Achim Aigner

Background: Glioblastomas (GBMs) are highly invasive and heterogeneous in nature which strongly contributes to poor survival of patients, averaging 15 months after diagnosis. With little treatment options being available in particular for addressing GBM invasiveness, there is a high need for supplementary therapies in addition to classical cytostatics like Temozolomide (TMZ). Methods: The aim of this study was to examine promising compounds, the natural indole scaffold RAD114 and three semisynthetic analogs (RAD118, RAD119, RAD120) as well as the aryl sulfonamide AqB011, for their inhibitory potential specifically on tumor cell invasion. For this, GBM xenograft tissue slice tandem co-culture (XTCC) models were employed, closely reflecting the in vivo patient situation. In particular, these ex vivo tissue slice air-liquid interface (ALI) models also allow for distinguishing between inhibitory effects on bulk tumor growth vs tumor cell invasion into the normal brain tissue. Results: Profound anti-invasive effects of the natural indole scaffold RAD114, three semisynthetic analogs (RAD118, RAD119, RAD120), and the aryl sulfonamide AqB011 were observed in XTCC models based on G55T2 and U87-MG GBM xenografts co-cultured with mouse brain slices in ALI tandem co-culture. While effects on bulk tumor growth were rather moderate, profound inhibition or even abolishment of tumor invasion was observed, dependent on the compound, its dosage and the model. At a non-toxic concentration (10 µM), RAD118 and AqB011 showed the highest effectiveness in reducing tumor invasion. Combination effects at sub-optimal compound dosages were observed as well. Conclusions: Both compound classes - the tested semisynthetic indole analogs and the sulfonamide derivative – may be promising lead candidates for drugs useful to specifically address the issue of GBM invasiveness. They strongly restrained GBM cell invasion, alone and in combination. Rather than acting through cytotoxic effects, they specifically inhibit cell migration into native brain tissue.

Ferroptosis in gastrointestinal stromal tumors

Gastric cancer remains one of the most lethal malignancies worldwide, with high relapse rates and limited survival for patients with advanced disease. Despite advances in targeted therapies and immune checkpoint inhibition, intrinsic tumor heterogeneity poses challenges for effective treatment. The HER3 receptor (ERBB3) has emerged as an important player in cancer progression, contributing to aggressive tumor behavior and poor prognosis. Recent evidence indicates that activating ferroptosis—an iron-dependent, non-apoptotic form of cell death—offers a promising strategy to inhibit cancer growth. In gastric cancer, ferroptosis plays a crucial role, and promoting this process may open new avenues for therapeutic intervention. Ferroptosis is characterized by iron-mediated lipid peroxidation of cell membranes and is critically regulated by the cystine/glutamate antiporter system (SLC7A11) and glutathione peroxidase 4 (GPX4). Our study aimed to investigate the relationship between ERBB3 and ferroptosis in gastric cancer. We found that high ERBB3 expression correlated with resistance to ferroptosis-inducing agents, including GPX4 and SLC7A11 inhibitors, across multiple cell lines. Vice versa, ERBB3 inhibition with TX1-85-1 induced lipid peroxidation in gastric cancer cells, with effects most pronounced in cell lines expressing higher SLC7A11 levels. Knockdown of ERBB3 reproduced these effects, suggesting SLC7A11 as a predictive marker. Importantly, combined inhibition of ERBB3 and GPX4 significantly enhanced lipid peroxidation and cytotoxicity, while ERBB3 activation by co-treatment with the ERBB3 ligand heregulin reduced lipid peroxidation in cells with lower baseline SLC7A11 expression. Analysis of glutathione levels and SLC7A11 expression further supported the role of ERBB3 in modulating ferroptosis sensitivity. These findings identify ERBB3 as a critical regulator of ferroptosis and a promising target for enhancing ferroptosis-mediated cell death. Its inhibition in combination with ferroptosis inducers may thus represent a particularly promising and efficacious therapeutic strategy in gastric cancer.

