Refractory CLL Patients Research Articles

Rapid Tumor Debulking of Relapsed/Refractory CLL Patients By PI3Kδ Inhibition and Anti-CD20 Monoclonal Antibody Treatment

Introduction Effective multi-targeted drug combinations could rapidly achieve deep remissions in CLL allowing for defined duration treatment. The combination of a novel highly-specific phosphoinositide 3-kinase delta (PI3Kδ) inhibitor and casein kinase 1 epsilon (CSK1ε) inhibitor, umbralisib (UMB), and a glycoengineered chimeric monoclonal antibody (mAb) targeting a unique epitope on CD20, ublituximab (UBL), with the BCL2 inhibitor venetoclax (VEN) may decrease drug resistance, reduce risk of tumor lysis syndrome (TLS) and achieve higher levels of undetectable minimal residual disease (MRD). This triple agent therapy is being evaluated in relapsed or refractory CLL patients (U2-VEN, ClinicalTrials.gov NCT03379051). Patients received oral 800 mg UMB once-daily. For Cycle 1, UBL was given via IV on Day 1 (150 mg), Day 2 (750 mg), Day 8 (900 mg), Day 15 (900 mg), and then on Day 1 in Cycles 2 and 3 (900 mg); for 3 cycles of 28 days, followed by consolidation with UMB and VEN for cycles 4 to 24. Patient (n = 25) blood samples were collected Pre- and Post-treatment on days of UBL administration during cycles 1-3. Our studies focused on UMB and UBL treatment effects on: 1) The decrease in the circulating CLL tumor burden and thus the subsequent risk of TLS from the addition of VEN to the regimen; and 2) CD20 antigen loss on CLL cells, a major mechanism of anti-CD20 mAb resistance. Methods Blood was analyzed by clinical laboratory automated cell counter and then stained with antibodies against CD3, CD5, CD19, and CD20 for flow cytometry analysis. CLL cell counts were calculated from flow cytometry and clinical laboratory results. To quantitate CD20 surface molecules, peripheral blood mononuclear cells were isolated and stained for flow cytometry analysis with the addition of a Molecules of Equivalent Soluble Fluorochrome standard curve. All human specimen collection and usage was conducted with written informed consent after approval of our Institutional Research Subjects Review Board according to the ethical guidelines of the Declaration of Helsinki. Results Average CLL cell counts (47 x 109/L) decreased rapidly after administration of the first dose of UMB and UBL (C1D1 Post, 26 x 109/L), which rebounded the next day (C1D2 Pre, 43 x 109/L) and then decreased after the second treatment (C1D2 Post, 24 x 109/L). By Day 15, the average CLL cell count had dropped to 9 x 109/L, which decreased further to 5 x 109/L by the start of cycle 3 (Fig. 1). Notably, the variation in CLL cell counts (error bars) decreases over treatment time, suggesting that most CLL patients' tumor burdens are decreasing to a similar low level independent of starting CLL cell count. The change in average CLL cell count between Pre and Post measurements is initially very high (45% decrease C1D1) with decreasing effectiveness over treatment time (8% decrease C3D1). This change could be due to the loss of CLL cell membrane CD20 levels leading to mAb resistance. Pre-treatment (C1D1 Pre) average CLL cell membrane CD20 levels (n=18) were 29,000 CD20 molecules/cell. This decreased to 9,000 molecules/cell after the first treatment (C1D1 Post), stabilized at 9,100 molecules/cell on the next day (C1D2 Pre) and then decreased further after the second treatment (C1D2 Post) to 1600 molecules/cell. After day 8 treatment, CD20 levels remained at 1,000 molecules/cell or less until the end of this study (C3D1). Discussion UMB and UBL rapidly decrease circulating CLL counts over three cycles of treatment, reducing most CLL patient tumor burden to similar low levels. This clinical response could facilitate addition of VEN in the 4thcycle of therapy by decreasing the risk of TLS. The rapid loss of surface CD20 from CLL cells to 1,000 CD20 molecules/cell or less by C1D8 Post has the potential to cause acquired resistance to anti-CD20 mAb therapy. This rapid and sustained loss of CD20 levels is consistent with that reported at standard doses for other anti-CD20 mAbs. However, an effect of concomitant UMB on CD20 levels cannot be ruled out. Previous work suggests that lower anti-CD20 mAb doses can prevent these large decreases in CD20 levels and increase UBL efficacy. In conclusion, our initial data show that treatment with a selective PI3Kδ and CSK1ε inhibitor (UMB) combined with an anti-CD20 mAb (UBL) is effective in decreasing CLL cell counts in all patients assessed in the U2-VEN clinical trial. However, treatment efficacy could be limited by large decreases in the CLL surface CD20 levels. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Lipase-triggered drug release from BCL2 inhibitor ABT-199-loaded nanoparticles to elevate anti-leukemic activity through enhanced drug targeting on the mitochondrial membrane

Selective BCL2 inhibitor ABT-199 has been approved to treat hematological malignancies including acute myeloid leukemia (AML). However, acquired drug resistance and severe side effects occur after extended treatment limiting the clinical usage of ABT-199. Here, we successfully encapsulated pure ABT-199 in amphiphilic mPEG-b-PTMC169 block copolymer, forming mPEG-b-PTMC169@ABT-199 nanoparticles (abbreviated as PEG-ABT-199), which presented better aqueous dispersion and higher efficiency of loading and encapsulation than pure ABT-199. We then compared the anti-leukemic ability of pure ABT-199 and PEG-ABT-199 in vitro and in vivo. PEG-ABT-199 had a lower IC50 value compared with pure ABT-199 in MV4-11 and MOLM-13 cell lines. In addition, PEG-ABT-199 significantly induced apoptosis and decreased colony number than pure ABT-199. Most importantly, PEG-ABT-199 markedly reduced leukemic burden, inhibited the infiltration of leukemic blasts in the spleen, and extended the overall survival (OS) in MLL-AF9-transduced murine AML compared with free ABT-199. Meanwhile, the blank PEG169 NP was non-toxic to normal hematopoiesis in vitro and in vivo, suggesting that PEG169 NP is a safe carrier. Mechanistically, PEG-ABT-199 enhanced mitochondria-targeted delivery of ABT-199 to trigger the collapse of mitochondrial membrane potential (MMP), the release of cytochrome c (cyt-c), and mitochondria-based apoptosis. In conclusion, our results suggest that PEG-ABT-199 has more vital anti-leukemic ability than pure ABT-199. PEG-ABT-199 has potential application in clinical trials to alleviate side effects and improve anti-leukemia ability. Statement of significanceATB-199, an orally selective inhibitor for BCL2 protein, presents marked activity in relapsed or refractory AML, T-ALL, and CLL patients. However, ABT-199 resistance severely limits the further clinical usage because of off-target effects, non-specific toxicities, and low delivery of drugs. To reduce the side-effects and improve the solubility and bioavailability, ABT-199 was encapsulated into the amphiphilic mPEG-b-PTMC block copolymer by co-assembly method to obtain mPEG-b-PTMC@ABT-199 nanoparticles (PEG-ABT-199). PEG-ABT-199 has several advantages compared with pure ABT-199. 1.PEG-ABT-199 presents better aqueous dispersion and higher efficiencies of loading and encapsulation than pure ABT-199. 2. PEG-ABT-199 substantially enhances the anti-leukemic ability in vitro and in vivo compared with pure ABT-199. 3. PEG-ABT-199 has little effects on normal cells. 4. PEG-ABT-199 can reduce treatment cost.

