GPR3, GPR6, and GPR12 are three orphan receptors that belong to the Class A family of G-protein-coupled receptors (GPCRs). These GPCRs share over 60% of sequence similarity among them. Because of their close phylogenetic relationship, GPR3, GPR6, and GPR12 share a high percentage of homology with other lipid receptors such as the lysophospholipid and the cannabinoid receptors. On the basis of sequence similarities at key structural motifs, these orphan receptors have been related to the cannabinoid family. However, further experimental data are required to confirm this association. GPR3, GPR6, and GPR12 are predominantly expressed in mammalian brain. Their high constitutive activation of adenylyl cyclase triggers increases in cAMP levels similar in amplitude to fully activated GPCRs. This feature defines their physiological role under certain pathological conditions. In this review, we aim to summarize the knowledge attained so far on the understanding of these receptors. Expression patterns, pharmacology, physiopathological relevance, and molecules targeting GPR3, GPR6, and GPR12 will be analyzed herein. Interestingly, certain cannabinoid ligands have been reported to modulate these orphan receptors. The current debate about sphingolipids as putative endogenous ligands will also be addressed. A special focus will be on their potential role in the brain, particularly under neurological conditions such as Parkinson or Alzheimer’s disease. Reported physiological roles outside the central nervous system will also be covered. This critical overview may contribute to a further comprehension of the physiopathological role of these orphan GPCRs, hopefully attracting more research towards a future therapeutic exploitation of these promising targets.

Background: The glyoxalase system including two thiol-dependent enzymes, glyoxalase I (Glo-I) and glyoxalase II, plays an important role in a ubiquitous metabolic pathway involved in cellular detoxification of cytotoxic 2-oxoaldehydes. Tumor cells have high glycolytic activity, leading to increased cellular levels of these toxic metabolites. The increased activity of the detoxification system in cancerous cells makes this pathway a viable target for developing novel anticancer agents. In this study, we examined the potential utility of non-glutathione-based inhibitors of the Glo-I enzyme as novel anticancer drugs. Methods: Computer-aided drug design techniques, such as customized pharmacophoric features, virtual screening, and flexible docking, were used to achieve the project goals. Retrieved hits were extensively filtered and subsequently docked into the active site of the enzyme. The biological activities of retrieved hits were assessed using an in vitro assay against Glo-I. Results: Since Glo-I is a zinc metalloenzyme, a customized Zn-binding pharmacophoric feature was used to search for selective inhibitors via virtual screening of a small-molecule database. Seven hits were selected, purchased, and biologically evaluated. Three of the seven hits inhibited Glo-I activity, the most effective of which exerted 76.4% inhibition at a concentration of 25 µM. Conclusion: We successfully identified a potential Glo-I inhibitor that can serve as a lead compound for further optimization. Moreover, our in silico and experimental results were highly correlated. Hence, the docking protocol adopted in this study may be efficiently employed in future optimization steps.

Wheldone (1) was isolated and elucidated from a coculture of Aspergillus fischeri (NRRL 181) and Xylaria flabelliformis (G536), where secondary metabolite biosynthesis was stimulated by antagonism between these fungi. First observed via in situ analysis between these competing fungal cultures, the conditions were scaled to reproducibly generate 1, whose novel structure was elucidated by one- and two-dimensional NMR and mass spectrometry. Compound 1 displayed cytotoxic activity against breast, ovarian, and melanoma cancer cell lines.

