Geissospermum species are medically important plants due to their health‐promoting effects. The objective of this study was to determine the antioxidant ability and antiproliferative and cytotoxic effects of infusions, tinctures, and ethanolic extracts of Geissospermum reticulatum barks in relation to the contents of total phenolics and flavonoids. Seven samples of barks were collected in various regions of Peruvian Amazonia. We found that the amount of total phenolics in the studied products varied from 212.40 ± 0.69 to 1253.92 ± 11.20 mg GAE/kg. In our study there is a correlation (R² = 0.7947) between the results of antioxidants assays: FRAP and ORAC for tinctures, infusions, and ethanolic extracts of G. reticulatum barks. We have also observed antiproliferative activities of the ethanolic extracts on normal T‐cells. These extracts have caused death on malignant cell lines (THP‐1 and HL‐60) and this data correlates well with their antioxidant capacity measured by ORAC method. Interestingly, the highest concentration of the ethanolic extract was not toxic in the zebrafish embryo developmental assay. Our results indicate that G. reticulatum is rich in antioxidants and have cytotoxic and antiproliferative properties. The data suggests potential immunosuppressive role of the extracts. This is the first study presenting the results of chemical and biological analysis of multiple preparations from G. reticulatum.

Background and Purpose: Wnt binding to Frizzleds (FZD) is a crucial step that leads to the initiation of signalling cascades governing multiple processes during embryonic development, stem cell regulation and adult tissue homeostasis. Recent efforts have enabled us to shed light on Wnt–FZD pharmacology using overexpressed HEK293 cells. However, assessing ligand binding at endogenous receptor expression levels is important due to differential binding behaviour in a native environment. Here, we study FZD paralogue, FZD₇, and analyse its interactions with Wnt‐3a in live CRISPR‐Cas9‐edited SW480 cells typifying colorectal cancer.Experimental Approach: SW480 cells were CRISPR‐Cas9‐edited to insert a HiBiT tag on the N‐terminus of FZD₇, preserving the native signal peptide. These cells were used to study eGFP‐Wnt‐3a association with endogenous and overexpressed HiBiT‐FZD₇ using NanoBiT/bioluminescence resonance energy transfer (BRET) and NanoBiT to measure ligand binding and receptor internalization.Key Results: With this new assay the binding of eGFP‐Wnt‐3a to endogenous HiBiT‐FZD₇ was compared with overexpressed receptors. Receptor overexpression results in increased membrane dynamics, leading to an apparent decrease in binding on‐rate and consequently in higher, up to 10 times, calculated K_d. Thus, measurements of binding affinities to FZD₇ obtained in overexpressed cells are suboptimal compared with the measurements from endogenously expressing cells.Conclusions and Implications: Binding affinity measurements in the overexpressing cells fail to replicate ligand binding affinities assessed in a (patho)physiologically relevant context where receptor expression is lower. Therefore, future studies on Wnt–FZD₇ binding should be performed using receptors expressed under endogenous promotion.

Bioluminescence- and fluorescence-based resonance energy transfer assays have gained considerable attention in pharmacological research as high-throughput scalable tools applicable to drug discovery. To this end, G protein-coupled receptors represent the biggest target class for marketed drugs, and among them, orphan G protein-coupled receptors have the biggest untapped therapeutic potential. In this review, the cases where biophysical methods, BRET and FRET, were employed for deorphanization and ligand discovery studies on orphan G protein-coupled receptors are listed and discussed.

GPR61 is an orphan receptor that belongs to Class A of G-protein-coupled receptors. It has been reported that GPR61 has a constitutive activity and couples to Gαs. In the present study, we characterized GPR61 function and ligand binding by experimental and molecular docking studies. We demonstrated that heterologous expression of GPR61 in HEK293 cells enhanced the cAMP synthesis response to forskolin, whereas the basal cAMP synthesis was unaffected. 5-(Nonyloxy)tryptamine inhibited forskolin-stimulated cAMP production in GPR61-expressing HEK293 cells. These studies highlight that the intrinsic activity of this receptor is only measurable following its synergy with Gαs.

Fibrillarin (FBL), a core component of the C/D box small nucleolar ribonucleoprotein (snoRNP) complex, catalyzes the 2'-O-methylation (Nm) of the ribose 2'-hydroxyl moiety in ribosomal RNA (rRNA). Distinct Nm patterns contribute to ribosome heterogeneity, which is linked to selective translation of oncogenes. FBL dysregulation generates an aberrant Nm signature in triple-negative breast cancer (TNBC), the most aggressive breast cancer subtype. This study investigated the role of FBL in TNBC via translation-driven mechanisms. Our findings show that FBL knockdown impairs oncogenic traits, triggers metabolic stress, and reduces the translation efficiency of oncogenes, such as metastasis-associated protein 1 (MTA1), interleukin-1 receptor-associated kinase 1 (IRAK1), and thymosin beta 10 (TMSB10). RiboMethSeq confirmed that the rRNA Nm sites exhibited differential sensitivity to FBL depletion. Additionally, FBL knockdown led to alterations in 18S ribosome structure confirmed by SHAPE and specifically reduced RPS28 incorporation into ribosomes. Notably, silencing RPS28 also disrupted both the oncogenic phenotype and downregulated MTA1, IRAK1, and TMSB10 expression. These findings reveal a complex interplay between FBL, rRNA Nm modifications, and RPS28 in shaping oncogenic protein pools and ribosomal composition in TNBC, offering promising insights into therapeutic approaches targeting this aggressive cancer subtype.