ALBERT

Description: The tumor microenvironment is characterized by multiple interactions of transformed malignant cells with non-transformed stroma or immune cells. Particularly macrophages play a pivotal role in this network determining disease progression and therapeutic response. In previous work we could show that macrophages are an essential mediator of therapeutic response in the synergistic response to the administration of the chemoimmunotherapy. The combination treatment strongly increases tumor clearance by repolarization of tumor-associated macrophages from a suppressive to an activated phenotypic state. Here, se analyzed the functional implications of the DNA damage response pathway for the generation of the ASAP and synergy in chemoimmunotherapy. We attempted to disrupt DNA damage response pathway in lymphoma cells generated from the hMB humanized Double-Hit-Lymphoma model by knock-down of key elements like ATM, DNA-PK or p53. We could prevent the formation of the stimulatory cytokine release effect on macrophage phagocytic capacity. Here, p53 status displays a key regulatory role on macrophage mediated malignant cell depletion. TP53 activation via Nutlin-3A treatment of lymphoma cell enhances ADCP in in p53 wild-type cells, while not displaying enhancement in p53-deficient lymphoma cells. Addressing the treatment in vivo using the hMB model for modeling of Double-Hit Lymphoma bearing mice we could demonstrate diminished ASAP and ADCP for p53-deficient lymphoma treated with cyclophosphamide in vivo. Using primary human CLL patient cells comparing both wild-type and p53-deficient status, the p53-deficient CLL cells failed to induce the stimulatory, cytokine-mediated effect on macrophage phagocytosis in response to combination treatment as seen with the p53 proficient CLL cells. Using a CLL mouse model by treating Eµ-TCL1/p53wt/wt as well as Eµ-TCL1p53-/- mice we could show that low-dose cyclophosphamide treated Eµ-TCL1p53-/- mice failed to induce an antibody mediated stimulatory effect on macrophage phagocytosis capacity as seen with Eµ-TCL1/p53wt/wt mice. A similar effect was seen for primary multiple myeloma cells in response to daratumumab displaying significantly less ADCP of p53-deficient multiple myeloma cells. As for the mechanism of p53-defined interaction within the tumor microenvironment we subjected p53-wild-type and p53-deficient lymphoma cells for proteomic analysis. Here we could identify a significantly deregulated protein expression profile for exosome release in p53 deficient lymphoma cells. Verifying this finding by assessing size and frequency exosomes released by respective cell populations we reveal profound changes induced by p53 loss. Furthermore we could identify up-regulation of PD-L1 in p53-deficient cells. Blocking this checkpoint in the ADCP assay could significantly restore phagocytic capacity of macrophages and overall therapeutic response. In this work, we indicate that p53 functional status determines phagocytic function and therapeutic response to monoclonal antibodies. We can verify this finding in independent models in vitro and in vivo as in primary CLL and myeloma patient cells. We furthermore identify altered exosome profiles and checkpoint inhibitor expression in lymphoma cells as underlying mechanism of macrophage modulation. Finally our ongoing research offers possibility to reveal and tailor new combinatorial treatment approaches for chemo-refractory patients. Disclosures Wendtner: Genetech: Consultancy, Honoraria, Other: travel support, Research Funding; GlaxoSmithKline: Consultancy, Honoraria, Other: travel support, Research Funding; Gilead: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Other: travel support, Research Funding; Pharmacyclics: Consultancy, Honoraria, Other: travel support, Research Funding; Abbvie: Consultancy, Honoraria, Other: travel support, Research Funding; MorphoSys: Consultancy, Honoraria, Other: travel support, Research Funding; Gilead: Consultancy, Honoraria, Other: travel support, Research Funding; Roche: Consultancy, Honoraria, Other: travel support, Research Funding; Mundipharma: Consultancy, Honoraria, Research Funding. Hallek:Janssen: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Mundipharma: Honoraria, Research Funding; Pharmacyclics: Honoraria, Research Funding; Roche: Honoraria, Research Funding; Gilead: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding. Pallasch:Gilead: Research Funding.