Obinutuzumab for CLL Relapsed and Refractory Patients When New Oral Drugs an Not Available

Introduction:The management of relapsed or refractory chronic lymphocytic leukemia (r/r CLL) has undergone profound changes due to a better understanding of the disease's biology and the new drugs. The choice of the best treatment at any time after the first line therapy, for a single patient, during the course of the disease depends on age, performance status, comorbidities, renal function, and effectiveness of the previous treatment received. In many countries, although approved by regulatory agencies, new therapeutic options are not available for the patients or can be offered just for a restricted group. In Brasil, for relapsed or refractory CLL patients, ibrutinib is not funded by the public nor private health insurance. This real world scenario imposes a limitation on standard international guidelines CLL treatment recommendations.Objective:To describe the acute toxicity and responses to treatment with Obinutuzumab, outside of a clinical trial, for CLL relapsed and refractory patients.Methods:Descriptive, real word, case series report of unfit refractory or relapsed CLL patients treated with obinutuzumab with or without chlorambucil as the second line and beyond. Data were obtained from a questionnaire collected through patient-record forms completed by attending physicians.Results:A total of 14 patient-completed questionnaires were analyzed. Median age was 72 (51-84) years, 10 pts (71%) had ECOG status ≥ 0-2. Four pts (28%) had ≥ 3 comorbidities and 11 pts (78%) an impaired renal function at baseline with a creatinine clearance of 30-70ml/min. No data on cytogenetic or IGHV status was collected. Previous treatments were FC or FCR in 7 pts (50%) and all patients were treated with rituximab in the previous line therapy. Prior obinutuzumab infusion, 3 pts (23%) presented thrombocytopenia grade 3 CTC, and median ALC was 49.938 (1.890- 194.000). Seven (50%) patients experienced infuse reaction in cycle one (6 pts CTC 2, pt 1 CTC 2). Some of the cycles were delayed in 7 pts (50%) and interrupted in 2 (15%) due to hematologic toxicity. Chlorambucil was associated in 12 (85%) and 2 pts (15%) were treated with obinutuzumab in monotherapy. Neutropenia 3/4 CTC was observed in 9 pts (64%) and thrombocytopenia 3 CTC in one patient. Median obinutuzumab cycles were 5 (1-6), and the overall response was achieved by 13 pts (9 CR + 3 PR, Hallek 2008). As of July 2017, one death was described in a patient with a severe and prolong hematologic toxicity after the first cycle of Clb+obinutuzumab in which bone marrow biopsy demonstrated persistent CLL infiltration. This patient started ibrutinib treatment but died of H1N1 infection. So far, 13 pts were alive, without further serious adverse event.Conclusion:New drugs for r/r CLL treatment are not available in some countries. Obinutuzumab demonstrated a superior efficacy over Rituximab in CLL patients. Although the administration of Obinutuzumab to CLL relapsed or refractory patients was not tested in large clinical trials, it can be an alternative for unfit patients. The results of this descriptive, real world data demonstrated that obinutuzumab was effective, with no unexpected AE/SAE. The r/r CLL unfit population, previously exposed to fludarabine, needs a special attention considering the safety aspects. Acknowledgments for Matheus Golçalves and Celso Rodrigues on behalf of Brazilian CLL Group. DisclosuresFogliatto:Roche: Honoraria, Other: Travel Support; accomodations; Novartis: Honoraria, Other: Travel support; AbbVie: Other: Travel Support.

Ibrutinib Induces Durable Remissions in Treatment-Naïve CLL Patients with 17p Deletion/TP53 Mutations: Five Year Follow-up from a Phase 2 Study

Ibrutinib Induces Durable Remissions in Treatment-Naïve CLL Patients with 17p Deletion/TP53 Mutations: Five Year Follow-up from a Phase 2 Study

A Phase 1/2 Study of Umbralisib Ublituximab and Venetoclax in Patients with Relapsed or Refractory Chronic Lymphocytic Leukemia (CLL)

A Phase 1/2 Study of Umbralisib Ublituximab and Venetoclax in Patients with Relapsed or Refractory Chronic Lymphocytic Leukemia (CLL)

UK CLL Forum 5 Year Update on 315 Relapsed Refractory CLL Patients Treated with Ibrutinib in 66 UK and Ireland Centres

Introduction UK and Ireland clinicians gained non-trial access to ibrutinib for relapsed / refractory CLL through a named patient scheme (NPS) for 7 months in 2014, and the UK CLL Forum has been following a cohort of 315 NPS patients treated with ibrutinib at 66 UK and Ireland hospitals. We now update this dataset with up to 5 years follow-up. Results The primary end-points of the study were Discontinuation Free Survival (DFS) and Overall Survival (OS). Of the original 315 patients, 213 have stopped ibrutinib and 159 patients have died, with all living patients having a survival update in 2019. With 55.6 months median follow-up, median DFS was 33.5 months (95% CI: 28.4-29.3) and median OS was 55.9 months (95% CI: 42.5-N/A) (Figure 1A and 1B). Four years after starting ibrutinib, 37.0% of patients were still taking ibrutinib and 52.4% were still alive. Both DFS and OS were inferior for older (age > median) and PS 2+ patients. Patients with 17p- have inferior DFS and OS compared to patients with other defined cytogenetic aberrations (13q-, 11q-, t(12)) and more patients with 17p- compared with non-17p- stopped ibrutinib for progressive CLL (PD) (26.4% vs 18.8%; p=0.01) and Richter's Syndrome (12.1% vs 8.3%; p=0.05). First relapse patients had more than a 3 fold lower rate of ibrutinib discontinuation due to CLL progression (7.1% versus 24.5%; two-tailed Fisher's exact test; p=0.001) compared with patients receiving ibrutinib at later lines of therapy, although DFS and OS for these groups were similar. With multivariable analysis of pre-treatment factors, older age, PS2+, 17p- and >1 prior line of therapy retained significance for earlier ibrutinib discontinuation and age, PS2+ and 17p- associated with inferior OS. To analyze any potential impact of reduced ibrutinib dosing during the first year of therapy, we originally classified all patients alive at 12 months into 4 groups, depending on first-year ibrutinib treatment. Group A: No dose reductions or breaks >14 days; Group B: Dose reductions, but no breaks > 14 days; Group C: Breaks > 14 days but back on ibrutinib by 1 year; Group D: stopped ibrutinib permanently but still alive at 1 year. With extended follow-up, patients dose reduced in year 1 are very likely to have on-going dose reductions beyond the first year (86% patients in group B are still dose reduced beyond 4 years, compared with 12% in group A). Groups A and B retain similar DFS and OS initially, although beyond 2 years the curves separate (Figure 1C and 1D) (DFS HR: 1.62 (1.05-2.50), OS HR: 1.52 (0.86-2.70)). Interestingly, the dose-reduced group do not have a higher incidence of disease progression, (PD A:24.8%, PD B:23.8%), but have a higher chance of discontinuation for all non-PD causes (A:23.6%, B:42.9%). With longer follow-up, patients from groups C and D had significantly impaired OS compared with group A, with 4 year OS 39.4% and 16.7% respectively compared with 76.4% for group A (p<0.001). Considering all patients who discontinued ibrutinib, stopping for AEs decreased over time (year 1: 57%, 2: 53%, 3: 30%, 4: 21%). Stopping for infection was much more common in the first year (19.5%) compared with years 2 to 4 (8.4%). The risk of stopping ibrutinib due to PD-CLL increases with time (year 1: 11%, 2: 18.6%, 3: 42.5%, 4: 57.6%). Of the PD-CLL patients, treatment with venetoclax had the best OS (n=43; 1 year OS: 77.5%) compared with the 7 treated with rituximab / idelalisib (1 year OS: 42.9%) and 14 treated with other strategies (including palliative care) (1 year OS: 10%). Although this supports the use of venetoclax for disease progression post ibrutinib, there are two potentially significant biases; more palliative patients did not receive venetoclax and patients who relapsed within 1 year of starting ibrutinib, who were potentially more unstable biologically, did not receive venetoclax (unavailable in the UK pre-2016) Conclusions With approaching 5 years of follow-up, around one third of 'real-world' UK / Ireland patients treated with ibrutinib for relapsed / refractory CLL remain on ibrutinib. Our data suggest that younger, better performance status patients who have had fewer prior lines of therapy and no evidence of 17p deletion seem most likely to remain on ibrutinib long-term. Likewise, patients who remain free of adverse events that often associate with dose reductions and treatment breaks, have the best chances of remaining on ibrutinib. Treatment of PD-CLL with venetoclax post ibrutinib shows encouraging results. Disclosures CLL Forum: Roche: Other: Sponsorship of the UK CLL Forum; Abbvie: Other: Sponsorship of the UK CLL Forum; Janssen: Other: Sponsorship of the UK CLL Forum; Gilead: Other: Sponsorship of the UK CLL Forum. Follows:Janssen: Consultancy, Honoraria, Speakers Bureau; Abbvie: Consultancy, Honoraria, Speakers Bureau; Roche: Consultancy, Honoraria, Speakers Bureau; AZ: Consultancy, Honoraria, Speakers Bureau.

