Research Paper Volume 14, Issue 5 pp 2081—2100
Modulation of RNA splicing associated with Wnt signaling pathway using FD-895 and pladienolide B
- 1 Moores Cancer Center, University of California San Diego, La Jolla, CA 92093, USA
- 2 ThermoFisher Scientific, Carlsbad, CA 92008, USA
- 3 Amity Stem Cell Institute, Amity Medical School, Amity University Haryana, Panchgaon (Manesar), Haryana 122413, India
- 4 Department of Chemistry and Biochemistry, University of California San Diego, La Jolla, CA 92093, USA
- 5 CLL Research Consortium and Department of Medicine, University of California San Diego, La Jolla, CA 92093, USA
- 6 Hematology-Oncology Division, Mayo Clinic, Phoenix, AZ 85054, USA
Received: December 28, 2021 Accepted: February 22, 2022 Published: March 1, 2022
https://doi.org/10.18632/aging.203924How to Cite
Copyright: © 2022 Kumar et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Abstract
Alterations in RNA splicing are associated with different malignancies, including leukemia, lymphoma, and solid tumors. The RNA splicing modulators such as FD-895 and pladienolide B have been investigated in different malignancies to target/modulate spliceosome for therapeutic purpose. Different cell lines were screened using an RNA splicing modulator to test in vitro cytotoxicity and the ability to modulate RNA splicing capability via induction of intron retention (using RT-PCR and qPCR). The Cignal Finder Reporter Array evaluated [pathways affected by the splice modulators in HeLa cells. Further, the candidates associated with the pathways were validated at protein level using western blot assay, and gene-gene interaction studies were carried out using GeneMANIA. We show that FD-895 and pladienolide B induces higher apoptosis levels than conventional chemotherapy in different solid tumors. In addition, both agents modulate Wnt signaling pathways and mRNA splicing. Specifically, FD-895 and pladienolide B significantly downregulates Wnt signaling pathway-associated transcripts (GSK3β and LRP5) and both transcript and proteins including LEF1, CCND1, LRP6, and pLRP6 at the transcript, total protein, and protein phosphorylation’s levels. These results indicate FD-895 and pladienolide B inhibit Wnt signaling by decreasing LRP6 phosphorylation and modulating mRNA splicing through induction of intron retention in solid tumors.
Abbreviations
CCND1: Cyclin D1; CTNNB1: Catenin, beta-1; CLL: Chronic lymphocytic leukemia; DLT: Dose limiting toxicity; FBS: Fetal bovine serum; FN1: Fibronectin 1; GeneMANIA: Gene Multiple Association Network Integration Algorithm; GSK3: Glycogen synthase kinase-3; IR: Intron retention; LEF1: Lymphoid enhancer-binding factor-1; LRP6: LDL Receptor Related Protein 6; MCL: Mantle cell lymphoma; MTD: Maximum tolerated dose; PBMCs: Peripheral blood mononuclear cells; RT-PCR: Reverse transcriptase PCR; SF3B1: Splicing Factor 3b Subunit 1; SPLMs: Splicing modulators; SRSF2: Serine And Arginine Rich Splicing Factor 2; U2AF1: U2 Small Nuclear RNA Auxiliary Factor 1.