Orphan G Protein-Coupled Receptor GPR37 as an Emerging Therapeutic Target

Abstract

G protein-coupled receptors (GPCRs) are successful druggable targets, making up around 35% of all FDA-approved medications. However, a large number of receptors remain orphaned, with no known endogenous ligand, representing a challenging but untapped area to discover new therapeutic targets. Among orphan GPCRs (oGPCRs) of interest, G protein-coupled receptor 37 (GPR37) is highly expressed in the central nervous system (CNS), particularly in the spinal cord and oligodendrocytes. While its cellular signaling mechanisms and endogenous receptor ligands remain elusive, GPR37 has been implicated in several important neurological conditions, including Parkinson’s disease (PD), inflammation, pain, autism, and brain tumors. GPR37 structure, signaling, emerging physiology, and pharmacology are reviewed while integrating a discussion on potential therapeutic indications and opportunities.

Graphical Abstract

1. INTRODUCTION

The G protein-coupled receptor (GPCR) superfamily comprises a diverse group of transmembrane receptors that detect extracellular molecules and initiate cellular responses.¹ GPCRs are extensively studied and represent a significant focus for drug development, with approximately 35% of FDA-approved medications targeting these receptors.^{2, 3} Their crucial role in regulating normal physiological functions, involvement in human pathophysiology, and suitability as pharmacological targets make them highly appealing.⁴ Despite the identification of around 800 different GPCRs in mammals, only a quarter of them have been utilized for therapeutic purposes, and the function of ~200 GPCRs is unknown.⁴ In the nervous system, neurotransmitters exert their effects through either individual GPCRs or a group of GPCRs known as neuromodulator receptor families.^{5, 6} The importance of GPCR signaling in maintaining proper brain function is evident from the fact that numerous neuropsychiatric disorders and neurological diseases arise from dysfunctional GPCRs. Consequently, the primary focus of drugs designed to treat nervous system disorders lies in targeting GPCRs.⁷

The rhodopsin, or Class A, family of GPCRs offers the largest pool of therapeutic targets, encompassing 719 receptors in humans.⁸ Notably, 29% of FDA-approved drugs target these rhodopsin-like GPCRs, and approximately 500 novel GPCR drug candidates are presently undergoing clinical trials.^{9, 10} Despite advancements, a substantial number of rhodopsin-like receptors are categorized as “orphans”. These orphan receptors, around 120 in number, still possess mostly unknown physiological functions and lack identified endogenous ligands.¹¹ As a result, our understanding of the role of orphan GPCRs (oGPCRs) in mediating signal transduction within the central nervous system (CNS) remains significantly limited. However, evidence from animal studies, post-mortem transcriptomic analysis of patients, and genome-wide association studies have unveiled the critical involvement of oGPCRs in the realms of neuropharmacology and psychopharmacology.^12–14 Despite their potential significance, oGPCRs have yet to be extensively explored as viable targets for the treatment of CNS diseases, but progress has been made in recent years to shed light on their roles and potential therapeutic implications.^15–17

The Rhodopsin-like orphan G-protein coupled receptor 37 (GPR37), also known as parkin-associated endothelin receptor-like receptor (PaelR), has garnered attention due to its association with Parkinson’s disease (PD) pathology as a substrate of the E3 ubiquitin ligase parkin, which is further reviewed in subsequent sections.^18–20 Recent studies have focused more on the implication of GPR37 in chronic inflammation, which underlies various diseases such as autoimmune, metabolic, neurodegenerative, and cancer conditions.^{21, 22} In-depth RNA sequencing of over fifty human tissues conducted by the Genotype-Tissue Expression (GTEx) project has revealed that GPR37 is predominantly expressed in the CNS, including regions such as the amygdala, basal ganglia (caudate, putamen, and nucleus accumbens), substantia nigra, hippocampus, frontal cortex, hypothalamus, and notably, exhibits exceptionally high expression in the spinal cord.²³ This specific expression pattern suggests a potential regulatory role of GPR37 in neuronal function and CNS-related processes.^{24, 25} Further investigations utilizing in situ hybridization have revealed the specific expression of GPR37 in astrocytes and its role in regulating postnatal cerebellar granule neuron proliferation/differentiation, as well as the maturation of Bergmann glia and Purkinje neurons (Figure 1).^26–30 Transcriptomic analysis conducted in mice and humans has indicated that GPR37 is enriched in glial cells, with mature astrocytes and myelinating oligodendrocytes (OLs) displaying the highest expression levels.^{13, 31, 32} Notably, GPR37 expression declines in newly formed OLs and is absent in OL progenitor cells, showing negligible expression in other cell types.^{31, 33} As the molecular mechanisms governing OL differentiation, a crucial process for efficient neuronal signaling in the CNS, remain largely unknown, the presence of GPR37 in OLs has garnered significant interest in unraveling its involvement in OL differentiation and the myelination process.