Abstract 29: Characterization and treatment of a novel adoptive transfer model of Sf3b1mut/Atmdel chronic lymphocytic leukemia

Abstract The spliceosome component SF3B1 is among the most frequently mutated genes in CLL (Landau et al. Nature 2015), and its mutations are associated with transcriptomic changes in numerous pathways involved in the regulation of CLL-cell survival (Wang et al. Cancer Cell 2016). Across studies, the presence of SF3B1 mutations has been associated with poorer prognosis, and expansion of SF3B1-mutated subclones has been associated with resistance to current CLL therapies. Therefore, therapeutic options that selectively interfere with spliceosome components may represent an attractive therapeutic strategy for selected CLL patients. We recently developed a novel mouse model of human CLL, based on the B-cell restricted coexpression of mutated Sf3b1 and deletion of Atm (Sf3b1mut/Atmdel), two commonly co-occurring genetic alterations in CLL. One of the main advantages of this model is its transplantability in either immunocompetent C57BL/6 or immunocompromised NSG hosts, with the engrafted mice rapidly succumbing to disease (3-4 months for C57BL/6, 4-6 weeks for NSG), thus providing a time frame for informative treatment studies. Sf3b1mut/Atmdelcells express unmutated IGHV B cell receptors (BCR), they show high BCR signaling responsiveness, and express high levels of the CD49d integrin and the chemokine receptor CXCR4, all phenotypic and functional features associated with aggressive human CLL. Sf3b1mut/Atmdel also express high levels of the aberrantly spliced isoforms of Tnpo3, Aurke, and Narfl1 genes, indicative of alternative splicing dysregulation. We took advantage of the spliceosome modulator E7107, to test the effects of splicing inhibition in NSG mice challenged intravenously (i.v.) with 5 x 106 Sf3b1mut/Atmdel splenocytes. Fifteen days post-transplant, 40-50% circulating B220+CD5+Igk+ cells were detectable in the peripheral blood (PB) of recipient mice by flow cytometry. Mice were then randomized and treated i.v. for 5 days with either 4 mg/kg E7107 or vehicle control (3 controls/4 treated), then immediately euthanized. Three hours after the first E7107 dose, we observed significant reduction in Slc25a19 and Dph2 mature-RNA levels, two known targets of E7107, as analyzed by RT-PCR of PB cells collected from the treated animals. After the 5-day treatment, we observed a significant reduction in leukemia burden in the PB, bone marrow (BM), peritoneum (PLC), and spleen (SP) of the treated animals, associated to the presence of apoptotic AnnexinV+ Sf3b1mut/Atmdel cells. Interestingly, MCL1 protein and RNA levels were significantly reduced after the 5-day E7107 administration, as tested by RT-PCR and Western blot analysis of splenocytes harvested at sacrifice. Depletion of MCL1 total protein levels was also observed after 12 hours of in vitro treatment of Sf3b1mut/Atmdel cells with up to 1 μM E7107. In conclusion, we propose a novel and valuable preclinical platform of Sf3b1mut/Atmdel CLL, with phenotypic and functional features similar to human CLL. We demonstrate that splicing modulation by E7107 treatment can reduce leukemia burden, not only in the PB, but also in the SP, BM, and PLC of these mice, by inducing CLL-cell apoptosis. Apoptosis induction is associated with depletion of MCL1 protein, an important antiapoptotic factor in CLL, which is commonly upregulated by microenvironmental interactions in lymphoid organs, and frequently associated with treatment resistance. Studies aimed at further evaluating E7107 selectivity towards Sf3b1mut as compared to Sf3b1wt cells are under way, along with combination treatments with standard or novel CLL therapeutics. These studies may provide the basis for personalized treatment strategies for patients harboring SF3B1 mutations, and novel therapeutic opportunities for refractory CLL patients carrying SF3B1mut subclones. Citation Format: Elisa ten Hacken, Lili Wang, Fara Faye Regis, Michael Thomas, Jing Deng, Kaitlyn Baranowski, Zachary Cartun, Silvia Buonamici, Donna Neuberg, Anthony Letai, Ruben Carrasco, Catherine J. Wu. Characterization and treatment of a novel adoptive transfer model of Sf3b1mut/Atmdel chronic lymphocytic leukemia [abstract]. In: Proceedings of the Second AACR Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; May 6-9, 2017; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(24_Suppl):Abstract nr 29.

Very Early Effects of Ibrutinib on Tumor and Immune Cells in Blood and Lymph Nodes in Relapsed or Refractory Chronic Lymphocytic Leukemia (CLL) Patients

In this study we analyzed the very early effects of the Bruton’s tyrosine kinase (BTK) inhibitor ibrutinib on tumor and immune cells in 7 symptomatic, relapsed or refractory CLL patients during the first four weeks of treatment. Five of the patients had Rai stage IV at study entry and four patients had 17p deletion or TP53 mutation. Median number of previous treatment regimens was 2 (range 1-4). Peripheral blood (PB) samples were collected before (< 1 week) treatment start and at six different time points during the treatment (9 hours after treatment start on day 1; on day 2; day 4; day 8; day 15 and day 29). Fine needle aspiration of two pathological lymph nodes (LN) identified by ultrasound was performed before (< 1 week) treatment start and at day 2, day 8 and day 29. Flow-cytometry was performed to analyze the changes in the peripheral blood mononuclear cell populations in PB including CLL cells, Natural Killer (NK) cells, monocytes, dendritic cells (DC) and T cells memory subsets, helper subpopulations (Ths) and regulatory T cells (Tregs). Moreover, changes in expression of 18 B-cell activation and migration markers on CLL cells as well as T-cell activation and proliferation markers, were analyzed in paired LN and PB samples. Finally, plasma levels of 92 inflammation-related protein biomarkers were assessed by Multiplex Proximity Extension Assay (PEA).In six out of seven patients the size of the LN decreased gradually during the four weeks of observation, achieving complete clinical remission in three patients and partial remission in three other. In 5 evaluated patients, the CLL cell counts in PB increased already 9 hours after treatment start (fold increase 1.36-2.84). In all the patients, CLL cells were higher at day 2 and 8 compared to baseline (p=0.02), and decreased by day 29 to levels not significantly different from the baseline. Over the four weeks period CD4+ cells increased (p=0.03), while CD8+, NK, and NKT cells remained stable. The distribution of the CD4+ and CD8+ memory cell subsets remained unchanged. Th1 cells increased (p=0.01) while Th2, Th17 and Tregs were stable.Expression of Programmed cell death protein 1 (PD-1) and Cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) in T cells was unaffected. No effect was seen on CLL apoptosis. Proliferating (Ki67+) CLL cells, which at baseline varied between 1.2-10.8%, were not detected in blood after 1 week of treatment (p=0.0006). Ki67+ T cells were also eliminated (p=0.0007), but with a delayed kinetics (4 weeks).Paired LN and PB samples were screened for changes in 18 surface markers relevant for B-cell activation and migration. With the exception of CD23, whose expression was lower in PB compared to LN (p=0.02), no difference was observed in the expression of the other analyzed surface markers on CLL cells from the two different tumor compartments at baseline. CD5 expression remained stable during treatment, albeit decreasing significantly by day 29 in PB (p=0.02). Expression of CD20 decreased in both compartments at day 8 (p=0.02) and CD40 decreased at day 8 only in PB (p=0.02). CD69 expression (MFI) decreased both in PB and LN by day 8 (p=0.02), while the percentage of positive cells significantly dropped already at day 2 in PB (p=0.02) and from day 8 in LN (p=0.03). CD23 MFI decreased in the LN compartment already at day 2 (p=0.02), but the percentage of positive cells was significantly lower only at day 8 (p=0.03). Compared to LN, CD23 expression was lower in PB before treatment and therefore the significant drop both in MFI and percentage of positive cells was seen only after 4 weeks (p=0.02). No change in the expression of CD49d was observed.In 5 evaluable patients, the plasmacytoid DCs, undetectable at baseline, increased during treatment (p=0.06), while CD16+SLAN+monocytes decreased (p=0.06).Finally, plasma levels of 50 molecules were significantly changed at ≥ 1 time point during treatment. With the exception of CST5 and IL-17C, which were increased, the residual 48 proteins were all down-regulated, some of them (e.g. CCL3 and CCL4) already by 9 hours. The majority of the cytokines with significantly reduced levels are pro-inflammatory molecules.In conclusion, these data indicate that ibrutinib causes major changes both in CLL and bystander cells as well as in the levels of several inflammation-related protein biomarkers already shortly after treatment initiation. DisclosuresLundin:Janssen: Research Funding. Kimby:Gilead: Honoraria, Other: honoraria for educational lecture in meeting sponsored by Gilead; Roche: Other: Honoraria for lecture in educational meetings; Pfizer: Other: Research grant; Celgene: Other: Honoraria for lecture. educational meeting; Jansen: Membership on an entity’s Board of Directors or advisory committees, Other: Honoraria for lecture at educational session. Österborg:Janssen: Honoraria, Research Funding.

