Enhancement Of Antibody-dependent Cellular Cytotoxicity Research Articles

SGNBCMA-001: A phase 1 study of SEA-BCMA, a non-fucosylated monoclonal antibody, in subjects with relapsed or refractory multiple myeloma.

TPS8054 Background: Despite recent advances in treatment, multiple myeloma (MM) remains incurable in most patients. The standard of care for MM includes combinations of agents with different mechansims of action, e.g. proteasome inhibitors (PIs), immunomodulatory drugs (IMiDs), monoclonal antibodies, alkylating agents, and corticosteroids. Sequential lines of treatment typically yield shorter durations of disease control with each subsequent relapse. Frail patients face a particularly high unmet need, given the toxicity profiles of many agents. SEA-BCMA is a humanized non-fucosylated IgG1 monoclonal antibody targeting BCMA, a plasma cell-specific protein that is expressed on MM cells of most patients. Based on preclinical data, SEA-BCMA displays enhanced antibody dependent cellular cytotoxicity through increased FcγRIII binding, antibody dependent cellular phagocytosis, and blocking of BCMA-mediated pro-survival and proliferative signaling. SEA-BCMA is active at 0.03 mg/kg in xenograft models and does not cause adverse effects in preclinical models, supporting clinical investigation for MM. Methods: This is a phase 1, open-label, multicenter, dose-escalation study to evaluate the safety, tolerability, and antitumor activity of SEA-BCMA in adults with relapsed/refractory MM. Enrollment began in November 2018 for adults aged ≥18 years with histologically confirmed MM, Eastern Cooperative Oncology Group performance status of ≤1, and no other therapeutic options available. Prior therapies must include a PI, an IMiD, and an anti-CD38 antibody. Prior receipt of BCMA targeted agents is prohibited. BCMA expression is not required for study entry, but will be tested retrospectively. Dose-escalation is being conducted in ~25 subjects using the modified toxicity probability interval method. During dose-expansion, ~40 subjects will be treated at the maximum tolerated or optimal dose. Responses are assessed per the 2016 International Myeloma Working Group criteria. Biomarker and pharmacokinetic evaluations will be performed on peripheral blood and bone marrow. Clinical trial information: NCT03582033.

Glyco-Engineered Anti-Human Programmed Death-Ligand 1 Antibody Mediates Stronger CD8 T Cell Activation Than Its Normal Glycosylated and Non-Glycosylated Counterparts.

The programmed death 1 (PD-1)/programmed death-ligand 1 (PD-L1) axis plays a central role in suppression of anti-tumor immunity. Blocking the axis by targeting PD-L1 with monoclonal antibodies is an effective and already clinically approved approach to treat cancer patients. Glyco-engineering technology can be used to optimize different properties of monoclonal antibodies, for example, binding to FcγRs. We generated two glycosylation variants of the same anti-PD-L1 antibody: one bearing core fucosylated N-glycans in its Fc part (92%) and its de-fucosylated counterpart (4%). The two glycosylation variants were compared to a non-glycosylated commercially available anti-PD-L1 antibody in various assays. No differences were observed regarding binding to PD-L1 and blocking of this interaction with its counter receptors PD-1 or CD80. The de-fucosylated anti-PD-L1 antibody showed increased FcγRIIIa binding resulting in enhanced antibody dependent cellular cytotoxicity (ADCC) activity against PD-L1+ cancer cells compared to the “normal”-glycosylated variant. Both glycosylation variants showed no antibody-mediated lysis of B cells and monocytes. The non-glycosylated reference antibody showed no FcγRIIIa engagement and no ADCC activity. Using mixed leukocyte reaction it was observed that the de-fucosylated anti-PD-L1 antibody induced the strongest CD8 T cell activation determined by expression of activation markers, proliferation, and cytotoxicity against cancer cells. The systematic comparison of anti-PD-L1 antibody glycosylation variants with different Fc-mediated potencies demonstrates that our glyco-optimization approach has the potential to enhance CD8 T cell-mediated anti-tumor activity which may improve the therapeutic benefit of anti-PD-L1 antibodies.

Abstract CT158: Phase II study of monalizumab, a first-in-class NKG2A monoclonal antibody, in combination with cetuximab in previously treated recurrent or metastatic squamous cell carcinoma of the head and neck (R/M SCCHN): Preliminary assessment of safety and efficacy

Abstract Background Monalizumab is an immune checkpoint inhibitor targeting NKG2A receptors expressed on subsets of tumor-infiltrating cytotoxic CD8 T cells and Natural Killer (NK) cells. NKG2A ligand is HLA-E, a non-classical HLA class I molecule often upregulated in cancer. Preclinical experiments have shown that blocking NKG2A binding to HLA-E may promote NK and T cell anti-tumor responses. NK cell stimulation with a checkpoint inhibitor might also enhance antibody dependent cellular cytotoxicity (ADCC) induced by cetuximab. Although approved in SCCHN after platinum-based therapy, cetuximab has limited activity in that setting (12% response rate). Methods This is a multicenter non-randomized study (NCT02643550). After previous exploration of 5 dose levels of monalizumab (0.4, 1, 2, 4 or 10 mg/kg every 2 weeks) in combination with fixed doses of cetuximab (400 mg/m² load then 250 weekly) using a 3+3 design, the cohort expansion used monalizumab at the highest dose tested (10 mg/kg) and included a futility analysis after the first 11 patients (pts). The trial was open to pts ≥ 18 years old with SCCHN progressing after platinum-based therapy with no more than 2 previous lines, regardless of HLA-E or human papilloma virus status. The primary endpoint for anti-tumor activity was overall response rate per RECIST, assessed every 8 weeks. Pts were treated until disease progression or unacceptable toxicity. Results As of 12/19/2017, 26 pts were enrolled in the expansion part, and 16 pts had a minimum of 16 weeks of follow-up to be evaluable for efficacy. The safety profile was as expected, similar to the single agent experience with either agent. The majority of adverse events (AE) were of Grade 1-2 severity, rapidly reversible and easily manageable, with 3 treatment-related grade 3-4 AE and 1 pt stopped monalizumab due to safety. Median age was 62 years (range: 34-77); 56 % were male; PS was 0 or 1; 4 were HPV+. All 16 pts had received prior platinum-based therapy, 8 prior immune therapy, 2 prior cetuximab with radiation. There were 6 pts with partial responses (PR) (4 confirmed; 2 not yet confirmed) of whom 2 were previously treated with immune therapy and 1 had disease deemed resistant to cetuximab. Median treatment duration for confirmed PR is 25+ weeks (16, 23+, 28+, 35+), 9 pts had stable disease (SD). The study was not stopped for futility and is planned to enroll up to 40 pts. Further follow-up is needed to evaluate duration of response, progression-free and overall survival. Conclusion Preliminary data suggest promising antitumor activity of the combination of monalizumab and cetuximab compared to historical data with single agent cetuximab, with acceptable safety. These encouraging results will need to be confirmed on larger sample size with longer follow up. Citation Format: Roger Cohen, Jérôme Fayette, Marshall Posner, Gautier Lefebvre, Jessica Bauman, Sébastien Salas, Caroline Even, Tanguy Seiwert, Dimitrios Colevas, Antonio Jimeno, Esma Saada, Barbara Burtness, Pascale André, Carine Paturel, Cécile Bonnafous, Anne-Marie Soulié, Anne Tirouvanziam-Martin, Robert Zerbib, Agnès Boyer-Chammard. Phase II study of monalizumab, a first-in-class NKG2A monoclonal antibody, in combination with cetuximab in previously treated recurrent or metastatic squamous cell carcinoma of the head and neck (R/M SCCHN): Preliminary assessment of safety and efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr CT158.

