Abstract 2055: Delivery of oligonucleotide therapeutic drugs: Universal detection reagents specific for nucleic acid modifications

Abstract Nucleic acid therapies, including oligonucleotide therapeutics (ONTs) and vaccines, have increased in development, respectively, since their inception as an emerging drug modality and after the first mRNA COVID-19 vaccine was granted EUA approval in 2020. RNA-based therapies, such as antisense oligonucleotides (ASO), RNAi, aptamers, and mRNA, are designed to modulate protein synthesis in patients and have applications in oncology, gene therapy, and infectious disease prevention. Strategies for ONT drug delivery have advanced over recent years with the goal of overcoming limitations when targeting extrahepatic tissues. Improving the “drug-likeness” of ONTs is facilitated by chemical modifications to the sugar-phosphate backbone and/or nitrogenous bases. Modifications are designed to increase ONT drug stability, uptake, and efficacy, but may increase the likelihood of toxicity. First generation chemical modifications include phosphorothioate (PS) modification of the phosphate portion of the backbone, whereas second generation modifications include adding substituent groups to the 2’ position of the ribose portion of the backbone, specifically 2’-O-methyl (OME), 2’-O-methoxy-ethyl (MOE), and 2’-fluoro modifications of RNA. Third generation modifications include peptide nucleic acids (PNA), locked nucleic acids (LNA), morpholino phosphoroamidate (PMO) modifications, and others. We have developed, optimized, and validated a library of monoclonal antibody reagents that detect these chemical modifications independent of nucleic acid composition, structure, strandedness, configuration, or platform. These “universal detection reagents” can be used in (1) immunohistochemistry (IHC) to determine biodistribution, (2) immunofluorescence (IF) to determine intracellular localization, (3) ELISA to determine anti-drug antibody binding, immunogenicity, and drug ranking, (4) immunoprecipitation to determine protein binding in serum and/or tissue lysates, (5) in vivo cell culture studies to determine potency, and (6) other immunoassays. While in situ hybridization (ISH) is an established method for the detection and localization of ONTs in cells and tissues, this method has limitations that include lack of utility (as unique probes must be designed individually for each ONT), high costs in time and materials to produce probes, the inability of ISH to bind and detect short ONT drug sequences, and difficulties encountered when repeated sequences are present within the ONT sequence. Here we report differential sensitivity and specificity of panels of antibodies specific for PS and MOE modifications. We demonstrate the performance of these antibodies by various in vitro assays that show the utility of these specialty reagents to facilitate the collection of analytical data to support ADMET studies useful to ensure ONT approval by regulatory agencies. Citation Format: Todd Giardiello, Amy Anderson, Renee Salvo, Carl Ascoli. Delivery of oligonucleotide therapeutic drugs: Universal detection reagents specific for nucleic acid modifications abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 2055.