Case Studies

The C5a anaphylatoxin chemotactic receptor 1 (C5aR, also known as CD88) belongs to the rhodopsin family of GPCRs and has been a topic of interest in the last decades due to its relevance in several inflammatory pathologies, such as asthma, arthritis, sepsis, and more recently Alzheimer’s disease and cancer. Even though the interaction between C5a and C5aR is of high therapeutic value, their molecular binding mechanism remains elusive, and several efforts are ongoing to find new drugs acting on C5aR.

One promising way of speeding up the identification of potential drugs acting on C5aR (being small molecules or biologicals as antibodies), is using fluorescence-based assays, like flow cytometry or fluorescence polarization. This kind of assays can be adapted to medium and high throughput screening with a read out easy to scale up. However, there is a major bottleneck in this approach: the availability of optimal fluorescent probes.

 In this case study we describe how, following the demand of the US-based biotech company Twist Bioscience (https://www.twistbioscience.com), we applied Celtarys technology to the development of two new fluorescent ligands for C5AR, opening the way to the use of fluorescence-based assays to study this promising target.

Among controlled substances, opioids have always been very well known. These substances act at the opioid receptors, which belong to the G protein-coupled receptors (GPCRs). μ -OR (or hMOR) is a class A G-protein-coupled receptors (GPCRs), which is considered the main responsible for both the analgesic and side-effects of the ligands. Thus, understanding the mechanism through which each of these effects happens is key in developing safer pain management drugs.

One way to improve understanding is by finding non-subjective methods of testing pain response. Such is the case with Prof. M. Galko’s work on the Drosophila larvae. They have identified fentanyl-induced anesthesia in these larvae, that happens through the activation of a fly receptor similar to the hMOR. Thus, a fluorescent probe for binding assays was required, with specific characteristics.

 In this case study we describe how, following the demand of Prof. M. Galko's group from the University of Texas (https://faculty.mdanderson.org/profiles/michael_galko.html), we applied our proprietary technology to the development of new fluorescent hMOR ligands, facilitating binding and microscopic studies for the receptor.

When measuring molecular interactions it is key to have simple and accurate assays. Fluorescence-based assays are among the most widely preferred by scientist, thanks to their versatility and sensitivity. Celtarys and Nanotemper are both specialized in this kind of assays: Celtarys develops unique fluorescent ligands for therapeutical targets such as GPCRs and Nanotemper develops advanced lab equipment for assessment of protein stability or high-throughput screening.

In this collaboration between Celtarys and Nanotemper, we have combined Celtarys Adenosine A2A fluorescent ligand, CELT-300, with Nanotemper Monolith X developing an easy to perform and reproducible competitive binding assay with unpurified GPCR

Cannabinoid receptors, CB1 and CB2, lead to diverse cellular events mediated through different cell signaling pathways, and therefore cannabinoids are potential tools to combat a variety of diseases including neurological or metabolic conditions. CB1 receptors are mainly found in the brain and central nervous system, where they play a role in regulating mood, memory, pain, appetite and motor function. This interest is driving efforts to understand the pharmacology of naturally occurring and synthetic cannabinoids at the receptor level.

Here, we report a TR-FRET assay combining the PHERAstar FSX microplate reader from BMG Labtech and the CELT- 335 fluorescent ligand developed by Celtarys Research, that allows to accurately measure binding affinities for cannabinoid and other GPCR receptors