Biology subjects

Scharner, J.

Publications and source records attributed to Scharner, J..

3 recordsLinked to original sources

bioRxiv2020

ASO-based PKM Splice-switching Therapy Inhibits Hepatocellular Carcinoma Cell Growth

The M2 pyruvate kinase (PKM2) isoform is upregulated in most cancers and plays a crucial role in the Warburg effect, which is characterized by the preference for aerobic glycolysis for energy metabolism. PKM2 is an alternative-splice isoform of the PKM gene and is a potential therapeutic target. Previously, we developed antisense oligonucleotides (ASOs) that switch PKM splicing from the cancer-associated PKM2 to the PKM1 isoform and induce apoptosis in cultured glioblastoma cells. Here, we explore the potential of ASO-based PKM splice-switching as a targeted therapy for liver cancer. We utilize a more potent lead cEt/DNA ASO and demonstrate that it induces PKM splice-switching and inhibits the growth of cultured hepatocellular-carcinoma (HCC) cells. This PKM isoform switch increases pyruvate-kinase activity and alters glucose metabolism. The lead ASO and a second ASO targeting a non-overlapping site inhibit tumorigenesis in an orthotopic-xenograft HCC mouse model. Finally, a surrogate mouse-specific ASO induces Pkm splice-switching and inhibits HCC growth, without observable toxicity, in a genetic HCC mouse model. These results lay the groundwork for a potential ASO therapy for HCC. Statement of significanceAntisense oligonucleotides are used to force a change in PKM isoform usage in HCC, reversing the Warburg effect and inhibiting tumorigenesis.

cancer biology↗

bioRxiv2018

Delivery of GalNAc-conjugated splice-switching ASOs to non-hepatic cells through ectopic expression of asialoglycoprotein receptor

Splice-switching antisense oligonucleotides (ASOs) are promising therapeutic tools to target various genetic diseases, including cancer. However, in vivo delivery of ASOs to orthotopic tumors in cancer mouse models or to certain target tissues remains challenging. A viable solution already in use is receptor-mediated uptake of ASOs via tissue-specific receptors. For example, the asialoglycoprotein receptor (ASGP-R) is exclusively expressed in hepatocytes. Triantennary GalNAc (GN3)-conjugated ASOs bind to the receptor and are efficiently internalized by endocytosis, enhancing ASO potency in the liver. Here we explore the use of GalNAc-mediated targeting to deliver therapeutic splice-switching ASOs to cancer cells that ectopically express ASGP-R, both in vitro and in tumor mouse models. We found that ectopic expression of the major isoform ASGP-R1 H1a is sufficient to promote uptake and increase GN3-ASO potency to various degrees in all tested cancer cells. We show that cell-type specific glycosylation of the receptor does not affect its activity. In vivo, GN3-conjugated ASOs specifically target subcutaneous xenograft tumors that ectopically express ASGP-R1, and modulate splicing significantly more strongly than unconjugated ASOs. Our work shows that GN3-targeting is a useful tool for proof-of-principle studies in orthotopic cancer models, until endogenous receptors are identified and exploited for efficiently targeting cancer cells.

molecular biology↗

bioRxiv2017

A human-specific switch of alternatively spliced AFMID isoforms contributes to TP53 mutations and tumor recurrence in hepatocellular carcinoma

Pre-mRNA splicing can contribute to the switch of cell identity that occurs in carcinogenesis. Here we analyze a large collection of RNA-Seq datasets and report that splicing changes in hepatocyte-specific enzymes, such as AFMID and KHK, are associated with HCC patients survival and relapse. The switch of AFMID isoforms is an early event in HCC development, and is associated with driver mutations in TP53 and ARID1A. Finally, we show that the switch of AFMID isoforms is human-specific and not detectable in other species, including primates. The integrative analysis uncovers a mechanistic link between splicing switches, de novo NAD+ biosynthesis, driver mutations, and HCC recurrence.

bioinformatics↗