Abstract 691: Antibody conjugated to a bispecific RNA molecule targeting RIG-I and PLK1

Abstract

Abstract Simultaneous access to several targets has become the subject of intense studies in immuno-oncology. In order to benefit from the synergies provided by the activation of different signaling pathways in immunology and the knockdown of proteins involved in cancer cell survival, we established a bispecific approach. The activation of the innate immune response by delivering agonists of pattern-recognition receptors (PRR) such as RIG-I (retinoic acid-inducible I) represents a promising strategy. RIG-I detects short double-stranded RNA molecules ended by a 5'-di/triphosphate moiety (5'ppp-dsRNA). RIG-I activation promotes type I IFN secretion and cancer-cell selective apoptosis. To obtain a bifunctional molecule, the 5'ppp-dsRNA sequence was designed to silence PLK1 (polo-like kinase 1). Suppressing PLK1 expression with small interfering RNAs (siRNA) leads to cell cycle arrest and retards cancer cell growth. This concept of bifunctional RNAs has been validated by using non-targeted systems.1 To enhance this synergy, we conjugated this 5'ppp-siPLK1 to an antibody for a specific delivery to cancerous cells that overexpress erythropoietin-producing hepatocellular receptor A2 (EphA2) at their surface. Upon binding to EphA2 receptor, the antibody is well internalized, thus making it a good vehicle to deliver the bispecific 5'ppp-siPLK1. After EphA2-positive cells treatment, we observed RIG-I specific activation as well as PLK1 depletion. Both effects were correlated with cellular apoptosis and the mode of action was further confirmed with mechanistic and kinetic studies. Finally, while non-modified unconjugated siRNA has a very short half-life in plasma, we observed an increase in stability for the antibody-5'ppp-siPLK1 conjugates. These data suggest that anti-EphA2 receptor antibody could be used to deliver a bispecific RNA molecule.