transMed is an integrated training network funded by the European Union to train the next generation of scientists in translational medicine. The consortium focuses on bench-to-bedside development of treatments for inherited retinal degenerative diseases (IRDs) and has academic as well as non-academic partners which collaborate in all aspects of translational research, including disease mechanisms, identification of drug candidates, drug development and delivery. Akanksha Roy, transMed PhD candidate on behalf of PamGene International, and François Paquet-Durand, Professor at the University of Tübingen’s Institute for Ophthalmic Research recently published a review paper which described key pathways involved in IRDs and their study by transcriptomic, proteomic, and metabolomic techniques . The paper also discussed the relevance of PamGene’s multiplex peptide microarray technology for the field of IRDs; a technology that could not only generate hypotheses for specific kinases active in retinal degeneration but also identify novel druggable targets.
Common targets and mutation-independent treatments
The high genetic diversity of IRDs, with mutations in over 270 genes leading to vision loss, is impeding development of treatment options. However, analysis of the involved genetic defects revealed that distinct mutations may converge on a common cellular signaling pathway. “From the literature and through our own research we know that the pathways triggering photoreceptor cell death involve cGMP signaling and its major target – Protein Kinase G (PKG),” tells Prof. Paquet-Durand.
“Closer to real-life”
“In this paper, we described different techniques that are applied to unravel affected pathways in IRDs, with the objective to identify druggable targets that could benefit larger subgroups of patients,’’ tells Roy. Amongst the technologies mentioned in this publication were gene expression assays, mass-spectrometry based protein identification methods, in situ enzyme activity assays, and multiplex microarray technology. “The concordance between gene and protein expression studies is low as the mRNA detected might not be converted to proteins. Therefore, it is important to study signaling pathways at the protein level. However, protein detection techniques such as Western Blot and enzyme-linked immunosorbent assays detect protein presence but not protein activity. PamGene’s multiplex peptide microarrays allow real-time detection of kinase activity of both recombinant enzymes and complex mixtures of kinases,” explains Roy. “PamGene’s technology proved to be very handy as genomic technologies are unable to inform what the level of protein activity is. PamGene’s technology brought us much closer to what actually is happening, closer to real-life,” adds Paquet-Durand.
The power of joining forces
In an earlier publication, Roy et al. applied recombinant PKGs on PamGene’s multiplex peptide microarrays to identify about 50 novel PKG targets . Next, in collaboration with Paquet-Durand’s group, PamGene’s technology was used to identify peptides whose phosphorylation significantly decreased in murine retinal explants of IRD when treated with a novel PKG inhibitor. Ex-vivo analysis of key affected proteins corresponding to the differentially phosphorylated peptides, confirmed their expression in different cell layers of retina tissue. The preprint of this paper is available on bioRxiv .
“The power of combining knowledge, results, and technologies via projects like transMed is immense for rare diseases like IRDs. It not only results in potential new treatment options but also trains the next generation of highly skilled scientists about how joining a broad scientific perspective and collaboration can translate basic research into new medicines for society. PamGene’s technology has and will play an important role in this,” concludes Paquet-Durand.
- Roy A, Hilhorst R, Groten J, Paquet-Durand F, Tomar T. Technological advancements to study cellular signaling pathways in inherited retinal degenerative diseases. Current Opinion in Pharmacology. 2021 Oct 1;60:102-10.
- Roy A, Groten J, Marigo V, Tomar T, Hilhorst R. Identification of novel substrates for cGMP dependent protein kinase (PKG) through kinase activity profiling to understand its putative role in inherited retinal degeneration. International journal of molecular sciences. 2021 Jan;22(3):1180.
- Roy A, Tolone A, Hilhorst R, Groten J, Tomar T, Paquet-Durand F. .Kinase activity profiling identifies putative downstream targets of cGMP/PKG signaling in inherited retinal neurodegeneration