Description: Background: Chromosome 14q32 rearrangements involving the immunoglobulin heavy chain gene (IGH) affect less than 5% of chronic lymphocytic leukemia (CLL) patients. Their clinical course is aggressive and the outcome, worse than other CLL subtypes (Cavazzini et al, 2008; Gerrie et al, 2012). However, the biology of CLL showing IGH rearrangements (CLL-IGHR) is not completely defined. The identification of novel recurrent mutations in CLL by next generation-sequencing (NGS) has offered a more comprehensive view into the genomic landscape of the disease and improved the prognostication of CLL. Thus, mutational analysis might be especially useful in those patients with uncertain prognosis, such as those carrying IGH rearrangements. Aim: To analyze the mutational profile of CLL-IGHR patients by targeted NGS in order to improve our understanding of the genetic underpinnings of this subgroup. Methods: The study was based on 899 CLL patients, well characterized at cytogenetic, biological and clinical level, forty-two of them (4.7%) showing IGH rearrangements. Targeted NGS was performed in 231 CLL samples: 117 with 13q deletion, 27 with 11q deletion, 26 trisomy 12, 42 showing IGH rearrangements and the remaining 19 without any cytogenetic alteration. CD19+ B cells were isolated and DNA extracted. SureSelectQXT targeted enrichment technology and a custom-designed panel (MiSeq, Illumina), including 54 CLL-related and recurrent mutated genes, was carried out. The panel yielded 100x or greater coverage on 97% of the genomic regions of interest and the mean coverage obtained was 600x. Mutations were detected down to 3% allele frequency. Results: The mutational analysis of CLL-IGHR patients identified a total of 72 mutations in 32 genes. Seventy-one percent of patients (30/42) harbored at least one mutation. The most frequently mutated genes in this cohort were NOTCH1 (28.6%), POT1 (14.3%), TP53 (9.5%), SF3B1 (7%), BRAF (7%), EGR2 (7%), IGLL5 (7%) and MGA (7%), followed by BCL2, HIST1H1E and FBXW7 (4.8%), uncommonly mutated genes in CLL at these frequencies (Table 1). In fact, mutations in NOTCH1, BRAF, EGR2, BCL2, HIST1H1E and FBXW7 were significantly associated with CLL-IGHR patients (p=0.013, p=0.003, p=0.021, p=0.038, p=0.038 and p=0.021 respectively). In terms of time to the first therapy (TFT), CLL-IGHR had an intermediate-negative impact (median TFT=24 months) compared to the presence of cytogenetic alterations associated with good prognosis such as 13q deletions (median TFT〉120 months; p

Description: Background AFM13 is a bispecific, tetravalent NK cell-engaging antibody construct binding to CD30 on CD30+ tumor cells and CD16A on NK cells. By engaging CD16A-positive NK cells, AFM13 leads to NK cell-mediated killing of CD30-positive lymphoma cells (Reusch et al., 2014) making it an attractive agent to target classical Hodgkin lymphoma (HL). Pembrolizumab is a PD-1 blocking antibody which has shown high single-agent response rates in patients (pts) with relapsed/refractory HL (RRHL; Armand et al., 2016, Chen et al., 2017). AFM13 has shown clinical activity in RRHL as a single agent in a preceding Phase 1 study (Rothe et al., 2015). Preclinical in vivo data of the combination of AFM13 with PD-1 blockade showed synergistic activity and the potential for induction of cross-talk between innate and adaptive immunity (Zhao et al., 2016). We hypothesize that the combination of the two agents could improve outcomes in pts with RRHL. Methods This Phase 1b study is evaluating the safety and tolerability of the combination of AFM13 with pembrolizumab (Keytruda) as salvage therapy after failure of standard therapies including brentuximab vedotin (BV) in HL (NCT02665650). Pts receive escalating doses of AFM13 in combination with pembrolizumab at a dose of 200 mg flat administered every 3 weeks following a classical 3+3 design, followed by enrollment into an extension cohort at the maximum tolerated dose (MTD)/maximum administered dose (MAD). Response assessment is performed every 12 weeks by PET/CT according to the Lugano Classification (Cheson et al., 2014). The main objectives of the study is to ascertain the MTD/MAD along with the preliminary efficacy of the