Abstract A88: Targeting 4-1BB costimulation to the tumor stroma with bispecific aptamer conjugates enhances the therapeutic index of tumor immunotherapy
Authors: Schrand Brett , Berezhnoy Alexey , Brenneman Randall , Williams Anthony , Levay Agata , Kong Ling-Yuan , Rao Ganesh , Zhou Shouhao , Heimberger Amy , Gilboa Eli 
Published: October 2015

Abstract
Preclinical studies in mice and recent clinical trials have highlighted the promise of immune costimulation in cancer immunotherapy. Not unexpectedly, murine studies have also shown that systemic administration of immune stimulatory antibodies can be associated with adverse effects, mostly on-target autoimmune pathologies resulting from the activation of autoreactive T cells. This was underscored in a recent phase I clinical trials of an agonistic 4-1BB antibody that was associated with high frequencies of objective responses but also accompanied by adverse effects that became significant at the highest dose tested causing liver toxicity resulting in two fatalities, by-and-large precluding the dose escalation necessary to fully exploit the therapeutic potential of this otherwise promising drug. Arguably, targeting costimulatory ligands to the disseminated tumor lesions of the patient would reduce drug associated toxicities. With the goal of developing a clinically feasible and broadly applicable approach to reduce the aforementioned dose limiting toxicities we are developing an oligonucleotide aptamer platform consisting of bi-specific aptamer conjugates whereby costimulatory aptamer ligands (the therapeutic agents) are conjugated to aptamers that bind to tumor-specific aptamers (the targeting ligands). The chemically synthesized oligonucleotide aptamers offer significant advantages over antibodies in terms of synthesis, cost, conjugation chemistry, and reduced risk of neutralizing immunogenicity.
We have previously shown that systemic delivery of a bi-specific aptamer composed of an agonistic 4-1BB binding aptamer conjugated to a an aptamer that bound to a product expressed on the surface of a tumor cell, led to inhibition of tumor growth, and exhibited a superior therapeutic index compared to nontargeted costimulation with 4-1BB antibodies or 4-1BB aptamers. Nonetheless, the main limitation of tumor targeted costimulation in its current form stems from the fact that the costimulatory (aptamer) ligand, has to be displayed on the surface of the targeted tumor cells. Consequently one is limited to target receptors that do not internalize upon ligand binding. While such receptor-ligand interactions exist, most receptor-ligand complexes are internalized, and consequently the targeting choices are severely limited.
To address this limitation, and broaden the scope of tumor-targeted costimulation, the 4-1BB aptamers were targeted to products secreted in the tumor stroma by conjugation to aptamers that bind to either VEGF or osteopontin (OPN). This approach was predicated on the premise that by targeting the costimulatory ligands to products secreted into the tumor stroma the T cells will be costimulated prior to their engagement of the MHC/peptide complex on the tumor cell, thereby obviating the need to target the costimulatory ligands to non-internalizing cell surface products expressed on the tumor cells. We have shown that the stroma targeted 4-1BB aptamers exhibited a superior therapeutic index compared to an agonistic 4-1BB antibody, and engendered potent antitumor immunity against multiple unrelated tumors in subcutaneous, post surgical metastasis, and carcinogen-induced tumor models.
We are currently developing increasingly potent stroma targeted platforms using multi-valent dendrimers and are exploring the use of peptides as alternative targeting ligands. In addition we are developing stroma targeted strategies with stimulatory CD27 and OX40 aptamers and blocking PD-1 aptamers.
Citation Format: Brett Schrand, Alexey Berezhnoy, Randall Brenneman, Anthony Williams, Agata Levay, Ling-Yuan Kong, Ganesh Rao, Shouhao Zhou, Amy Heimberger, Eli Gilboa. Targeting 4-1BB costimulation to the tumor stroma with bispecific aptamer conjugates enhances the therapeutic index of tumor immunotherapy. [abstract]. In: Proceedings of the AACR Special Conference: Tumor Immunology and Immunotherapy: A New Chapter; December 1-4, 2014; Orlando, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2015;3(10 Suppl):Abstract nr A88.