Abstract 3833: SEA-BCMA: A highly active enhanced antibody for multiple myeloma

Abstract Despite a number of new therapies for multiple myeloma (MM) most patients relapse, require multiple lines of therapy, and ultimately succumb to disease. Therapies that are well tolerated and active remain an unmet need. Currently, successful MM therapies combine agents with different mechanisms of action and safety profiles. Therapeutic antibodies have recently altered the MM treatment paradigm, allowing patients to achieve deeper responses with minimal added toxicity. SEA-BCMA is a humanized afucosylated IgG1 antibody targeting BCMA, which shows preclinical evidence of encouraging activity and tolerability. BCMA is expressed at the surface of plasma cells, and induces proliferative signals through the binding of its ligands APRIL and BAFF. SEA-BCMA acts through three mechanisms of action. One, it engages in increased binding to FcγRIII through SEA technology leading to enhanced antibody dependent cellular cytotoxicity. Two, it mediates antibody dependent cellular phagocytosis. Three, it blocks the proliferative signals from BCMA ligand binding. This antibody shows activity in all seven tumor xenograft models tested, inducing tumor delays at doses as low as 0.1mg/kg, and generating prolonged survival and durable regressions with repeat dosing. SEA-BCMA is active on tumor xenografts expressing as few as 2000 surface copies of BCMA antigen. Both effector function and ligand blocking contribute to overall in vivo activity. In the absence of effector cell recruitment, ligand blocking alone can induce prolonged durable regressions. In addition, SEA-BCMA can target MM cells in the presence of soluble BCMA. SEA-BCMA is tolerated up to 100mg/kg in cynomolgus monkey, reflecting lack of toxicity from cellular interactions with the Fc portion of the Ab, since SEA-BCMA does not bind to the cynomolgus monkey antigen. SEA-BCMA displays a 12-day half-life in these animals. In vitro testing with human PBMCs or bone marrow mononuclear cells induced minimal cytokine production in the presence of BCMA target. In summary, SEA-BCMA is highly active and well tolerated in preclinical models and is a strong candidate for treatment of MM patients. Citation Format: Heather Van Epps, Martha Anderson, Changpu Yu, Kerry Klussman, Lori Westendorf, Chris Carosino, Luke Manlove, Julia Cochran, Jason Neale, Dennis Benjamin, Maureen Ryan. SEA-BCMA: A highly active enhanced antibody for multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3833.

Augmentation of antibody-dependent cellular cytotoxicity with defucosylated monoclonal antibodies in patients with GI-tract cancer.

Enhancement of antibody-dependent cellular cytotoxicity (ADCC) with some modalities may be a promising approach to enhance the efficacy of therapeutic monoclonal antibodies (mAbs). It has previously been demonstrated that the removal of fucose from antibody oligosaccharides (defucosylation) leads to augmentation of ADCC activity. To establish clinically relevant evidence of this procedure, the present study evaluated trastuzumab- and cetuximab-mediated ADCC by comparing defucosylated mAbs with conventional mAbs using peripheral blood mononuclear cells (PBMCs). PBMCs were isolated from 20 patients with gastrointestinal tract cancer and 10 healthy volunteers. ADCCs were measured using PBMCs as effector cells and two gastric cancer cell lines as target cells. ADCCs were significantly enhanced with defucosylated mAbs compared with conventional mAbs using PBMC from the healthy donors and patients with cancer. The results confirmed that the cetuximab- and trastuzumab-mediated ADCCs in advanced disease were impaired in comparison to those in early disease or healthy individuals. However, when the defucosylated mAbs were used instead of the conventional mAbs, the ADCC activities in the advanced cases were almost comparable with those in early disease or healthy individuals. Furthermore, the expression of ADCC associated molecules were modified toward immunosuppressive status with a mitogen-activated protein kinase inhibitor in vitro, the conventional cetuximab- and trastuzumab-mediated ADCC was downregulated, and the defucosylated mAbs overcome the downregulation of ADCC. In conclusion, defucosylated therapeutic mAbs may enhance ADCC activities in patients with cancer, which may lead to more effective anti-cancer treatments.

Enhancement of antibody-dependent cellular cytotoxicity of cetuximab by a chimeric protein encompassing interleukin-15