Figure 1:

GPR37 is highly expressed in the CNS, specifically in myelinating oligodendrocytes and astrocytes.

GPR37 is suspected to function as a receptor for neuropeptides.^34–37 However, the precise ligand pairings of GPR37 have proven challenging to validate due to GPR37’s specific expression in CNS tissue, which limits the use of cell culture lines.^{38, 39} As a result, GPR37 currently remains categorized as an orphan receptor. Utilizing homology modeling techniques may provide valuable insights into the three-dimensional structure of GPR37, enabling the prediction of potential ligand binding sites and interactions.⁴⁰ However, GPR37’s closest homolog is the GPR37 Like 1 (GPR37L1), exhibiting 68% similarity and 48% identity, which is also an oGPCR.⁴¹ GPR37 also shares significant homology with peptide-specific GPCRs, including human endothelin-type B (ET_BR; 48% similarity and 29% identity), bombesin-BB1, and bombesin-BB2 receptors.^{42, 43} Homology models hold promise for expediting the discovery of novel ligands. This review focuses on the pharmacology and physiology of GPR37, emphasizing the potential therapeutic benefits associated with targeting this receptor using small molecule ligands.

2. GPR37 SIGNALING AND FUNCTION

GPR37 is proposed to engage in cellular signaling by coupling with heterotrimeric Gα_i/o G proteins, potentially involving cAMP signaling, calcium signaling and β-arrestin proteins (Figure 2).⁴⁴ Studies investigating proposed agonist activators of GPR37 have indicated that GPR37 signals through a mechanism sensitive to pertussis toxin, highlighting the requirement of Gα_i/o proteins for GPR37 signaling.^{32, 44, 45} However, multiple downstream mechanisms have been suggested for GPR37-Gα_i/o signaling, including anticipated inhibition of adenylyl cyclase to decrease cAMP levels, inhibition of exchange protein activated by cAMP (EPAC)-mediated signaling, activation of a membrane calcium channel by GPR37-Gα_i/o to facilitate extracellular Ca²⁺ influx, enhanced extracellular signal-regulated kinases (ERK) phosphorylation, and activation of the mitogen-activated protein kinase (MAPK) pathway.^{32, 37, 45–48} GPR37 signals through heterotrimeric G and β-arrestin proteins, leading to receptor endocytosis and the activation of ERK through distinct pathways.^{49, 50} GPR37 demonstrated the first channel rhodopsin 2 (ChR2)-GPCR approach, enabling the selective activation of the oGPCR signal through optogenetics.⁵¹ Optogenetic activation of ChR2/opto-GPR37 allows for the causal analysis of GPR37 activity in specific cells and the behavioral responses of freely moving animals. The specificity of opto-GPR37 signaling is confirmed by the reduction of cAMP levels, enhanced ERK phosphorylation, and increased motor activity. While GPR37’s Gα_i/o signaling has been recognized as an important mode of action, further studies are crucial to validate the activation of other G protein signaling pathways.

Figure 2:

Schematic presentation of the plausible potential intracellular signaling mechanisms of GPR37, including downstream signaling events following receptor activation by a GPR37 ligand. The three secondary signaling events shown are Gα_i/o-mediated inactivation of adenylyl cyclase, β-arrestin recruitment, and membrane calcium channel activation that eventually lead to changes in the Raf-MAPK-Erk 1/2 signaling cascade to alter gene expression or neuronal plasticity.

2.1. GPR37 GAIN AND LOSS OF FUNCTION STUDIES: RELEVANCE FOR PARKINSON’S DISEASE, NEUROPROTECTION AND OLIGODENDROCYTE FUNCTION

Much of our understanding of GPR37 and GPR37L1, in the absence of a defined ligand, has come from studies utilizing animal models that either overexpress or lack these receptors.^{18, 52–55} A major focus of prior GPR37 research has primarily been related to Parkinson’s disease (PD). PD is a complex neurodegenerative disorder with multifaceted mechanisms contributing to its development, including impaired protein clearance pathways, such as the ubiquitin-proteasome system and autophagy leading to the accumulation of aggregated misfolded proteins called Lewy bodies, the loss of dopamine-producing neurons, an imbalance in oxidative stress response, impaired mitochondrial function, overactive immune cells such as microglia and astrocytes contributing to inflammation and neuronal damage, and certain genetic mutations and variations associated with an increased risk of developing PD.⁵⁶ A major finding relevant to PD is that misfolded GPR37 is prominently found within Lewy bodies, indicating GPR37 is a biomarker for PD pathology.^57–61 Mutations in GPR37 are also implicated in endoplasmic reticulum (ER) stress, leading to loss-of-function (LOF) effects that exacerbate the death of dopaminergic neurons by promoting the accumulation and aggregation of misfolded proteins.^{18, 62, 63} In fact, LOF mutations are associated with the early-onset form of PD known as autosomal recessive juvenile PD.⁵³ In cell culture models of PD with GPR37 overexpression, indole-3-propionic acid (IPA) effectively prevented β-amyloid aggregation and ER stress, resulting in reduced neuronal cell death.⁶⁴ Conversely, exposure to buprenorphine was found to increase GPR37 accumulation, but coadministration with dextromethorphan inhibited this aggregation, thus mitigating the associated proapoptotic ER stress response.⁶⁵ These findings suggest the potential for treating neurodegenerative diseases and conditions that are characterized by protein misfolding and ER stress. Furthermore, the identification of peptides from the N-terminus-cleaved domain of GPR37 (ecto-GPR37) through mass spectrometry has revealed a notable increase in PD patients’ cerebrospinal fluid (CSF) levels, with no significant changes observed in patients with Alzheimer’s disease.⁶¹ These findings provide strong support for conducting further clinical studies to validate and assess the potential utility of ecto-GPR37 as a biomarker for PD.

