Jaime Bald

Daratumumab depletes CD38+ immune-regulatory cells, promotes T-cell expansion, and skews T-cell repertoire in multiple myeloma

Daratumumab targets CD38-expressing myeloma cells through a variety of immune-mediated mechanisms (complement-dependent cytotoxicity, antibody-dependent cell-mediated cytotoxicity, and antibody-dependent cellular phagocytosis) and direct apoptosis with crosslinking. These mechanisms may also target nonplasma cells that express CD38, which prompted evaluation of daratumumabs effects on CD38-positive immune subpopulations. Peripheral blood (PB) and bone marrow (BM) from patients with relapsed/refractory myeloma from 2 daratumumab monotherapy studies were analyzed before and during therapy and at relapse. Regulatory B cells and myeloid-derived suppressor cells, previously shown to express CD38, were evaluated for immunosuppressive activity and daratumumab sensitivity in the myeloma setting. A novel subpopulation of regulatory T cells (Tregs) expressing CD38 was identified. These Tregs were more immunosuppressive in vitro than CD38-negative Tregs and were reduced in daratumumab-treated patients. In parallel, daratumumab induced robust increases in helper and cytotoxic T-cell absolute counts. In PB and BM, daratumumab induced significant increases in CD8(+):CD4(+) and CD8(+):Treg ratios, and increased memory T cells while decreasing naïve T cells. The majority of patients demonstrated these broad T-cell changes, although patients with a partial response or better showed greater maximum effector and helper T-cell increases, elevated antiviral and alloreactive functional responses, and significantly greater increases in T-cell clonality as measured by T-cell receptor (TCR) sequencing. Increased TCR clonality positively correlated with increased CD8(+) PB T-cell counts. Depletion of CD38(+) immunosuppressive cells, which is associated with an increase in T-helper cells, cytotoxic T cells, T-cell functional response, and TCR clonality, represents possible additional mechanisms of action for daratumumab and deserves further exploration.

Monitoring multiple myeloma patients treated with daratumumab: teasing out monoclonal antibody interference

Abstract Background: Monoclonal antibodies are promising anti-myeloma treatments. As immunoglobulins, monoclonal antibodies have the potential to be identified by serum protein electrophoresis (SPE) and immunofixation electrophoresis (IFE). Therapeutic antibody interference with standard clinical SPE and IFE can confound the use of these tests for response assessment in clinical trials and disease monitoring. Methods: To discriminate between endogenous myeloma protein and daratumumab, a daratumumab-specific immunofixation electrophoresis reflex assay (DIRA) was developed using a mouse anti-daratumumab antibody. To evaluate whether anti-daratumumab bound to and shifted the migration pattern of daratumumab, it was spiked into daratumumab-containing serum and resolved by IFE/SPE. The presence (DIRA positive) or absence (DIRA negative) of residual M-protein in daratumumab-treated patient samples was evaluated using predetermined assessment criteria. DIRA was evaluated for specificity, limit of sensitivity, and reproducibility. Results: In all of the tested samples, DIRA distinguished between daratumumab and residual M-protein in commercial serum samples spiked with daratumumab and in daratumumab-treated patient samples. The DIRA limit of sensitivity was 0.2 g/L daratumumab, using spiking experiments. Results from DIRA were reproducible over multiple days, operators, and assays. The anti-daratumumab antibody was highly specific for daratumumab and did not shift endogenous M-protein. Conclusions: As the treatment of myeloma evolves to incorporate novel monoclonal antibodies, additional solutions will be needed for clinical monitoring of patient responses to therapeutic regimens. In the interim, assays such as DIRA can inform clinical outcomes by distinguishing daratumumab from endogenous M-protein by IFE.

Abstract 3636: Potent antitumor activity of duvortuxizumab, a CD19 x CD3 DART® molecule, in lymphoma models

Duvortuxizumab (JNJ-64052781 or MGD011) is a CD19 x CD3 DART® protein designed to engage and redirect CD3+ T-cells to eliminate CD19+ B-cells through T-cell-mediated cytotoxicity. Duvortuxizumab displays potent killing activity in lymphoma cell lines and animal models and is currently in clinical development for the treatment of B-cell malignancies. Here we examined duvortuxizumab activity alone and in combination with standard chemotherapy regimens in preclinical lymphoma models. Duvortuxizumab plus bendamustine increased T-cell mediated cytotoxicity of CD19+ Raji and DOHH2 lymphoma cells in the presence of isolated human pan T-cells (effector:target = 10:1). Minimal inhibition of T-cell activation was observed at bendamustine concentrations up to 40 µM. A Burkitt’s lymphoma model was used to evaluate the in vivo effects of duvortuxizumab plus bendamustine. NOD scid gamma mice were subcutaneously implanted with Daudi tumor cells and inoculated with either peripheral blood mononuclear cells or purified, activated human pan T-cells. Duvortuxizumab was dosed at 0.5 mg/kg (x7, twice weekly); bendamustine was dosed once at 25 mg/kg on day 1. Dosing with bendamustine alone inhibited tumor growth by 79% (p=0.0002) whereas duvortuxizumab alone resulted in complete tumor regression with no signs of relapse for up to 90 days. Dosing with duvortuxizumab plus bendamustine resulted in complete and durable tumor regression and elimination. Analysis of tumor-infiltrating lymphocytes (TILs) 3 days after the start of therapy showed that bendamustine inhibited CD3+ T-cell infiltration and activation whereas duvortuxizumab increased both. The combination showed greater activation of CD4+ and CD8+ T-cells. At 48 days after the last duvortuxizumab dose, TILs had a large percentage of CD3+ CD45RA- CCR7- cells, indicative of effector memory T-cells. This activation was accompanied by higher expression of PD-1 on T-cells and PD-L1 on tumor cells. Similar effects were seen when duvortuxizumab was combined with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP). In vitro, a decrease in T-cell activation was observed with duvortuxizumab/CHOP, which may have been caused by inclusion of prednisone; however, cytotoxicity against tumor cells was maintained. In a patient sample-derived model of diffuse large B-cell lymphoma, duvortuxizumab (twice weekly for 4 consecutive weeks) in combination with CHOP (given once) resulted in rapid tumor regression that was more sustained than CHOP treatment alone. In summary, duvortuxizumab has potent anti-tumor activity as a single agent and in combination with standard chemotherapy in lymphoma preclinical models. Duvortuxizumab-mediated tumor killing and T-cell activation were maintained or increased in the presence of multiple chemotherapeutics, suggesting the potential clinical utility of combining duvortuxizumab with standard therapies in the treatment of B-cell malignancies. Citation Format: Liat Izhak, Dana E. Cullen, Maha Elgawly, Leopoldo Luistro, Syd Johnson, Jaime Bald, A Kate Sasser, Sriram Balasubramanian. Potent antitumor activity of duvortuxizumab, a CD19 x CD3 DART® molecule, in lymphoma models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3636. doi:10.1158/1538-7445.AM2017-3636