combination. Results As of June 29, 2018, 30 pts have been enrolled into the study. The median age is 34 years (range, 18-73), with a median of 4 (range 3-7) prior lines of therapy. All pts had relapsed or refractory disease (43% relapsed, 57% refractory) and had failed standard treatments including BV and 43% of pts (13/30) had BV as their latest therapy. Thirty seven percent (11/30) had undergone prior autologous stem cell transplantation. All 30 pts have completed the 6-week dose-limiting toxicity (DLT) observation period. Twelve pts were enrolled into the dose escalation cohorts (Cohorts 1 (n=3), 2 (n=3), and 3 (n=6)) and 18 into the Extension Cohort, with a total of 24 patients treated at the MAD (dose level 3). One DLT was observed in Cohort 3 (missing ≥25% of AFM13 during the DLT period) and another observed in the Extension Cohort (G4 infusion-related reaction; IRR). The most common related adverse events (AEs) were IRRs (80%), rash (30%), pyrexia (23%), nausea (23%), diarrhea (20%), fatigue (17%), headache (17%), increased aspartate aminotransferase (13%), and increased alanine aminotransferase (10%). Treatment related G3/4 AEs included IRRs (13%), elevated AST (3%), gastritis (3%), hypotension (3%), nausea (3%), neutropenia (3%), and vomiting (3%). The majority of IRRs were manageable with standard of care measures and did not lead to treatment discontinuations. Included in the efficacy analysis were the best response from 29 evaluable pts who had at least one post-baseline disease assessment as of the data cutoff on June 29, 2018. The overall response rate (ORR) and complete response (CR) rate for evaluable pts treated at the dose and schedule chosen for expansion (n=23; Cohort 3 and Extension Cohort) were 87% and 35% by the investigator-confirmed assessment, respectively. Independent assessment resulted in an ORR of 87% and CR rate of 39% for these pts. Updated data for all 30 patients will be presented at the meeting. Conclusions The combination of AFM13 and pembrolizumab is a well-tolerated salvage therapy in pts with RRHL. IRRs were the most frequently observed adverse events; however, most of these events were of mild or moderate severity and manageable. Both the ORR and CR rate compare favorably to monotherapy pembrolizumab in a similar RRHL population (Chen et al., 2017). The combination of AFM13 and pembrolizumab could be a potential new therapeutic option for HL patients. Disclosures Bartlett: Immune Design: Research Funding; Affimed: Research Funding; Bristol-Meyers Squibb: Research Funding; Merck & Co: Research Funding; Pharmacyclics: Research Funding; Celgene: Research Funding; Pharmacyclics: Research Funding; Genentech: Research Funding; Forty Seven: Research Funding; Novartis: Research Funding; Novartis: Research Funding; Millennium: Research Funding; ImaginAB: Research Funding; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Research Funding; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Seattle Genetics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astra Zeneca: Research Funding; Acerta: Membership on an entity's Board of Directors or advisory committees. Chen:Affimed: Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Genentech Inc.: Consultancy; Seattle Genetics: Consultancy, Honoraria, Research Funding, Speakers Bureau; Merck & Co., Inc.: Consultancy, Research Funding, Speakers Bureau; Millennium Pharmaceuticals: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding. Domingo-Domenech:Affimed: Research Funding. Forero-Torres:Affimed: Research Funding. Garcia-Sanz:Affimed: Research Funding. Devata:Affimed: Research Funding. Rodriguez Izquierdo:Affimed: Research Funding. Lossos:Affimed: Research Funding. Reeder:Affimed: Research Funding. Sher:Affimed: Research Funding. Choe-Juliak:Affimed: Employment. Prier:Affimed: Research Funding. Schwarz:Affimed: Employment. Strassz:Affimed: Employment. Alland:Affimed: Employment. Ansell:Bristol-Myers Squibb: Research Funding; Celldex: Research Funding; LAM Therapeutics: Research Funding; Trillium: Research Funding; Pfizer: Research Funding; Regeneron: Research Funding; Seattle Genetics: Research Funding; Merck & Co: Research Funding; Affimed: Research Funding; Takeda: Research Funding.