A Dual Bcl-2/xL Inhibitor Induces Tumor Cell Apoptosis in a Hematopoietic Xenograft Model

Over-expression of the antiapoptotic proteins Bcl-2 and Bcl-xL provides a common mechanism through which cancer cells gain a survival advantage and become resistant to conventional standard of care therapies. Because Bcl-2 family members play key, but partially overlapping, roles as regulators of apoptosis, simultaneous inhibition of both Bcl-2 and Bcl-xL could serve as a promising anticancer strategy. Clinical validation of this concept has been demonstrated with multiple agents targeting different Bcl2 family members. For example, recent Phase I/II trials of the dual Bcl-2 / Bcl-xL inhibitor Navitoclax demonstrated a 20% response rate in relapsed, refractory CLL patients. However, in this study dosing was often limited by chronic thrombocytopenia, a well established side effect of inhibiting Bcl-xL.Here we disclose for the first time a novel and potent BH3 mimetic (Bcl2-32), with nanomolar binding affinity for Bcl-2 and Bcl-xL, (Ki = 3.3 and 8.5nM respectively). In contrast to Navitoclax which requires daily oral administration for activity, Bcl2-32 has shown promising efficacy on an intermittent dosing schedule that allows full platelet recovery between doses. In this poster, we describe the in vitro and in vivo activity profile of Bcl2-32 in a variety of cell types that highlights its potential as a dual inhibitor of Bcl2 and BclXL. In addition to exhibiting potent single agent anti-cancer activity in a sensitive Acute Lymphocytic Leukemia (RS4; 11) model with (TGI >100%, P < 0.001), Bcl2-32 also potentiates the effectiveness of standard chemotherapeutic agents in less sensitive DLBCL xenograft models. Taken together, these data suggest that Bcl2-32 represents an exciting development opportunity as single agent or in combination in a broad range of hematopoietic malignancies. DisclosuresAdam:Astrazenenca: Employment. Secrist:AstraZeneca: Employment.

Results from a Phase IIa Study of the Anti-CXCL12 Spiegelmer Olaptesed Pegol (NOX-A12) in Combination with Bendamustine/Rituximab in Patients with Chronic Lymphocytic Leukemia

BackgroundOlaptesed pegol (NOX-A12) is a novel, potent, L-stereoisomer RNA aptamer that binds and neutralizes CXCL12/SDF-1, a chemokine which attracts and activates immune- and non-immune cells via interaction with the receptors CXCR4 and CXCR7. Signaling of CXCL12 is pivotal to the interactions of leukemic cells with bone marrow microenvironment. The therapeutic concept of olaptesed is to inhibit such tumor-supporting pathways and thereby mobilizing and sensitizing CLL cells to therapy.Here we aim to assess the activity and safety of olaptesed in combination with bendamustine and rituximab (BR) in patients with relapsed / refractory CLL.MethodsTwenty-eight relapsed or refractory CLL patients were enrolled and treated in this open-label, single-arm Phase IIa study. To investigate PK/PD, a pilot dose of 1 to 4 mg/kg olaptesed alone was administered to 3 patients per dose group (plus one additional replacement pt) before start of the regular treatment regimen (pilot group). Patients were treated using a dose titration design with intravenous (IV) olaptesed at doses increasing from 1 mg/kg to 2 mg/kg and 4 mg/kg at cycles 1, 2 and 3, respectively, at 1 hour before rituximab treatment. During cycles 4 to 6, olaptesed was dosed at the highest individually titrated dose. Rituximab was administered IV at doses of 375 mg/m² on day 1 of the 1st28-day cycle and 500 mg/m² on day 1 of subsequent cycles. Bendamustine (70 - 100 mg/m²) was given IV on days 2-3 (cycle 1) or days 1-2 (cycles 2-6) of each 28-day cycle following administration of rituximab. Clinical response was assessed according to NCI-WG Guidelines (Hallek M et al. Blood 111; 2008: 5446-56).ResultsTo date, 24 patients completed treatment (12 women, 12 men) with a median age of 68.5 years (range 52 to 79). At screening 5, 9 and 10 patients presented with Binet stage A, B and C, respectively. The median number of prior treatment lines was 1 (range 1-2). Seven high-risk patients presented an unfavorable disease state being relapsed within 24 months after fludarabine/bendamustine treatment (5 pts) and/or presenting a deletion/mutation of the TP53 gene (3 pts). Most patients (19 of 24) were previously treated with fludarabine or bendamustine.A flow cytometric analysis of CD19+/CD5+CLL cells showed a rapid mobilization into the peripheral blood by a single dose of olaptesed which lasted throughout the observational time of 72h. Interestingly, CXCR4 expression levels increased on CLL cell surface in the periphery after olaptesed treatment. This increase, which peaked at 24h, likely reflects the extended circulation of CLL cells in the periphery due to the sustained blockade of CXCL12 by olaptesed. Reduction of lymphadenopathy by ≥ 50% was achieved in 14 out of 21 evaluable patients with reported enlarged lymph nodes by the end of treatment. Concomitantly, rapid reduction of lymphocytosis in peripheral blood with normalization by treatment cycle 2 – 3 was observed and the CLL to leukocyte ratio significantly improved.Efficacy was assessed at the end of cycles 3 and 6. In the full-analysis-set, which excludes two non-evaluable patients (drop-out after the 1st cycle due to adverse events), the overall response rate was 96%: Three patients (14%) achieved a complete response at end of cycle 6 (2 confirmed, 1 investigator reported) and eighteen patients (82%) achieved a partial response (fifteen at end of cycle 6 and three at end of cycle 3). Notably, all seven high-risk patients (defined as relapsed within 24 months after fludarabine/bendamustine treatment and/or presenting a deletion/mutation of the TP53 gene) responded to treatment with olaptesed + BR with a partial response. One patient had a progressive disease. Olaptesed at 1, 2 and 4 mg/kg at a single dose and in combination with BR was safe and well tolerated. The observed adverse reactions were qualitatively and quantitatively as expected for patients treated with BR.ConclusionOlaptesed in combination with BR was safe and well tolerated. Compared with historical data, olaptesed showed superiority over baseline therapy with regards to overall response rate and increasing rates of complete remission, warranting further development of this Spiegelmer in CLL. DisclosuresMontillo:Janssen: Honoraria. Kruschinski:NOXXON Pharma AG: Employment. Dümmler:NOXXON Pharma AG: Employment. Riecke:NOXXON Pharma AG: Employment.

Bendamustine, Ofatumumab and High-Dose Methylprednisolone (BOMP) in Relapsed/Refractory CLL: Results of a Planned Interim Analysis of the French CLL Intergroup ICLL01 Phase II Trial