ABSTRACTEnhancement of antibody-dependent cellular cytotoxicity (ADCC) may potentiate the antitumor efficacy of tumor-targeted monoclonal antibodies. Increasing the numbers and antitumor activity of NK cells is a promising strategy to maximize the ADCC of standard-of-care tumor-targeted antibodies. For this purpose, we have preclinically tested a recombinant chimeric protein encompassing the sushi domain of the IL15Rα, IL-15, and apolipoprotein A-I (Sushi-IL15-Apo) as produced in CHO cells. The size-exclusion purified monomeric fraction of this chimeric protein was stable and retained the IL-15 and the sushi domain bioactivity as measured by CTLL-2 and Mo-7e cell proliferation and STAT5 phosphorylation in freshly isolated human NK and CD8+ T cells. On cell cultures, Sushi-IL15-Apo increases NK cell proliferation and survival as well as spontaneous and antibody-mediated cytotoxicity. Scavenger receptor class B type I (SR-B1) is the receptor for ApoA-I and is expressed on the surface of tumor cells. SR-B1 can adsorb the chimeric protein on tumor cells and can transpresent IL-15 to NK and CD8+ T cells. A transient NK-humanized murine model was developed to test the increase of ADCC attained by the chimeric protein in vivo. The EGFR+ human colon cancer cell line HT-29 was intraperitoneally inoculated in immune-deficient Rag2−/−γc−/− mice that were reconstituted with freshly isolated PBMCs and treated with the anti-EGFR mAb cetuximab. The combination of the Sushi-IL15-Apo protein and cetuximab reduced the number of remaining tumor cells in the peritoneal cavity and delayed tumor engraftment in the peritoneum. Furthermore, Sushi-IL15-Apo increased the anti-tumor effect of a murine anti-EGFR mAb in Rag1−/− mice bearing subcutaneous MC38 colon cancer transfected to express EGFR. Thus, Sushi-IL15-Apo is a potent tool to increase the number and the activation of NK cells to promote the ADCC activity of antibodies targeting tumor antigens.

Frequency and proliferative response of circulating invariant natural killer T cells in pediatric patients with malignant solid tumors.

Invariant natural killer T (iNKT) cells play an important role in tumor immunity, enhancing both innate and acquired immunity. We have previously shown the enhancement of antibody-dependent cellular cytotoxicity against neuroblastoma by activated iNKT cells. As a first step towards clinical application, we studied the frequency and proliferative response of circulating iNKT cells in children with and without cancer. Blood samples were collected from 10 patients with pediatric malignant solid tumors and 11 patients with non-neoplastic diseases (control). The frequency of circulating iNKT cells was quantified by flow cytometry. Whole peripheral blood mononuclear cells were then stimulated with α-galactosylceramide (α-GalCer) for 7 days, and the expansion rate of the iNKT-cell fraction was assessed. The frequency of iNKT cells in the patients of the cancer and control group did not differ to a statistically significant extent. The iNKT-cell population increased after α-GalCer stimulation in all cases. The iNKT cells of patients who had undergone intensive chemotherapy also had the potential to expand in vitro. Unlike adult cancer patients, the numbers of circulating iNKT cells were not decreased in pediatric cancer patients. α-GalCer stimulation induced a proliferative response in all of the patients.

Antibody-dependent NK cell control of Plasmodium falciparum infection

Abstract Plasmodium falciparum (P.f) infection is a major cause of morbidity and mortality world-wide where symptoms and death occur as a result of the blood stage of the P.f. life cycle. To date, there is no effective blood stage malarial vaccine. Natural Killer (NK) cells are key players in the control of hematopoietic cancers and viral infections, however their role in blood stage malaria is unknown. We undertook a comprehensive analysis of NK cell phenotype and function in a cohort of subjects from a malaria clinical study in Mali, Africa. Using an unbiased analysis of different NK cell subsets, we found that ‘adaptive’ NK cells (lacking FcR gamma chain) increased with age and correlated with reduced malaria risk. Adaptive NK cells had enhanced antibody dependent cellular cytotoxicity (ADCC) function relative to other NK cell subsets. Antibodies are known to be protective in malaria, however the mechanism of this protection is still being elucidated. We hypothesized the protective mechanism of adaptive NK cells may be adaptive NK cells pairing with malaria antibodies to clear infected RBCs via ADCC. In a parallel study, we have shown that NK cells from healthy US donors can inhibit the growth of Plasmodium falciparum 3D7 strain in vitro in the presence of IgG from malaria-exposed adults. Adaptive NK cells from Mali also have enhanced ADCC to infected RBCs. This work indicates that protective antibodies may pair with the strong ADCC activity of adaptive NK cells to protect human subjects from malaria symptoms. These findings thus open a new avenue for vaccine development exploring antibody-dependent cell mediated killing of malaria parasites.

Updated Results from a Phase II Study of the Fc Engineered CD19 Antibody MOR208 in Combination with Lenalidomide for Patients with Chronic Lymphocytic Leukemia (CLL) and Richter's Transformation or Ibrutinib for Patients with Ibrutinib-Resistant Clones