Purpose Human epidermal growth factor receptor 2 (HER2) inhibition represents a therapeutic approach with proven clinical efficacy in gastric cancer. However, resistance against HER2-directed therapeutics highlights the need for alternative approaches or drug combinations. Histone deacetylase inhibitors (HDACi) display a broad spectrum of antitumor properties, which may include effects on receptor tyrosine kinases.Materials and Methods We analyzed the effects of the class I HDACi entinostat in a panel of HER2-amplified and non-amplified gastric adenocarcinoma cells in 2D cell culture as well as in tumor slice models ex vivo and in patient-derived xenografts in vivo. Effects on protein expression/signal transduction were evaluated by immunoblotting and quantitative reverse transcription polymerase chain reaction.Results HDAC inhibition reduced HER2 protein expression independently of initial HER2 expression levels. This was associated with the upregulation of the HER2-inhibiting microRNA miR-205. The downregulation of HER2 resulted in reduced AKT phosphorylation, apoptosis induction and antiproliferative effects, with particularly high efficiency in HER2-amplified gastric cancer cells. Inhibiting HER2 by a specific kinase inhibitor in gastric cancer cells with low basal HER2 expression led to HER2 upregulation. This was reversed by entinostat treatment and provided the basis for synergistic cell inhibition upon double treatment.Conclusion We describe the downregulation of HER2 in gastric carcinoma cells upon HDACi treatment. Concomitantly, cells with high basal or treatment-induced HER2 expression showed most profound sensitivities towards HDACi. These findings may thus provide the basis for HDACi treatment as a therapeutic option particularly valuable in HER2-amplified gastric cancer and particularly useful in combination therapies with HER2 inhibitors.

Background: Polymeric nanoparticles have been explored as efficient tools for siRNA delivery to induce RNAi-mediated gene knockdown. Chemical modifications of polyethylenimines (PEI) enhance nanoparticle efficacy and biocompatibility. Their in vivo use, however, benefits from prior analyses in relevant in vitro 3D conditions. Methods: We utilize a 3D ALI cell culture model for testing the biological activities and toxicities of a set of different PEI-based nanoparticles with different chemical modifications. This also includes a novel, fluoroalkyl-modified PEI. Reporter gene knockdown is directly compared to 2D cell culture. In parallel, biocompatibility is assessed by measuring cell viability and lactate dehydrogenase (LDH) release. Results: Knockdown efficacies in the 3D ALI model are dependent on the chemical modification and complex preparation conditions. Results only correlate in part with gene knockdown in 2D cell culture, identifying nanoparticle penetration and cellular internalization under 3D conditions as important parameters. The 3D ALI cell culture is also suitable for the quantitative determination of nanoparticle effects on cell viability and acute toxicity, with biocompatibility benefitting from PEI modifications. Conclusions: The 3D ALI cell model allows for a more realistic assessment of biological nanoparticle effects. A novel fluoroalkyl-modified PEI is described. Optimal preparations of PEI-based nanoparticles for siRNA delivery and gene knockdown are identified.

The development of in vitro hepatocyte cell culture systems is crucial for investigating drug-induced liver injury (DILI). One prerequisite for monitoring DILI related immunologic reactions is the extension of primary human hepatocyte (PHH) cultures towards the inclusion of macrophages. Therefore, we developed and characterized an autologous co-culture system of PHH and primary human hepatic macrophages (hepM) (CoC1). We compared CoC1 with a co-culture of the same PHH batch + M0 macrophages derived from THP1 cells (CoC2) in order to represent a donor independent macrophage reaction. Then, we treated the mono- and co-cultures with drugs that cause DILI—menadione (MEN, 1 or 10 µM, 3 h), diclofenac (DIC, 0.5 or 5 mM, 6 h), or acetaminophen (APAP, 0.5 or 5 mM, 6 h)—and assessed culture stability, cell activity, macrophage differentiation, cytokine production and cell viability. Without drug treatment, CoC1 was the most stable over a culture time of up to 60 h. Cytokine array analysis revealed a proinflammatory profile of PHH mono-cultures due to isolation stress but showed different influences of hepM and M0 on the cytokine profile in the co-cultures. MEN, DIC and APAP treatment led to donor-dependent signs of cell stress and toxicity. HepM can either promote or reduce the DILI effects donor dependently in CoC1. CoC2 are slightly less sensitive than CoC1 in representing DILI. In summary, we present a new autologous co-culture system that can mimic DILI in a donor-dependent manner. This cellular system could be useful for new drug testing strategies and reducing animal testing.