Introduction:For relapsed or refractory (R/R) CLL patients (pts), combination of bendamustine and rituximab appears safe and effective (Fischer 2011). Ofatumumab monotherapy gives 58% ORR in heavily pre-treated (median 4 prior lines) R/R CLL pts (Wierda JCO 2010). High doses (HD) steroids are also active in poor prognosis pts with bulky nodal involvement or p53 impairment (Castro 2008, Xu 2010). We report a planned interim analysis of the ICLL01-BOMP phase II trial evaluating the association of Bendamustine, Ofatumumab and high-dose MethylPrednisolone for fit R/R CLL pts after 1-3 previous lines (NCT01612988).Patients and Methods:Primary endpoint was CR rate after 6 cycles (cy) of the BOMP regimen [i.e. bendamustine (70 mg/m2 d1, d2), ofatumumab (1000 mg d1;15 on cy#1-2 and d1 on cy#3-6) and HD methylprednisolone (1 g/m2 d1-3)]. The c#1 was preceded by an ofatumumab (300 mg) prephase. Response evaluation (IWCLL 2008) was done 3 months (m) after the last cy along with blood and bone marrow 10-color flow MRD analysis.Results:Among the 55 pts of this analysis, median age was 64 years (44-76). CIRS-G comorbidity score was 2-6 in 61% and pts had received 2-3 lines in 37% of the cases. Prior FCR-like regimens (50 (91%) patients) had been followed by relapse within 2y in 22/55 and 5/55 pts were fludarabine-refractory (FR). IGVH was unmutated (UM) in (47/52) 90.4%. Karyotypes were complex in 18/46 (39%) cases. Distribution according to FISH hierarchical model was: del(17p) in 15 (27%), del(11q) in 14 (26%), trisomy 12 in 4 (7%), del(13q) in 17 (31%) and normal in 5 (9%). Mutations on the TP53, SF3B1 and NOTCH1 genes occurred in 17 (31%), 14 (26%) and 5 (9%) pts, respectively.According to published risk stratification (Zenz, 2012), 34/55 pts (62%) belonged to the “highest-risk” group with either TP53 disruption (deletion and/or mutation) (n=19) and/or early relapse within 2 years post-FCR (n=22). The remaining patients belonged to either the “high-risk” group (UM-IGVH and/or Highb2mic and/or del11q) accounting for 17 pts (31%) or to the “low-risk” group (or non evaluable) accounting for 4 (7%) pts.Overall, 292 BOMP cy (mean 5.3 cy/pts) were delivered. Safety analysis recorded 158 grade 3-4 adverse events (G3-4/AE) with according to cy: neutropenia: 20.8%, thrombocytopenia: 11.3%, anemia: 2.4%, infection: 5,8%, hyperglycemia: 7,5%, liver enzyme elevation: 1,4%, cutaneous reaction: 1,4%, ofatumumab infusion related reaction: 0,3% and other AE: 3,4%. Overall 43 out of 55 pts (78.2%) had at least one G3-4/AE. Twenty-eight severe adverse events were reported in 20 pts. Treatment interruption before planned 6 cy occurred for pts’ decision (n=3), excessive toxicity (n=5) or early progressive disease (PD) (n = 4).Response in the ITT population was 76.4% ORR with 20% CR (n=11), 56.4% PR (n=31 including 5 nPR and 1 CRi), 9.1% stable disease (n=5), 10.9% PD (n=6) and 3.6% (n=3) non evaluable. Blood MRD obtained in 45 pts was negative (<10-4) in 13 (28.8%) cases. Following evaluation, 5 responding pts (9%) had RIC allogeneic (RIC-Allo) transplantation with a persistent remission.With median follow-up of 16.2 (5.1-23.6) months (m) we observed 9 deaths, related to PD (n=5), EBV-induced lymphoproliferation (n=1), PML encephalitis (n=1), sepsis/pancytopenia (n=1) or unknown origin (n=1). We recorded 22 relapses (including 4 Richter Syndromes) resulting in treatment in 17 cases, with a BTK inhibitor in 8 cases.The median OS has not been reached (estimation 84% at 18 m) (Fig 1B). The median PFS was 18.4 m (95%CI, 14.6-22.2) and the median time to next treatment 17.6 m (95%CI, 12.9-22.4). With 5 cases censored at time of RIC-Allo, the PFS (censored analysis) was 17.5 m (95%CI, 13.2-21.8). (Fig 1A) After univariate analysis, ORR was lower in the “highest-risk” (64,6%, p=0.01), del(17p) (40%, p=0.003), TP53 mutation (47.1%, p=0.01) and complex karyotype (61.2%, p=0.024) groups. PFS was shorter in the “highest-risk” (14 m, p=0.046), FR (4.96 m, p<0.001), del(17p) (9.5 m, p=0.017) and TP53 mutation (9.5 m, p=0.007) groups.Conclusion:Relapse treatment of CLL is a challenge especially after prior FCR-like treatments, accounting for >90% of this trial population. These results in terms of response and survival appear noteworthy considering that >60% are “highest-risk” pts. This study provides important information for forthcoming comparison with next emerging CLL therapies. [Display omitted] Disclosuresde Guibert:Roche: Honoraria. Feugier:Roche: Honoraria. Schuh:Roche, Gilead, GSK, NAPP, Celgene: Honoraria. Leblond:Roche: Honoraria, Speakers Bureau. Tournilhac:mundipharma: Honoraria, Other, Research Funding; GSK: Honoraria, Other, Research Funding; Roche: Honoraria, Other, Research Funding.

Complex Karyotype, Rather Than Del(17p), Is Associated with Inferior Outcomes in Relapsed or Refractory CLL Patients Treated with Ibrutinib-Based Regimens

▪Background.Ibrutinib is active in relapsed/refractory (R/R) CLL, including patients (pts) with del(17p); pts with del(17p) have similar response rate to pts without, but have shorter progression-free survival and a pattern of continuous relapses. (Byrd NEJM 2013). It is unknown whether subpopulations of pts with del(17p) with distinct clinical courses can be identified. Del(17p) is frequently associated with a complex metaphase karyotype (CKT), defined as ≥3 distinct chromosomal abnormalities. CKT has been associated with inferior outcomes in treatment-naïve and R/R CLL, but its prognostic significance in ibrutinib-treated pts is unknown.Methods.We reviewed 100 pts treated for R/R CLL at MD Anderson Cancer Center with investigational ibrutinib-based regimens from 2010-2013. 50 pts received ibrutinib (Ib) monotherapy, 36 Ib plus rituximab and 14 Ib + bendamustine and rituximab (BR). All pts provided informed consent and studies were conducted according to the declaration of Helsinki. Pre-treatment FISH and CpG-stimulated metaphase cytogenetic analysis was performed on bone marrow.Results.Pt characteristics are shown below:TableCharacteristicAge, median (range)65 (35-83)# prior therapies, median (range)2 (1-12)FISH hierarchy, n (%) (n=95)del(11q)26 (28)del(17p)46 (49)Complex karyotype, n (%) (n=72)26 (36)Unmutated IGHV gene, n (%) (n=98)80 (81)Fludarabine-refractory, n19β2-microglobulin ≥4.0, n (%)48 (56)22/26 pts with CKT had del(17p), [OR 19.2 (4.9-76.4) compared to non-del(17p) pts, p<0.001]; 3 had del(11q). 1 did not have FISH results available. There was no association between CKT and other baseline characteristics. 35/46 pts with del(17p) had metaphase cytogenetics performed. Karyotype was abnormal in 33/35. Mechanisms of TP53 loss were: unbalanced translocations, -17, del(17p) and i(17q) in association with CKT (n=22), del(17p) in the absence of CKT (n=3) and TP53 loss identified by FISH only (n=10).The overall response rate (ORR) for the entire population was 95%, with 16% complete remission (CR). ORR was not different according to baseline characteristics. CR rate was 50% among pts who received Ib + BR versus 10.7% in those combined who received Ib or Ib + rituximab [OR 40.1 (3.0-538.5), p=0.005]. There was a trend toward lower CR rate on multivariable analysis (MVA) for pts with B2M ≥4.0 (p=0.055).Median follow-up in surviving pts was 27mo. On univariable analyses (UVA), the following were significantly associated with event-free survival (EFS): fludarabine-refractory CLL [median EFS 27mo vs. not reached (NR), p=0.025], del(17p) (27mo vs. NR, p=0.008) and CKT (16 vs. 39mo, p<0.0001). There was no association between del(17p) and EFS when pts with CKT were excluded. On MVA, only CKT was significantly associated with EFS [HR 4.1 (1.3-13.4), p=0.018]. On UVA, the following were significantly associated with survival (OS): fludarabine-refractory CLL (28mo vs. NR, p=0.009), del(17p) (33mo vs. NR, p=0.024) and CKT (24mo vs. NR, p=0.003), with no association between del(17p) and OS when pts with CKT were excluded (Figure). There was a trend toward inferior OS in pts with baseline B2M ≥4.0 (42mo vs. NR, p=0.07). On MVA, fludarabine-refractory CLL [HR 5.6 (1.7-17.9), p=0.004], CKT [HR 5.1(1.5-17.5), p=0.009] and B2M ≥4.0 [HR 3.3 (1.0-10.4), p=0.044] were all significantly associated with inferior OS. There was no association between del(17p) and OS. [Display omitted] EFS and OS were significantly longer in pts who achieved CR compared with PR (p<0.001). Pts with primary refractory disease had poor outcomes. 5 pts developed Richter Syndrome, of whom 4 had CKT.Discussion.The presence of CKT is independently associated with inferior EFS and OS in pts with relapsed/refractory CLL treated with Ib, while del(17p) is not. CKT is strongly associated with del(17p) and may be a key determinant of biological behavior in del(17p) CLL. This has important implications for treatment of pts with del(17p); those without CKT appear to have equivalent outcomes with Ib compared to pts without del(17p) and could potentially be managed with long-term Ib and close monitoring. In contrast, the inferior outcomes after initial response in pts with CKT make them ideal candidates for treatment-intensification strategies after initial Ib-based treatment, either with novel drug combinations or with allogeneic stem cell transplant, ideally in the context of well-designed clinical trials. DisclosuresWierda:Pharmacyclics: Consultancy. O’Brien:Pharmacyclics: Research Funding. Burger:Pharmacyclics: Research Funding.