MOR208 is an Fc engineered CD19 monoclonal antibody with preliminary efficacy in CLL as a single agent (Woyach et al, Blood 2014). Compared to non-engineered antibodies, MOR208 has significantly enhanced antibody dependent cellular cytotoxicity (ADCC), which can be further augmented in vitro with the addition of lenalidomide. Given the potential synergy of these agents, acceptable individual safety profiles, and efficacy of each as a single agent, we chose to combine MOR208 and lenalidomide in patients (pts) with previously treated and previously untreated CLL and in pts who have undergone Richter's Transformation (RT). Because recent data show poor outcomes in pts who relapse after the Bruton's Tyrosine Kinase (BTK) inhibitor ibrutinib and the presence of mutations in BTK prior to relapse, we included a cohort of ibrutinib-treated pts with identified resistance mutations but no clinical relapse where MOR208 was added to ibrutinib.This study is a single institution phase II trial of MOR208 in combination with lenalidomide or MOR208 in combination with ibrutinib with an initial safety run-in as part of each cohort. In combination with lenalidomide, MOR208 was given at a dose of 1 mg/kg on cycle 1 day 1, 9 mg/kg on days 2, 8, 15, and 22 of cycle 1, and day 1 of cycles 2-12. Lenalidomide was started at a dose of 2.5 mg daily on cycle 1 day 8 and given continuously. The dose of lenalidomide could be escalated up to 10 mg daily in pts without toxicity. After 12 cycles, lenalidomide could be continued indefinitely in responding pts. In combination with ibrutinib, ibrutinib was continued at a dose of 420 mg daily, and MOR208 was given at a dose of 1 mg/kg on cycle 1 day 1, 12 mg/kg on days 2, 8, 15, and 22 of cycle 1, then weekly during cycles 2 and 3, and every other week through cycle 12. Toxicity was assessed using the National Cancer Institute's Common Criteria for Adverse Events (AE) v4.0 for non-hematologic toxicity, and IWCLL 2008 guidelines for hematologic toxicity. For pts on MOR208 plus ibrutinib, variant allele frequency of mutant BTK was measured every 3 cycles.In the previously untreated cohort, 11 pts have been enrolled, with a median age of 62 (range 44-75). 1 pt had both del(17p) and del(11q) on FISH. Grade 3/4 adverse events regardless of attribution have been uncommon and have included hypertension (3 pts), infusion reaction (2 pts), and fatigue, neutropenia, colitis, hyperglycemia, dyspnea, and sinusitis (1 pt each). After a protocol amendment augmenting steroid premedication, no further grade 3 infusion reactions were observed.In the previously treated cohort, 11 pts have been enrolled, with a median age of 70 (range 62-75). 2 pts each had del(17p) and del(11q) on FISH, and 8 have Zap-70 methylated disease. Grade 3/4 toxicities regardless of attribution have included neutropenia (6), hyperglycemia (2), hypertension (2), and thrombocytopenia, fatigue, anemia, upper respiratory infection, catheter related infection, hypercalcemia, hypophosphatemia, infection, respiratory failure, and sepsis in 1 pt each.In the RT cohort, 5 pts have been enrolled, with a median age of 60 (range 55-70). Four had previously treated CLL, and one was previously untreated. Three had del(17p) and 2 had del(11q) on FISH. Grade 3/4 toxicities included hyperglycemia (3) and hyponatremia, thrombocytopenia, and neutropenia in 1 pt each.In the cohort of pts with molecular progression on ibrutinib, 7 pts have been enrolled, with a median age of 62 (range 45-77). Five have del(17p), 1 of whom also has del(11q). Grade 3/4 toxicities have included hypertension (2) and hyperglycemia and hyperuricemia in 1 pt each. Four pts have been on study for at least 3 cycles. One pt had an increase in BTK C481S variant allele frequency (VAF) from 66.3% to 78.8% while the others all have decreased (15.1% to 3.7%, 46.4% to 34.9%, and 67.2% to 18.1%). No pt has developed progressive disease.In conclusion, this Phase II trial in progress demonstrates preliminary safety and activity of the combination of MOR208 and lenalidomide in pts with CLL. This combination also has the potential to positively modulate the immune system, and detailed correlative studies will evaluate T- and NK-cell function. MOR208 appears safe in combination with ibrutinib, and preliminary evidence of activity against CLL cells with BTK C481S has been observed. Trial accrual is ongoing and updated results will be presented at the meeting. DisclosuresWoyach:Morphosys: Research Funding; Karyopharm: Research Funding; Acerta: Research Funding. Awan:Innate Pharma: Research Funding; Novartis Oncology: Consultancy; Pharmacyclics: Consultancy. Jones:Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics, LLC, an AbbVie Company: Membership on an entity's Board of Directors or advisory committees, Research Funding. Andritsos:Hairy Cell Leukemia Foundation: Research Funding. Lozanski:Genentech: Research Funding; Stemline Therapeutics Inc.: Research Funding; Beckman Coulter: Research Funding; Boehringer Ingelheim: Research Funding.

Harnessing the innate immune system to treat cancer: enhancement of antibody-dependent cellular cytotoxicity with anti-CD137 Ab.

Background: Monoclonal antibodies are an invaluable therapeutic class when it comes to cancer therapy. One of the primary mechanisms of anti-tumor activity of monoclonal antibodies is antibody dependent cell-mediated cytotoxicity (ADCC) mediated by natural killer (NK) cells. Innate immune effector cells play important roles in generating and maintaining antitumor immunity to enhance ADCC. Methods: Efficacy of anti-CD137 mAb was examined by in vitro and in vivo experiments. Clinical trials of anti-CD137 mAb are also on-going. Results: CD137 is a costimulatory receptor that belongs to the tumor necrosis factor receptor (TNFR) superfamily. CD137 is expressed on T cell, activated NK cells and other immune cells. We recently showed that an agonistic anti-CD137 mAb activates NK cells, resulting in increased ADCC against cancer cells. Anti-CD137 agonistic mAb therapy may both increase ADCC due to mAb-activated NK cells and promote the proliferation and cytotoxicity of antigen-specific T cells induced by mAb treatment. Conclusions: We discuss some of the promising strategies that could potentially enhance ADCC with anti-CD137 mAbs.

Abstract B144: Reduction of minimal residual disease in pediatric B-precursor acute lymphoblastic leukemia by an Fc-optimized CD19 antibody

Abstract B-precursor acute lymphoblastic leukemia (BCP-ALL) is the most common childhood cancer. Although this disease can be successfully treated in 80 % of patients by chemotherapy, prognosis for primary refractory or relapsed patients is very poor. Even after allogeneic stem cell transplantation (SCT), relapse rates are considerable and correlate significantly with persistent minimal residual disease (MRD) prior to or after SCT. In an MRD constellation favorable effector-target ratios prevail and thus it is well suited for immunotherapy with therapeutic antibodies. We developed a chimeric Fc-optimized third-generation CD19 antibody (4G7SDIE) and produced it in pharmaceutical quality. This antibody mediates markedly enhanced antibody dependent cellular cytotoxicity (ADCC) through its improved capability to recruit FcγRIIIa-bearing effector cells. In this study, 4G7SDIE was applied on a compassionate need basis to pediatric patients with relapsed or refractory BCP-ALL in an MRD situation (n=14) or in relapse (n=2) (initial refractory disease, n=3; 1st relapse, n=7; ≥ 2nd relapse, n=6; 12/16 patients had previous SCT). Side effects were negligible. In all patients complete CD20+ B-cell depletion was observed during therapy. After discontinuation of 4G7SDIE therapy B cell counts recovered rapidly to normal levels. Both patients in relapse did not respond to 4G7SDIE treatment. In 9/14 MRD-positive patients, MRD was reduced by ≥ 1 log or fell below MRD-detection threshold of 10-4 over the course of treatment. 2/9 responders were receiving additional treatment. In CD107a assays primary NK cells and γδ T cells were identified as main effector cell populations. The FcγRIIIA-V158F polymorphism had no influence on ADCC mediated by 4G7SDIE. Cytotoxicity assays confirmed sustained functionality of patient effector cells over the course of 4G7SDIE treatment. In vitro cytotoxicity assays were performed using PBMC from transplanted patients obtained at different time points of 4G7SDIE treatment. Lysis of autologous leukemic blasts was increased when 4G7SDIE or autologous patient serum taken after antibody infusion was added. After infusion of 20 mg/m2 4G7SDIE serum half-life was 34 ± 13 hours (n=3) and serum levels of 4G7SDIE remained above saturating concentrations of ≥ 700 ng/ml (EC50=65 ng/ml) at day 13 and following treatment cycle, respectively. Notably, in 3/3 analyzed patients under 4G7SDIE therapy, a transient down modulation of CD19 surface expression on the leukemic blasts was observed. In vitro antigenic shift assays on primary leukemic blasts showed considerable but very heterogeneous shift of CD19 surface expression. Furthermore, a positive correlation between CD19 surface expression levels and 4G7SDIE mediated lysis was observed. These observations hint at in vivo tumor escape mechanisms and furthermore indicate selective pressure exerted by immunotherapy with 4G7SDIE, underlining its therapeutic potential, but also delineating possible limitations. In conclusion, promising anti-leukemic effects of the 4G7SDIE antibody have been observed in vitro and in vivo. We are currently preparing a phase I/IIa trial. Citation Format: Ursula Joerdis Eva Seidel, Ludger Grosse-Hovest, Patrick Schlegel, Martin Hofmann, Friedhelm R. Schuster, Roland Meisel, Kai-Erik Witte, Steffen Aulwurm, Elwira Pyz, Hans-Georg Rammensee, Gundram Jung, Rupert Handgretinger, Peter Lang. Reduction of minimal residual disease in pediatric B-precursor acute lymphoblastic leukemia by an Fc-optimized CD19 antibody. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B144.