GPR6 is an orphan G protein-coupled receptor that has been associated with the cannabinoid family because of its recognition of a sub-set of cannabinoid ligands. The high abundance of GPR6 in the central nervous system, along with high constitutive activity and a link to several neurodegenerative diseases make GPR6 a promising biological target. In fact, diverse research groups have demonstrated that GPR6 represents a possible target for the treatment of neurodegenerative disorders such as Parkinson’s disease, Alzheimer’s disease, and Huntington’s disease. Several patents have claimed the use of a wide range of pyrazine derivatives as GPR6 inverse agonists for the treatment of Parkinson’s disease symptoms and other dyskinesia syndromes. However, the full pharmacological importance of GPR6 has not yet been fully explored due to the lack of high potency, readily available ligands targeting GPR6. The long-term goal of the present study is to develop such ligands. In this paper, we describe our initial steps towards this goal. A human GPR6 homology model was constructed using a suite of computational techniques. This model permitted the identification of unique GPR6 structural features and the exploration of the GPR6 binding crevice. A subset of patented pyrazine analogs were docked in the resultant GPR6 inactive state model to validate the model, rationalize the structure-activity relationships from the reported patents and identify the key residues in the binding crevice for ligand recognition. We will take this structural knowledge into the next phase of GPR6 project, in which scaffold hopping will be used to design new GPR6 ligands.

Sodium–glucose cotransporter 2 (SGLT2) inhibitors are a relatively new class of antidiabetic agents that lower blood glucose independently of insulin by inhibiting renal SGLT2 in the proximal tubule, thereby increasing urinary glucose and sodium excretion. Empagliflozin (Empa), an FDA-approved SGLT2 inhibitor, exhibits antioxidant, anti-inflammatory, and additional metabolic effects beyond glycemic control. Given the high expression of SGLT2 in the central nervous system and the established link between cognitive impairment, chronic hyperglycemia, oxidative stress, and inflammation, this study investigated Empa’s neuroprotective potential on learning and memory deficits in streptozotocin-induced hyperglycemic male Wistar rats. Hyperglycemia was induced using streptozotocin (40 mg/kg/IP), followed by Empa treatment (10 mg/kg/day/PO). Cognitive performance was evaluated using the radial arm water maze, assessing learning and both short-term and long-term memory. Concurrently, hippocampal oxidative stress markers and key molecular mediators, including brain-derived neurotrophic factor (BDNF) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), were measured to elucidate possible underlying neuroprotective mechanisms. Hyperglycemic rats exhibited significant impairments in learning, short-term memory, and long-term memory compared to normoglycemic controls. Empa treatment significantly improved short-term memory, restoring performance to near-control levels. However, its impact on long-term memory was minimal. Unexpectedly, Empa induced only modest, non-statistically significant changes in hippocampal oxidative stress markers and BDNF and NF-κB levels. The findings underscore the complexity of oxidative stress and inflammatory pathways involved in hyperglycemia-associated cognitive impairment. The beneficial effects of Empa on short-term memory may involve alternative mechanisms unrelated to oxidative stress modulation. Further studies involving extended durations, higher dosages, or larger sample sizes are warranted to better elucidate the neuroprotective mechanisms of Empa.

Hematopoietic progenitor kinase 1 (HPK1), a negative regulator of T-cells, B-cells, and dendritic cells, has gained attention in antitumor immunotherapy research over the past decade. No HPK1 inhibitor has yet reached clinical approval, largely due to selectivity and drug-like limitations. Leveraging the available structural insights into HPK1, we conducted a rational hit identification using a structure-based virtual screening of over 600,000 drug-like molecules from ASINEX and OTAVA databases. A series of molecular docking studies, in vitro kinase assays, and molecular dynamics simulations were conducted to identify viable HPK1 inhibitor hits. This approach resulted in two promising novel hit scaffolds, 4H-Pyrido[1,2-a] thieno[2,3-d] pyrimidin-4-one (ISR-05) and quinolin-2(1H)-one (ISR-03), neither of which has previously been reported as an HPK1 inhibitor. ISR-05 and ISR-03 exhibited IC50 values of 24.2 ± 5.07 and 43.9 ± 0.134 µM, respectively, in kinase inhibition assays. These hits constitute tractable starting points for future hit-to-lead optimization aimed at developing more effective HPK1 inhibitors for cancer therapy.