Phase 2 Study Of Otlertuzumab (TRU-016), An Anti-CD37 ADAPTIRTM Protein, In Combination With Bendamustine Vs Bendamustine Alone In Patients With Relapsed Chronic Lymphocytic Leukemia (CLL)

BackgroundCD37 is a tetraspanin protein expressed on the surface of normal and transformed B-cells across a wide range of maturational stages and demonstrates death signaling via SHP1. Otlertuzumab is a CD37-specific therapeutic protein built on the ADAPTIRTM(modular protein technology) platform that has shown significantly greater direct killing of CLL cells than rituximab and higher levels of Fc mediated cellular cytotoxicity of CLL cells than either alemtuzumab or rituximab in pre-clinical models. Preclinical in vitro and in vivo models showed significant activity of otlertuzumab with bendamustine (benda). In phase 1 of this study involving 12 relapsed and/or refractory CLL patients (pts), otlertuzumab plus benda was well tolerated and showed a positive response. This phase 2 trial was conducted to investigate the activity of this combination compared to benda alone. MethodsPts with relapsed CLL who had 1-3 prior treatments, adequate organ function, ECOG performance status ≤2, ANC ≥1200/μL and CrCl >40 mL/min were eligible. Pts were stratified at randomization based on Cumulative Illness Rating Score, CrCl and deletion or mutation of 17p13.1. Pts in the study drug arm received otlertuzumab (20 mg/kg) weekly by IV infusion for two 28-day cycles then every 14 days for four 28-day cycles. In both arms, benda (70 mg/m2) was administered IV on Days 1 and 2 of each cycle for up to six 28-day cycles. Safety was evaluated using CTCAE and IWCLL 2008 Grading scale for Hematologic Toxicity in CLL Studies. Response was determined using the 1996 NCI and 2008 IWCLL Criteria. Flow cytometry was performed on peripheral blood to quantitate and characterize lymphocyte populations. Results65 pts have been treated in the randomized portion of the study, including 32 with otlertuzumab plus benda and 33 with benda alone. Pt characteristics and adverse events (AEs) are shown in the table. To date, 44 patients are evaluable for response assessment by the IWCLL criteria assessed 2.5 months after completion of therapy utilizing investigator assessment of CT scan and bone marrow data. The ORR for the combination of otlertuzumab and benda (n=20) was 80% (16) with a CR rate of 20% (4). For benda alone (n=24) the ORR was 42% (10) with a CR rate of 4% (1). Patients are still in treatment and follow-up but only 10% (2) have progressed on the combination of otlertuzumab and benda while 46% (11) have progressed on benda. The best response at anytime by NCI criteria using available data for the combination of otlertuzumab and benda (n=32) was ORR 78% and CR 16% and for benda alone (n=33) ORR 52% and CR 9%. There were no responses among pts with del17p; among pts with 17p mutation both pts in the combination arm had a response; no pts in the benda alone arm had a response. The frequency of all AEs was similar between treatment arms; hematologic and infection AEs were the most frequent in both arms. There were 13 serious AEs reported by 10 pts in the otlertuzumab+benda arm and 25 reported by 12 pts in the benda alone arm. The half-life of otlertuzumab is 11 days and there was no antidrug antibody in the 14 pts tested to date. ConclusionsPreliminary response rate with otlertuzumab in combination with benda was higher than benda alone by IWCLL or NCI response criteria. The overall incidence of AEs was similar between the 2 treatment cohorts, with a higher incidence of pyrexia, neutropenia and thrombocytopenia with the combination. However, the addition of otlertuzumab did not appear to increase the number of serious adverse events. Pts are in the process of completing treatment and obtaining CT scans and bone marrows; the IWCLL and NCI response rate will be updated. Disclosures:Robak:Gilead Sciences (Advisory board, research support): Consultancy, Honoraria, Research Funding; Emergent BioSolutions (Advisory board, research support): Consultancy, Honoraria, Research Funding; Celgene (Research support): Consultancy, Honoraria, Research Funding; GlaxoSmithKline (Research support): Consultancy, Honoraria, Research Funding; Roche (Advisory board, research support): Consultancy, Honoraria, Research Funding; Johnson & Johnson (Advisory board, research support): Consultancy, Honoraria, Research Funding. Off Label Use: Otlertuzumab is a CD37-specific therapeutic protein that has shown significant activity in CLL in preclinical and clinical studies. Mato:Emergent Product Development: Honoraria. Awan:Lymphoma Research Foundation - Reseach Funding: Research Funding, Speakers Bureau; Spectrum Pharmaceuticals, Inc. - Speakers bureau: Research Funding, Speakers Bureau. Hill:Teva Pharmaceuticals: Honoraria, Research Funding; Emergent Biosolutions: Honoraria, Research Funding. Stromatt:Emergent Product Development: Employment. Jaeger:Emergent: Honoraria, Research Funding.

TP53 Mutation or Deletion and Efficacy with Single-Agent Lenalidomide in Relapsed or Refractory Chronic Lymphocytic Leukemia (CLL) (CC-5013-CLL-009 Study)

IntroductionRelapsed and refractory (rel/ref) CLL patients have a poor prognosis and limited therapeutic options. The oral immunomodulatory agent lenalidomide has shown single-agent activity in untreated and rel/ref CLL patients, including patients with TP53 deleted/mutated CLL, which is associated with poor outcomes in response to chemotherapy. Lenalidomide targets the microenvironment to restore functional immunity to drive the elimination of leukemic cells. Here we report the efficacy of lenalidomide monotherapy in high-risk rel/ref CLL patients. MethodsThis phase 2, multicenter, double-blind, parallel-group, adaptive-design study evaluated the safety and efficacy of lenalidomide in rel/ref CLL. Patients had active disease per iwCLL guidelines and received ≥1 prior line of therapy, including purine-analogs or bendamustine. Patients were randomized 1:1:1 to receive oral lenalidomide at a starting dose of 5, 10, or 15 mg daily at their initial 28-day cycle, followed by intra-patient dose escalation in 5-mg increments every 28 days, up to 25 mg/day as tolerated. Samples to determine genetic status were collected at entry and analyzed centrally at Ulm University. ResultsA total of 104 patients were enrolled; 103 patients were dosed and median age was 64.0 (32-81). Del(17p) and TP53 mutation were found in 23/93 (25%) and 36/96 (38%) patients, respectively; 20 patients with TP53 mutation did not have del(17p).Del(17p) patients were more likely to have TP53 mutation (77% vs. 26%), age >65 years (64% vs. 43%), high risk/Binet C (64% vs. 33%), and platelet count <150,000/mm3 (77% vs. 50%); del(17p) patients were less likely to have del(11q) (14% vs. 36%). TP53 mutation was significantly associated with del(17p) (47% vs. 8%), age >65 years (67% vs. 38%), high risk/Binet C (53% vs. 33%), and platelet count <150,000/mm3(75% vs. 45%).The median number of treatment cycles was 6.6 vs. 11.1 for patients with and without del(17p), and 8.7 vs. 10.2 for patients with and without TP53 mutations. At a median follow-up of 47.6 weeks, the overall response rate (ORR) was 44% for all patients. There was a trend toward significance for ORR in patients with vs. without del(17p) (22% vs. 46%; p = 0.051). The ORR was 36% vs. 42% for patients with and without TP53 mutation (p = 0.669). A multivariate analysis for overall response in all evaluable patients was completed on the baseline characteristics of del(11q), del(17p), TP53 mutation, unmutated IGHV, high-risk disease (Binet C or Rai high-risk), B2M (≤4 vs. >4 mg/dL), age (<65 vs. ≥65 years), number of prior treatments (<3 vs. ≥3), bulky disease, and refractory to purine-analogues in a previous regimen. None of the baseline characteristics showed statistically significant impact on response.Median progression-free survival (PFS) was 45.1 weeks (95% CI 24.1-89.3) in all patients, 21.4 weeks (95% CI 12.4-117.6) in patients with del(17p), and 66.3 weeks (95% CI 31.4-98.4) in patients without del(17p) (p = 0.157). Median PFS in patients with and without TP53 mutation was 47.6 weeks (95% CI 20.6-117.6) vs. 41.1 weeks (95% CI 21.4-85.1) (p = 0.584).Median overall survival (OS) was 151.6 weeks (95% CI 90.3-not reached [NR])in the overall population, 80.9 weeks (95% CI 66.6-NR) in patients with del(17p), and 151.6 weeks (95% CI 90.7-NR) in patients without del(17p) (p = 0.464). Median OS in patients with and without TP53 mutation was 90.3 weeks (95% CI 68.1-NR) vs. 151.6 (95% CI 92.1-NR) (p = 0. 274).A multivariate analysis for PFS and OS was performed, including number of prior treatments. Median PFS was 89.3 weeks (95% CI 31.4-117.6) for patients with <3 prior therapies, and 31.9 weeks (95% CI 20.6-77.6) for patients with ≥3 prior therapies (p = 0.116). OS was NR for patients with <3 prior therapies, and 92.1 weeks (95% CI 68.1-151.6) for patients with ≥3 prior therapies (p = 0.012). No other variable showed statistically significant impact on PFS or OS. ConclusionSingle-agent lenalidomide treatment led to similar responses, PFS, and OS in rel/ref CLL patients with or without TP53 mutations, indicating that lenalidomide activity in rel/ref CLL patients is not affected by loss of functional TP53. Despite differences in baseline characteristics (more high-risk features in the TP53-mutated subgroup), outcomes with lenalidomide treatment were comparable, and the ORR of 44% is a promising result for this heavily pretreated rel/ref CLL population with poor prognosis. Disclosures:Wendtner:Celgene: Consultancy, Honoraria, Research Funding. Hallek:Celgene: Consultancy, Honoraria, Research Funding. Kipps:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Hillmen:Celgene: Honoraria. Purse:Celgene Corporation: Employment, Equity Ownership. Zhang:Celgene: Employment, Equity Ownership. Mei:Celgene: Employment, Equity Ownership. Stilgenbauer:Celgene: Honoraria, Research Funding.

