Identification and Characterization of an Agonistic Aptamer Against the T Cell Costimulatory Receptor, OX40

Abstract

Induction of an effective immune response that can target and eliminate malignant cells or virus-infected cells requires the stimulation of antigen-specific effector T cells. A productive and long-lasting memory response requires 2 signals: a specific signal provided by antigen recognition through engagement of the T cell receptor and a secondary signal via engagement of costimulatory molecules (such as OX40) on these newly activated T cells. The OX40-OX40-ligand interaction is critical for the generation of productive effector and memory T cell functions. Thus agonistic antibodies that stimulate OX40 on activated T cells have been used as adjuvants to augment immune responses. We previously demonstrated that an aptamer modified to stimulate murine OX40 enhanced vaccine-mediated immune responses in a murine melanoma model. In this study, we describe the development of an agonistic aptamer that targets human OX40 (hOX40). This hOX40 aptamer was isolated using systematic evolution of ligands by exponential enrichment and binds the target purified protein with high affinity [dissociation constants (K(d))<10 nM]. Moreover, the hOX40 aptamer-streptavidin complex has an apparent binding affinity of ~50 nM for hOX40 on activated T cells as determined by flow cytometry and specifically binds activated human T cells. A multivalent version of the aptamer, but not a mutant version of the aptamer, was able to stimulate OX40 on T cells and enhance cell proliferation and interferon-gamma production. Future studies will assess the therapeutic potential of hOX40 aptamers for ex vivo stimulation of antigen specific T cells in conjunction with dendritic cell-based vaccines for adoptive cellular therapy.

Aptamers as targeted therapeutics: current potential and challenges

“…This includes RNA aptamers targeting human epidermal growth factor receptor-3 (HER3/ERBB3) 116 , OX40 (CD134) 117, 118 , 4-1BB (CD137) 119 , CD40 120 , CD28 121 , and DNA aptamers targeting human VEGFR-2 122 and the insulin receptor (IR) 123 . Several of the RNA aptamers targeting immune costimulatory receptors (CD28, CD40, OX40 and 4-1BB) have been engineered into multimeric versions to act as receptor agonists for improved cancer immunotherapy 124 .…”

“…The resulting bivalent aptamer activated the OX40 receptor on primed T cells in vitro , and systemic administration of this bivalent aptamer significantly enhanced anti-tumor responses generated by a dendritic cell-based vaccine in mice. Recently, two anti-human OX40 RNA aptamers containing a biotin group at the 5′-end were formulated into a bivalent via a streptavidin linker, which stimulated OX40 on human T cells, and enhanced cell proliferation as well as interferon-gamma production 118 .…”

Agonists of Co-stimulation in Cancer Immunotherapy Directed Against CD137, OX40, GITR, CD27, CD28, and ICOS

“…In 2013, the same group developed a human OX40 costimulatory aptamer by SELEX, proving that the multimer human OX40 aptamer elicited a costimulatory activation signal on human T cells as measured by cell proliferation and IFNȖ secretion (172).…”

Aptamers as Therapeutics

“…Aptamers have been selected against 0X40 (99, 100) and 4–1BB (75), both of which are costimulatory receptors that function to promote T cell responses. When assembled as bivalent structures (using an all-nucleic acid scaffold), these aptamers were shown to have agonistic activity on their receptors, stimulating T cell activation in vitro.…”