A Phase II Study of the Fc Engineered CD19 Antibody MOR208 in Combination with Lenalidomide for Patients with Chronic Lymphocytic Leukemia (CLL)

MOR208 is an Fc engineered CD19 monoclonal antibody which has been shown in a Phase I trial in patients with relapsed and refractory CLL to be generally well tolerated and have preliminary efficacy, with an overall response rate (ORR) of 30% by IWCLL 2008 guidelines (Woyach et al, Blood 2014). Compared to non-engineered CD19 monoclonal antibodies, MOR208 has significantly enhanced antibody dependent cellular cytotoxicity (ADCC), which can be further augmented in vitro with the addition of lenalidomide. Given the in vitro synergy of these agents, acceptable individual safety profiles, and efficacy of each as a single agent, we chose to combine MOR208 and lenalidomide in patients with both previously treated and previously untreated CLL.This study is a single institution phase II trial of MOR208 in combination with lenalidomide with an initial safety run-in as part of each cohort. MOR208 was given at a dose of 1 mg/kg on cycle 1 day 1, then 9 mg/kg on days 2, 8, 15, and 22 of cycle 1, and then on day 1 of cycles 2-12. Lenalidomide was started at a dose of 2.5 mg daily on cycle 1 day 8 and given continuously. The dose of lenalidomide could be escalated up to 10 mg daily in patients without toxicity. After 12 cycles, lenalidomide could be continued indefinitely in responding patients. Toxicity was assessed using the National Cancer Institute's Common Criteria for Adverse Events v4.0 for non-hematologic toxicity, and IWCLL 2008 guidelines for hematologic toxicity. Disease response by IWCLL 2008 guidelines was assessed at cycle 7 day 1 and at the end of cycle 12.This study will enroll 20 patients with treatment-naïve CLL and 20 patients with relapsed/refractory CLL. At this time, 7 patients with relapsed/refractory disease and 5 patients with treatment-naïve disease have been enrolled and evaluated. The most common toxicities observed related to protocol therapy have been infusion related reactions, fatigue, thrombocytopenia, and neutropenia. In patients with relapsed disease, all toxicities except neutropenia have been grade 1 or 2, and 2 patients experienced grade 3 neutropenia. Of the 5 patients with treatment-naïve CLL, two experienced significant infusion reactions on cycle 1 day 1 that prevented further administration of MOR208. After a protocol amendment escalating steroid premedication, no further grade 3 infusion reactions have been observed. While the majority of patients were able to escalate lenalidomide to either 5 or 10 mg, all patients had lenalidomide eventually dose reduced to 2.5 mg daily due to cytopenias, rash, or fatigue.This combination has shown preliminary efficacy. In the cohort of patients with relapsed disease, two experienced progressive disease during cycle 2 and cycle 5 respectively. The remaining 5 patients achieved stable disease (SD, n=3) or a partial response (PR, n=2) at cycle 7 day 1, with one patient converting to PR by cycle 12. Three patients completed 12 cycles of therapy, and the remaining two completed 12 cycles and now remain on lenalidomide alone at cycle 18 and cycle 19 respectively. In the cohort of patients with treatment-naïve disease, three patients completed more than 1 day of therapy. All of these patients achieved a PR at cycle 7 day 1, and are now in cycle 10 (n=1) or cycle 11 (n=2).In conclusion, this Phase II trial in progress demonstrates preliminary safety and activity of the combination of MOR208 and lenalidomide in patients with CLL. This combination also has the potential to positively modulate the immune system, and detailed correlative studies are evaluating T cell and NK cell function in these patients. Trial accrual is ongoing and updated results will be presented at the meeting. DisclosuresJones:AbbVie: Research Funding; Pharmacyclics LLC, an AbbVie Company: Consultancy, Research Funding. Byrd:Acerta Pharma BV: Research Funding.

Depleting MET-Expressing Tumor Cells by ADCC Provides a Therapeutic Advantage over Inhibiting HGF/MET Signaling

Hepatocyte growth factor (HGF) and its receptor MET represent validated targets for cancer therapy. However, HGF/MET inhibitors being explored as cancer therapeutics exhibit cytostatic activity rather than cytotoxic activity, which would be more desired. In this study, we engineered an antagonistic anti-MET antibody that, in addition to blocking HGF/MET signaling, also kills MET-overexpressing cancer cells by antibody-dependent cellular cytotoxicity (ADCC). As a control reagent, we engineered the same antibody in an ADCC-inactive form that is similarly capable of blocking HGF/MET activity, but in the absence of any effector function. In comparing these two antibodies in multiple mouse models of cancer, including HGF-dependent and -independent tumor xenografts, we determined that the ADCC-enhanced antibody was more efficacious than the ADCC-inactive antibody. In orthotopic mammary carcinoma models, ADCC enhancement was crucial to deplete circulating tumor cells and to suppress metastases. Prompted by these results, we optimized the ADCC-enhanced molecule for clinical development, generating an antibody (ARGX-111) with improved pharmacologic properties. ARGX-111 competed with HGF for MET binding, inhibiting ligand-dependent MET activity, downregulated cell surface expression of MET, curbing HGF-independent MET activity, and engaged natural killer cells to kill MET-expressing cancer cells, displaying MET-specific cytotoxic activity. ADCC assays confirmed the cytotoxic effects of ARGX-111 in multiple human cancer cell lines and patient-derived primary tumor specimens, including MET-expressing cancer stem-like cells. Together, our results show how ADCC provides a therapeutic advantage over conventional HGF/MET signaling blockade and generates proof-of-concept for ARGX-111 clinical testing in MET-positive oncologic malignancies.