Description: Background: Nivolumab, an immune checkpoint inhibitor, has been tested in patients with classical Hodgkin lymphoma (cHL) who failed standard treatment options and has demonstrated remarkable activity with acceptable safety profile in clinical trials. After the impressive results of nivolumab phase I study, a significant number of patients were granted early access to nivolumab through a Name Patient Program (NPP) or compassionate use in Spain. Demonstrating that results of nivolumab use in real-life are similar to those in clinical trials is of major clinical relevance. Objective: The aim of this retrospective study was to analyze the efficacy and safety profile of nivolumab for the treatment of relapsed/refractory (RR) cHL in a real-life context. Methods: We retrospectively collected data from 34 GELTAMO centers. Eligible patients included RR cHL patients treated with at least one cycle of nivolumab. The primary end-point was to describe the overall response rate (ORR). Secondary objectives were to assess the complete response rate (CR), safety of nivolumab, and clinical outcomes (overall survival [OS], and progression free survival [PFS]). Results: Between September 2015 and May 2018, 74 patients with RR cHL received nivolumab monotherapy dosed at 3mg/kg once every 2 weeks (97%). The median age was 38 years (range 17-78). Patients have received a median of 4 (1-15) prior therapy lines; all but 2 were previously treated with brentuximab vedotin (97%), and 38 (51%) of them underwent a hematopoietic stem cell transplantation (HSCT) (n=33 autologous, autoHSCT, and n=5 allogeneic, alloHCST). Median number of nivolumab cycles was 8 (1-65). Ten (14%) patients are still on treatment. Reasons for nivolumab discontinuation were disease progression in 23/64 (36%), referral to HSCT in 27/64 (42%), adverse events (AE) in 8/64 (13%), patient or physician's decision in 5/64 (8%), and unknown in 1/64 (1%). Treatment related AE were reported in 42/69 (61%). Half of them (21, 30%) were probably immune related AE: grade 1-2, 67% (cutaneous n=5, hepatitis n=3, hypothyroidism n=3, gastrointestinal n=3, suprarenal insufficiency n=1); grade 3-4, 24% (pneumonitis n=2, hepatitis n=1, encephalitis n=1, hypothyroidism n=1); grade 5, 3% (pneumonitis n=1, Stevens-Johnson syndrome + hepatitis + nephritis n=1). ORR was 58% (CR 21/72 patients, partial response [PR] 21/72). Stable disease (SD) was achieved in 9 patients (13%). After an initial response (4 PR and 3 SD), 7 patients developed lymphoma progression. A total of 40 (54%) patients finally underwent HSCT, 4 autoHSCT and 36 alloHCST. AlloHSCT was performed after a median of 63 days (41-115) and 8 patients received prior salvage therapy. Complications after alloHSCT consisted of non-infectious fever requiring steroid treatment in 13 (36%), acute graft-versus-host disease in 19 (53%) (2 of them grade 3-4, 1 death), hepatic venocclusive disease in 2 (6%, 1 death), and non-infectious pulmonary complications in 2 (6%). Five (14%) patients died due to transplant complications. At the last follow-up, all autoHSCT patients and 23/36 alloHSCT were in CR. The 2-year OS for the whole series (n=74) was 54% (median not reached). After a median follow-up of survivors patients of 12.5 months (1-31), 29 (39%) were alive in CR. Conclusions: Our real-life experience confirms the efficacy of nivolumab in very heavily pretreated cHL patients with an ORR of 58%. The safety profile of our cohort is comparable with that previously reported in clinical trials with manageable side effects and low treatment related mortality. In our study the percentage of patients who bridged to transplantation was significantly higher to that previously reported indicating this preference for Spanish physicians. AlloHSCT post-nivolumab showed encouraging results and toxicity seemed comparable to that previously described with other treatment regimens. Authors thank Bristol Myers Squibb for its support in this study. Disclosures Martinez: BMS: Research Funding; Takeda: Consultancy. García-Sanz:Janssen: Consultancy, Honoraria, Other: Travel, Accommodations, Expenses; Takeda: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Honoraria; Hospira: Research Funding; Pharmacyclics: Research Funding; Spanish Government: Research Funding; Gilead: Research Funding; Amgen Inc.: Research Funding; Incyte: Consultancy.

