Bispecific T cell engagers (BiTEs) are engineered proteins combining the variable regions isolated from two different antibodies. By binding to both T cells and cancer cells, BiTEs help endogenous T cells recognize and eliminate malignant cells. Indeed, if one of the antigens recognized at the T cell surface is CD3, a receptor initiating T cell activation, T cells will be activated following cancer cell engagement. The concept of BiTEs is a recent product of immunotherapy and represents an improvement over injection of single antibodies as they can directly harness immune effector cells for anti-cancer efficacy. Despite this progress, the therapy fails in many patients and little is known about the mechanisms behind these failures
Investigators Drs. Roland Walter and Christopher Mehlin (Clinical Research Division), analyzed the effect of combining two BiTEs to improve this approach. The method was named SMITE for “Simultaneous multiple interaction T cell engaging bispecifics”. The results of this study were recently published the journal Leukemia. Lead co-authors, Drs. Colin Correnti and George Laszlo, staff scientists, tested the approach in vitro using two BiTEs binding to one or two tumor antigens, and either to the CD3 or to the CD28 co-receptor. CD28 provides a co-stimulatory signal for T cell activation.
Human T cells were obtained from healthy adult volunteers. The T cells were incubated in vitro with target cell lines overexpressing the antigens of interest or control cell lines at a 1:1 ratio, in the presence or absence of a single BiTE molecule or a SMITE bispecific pair. T cell activation and proliferation were assessed by flow cytometry using CD69 and CD25 markers. Several BiTEs were generated targeting the cancer antigens CD19 or ROR1, both specifically expressed in leukemic B cells. The variable regions targeting these antigens were isolated from known and previously tested antibodies.
The T cell activation mediated by the BiTE is dependent on simultaneous cancer cell binding, thus avoiding non-specific T cell activation. Indeed, BiTEs targeting both CD3 and cancer antigen, do not activate T cells in the absence of the cancer antigen. Additionally, lysis of the antigen-expressing cells by activated T cells following CD3/cancer antigen BiTE engagement was significantly improved in the presence of the CD28/cancer antigen BiTE, demonstrating the synergistic effect of the SMITE pair. However, CD28 monoclonal antibody or CD28/cancer antigen BiTE alone did not activate T cells in the absence of the CD3 signal.
One of the mechanisms associated with immunotherapy failure is the expression of the inhibitory PD-L1 ligand expressed at the surface of the cancer cells. T cell binding to PD-L1 blocks T cell activation. However, this inhibition could be overturned by designing a CD28/PD-L1 BiTE to trigger the CD28 costimulatory signal instead of the inhibitory signaling pathway generally observed upon binding of the T cell to PD-L1 expressing cancer cells. The presence of CD28/PD-L1 BiTE did not impair T cell activation by CD3/cancer antigen BiTE in the absence of the PD-L1 antigen and significantly increased T cell activation in the presence of the PD-L1 -expressing cells relative to the CD3/cancer antigen BiTE alone.
As such, SMITE bispecifics could represent a promising “a la carte” therapy to target different kind of cancer antigens while transforming T cell inhibition signals into activation signals. “Our current focus is to develop a synergic SMITE pair that is ready for the clinic. We are now utilizing the Trianni™ mouse platform, which allows us to develop fully human antibodies specific for T cell co-receptors and cancer antigens, and leveraging our protein engineering capabilities to make hundreds of different BiTEs that we can then test in combination. We are incredibly excited about this approach and have all the pieces to make a dual targeting approach clinically feasible”, concluded Dr. Correnti.
Funding for this study was provided by the Leukemia & Lymphoma Society, Alex’s Lemonade Stand Foundation/Cure4Cam Childhood Cancer Foundation, Hyundai Hope on Wheels, Bezos Family Immunotherapy Initiative, the National Heart, Lung, and Blood Institute/ National Institutes of Health (NIH), National Institute of Diabetes and Digestive and Kidney Diseases/NIH (NIDDK/NIH, Co-operative Center of Excellence in Hematology).
Correnti CE, Laszlo GS, de van der Schueren WJ, Godwin CD, Bandaranayake A, Busch MA, Gudgeon CJ, Bates OM, Olson JM, Mehlin C, Walter RB. 2018. Simultaneous multiple interaction T-cell engaging (SMITE) bispecific antibodies overcome bispecific T-cell engager (BiTE) resistance via CD28 co-stimulation. Leukemia. 32:1239–1243.