Abstract 3630: RNA nanoparticles as a carrier for targeted drug delivery into cancer cells
Authors: Rychahou Piotr G. , Guo Sijin , Lee Eun Y. , Rychagov Nicole , Thorson Jon , Guo Peixuan , Evers B. Mark 
Published: August 2019

Abstract
Colorectal cancer (CRC) is the second leading cause of cancer deaths in the US; the inefficiency of chemotherapeutic treatments has severely limited the treatment options in patients with CRC metastasis. Polyvalent RNA nanoparticles have demonstrated metastatic tumor homing without accumulation in normal organ tissues surrounding metastatic tumors; the flexibility in constructing trimers also enables the assembly of polyvalent nanoparticles to carry drug molecules for therapeutic purposes. The purpose of our study was to: (i) demonstrate PI3K/mTOR inhibitor conjugation to RNA nanoparticles, and (ii) pH-sensitive intracellular drug delivery by folate receptor targeting (FA)-3WJ-PI-103 nanoparticles in CRC cells.
Methods. (1) PI-103-3WJ-Folate nanoparticle binding, internalization and drug release was evaluated in HT-29 (human CRC), SK-OV-3 (human ovarian cancer), JAR (human choriocarcinoma), HCT116 (human CRC), and Caco-2 (human CRC) cell lines. (2) Confocal microscopy was used to examine RNA nanoparticle binding and internalization by endocytosis. (3) Western blot was used to confirm drug delivery with PI-103-3WJ-Folate nanoparticle.
Results. (1) Drug release from fully assembled RNA nanoparticles was confirmed after 3WJ-PI-103 nanoparticles (5 µM) incubation in PBS pH 2.5, 3, 3.5 for 4h at 37°C and analysis of pAkt (Ser473) expression, a marker of PI3K inhibition; 1.5 mM 3WJ-PI-103 was minimum dose required to inhibit PI3K/Akt signaling in cancer cell lines. (2) Rapid folate-receptor mediated RNA nanoparticle internalization was observed within 2-4 h in HT29, HCT116, JAR and SK-OV-3 cell lines by confocal microscopy. (3) Cancer cells were treated with FA-3WJ-PI-103 nanoparticles directly to evaluate all steps of targeted receptor-mediated drug delivery-receptor binding, endocytosis, pH-triggered drug release and endosomal drug escape into the cytoplasm. HCT116 and JAR cells were treated with FA-3WJ-PI-103 in 0% and 1% folate-free media for 4h, media was replaced and evaluated 3h later for PI3K pathway activation. PI3K/Akt signaling was inhibited after direct treatment with FA-3WJ-PI-103 nanoparticles at 5 mM drug concentration; 3WJ-PI-103 particles were used as a control.
Conclusions. A critical finding of our current study was the ability to conjugate PI3K/mTOR inhibitor PI-103, via a pH-selective linker, to RNA nanoparticles and the demonstration of intracellular drug delivery into cancer cells and drug release in a pH-dependent fashion. Our results demonstrate the potential of receptor-selective drug delivery to cancer cells with high FRα expression and represents a promising approach for treatment of CRC metastases.
Citation Format: Piotr G. Rychahou, Sijin Guo, Eun Y. Lee, Nicole Rychagov, Jon Thorson, Peixuan Guo, B. Mark Evers. RNA nanoparticles as a carrier for targeted drug delivery into cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3630.