Emerging role of kinase-targeted strategies in chronic lymphocytic leukemia

AbstractChronic lymphocytic leukemia (CLL) is a malignancy of mature B cells that depend on host factors in the tissue microenvironment for survival and proliferation. In vitro, CLL cells rapidly undergo apoptosis unless microenvironmental factors are provided that support their survival. Signaling pathways activated in the microenvironment in vivo include the B-cell receptor (BCR) and NF-κB pathways. Thus, CLL is a disease “addicted to the host” and is dependent on pathways that promote normal B-cell development, expansion, and survival; this is particularly true in the case of the BCR signaling cascade. Small-molecule inhibitors of kinases that are essential for BCR signal transduction abrogate the stimulating effects of the microenvironment on CLL cells. The orally administered tyrosine kinase inhibitors fostamatinib and ibrutinib and the phosphatidylinositol 3-kinase inhibitor GS-1101 have induced impressive responses in relapsed and refractory CLL patients, mostly with moderate side effects. Reductions in lymphadenopathy and splenomegaly are seen within weeks and are frequently accompanied by a transient rise in absolute lymphocyte count that is asymptomatic and probably the result of changes in CLL cell trafficking. This review discusses the biologic basis for kinase inhibitors as targeted therapy of CLL and summarizes the exciting early clinical experience with these agents.

Phase 1b Study of TRU-016, an Anti-CD37 SMIP™ Protein, in Combination with Bendamustine Vs Bendamustine Alone in Relapsed Chronic Lymphocytic Leukemia

Abstract 1795 Background: CD37 is a tetraspanin protein expressed on the surface of normal and transformed B-cells across a wide range of maturational stages and demonstrates death signaling via SHP1. TRU-016 is a novel humanized anti-CD37 SMIP™ (mono-specific protein therapeutic) that has shown significantly greater direct killing of CLL cells than rituximab and greater Fc mediated cellular cytotoxicity of CLL cells than either alemtuzumab or rituximab in pre-clinical models. In preclinical in vitro and in vivo models significant activity of TRU-016 with bendamustine was observed. In a phase 1 study in 57 relapsed and/or refractory CLL patients treated with TRU-016, the maximum tested dose (20 mg/kg) was well tolerated. Given the single-agent clinical activity of TRU-016 and synergistic or additive effect of TRU-016 with multiple agents in preclinical models, this trial of TRU-016 with bendamustine was conducted to establish the maximum tolerated dose, overall safety, and clinical activity of TRU-016 in combination with bendamustine in patients with relapsed CLL. Methods: Patients with relapsed CLL who had 1–3 prior treatments (without prior bendamustine), adequate organ function, ECOG ≤2, and absolute neutrophil count ≥1200/μL were eligible. Patients received TRU-016 (15 or 20 mg/kg) weekly by IV infusion for two 28-day cycles then every 14 days for four 28-day cycles. Bendamustine (70 mg/m2) was administered on Days 1 and 2 of each cycle by IV infusion for up to six 28-day cycles. Safety was evaluated using CTCAE and IWCLL 2008 Grading scale for Hematologic Toxicity in CLL Studies. Response was determined using the 2008 IWCLL Criteria. Results: 12 patients have been treated to date (6 pts at each dose level). Patient characteristics: median age 67 yrs (range, 54–81), 7 patients received 1 and 2 patients received 2 prior regimens, 5 patients had bulky nodes ≥7 cm, 1 had del(17p13.1). All patients were intermediate to high-risk (Rai I/II=75%, III/IV=25%). The most frequent adverse events (&gt;2 pts) were: nausea (8); neutropenia (6); anemia, pyrexia, decreased appetite, hypokalemia, headache, cough (4 each); febrile neutropenia, constipation, diarrhea, dry mouth (3 each); and chills (2). The only grade 3/4 adverse events that occurred in &gt;2 patients were: neutropenia (6) and febrile neutropenia (3). There were 10 serious adverse events reported on 4 patients. Related serious events were febrile neutropenia in 2 patients and autoimmune hemolytic anemia (pre-existing) and urinary tract infection in 1 patient each. There was no apparent dose relationship to adverse event frequency. The best overall response rate at any time according to the investigator was 100% (4 CRs, 2 at each dose) with 9/12 responses occurring at cycle 1. CT scan data, which is necessary to determine IWCLL response, is pending and will be presented. There was no apparent dose relationship to response. Conclusions: TRU-016 in combination with bendamustine was well tolerated and showed a positive response. A randomized trial of TRU-016/bendamustine vs. bendamustine alone is ongoing. Disclosures: Jaeger: Emergent Product Development: Consultancy, Research Funding. Rifkin:Millennium Pharmaceuticals, Inc./Takeda: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; ONYX: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; INCYTE: Speakers Bureau; Amgen: Speakers Bureau. Stromatt:Emergent Product Development: Employment.

Abstract 2011: A novel bcl-2/xL inhibitor induces tumor cell apoptosis and potentiates the activity of standard chemotherapeutics

Abstract Tumor cells often evade apoptosis by tipping the balance of Bcl-2 family and BH3-only protein expression towards a more anti-apoptotic environment, thus providing a survival advantage as well as a means of resistance to anti-cancer therapies. Indeed, over-expression of Bcl-2 family members has been shown to confer chemo-resistance in multiple tumor types. One potential result of this survival adaptation, however, is that the tumor cells become dependent on said Bcl-2 family member and hence increasingly vulnerable to targeted pro-apoptotic BH3 mimetics. Clinical validation of this concept has been demonstrated with the Bcl-2/xL inhibitor Navitoclax, where as a single agent in phase I/II trials in relapsed, refractory CLL patients, a 20% response rate was observed. Due to the potential that directly targeting the apoptotic machinery holds in treating cancer, developing BH3 mimetics that easily and effectively combine with standard chemotherapies is an attractive proposition. With this in mind, we have discovered and characterized a novel small molecule BH3 mimetic with nanomolar affinity for Bcl-2 and Bcl-xL. Here we characterize the in vitro and in vivo activity of this inhibitor in a variety of cell types. In addition to exhibiting potent single agent anti-cancer activity, it also potentiates the effectiveness of standard chemotherapeutic agents. Importantly, it also induces regression of sensitive tumors in vivo after a single IV bolus dose. Overall, this novel Bcl-2/xL inhibitor represents an exciting tool to further explore the mechanistic rationale for using potent BH3 mimetics as a foundation therapy across a broad range of tumor types where resistance to standard therapies arises from deregulation of the Bcl-2 family of anti-apoptotic proteins. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2011. doi:1538-7445.AM2012-2011

Heat-Shock Protein Signature Is Associated with Refractory Chronic Lymphocytic Leukemia Cells From Different In Vivo Microenvironments,