A Phase II Open-Label, Multi-Center Study of Anti-CCR4 Monoclonal Antibody Mogamulizumab (KW-0761) in Patients with Previously Treated Peripheral T-Cell Lymphoma(PTCL)

Abstract Background CC chemokine receptor 4 (CCR4) is the receptor for macrophage-derived chemokine (MDC/CCL22) and thymus activation-regulated chemokine (TARC/CCL17). CCR4 is expressed on tumour cells in approximately 30-65% of patients with PTCL (Ogura, 2014). PTCL-NOS patients who are CCR4 positive have been reported to have a poorer prognosis compared to CCR4 negative patients (Ishida T CCR 2004). Mogamulizumab (KW-0761) is a defucosylated, humanized, IgG1 Mab with enhanced antibody dependent cellular cytotoxicity, that binds to CCR4. Mogamulizumab was evaluated in both phase 1 and 2 trials in Japanese patients. A phase II trial in PTCL and cutaneous T-cell lymphoma (CTCL) patients (Ogura, 2014) reported an overall response rate (ORR) of 35% in patients who relapsed after last systemic therapy (ORR was 34% in PTCL), leading to its approval in Japan in patients with previously treated CTCL and PTCL, in addition to its first indication, previously treated adult T-cell leukemia-lymphoma. Here we report the preliminary results of a European phase II trial of mogamulizumab in patients with relapsed/refractory PTCL. Methods A multi-center phase II study of mogamulizumab monotherapy was conducted to determine efficacy, safety and immunogenicity in patients with CCR4+ PTCL. The primary endpoint was ORR and secondary endpoints included duration of response, progression-free survival (PFS) and overall survival (OS). At least 34 evaluable patients were needed to detect a significant improvement over 15% assuming 80% power and a 0.0240 alpha significance level (assumes 35% ORR for alternative). Patients received mogamulizumab once weekly for 4 weeks and subsequently once every 2 weeks until progressive disease (PD) or unacceptable toxicity at a dose of 1.0 mg/kg. Responses were assessed every 8 weeks according to IWG criteria (Cheson et al 2007). Results Based on a preliminary analysis of the data, a total of 38 patients received at least one administration of mogamulizumab and were evaluable for safety analysis; (Male/female 23/15 ;Median age 58.5 years (range 19-87)). Three patients are still ongoing in the study (1 complete response (CR) and 2 stable disease (SD)). ECOG performance status at baseline was 0 (32%); 1 (29%); 2 (39%) and 92% of patients had stage III (32%) or IV (61%) disease. The median number of prior treatments was 2 (range 1-8). Only 17 patients (49%) responded to the last treatment administered prior to study entry. The median number of mogamulizumab administrations was 6 (range 1-32). The majority of adverse events (AEs) were CTCAE grade 1-2. Skin rash related to drug was observed in 32% of patients (12/38) and related AEs > grade 3 occurred in 32% (12) of patients. Infusion related reactions occurred in 3 patients (2 were CTCAE grade 2 and 1 was grade 3). Thirty-five patients were evaluable for efficacy. The ORR rate was 11% and the stable disease rate was 34% with a SD or better rate of 46%. The response by histological subtype is specified in the table below. The median duration of response (including SD) is 2.9 months. The median PFS is 2.1 months. Two patients (ALCL-ALK-neg and PTCL-NOS) proceeded to allogeneic SCT. Although the ORR in this study was less than seen in the Japanese study, the PFS was comparable. There were differences in patient population/study conduct between the Japanese study and this study, respectively, which included: inclusion of only relapsed patients (100% vs 49%), ECOG PS 2 (0% vs 39%) and response assessments (after 4 and 8 weeks versus 8 weekly from week 8). Conclusions Based on preliminary data, mogamulizumab demonstrates a SD or better rate of 46% and an ORR of 11% with an acceptable safety profile in this phase II study of heavily pre-treated relapsed/refractory PTCL patients. TableBest Overall Response by Histological subtypeNo of subjectsCR/PR n (%)SDn (%)>SD n (%)PTCL-NOS152* (13%)6 (40%)8 (53%)AITL122 (17%)3 (25%)5 (42%)TMF301 (33%)1 (33%)ALCL-ALK neg402 (50%)2 (50%)ALCL-ALK pos1000Efficacy Evaluable Subjects354 (11%)12 (34%)16 (46%) *One patient had CR by CT scan but did not have bone marrow done for confirmation of CR Disclosures Zinzani: Sandoz: Consultancy; Celgene International Sàrl: Advisory Board, Advisory Board Other, Honoraria, Speakers Bureau; MundiPharma International Ltd: Advisory Board, Advisory Board Other, Honoraria, Speakers Bureau; Pfizer Inc: Advisory Board Other, Honoraria, Speakers Bureau; Takeda Pharmaceutical Company Limited: Advisory Board Other, Honoraria, Speakers Bureau; F. Hoffmann-La Roche Ltd: Advisory Board Other, Honoraria; GlaxoSmithKline: Advisory Board, Advisory Board Other, Honoraria; Gilead: Advisory Board, Advisory Board Other; Bayer AG: Advisory Board Other, Consultancy. d'Amore:CTI Life Sciences: Speakers Bureau; Mundipharma: Speakers Bureau; Takeda/Seattle Genetics : Speakers Bureau; Sanofi-Aventis: Research Funding; Amgen: Research Funding; Roche: Research Funding; Kyowa-Kirin: Advisory Board Other. Haioun:Roche: Honoraria; Celgene: Honoraria; Takeda: Honoraria; Pfizer: Honoraria; Janssen: Honoraria. Morschhauser:Genentech: Honoraria; Bayer: Honoraria; Spectrum: Honoraria; Mundipharma: Honoraria; Gilead: Honoraria.

C-Abl modulates tumor cell sensitivity to antibody-dependent cellular cytotoxicity.