Description: Natural killer (NK) cells hold great promise as a source for allogeneic cell therapy against hematological malignancies, including acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). NK cell recognition of allogeneic tumors is strictly regulated by inhibitory killer cell immunoglobulin-like receptors (KIR) that bind to groups of HLA class I alleles. However, KIR expression on NK cells is highly diverse due to variation in gene content, polymorphism and copy number in combination with stochastic expression of the protein in individual cells. As a consequence, the number of efficacious allogeneic NK cells within a product isolated and expanded from random donors can vary a great deal and potentially be negligible. Our group has defined a repertoire of NK cells that is uniquely found in individuals with prior exposure to cytomegalovirus (CMV). Interestingly, these cells were shown to share many attributes usually reserved for adaptive immune cells including increased longevity, memory, and serial killing. We have previously described a 14-day protocol to enrich for adaptive NKG2C+CD57+ NK cells from CMV sero-positive donors with a homogenous expression of one single self-HLA specific KIR (self KIR). Here, we present new data on the GMP-transfer and clinical scale-up of this protocol, providing a route to off-the-shelf adaptive NK cell therapy for refractory high-risk AML/MDS. By screening 〉250 healthy donors, we first established the prerequisites for robust expansion of adaptive NK cells from peripheral blood of CMV+ donors and found that donors with 〉15% pre-existing adaptive NK cells showed efficient expansion of adaptive NK cells (Figure 1A-B). Apheresis products from a pool of pre-screened third-party donors are currently being collected for GMP freezing and use in an off-the-shelf setting intended for HLA mismatched patients to maximize alloreactivity by "missing" self. The GMP compatible protocol led to a robust expansion of clinical doses of self-KIR+ adaptive NK cells, with an average frequency of 60% self-specific KIR+ cells in the end product (Figure 1C-D). Based on the expression of self-KIR the expanded cells were educated, showing large dense-core granules and high levels of granzyme B. Further characterization in CyTOF using 36 phenotypic and functional markers revealed a highly activated state with high expression of DNAM-1 and CD2, which are critical for NK cell adhesion and function (Figure 1E). Notably, the expanded adaptive NK cells were negative for the HLA-E binding inhibitory receptor NKG2A, which is a major check point for T- and NK-cell based therapies. A microchip single-cell imaging platform revealed high serial killing capacity of the expanded adaptive NK cells. In flow cytometry-based killing assays and long-term killing assays this enhanced capacity for serial killing correlated with highly efficient targeting of mismatched PHA blasts (Figure 1F), tumor cell lines (Figure 1G), and MDS blasts. These pre-clinical data demonstrate the feasibility of off-the-shelf therapy with a non-engineered and yet highly specific NK cell population, representing the first route to clinical testing of missing self-recognition as it was originally defined over thirty years ago. Disclosures Valamehr: Fate Therapeutics Inc.: Employment. Alici:Vycellix: Consultancy, Equity Ownership, Patents & Royalties, Research Funding; Intellia: Membership on an entity's Board of Directors or advisory committees. Ljunggren:Fate Therapeutics: Patents & Royalties; Vycellix: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Malmberg:Fate Therapeutics Inc.: Consultancy, Research Funding.