed in AUC units (U) (normal range 98-180U). Controls included normal subjects (N=53). Consecutive samples before and during treatment were available in all patients. Statistical methods comprised t-Test and ANOVA using SAS. Results. Ristocetin-induced platelet aggregation was already diminished before ibrutinib treatment (median 51 RCoF U) when compared to normal controls (Table 1). This is likely due to lower platelet counts in CLL patients influencing overall platelet aggregability (Hanke AA et al., Eur J Med Res 2010, 15:214-219). During ibrutinib treatment, platelet aggregation was substantially impaired (median of 22U). A direct comparison of available paired samples in 5 patients showed a significant decrease after ibrutinib initiation (51 to 14.5U; p=0.0028). Of note, significantly lower values were measured at visits when bleeding events were documented (N=34) compared to patient visits without bleeding tendency (N=70) (median 13 vs. 42U; p 2 (N=10) vs.

Description: Maintenance therapy with the monoclonal anti-CD20 antibody Rituximab (R) improves clinical outcome in patients with follicular lymphoma in complete or partial remission (CR, PR) after induction therapy. However, optimal dosing schedule and duration of maintenance therapy are still under investigation. Commonly applied regimen include four doses of 375mg/m2 Rituximab once a week every 6 months, one dose every 3 months, or one dose every 2 months. Maintenance is usually given for 2 years. In addition to clinical trials, pharmacokinetic (PK) studies could help determining optimal dosing of Rituximab. PK values have been obtained during induction therapy in several trials and an association between drug trough levels and clinical effectiveness has been established (25μg/ml in responding patients). However, data on Rituximab PK during maintenance therapy are lacking. The Austrian Cooperative Study Group for Cancer Drug Therapy (AGMT) has conducted a phase II trial in 33 patients with follicular lymphoma in CS III/IV with an induction therapy with 6 cycles of Rituximab (375mg/m2 i.v., day 1), Fludarabine (30mg/m2 p.o., day 2–4)) and Mitoxantron (10mg/m2 i.v., day 1) every 4 weeks (R-FM) (NHL9). Thirty two patients obtaining a CR, CRu or PR received maintenance treatment with Rituximab 375mg/m2 every 2 months for 1 year. Pharmacokinetic data for Rituximab were obtained during induction and maintenance treatment in 16 of these patients. All patients converted from BCL2/IgH PCR positivity to negativity in peripheral blood after R-FM induction. PK serum concentrations during induction corresponded well to known data: A steady increase in pre- and post-dose levels was observed with increasing cycle number. Median, minimum and maximum Rituximab serum concentrations are given in the upper panel of Table 1: Table 1. Serum concentrations (μg/ml) during R-FM induction and two-monthly maintenance treatment with 375mg/m2 of Rituximab. R-FM Induction Cycle 1 2 4 6 (Q 4 weeks) min. med. max. min. med. max. min. med. max. min. med. max. Pre-dose N.A.