Monoclonal antibodies (mAb) can modulate cancer cell signal transduction and recruit antitumor immune effector mechanisms-including antibody-dependent cellular cytotoxicity (ADCC). Although several clinically effective antibodies can promote ADCC, therapeutic resistance is common. We hypothesized that oncogenic signaling networks within tumor cells affect their sensitivity to ADCC. We developed a screening platform and targeted 60 genes derived from an EGFR gene network using RNAi in an in vitro ADCC model system. Knockdown of GRB7, PRKCE, and ABL1 enhanced ADCC by primary and secondary screens. ABL1 knockdown also reduced cell proliferation, independent of its ADCC enhancement effects. c-Abl overexpression decreased ADCC sensitivity and rescued the effects of ABL1 knockdown. Imatinib inhibition of c-Abl kinase activity also enhanced ADCC-phenocopying ABL1 knockdown-against several EGFR-expressing head-and-neck squamous cell carcinoma cell lines by ex vivo primary natural killer cells. Our findings suggest that combining c-Abl inhibition with ADCC-promoting antibodies, such as cetuximab, could translate into increased therapeutic efficacy of mAbs.

Obinutuzumab may chart the way to improved QOL for CLL patients.

Obinutuzumab recently received accelerated approval from the US Food and Drug Administration with breakthrough therapy designation for use in combination with chlorambucil in patients with untreated chronic lymphocytic leukemia (CLL). Obinutuzumab is a CD20 targeting fully humanized, type II, IgG1 antibody. CD20 is weakly expressed on the surface of CLL cells but has been demonstrated to be an effective in vivo target as shown by the activity observed with the use of rituximab and ofatumomab.2-4 Obinutuzumab binds selectively to the extracellular domain of CD20 with reduced internalization and its structural modifications explain its enhanced effectiveness. The antibody is modified in the hinge region which allows for more potent direct cytotoxicity. More importantly, afucosylation in the Fc region allows for enhanced antibody dependent cellular cytotoxicity (ADCC) through robust engagement of Fc-gamma receptor type III on effector cells. Together, these modifications translate into a higher efficacy compared with rituximab both in preclinical and clinical studies.

B-1239, a Novel Anti-BAFF-R Afucosylated Human Antibody, Promotes Potent Natural Killer Cell- Mediated Antibody Dependent Cellular Cytotoxicity In Chronic Lymphocytic Leukemia Cells In- Vitro and Depletion Of Circulating Leukemic CLL B Cells In-Vivo

B-cell activating factor (BAFF) belongs to the TNF ligand superfamily of cytokines involved in B cell survival and maturation. BAFF is produced by diverse cell types including innate immune cells like monocytes and dendritic cells as well as T cells, activated B cells, and bone marrow stromal cells. BAFF binds to the BAFF receptor (BAFF-R) with high affinity compared to the other BAFF receptors, BCMA and TACI. While BAFF is known to regulate normal B-cell development and proliferation, it also contributes to survival in chronic lymphocytic leukemia (CLL). We observed expression of BAFF-R on virtually all B cells from CLL patients. B-CLL cells have strong up-regulation of BAFF and BAFF-R compared to normal healthy B cells. We describe here the in-vitro and in-vivo evaluation in CLL of B-1239, a fully human anti-BAFF-R monoclonal IgG1 antibody. B-1239 is devoid of fucose residues in its Fc domain, resulting in enhanced binding to FCgammaRIIIa activating receptor on Natural Killer (NK) cells. While B-1239 failed to induce direct or complement mediated cytotoxicity, binding of B-1239 to CLL cells resulted in enhanced antibody dependent cellular cytotoxicity (ADCC) with allogeneic or autologous NK effector cells in-vitro. Indeed, at a therapeutically relevant concentration of 10 ug/mL B-1239 shows more than 30% increased relative cytotoxic activity over current CLL antibody therapeutic Rituximab. Dilutions of B-1239 down to 0.01 ug/mL showed similar cytotoxicity to the 10 ug/mL concentration. At 0.0001 ug/mL B-1239 has a 40% cytotoxic effect on CLL cells in ADCC assays while antibody therapeutic controls, like Rituximab, show virtually no cytotoxic activity. Furthermore, B-1239 mediated antibody-dependent cellular phagocytosis (ADCP) by monocyte-derived macrophages and mediated activation of monocytes and macrophages as detected by TNF-alpha production. Consistent with the cross reactivity to murine BAFF-R, flow cytometric analysis revealed binding of B-1239 to CD5+CD19+ leukemic B cells from Eu-Tcl-1 transgenic mouse CLL cells. A single dose of B-1239 by i.v injection into Eu-Tcl-1 mice resulted in dramatic reduction in circulating CD5+CD19+ leukemic B cells in all three B-1239 injected mice. In contrast, we observed continued increase of leukemic CD5+CD19+ populations in the two vehicle treated mice. Ongoing studies are focused on determining how targeting BAFF-R on CLL B-cells depletes the leukemic population both in-vitro and in-vivo and the downstream effects of targeting through this receptor. Collectively, these results demonstrate that targeting BAFF-R on CLL cells provides a B-cell specific approach for rapid and robust depletion of leukemic CLL cells and provides evidence for a strong therapeutic advantage in BAFF-R targeted therapies in CLL. Disclosures:Huet:Novartis: Employment, Employment Related Perks Other. Gram:Novartis: Employment, Employment Related Perks Other. Baeck:Novartis: Employment, Employment Related Perks Other.

Tumour antigen targeted monoclonal antibodies incorporating a novel multimerisation domain significantly enhance antibody dependent cellular cytotoxicity against colon cancer

Tumour antigen targeted antibodies (mAbs) can induce natural killer (NK) cells to kill tumours through antibody dependent cellular cytotoxicity (ADCC) upon engagement of NK cell expressed FcγRIIIa. FcγRIIIa polymorphisms partially dictate the potency of the ADCC response. The high affinity FcγRIIIa-158-valine (V) polymorphism is associated with more potent ADCC response than the low affinity FcγRIIIa-158-phenylalanine (F) polymorphism. Because approximately 45% of patients are homozygous for the FcγRIIIa-158-F polymorphism (FF genotype), their ability to mount ADCC is impaired. We investigated whether a novel mAb capable of binding multiple antigen specific targets and engaging multiple low affinity FcγRIIIa receptors could further enhance ADCC against colon cancer in vitro. Specifically, we generated a novel anti-epidermal growth factor receptor (EGFR) antibody (termed a stradobody™) consisting of an unmodified Fab sequence and two Immunoglobulin G, subclass 1 (IgG1) Fc domains separated by an isoleucine zipper domain and the 12 amino-acid IgG2 hinge. The stradobody™ framework induced multimerisation and was associated with increased binding to the EGFR and FcγRIIIa. From a functional perspective, when compared to an unmodified anti-EGFR mAb with a sequence identical to cetuximab (a commercially available anti-EGFR mAb), stradobodies™ significantly enhanced ADCC. These effects were observed using both KRAS wild type HT29 and KRAS mutant SW480 colon cancer cells as targets, and by NK cells obtained from healthy donors and a cohort of patients with colon cancer. These data suggest that high avidity cross-linking of multiple tumour surface antigens and multiple NK cell associated FcγRIIIa molecules can enhance ADCC and partially overcome impaired ADCC by FF genotype individuals in vitro.