Abstract 3866Chronic Lymphocytic Leukemia (CLL) is a monoclonal B-cell disorder with accumulation of leukemic cells in peripheral blood, bone marrow and lymphoid organs. It presents with a heterogeneous clinical course. Many patients survive long periods of time without any need for treatment, whereas other patients show resistance to treatment or relapse soon after administration of therapy. Although some prognostic markers such as mutational status of immunoglobulin variable heavy chain, chromosomal abnormalities, CD38 levels, or ZAP-70 expression may help predict at initial diagnosis which patients will have more aggressive disease, the exact factors that can determine chances of remission in CLL are still not clear, making treatment challenging. Furthermore, CLL remains an incurable disease, necessitating a way for controlling its progression. Identifying novel molecular signatures associated with refractory CLL disease may help devise targeted treatment strategies and thus may prolong survival times and prevent the progression of CLL in relapsed patients. Considering this, we performed gene expression profiling (GEP) on peripheral blood (PB), bone marrow (BM) and lymph node (LN) samples collected at the time of diagnosis. We divided CLL samples into 3 groups based on their response to treatment; i) Stable CLL group: asymptomatic patients requiring no treatment, ii) Treated but stable CLL group: patients required treatment but had stable disease for at least one year after the end of the treatment cycle, and iii) Relapsed CLL: patients who relapsed within a year of end of the treatment cycle. Significance analysis of microarray (SAM) revealed that the heat-shock protein (HSP) signature (HSJ2, HSP70, HSP90, HSP60, HSP10, HSP 105, HSP40, HSP27, HSPA2, HSJ1, HSF4, HSPCA), BCR signaling pathway (JUN, NFATC4, NFKBIE, PPP3CB, TRAF3, CD81, CCT4), activation markers (CD81, CD83) and MMPs (MMP3, MMP9) were overexpressed in relapsed PB-CLL (n=3) compared to stable PB-CLL (n=6) and treated but stable PB-CLL (n=10). Overexpression of heat-shock protein signature genes were further observed in additional relapsed PB-CLL (n=6) group compared to other two PB-CLL (n=22) group. Interestingly, the HSP signature was consistently overexpressed in relapsed BM-CLL (n=6) and LN-CLL (n=12) compared to stable and treated but stable BM-CLL (n=11) and LN-CLL (n=3) groups. HSPs are considered chaperones of tumorigenesis and known to enhance survival, migration, and proliferation of tumor cells which may contribute to relapse in patients. Furthermore, the HSPs genes (HSP90 and HSP70) were significantly overexpressed in LN-CLL as compared to PB-CLL which implies important role of the microenviroment in rendering CLL refractory. To investigate the link between the expression of the individual genes with the aggressiveness of the disease, Kaplan-Meier log-rank tests were performed. We found that the higher expression of HSP90A, HSP90B, HSJ, and MMP9 were significantly (p<0.05) associated with shorter time to treatment. In summary, our study suggests that HSP genes are overexpressed in refractory CLL patients and thus are promising targets to improve clinical outcome. Disclosures:No relevant conflicts of interest to declare.

Improved Outcome in Unrelated Donor Recipients After Allogeneic Hematopoietic Stem Cell Transplantation for Patients with Advanced Relapse / Refractory Chronic Lymphocytic Leukemia

Abstract 4474Chronic lymphocytic leukemia (CLL) remains incurable with standard therapies and a large number of patients become refractory to treatment or develop toxicities that prevent further treatment and contribute to their mortality. Allogeneic hematopoietic stem cell transplantation (Allo-HSCT) is the only known curative treatment for CLL. However, management and prevention of toxicities as well as patient / donor selection remain as the most significant challenges that limit the success of this treatment.In this observational study we evaluated the efficacy and tolerability of Allo-HSCT in relapse / refractory patients with CLL and correlated the clinical outcome with prognostic markers as well as the donor type.All patients were enrolled at the University of California, San Diego - Moores Cancer Center between November 1998 and July 2009. Eighteen patients with progressive relapse / refractory CLL were included (9 males [8 Caucasian, 1 Hispanic], 9 females [8 Caucasian, 1 Hispanic]), nine patients received stem cells from related donors and the other nine from unrelated donors. The median age was 50 years (age range, 26–71), median interval between diagnosis and Allo-HSCT was 83 months, and the median number of regimens prior to transplant was 5.3 (range, 2–11).Five out of eighteen patients were fludarabine-refractory and one was not evaluable (NE). All patients had a high-risk disease at the time of transplantation, with advanced stages according to Rai classification, and high prevalence of IgVH unmutated state, as well a high expression of CD38 and ZAP-70. Six patients had cytogenetic abnormalities, ten were normal and two were not evaluated. The most common pre-transplantation cytogenetic alteration was deletion 17p (33.3%), followed by deletion 13q (22.2%) and deletion 11q (22.2%). Prior to Allo-HSCT, three were in CR, ten patients were in PR and five had PD. CMV serologic reactivity was present in 67% of patients. Seventeen patients received non-myeloablative conditioning regimens and one patient was treated with a myeloablative regimen.Disease status was assessed 30 and 100 days, 1, 2 and 3 years post Allo-HSCT. 13 out of 18 patients showed response to Allo-HSCT (6 PR and 7 CR) with the remaining patients having PD. Two patients had acute GVHD and 3 patients had extensive chronic GVHD. We observed no differences in the rate of GVHD in unrelated vs. related donor recipients (p=0.1). Five patients died, three due to progression of the disease, one due to post-transplant organ failure and one for acute GVHD. Survival analysis showed a median OS of 60.1 ± 3.9 months. We observed a significant difference in terms of OS in patients who had unrelated vs. related transplants (HR=3.1 [95% CI 1.5–18.7]). We observed no difference in survival of patients discriminated by the IgVH mutational status (HR =1.8 [95% CI 0.2–17.5]) or ZAP-70 expression (HR =1.0 [95% CI 0.9–1.1]). The median progression free survival was 56 months.Five of the 18 patients were retreated with donor lymphocyte infusions or a second Allo-HSCT after relapse or lost of chimerism. Three of these patients died of PD while the other two are still alive.In conclusion, Allo-HSCT induces long-term remission and survival in high-risk relapse / refractory CLL patients including patients with multiple previous treatments and progressive disease status at the time of transplantation. The response to Allo-HSCT is independent of prognostic markers such as IgVH and ZAP-70 expression. Patients that received stem cells from unrelated donors have a better clinical outcome including OS with no evidence of increased GVHD. Disclosures:No relevant conflicts of interest to declare.

Phase 1 Study of Tru-016, An Anti-CD37 SMIP™ Protein in Naïve and Relapsed and/or Refractory CLL Patients

Abstract 1792 Background:CD37 is a tetraspanin protein expressed on the surface of normal and transformed B-cells across a wide range of maturational stages. TRU-016 is a novel humanized anti-CD37 protein therapeutic that has demonstrated significantly greater direct killing of CLL cells than rituximab and greater Fc-mediated cellular cytotoxicity of CLL cells than either alemtuzumab or rituximab in pre-clinical models. A recent phase 1 study in 57 relapsed and/or refractory CLL patients treated with TRU-016 found the maximum tested dose (20 mg/kg) safe and tolerable. We now report an extension of that study to better characterize the safety and activity of this agent. Methods:Patients with naïve and relapsed/refractory CLL, adequate organ function and absolute neutrophil count >500/μL were eligible. Patients received 10, 20, or 30 mg/kg of TRU-016 administered IV once a week for 8 doses followed by 4 monthly doses. Patients treated in the dose escalation phase of the study were eligible for consideration as “roll-over” patients to be retreated in this expanded cohort study. Responses were determined using the 1996 NCI criteria. Results:26 patients (7 naïve, 16 relapsed/refractory, and 3 roll-over CLL) received TRU-016 in the expanded cohort with 12 patients in active follow up at the time of abstract submission. Patient characteristics: median age 69.5 yrs (range, 39–75), median prior regimens 2 (1–11), refractory to anti-CD20 therapy 33% (4/12), refractory to last regimen 35% (6/17) and bulky nodes ≥5 cm 50% (11/22).There were 3 subjects with dose limiting toxicities; none occurred more than once in a dose cohort. A maximum tolerated dose was not identified. The most frequent adverse events were neutropenia, chills, diarrhea, and fatigue at 21% each and thrombocytopenia, nausea, back pain, cough, and headache at 17% each. The most frequent Grade 3 or 4 adverse events were: infection in 4 patients, neutropenia in 3 patients and febrile neutropenia in 2 patients; 3 of the infections were in patients with G3/4 neutropenia or febrile neutropenia. There were 9 serious adverse events reported by 4 patients, with 2 patients having events considered possibly related to TRU-016 (febrile neutropenia in one patient; pneumonia, atrial fibrillation and diarrhea in a second).Lymphocyte reduction of ≥50% was observed in 81% (17/21) of naïve and relapse/refractory CLL and 33% (1/3) of roll-over CLL patients. Lymph node reduction of ≥50% by CT scan measurements was seen in 46% (6/13).The ORR was 86% (6/7 PR) in naive CLL and 17% (3/17 PR) in relapsed/refractory CLL. Response in relapsed CLL patients was limited to those with 1–2 prior treatments, 33% (3/9 PR), as was previously observed in the dose escalation phase of the study. Conclusions:TRU-016 treatment has a favorable safety profile and the MTD has not been reached. Clinical activity has been observed with a reduction in lymphocyte count and by a reduction in lymph node size by CT scans. The greatest response was seen in naïve CLL patients. Given the single-agent clinical activity of TRU-016 and synergistic or additive effect of TRU-016 with multiple agents in pre-clinical models, a combination trial of TRU-016 with bendamustine has been initiated in relapsed CLL patients. Updated results from ongoing patient follow-up will be presented at the meeting. Disclosures:Gopal:Millenium:. Stromatt:Emergent Product Development Seattle, LLC, Seattle, WA: Employment.