PhD Scientific Days 2025

Poster Session II. - B: Molecular Medicine

Utilizing miRNAscope™: A Novel Approach to Compare Transfection Efficiency of Antisense Oligonucleotides Across Cortical Cell Models

Előadó neve

Colar Zanjko Laura, MSc

Neptun code

G2NSTJ

Előadó munkahelye

BioTalentum Ltd, Gödöllő, Hungary

Előadó telefonszáma

+36705623866

Előadó e-mail címe

laura.colar@biotalentum.hu

Az előadás címe

Utilizing miRNAscope™: A Novel Approach to Compare Transfection Efficiency of Antisense Oligonucleotides Across Cortical Cell Models

Szerző(k) neve és munkahelye

Laura Colar Zanjko^1,2, Anna Meller¹, Andrea Balogh¹, János Farkas¹, Kriengkai Chessadangkul^1,3, Seppe Hermans⁴, Tobias Pöhlmann⁵, Willeke M. C. van Roon-Mom⁶, Melinda Zana¹, András Dinnyés^1,2

1: BioTalentum Ltd, Gödöllő, Hungary
2: Department of Physiology and Animal Health, Institute of Physiology and Animal Nutrition, Hungarian University of Agriculture and Life Sciences, Gödöllő, Hungary
3: Department of Biomedical Engineering, Faculty of Engineering, Chulalongkorn University, Bangkok, Thailand
4: BianoGMP GmbH, Gera, Thüringen, Germany
5: XNApharma GmbH, Gera, Thüringen, Germany
6: Leiden University Medical Centre, Leiden, The Netherlands

Bemutatás módja

Poszter

Szekció

Poster Session II. - B: Molecular Medicine

Language of the presentation

English

Preferred session

Molecular Medicine

Összefoglaló szövege

Antisense oligonucleotides (ASOs) are short, synthetic, single-stranded nucleic acid oligomers that bind with high specificity to target pre-mRNA or mRNA, thereby modulating gene expression through various mechanisms. ASOs represent a promising therapeutic approach for rare and ultra-rare genetic disorders, underscoring the need for sensitive and specific methods to evaluate their cellular uptake and functional efficacy. Traditional techniques for assessing transfection efficiency, such as qPCR or western blotting, provide only indirect or cumulative information. In contrast, RNAscope™, an advanced in situ hybridisation method, enables the visualisation of RNA at single-cell resolution using sequence-specific probes.
In this study, we assessed ASO transfection efficiency using miRNAscope™, a variant of RNAscope™ (Bio-Techne) optimised for the detection of small RNAs. Using the miRNAscope™ HD Reagent Kit in combination with fluorescence microscopy, we visualised and quantified the uptake of various ASOs, including gapmer and splice-switching variants. The cell models employed were derived from human induced pluripotent stem cells (iPSCs). These were monolayers of neural progenitor cells (NPCs), differentiated neural-astroglial co-cultures, microglial cultures, and neural-astroglial-microglial tri-cultures. To mimic the complexity of the developing brain, we incorporated neurospheroids into the experimental design. ASO delivery was tested via free uptake and lipid-mediated transfection.
Our results demonstrated dose-dependent transfection efficiency across all models, with higher uptake observed in differentiated cortical cultures than in NPCs. Quantitative analysis revealed lower signal intensity for gapmer ASOs than their splice-switching counterparts when detected with the same probe, likely reflecting reduced probe binding affinity to the gapmer structure. No differences were observed in the subcellular localisation of the ASOs; nuclear and cytoplasmic compartments were positively stained.
In conclusion, miRNAscope™ provides a robust and reliable platform for visualising ASOs in complex cortical cell models. This technique offers a sensitive and versatile tool for advancing ASO research and facilitating the development of RNA-based therapeutics.

This project is funded by the Horizon Europe MSCA scheme, MMM (No.101120256) and WhyNotDry (No.101131087).

University

Other, please specify in the next box

Other university, not listed above

Hungarian University of Agriculture and Life Sciences, Gödöllő, Hungary

Supervisor

Prof. Dr. András Dinnyés

Publication of my abstract

I give consent to the publication of my abstract on the website of the congress.

phd.section.field

in doctoral studies before complex exam (PhD)

Kind

Szabad

Status

elfogadva

Accepted presentation method

poszter

Előadás fájl jóváhagyás

nem rendelkezett róla

Előadó

9090

Start

18:18

End

18:24