Immunomodulatory effects of a dipeptidyl peptidase inhibitor, ARI-4175, and NK cell activation in a colon cancer model.

3058 Background: A possible mechanism of the antitumor effect of therapeutic antibodies is antibody-dependent cellular cytotoxicity (ADCC). We reported that an inhibitor of dipeptidyl peptidase (DPP)4-like serine proteases, Ari-4175, significantly slowed the growth of KRAS-mutated HCT-116 xenografts in nude mice, either as a single agent or with cetuximab (Ctx). Ari-4175 is not cytotoxic in vitro, and since it was able to overcome the resistance to Ctx due to the KRAS mutation in the HCT-116 line, we hypothesized that the antitumor activity involves the activation of NK cells and enhancement of ADCC. Methods: Ari-4175 was administered orally to nude mice at 200 µg q.d. x5 days/week. Peripheral blood or spleens were assayed for immune parameters, ex vivo, at various time points. Expression of surface markers on myeloid and NK cells were monitored by flow. Natural cytotoxicity and ADCC were assayed using HCT116 cells and Ctx using a flow based assay. Cytokines were measured using Luminex assays. Results: Ari-4175 induced upregulation of CD69 on NK cells on day 2, followed by upregulation of CD16 on day 7. Coordinate with these changes in peripheral blood, splenocytes from treated mice exhibited significantly increased in vitro cytotoxicity toward the HCT116 cells. Increased serum levels of inflammatory cytokines, including IL-1b, IL-6, MCP-1, GCSF, and IL-2 were seen. A significant expansion of a distinct CD45+CD14+Gr-1+MHC-II-CDllb+CD11cvariable myeloid cell population in both peripheral blood and spleens of mice was also seen after one week of treatment. Conclusions: Ari-4175 showed dramatic anti-tumor effect in KRAS-mutant CRC xengrafts when given alone or in combination with Ctx. In vitro data suggest that the therapeutic effect of 4175 might partially be due to the augmentation of ADCC through elevated expression of CD16 on NK cells. In addition, Ari-4175 appears, in vivo, to expand a unique myeloid cell population, which may be responsible for an inflammatory cytokine response and subsequent activation of NK cells. Our study provides a mechanistic rationale for testing Ari-4175 in a clinical trial and possible biomarker endpoints for evaluation in peripheral blood samples.

Abstract 494: Immunomodulatory effects of a small molecule dipeptidyl peptidase inhibitor, Ari-4175, are mediated through NK cell activation in a colorectal cancer model.

Abstract Introduction: A possible mechanism of the antitumor effect of many therapeutic monoclonal antibodies is antibody-dependent cellular cytotoxicity (ADCC) mediated by NK cells. We recently observed that an inhibitor of dipeptidyl peptidase (DPP)4-like serine proteases, Ari-4175, significantly slowed the growth of K-RAS mutated HCT-116 tumor xenografts in nude mice, either as a single agent or in combination with cetuximab. Ari-4175 is not directly cytotoxic in vitro, and since it was able to overcome, in vivo, the resistance to blockade of the epidermal growth factor receptor engendered by the KRAS mutation in the HCT-116 colorectal cancer (CRC) cell line, we tested the hypothesis that the mechanism of the antitumor activity involves the activation of NK cells, resulting in the enhancement of ADCC. Methods: Ari-4175 was administered orally to C57Bl/6 mice at 200 μg q.d. x5 days/week. Peripheral blood or spleens were assayed for immune parameters, ex vivo, at various time points. Expression of surface markers on myeloid and NK cells were monitored by flow cytometry. Natural cytotoxicity and ADCC were assayed using HCT116 cells and cetuximab using a flow cytometry-based assay. Cytokines were measured in mouse serum using Luminex assays. Results: Treatment of mice with Ari-4175 induced up-regulation of the activation marker CD69 on NK cells on day 2, followed by up-regulated expression of FcRIIIA (CD16, which mediates ADCC) on day 7. Coordinate with these changes in surface markers detected in peripheral blood samples, splenocytes from Ari-4175-treated mice exhibited significantly increased in vitro natural cytotoxicity responses toward the HCT116 colon cancer cell line. The treated mice exhibited increased serum levels of inflammatory cytokines, including IL-1, IL-6, MCP-1, GCSF, and IL-2. In addition to the impacts on NK cells, we also observed a significant expansion of a distinct CD45+CD14+Gr-1+MHC-II−CDllb+CD11cvariable myeloid cell population in both peripheral blood and spleens of mice after one week of treatment with Ari-4175. Conclusion: The prolyl peptidase inhibitor, Ari-4175, showed dramatic anti-tumor effect in KRAS mutant colorectal cancer xengrafts when given alone or in combination with cetuximab. In vitro data suggest that the therapeutic effect of 4175 might partially be due to the augmentation of ADCC through elevating expression of CD16 on NK cells. In addition, Ari-4175 appears, in vivo, to expand a unique myeloid cell population, which may be responsible for an inflammatory cytokine response and subsequent activation of NK cells. Our study provides a mechanistic rationale for testing Ari-4175 in a clinical trial and possible biomarker endpoints for evaluation in peripheral blood samples. Citation Format: Alexander W. MacFarlane, Jiping Zhang, Barry Jones, William Bachovchin, Kerry Campbell, Hossein Borghaei. Immunomodulatory effects of a small molecule dipeptidyl peptidase inhibitor, Ari-4175, are mediated through NK cell activation in a colorectal cancer model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 494. doi:10.1158/1538-7445.AM2013-494