While overexpression and misfolding of GPR37 can be cytotoxic, endogenous GPR37 is presumed to have a protective role under conditions of cellular stress when parkin expression is normal.^66–68 Additionally, GPR37 expression in astrocytes has shown protective effects in models of oxidative stress-induced toxicity.⁴⁸ Increased surface expression of GPR37 has been observed to protect N2a cells, a catecholaminergic neuronal-like cell line, against MPP⁺, rotenone, and 6-OHDA, which are known to mimic certain cellular aspects of PD.^69–71 Mitochondrial complex I activity has also been shown to be reduced in parkin knockout mice, suggesting that chronic unfolded protein stress may drive progressive neuronal death and oxidative stress in PD progression.⁷⁰ These findings suggest that properly functioning GPR37 may counteract the aging processes and oxidative stress arising in neurodegeneration. GPR37 also plays a role in controlling the response of progenitor cells to ischemic injury, offering new perspectives on the modulation of endogenous progenitor cells following stroke.^{72, 73} Genetic deletion of GPR37 has been found to alter the glial environment, leading to an increase in progenitor cells, a decrease in the activation of OL precursor cells, and a reduction in glia that are triggered by stroke injury. Additionally, increased apoptotic and autophagic cell death, along with caspase-3 activation and attenuated mechanistic target of rapamycin (mTOR) signaling, has been observed in GPR37 knockout mice. GPR37 deletion has also been shown to attenuate astrocyte activation and astrogliosis compared to wild-type stroke controls. Immunohistochemical staining has revealed an increased number of ionized calcium-binding adapter molecule (Iba) 1-positive cells in the ischemic cortex of GPR37 knockout mice, indicating altered microglia activation.⁷⁴ Moreover, RT-PCR analysis has demonstrated enrichment of M1-type microglia or macrophage markers in the GPR37 knockout ischemic cortex, and Western blotting experiments have confirmed elevated levels of inflammatory factors in GPR37 knockout mice following ischemia. Proteomic studies have shown that genetic deletion of GPR37, but not GPR37L1, leads to a significant decrease in brain expression of myelin-associated glycoprotein.⁷⁵ The molecular mechanisms underlying the formation of myelin by OLs remain largely unknown, despite its crucial role in the central nervous system.³² GPR37 is highly expressed in OLs and its expression increases during their differentiation into myelin-forming cells. GPR37 inhibits OL differentiation by suppressing the EPAC-dependent Raf-MAPK-ERK1/2 module and the nuclear translocation of ERK1/2. The loss of GPR37 does not cause increased loss of precursor or mature OLs, but it does result in increased susceptibility to demyelination and altered OL physiology after cuprizone treatment, suggesting that GPR37 may be a potential drug target for multiple sclerosis and PD.⁷⁵ The protective role of GPR37 under conditions of cell stress, its influence on the response of progenitor cells to ischemic injury, and its impact on OL differentiation and myelin formation indicate that modulating GPR37 activity could hold therapeutic implications for stroke and multiple sclerosis.

2.2. RELEVANCE OF GPR37 IN THE DEVELOPMENT OF PHYSIOLOGICAL SYSTEMS AND PATHOLOGICAL CONSEQUENCES

GPR37 has been implicated in epileptogenicity in focal cortical dysplasia patients and is a potential marker for localizing epileptogenic zones.⁷⁶ In general, genetic variants in GPR37 have strong associations with brain morphology, gene expression, methylation, and chromatin organization involved in various neuropsychiatric traits.⁷⁷ GPR37 plays a crucial role in Wnt/β-catenin signaling, a pathway essential for embryonic development, stem cell biology, and neurogenesis.⁷⁸ GPR37 is required for the protection of the Wnt co-receptor LRP6 from ER-associated degradation.⁷⁸ Studies in GPR37-deficient mice have shown impaired olfactory bulb nerve layer development, as well as attenuated or delayed maturation and migration of embryonic Gonadotropin-releasing hormone (GnRH) neurons involved in the hypothalamic-pituitary-gonadal axis.^{79, 80} It also shows high expression in ghrelin-positive cells and may play a role in metabolic regulation.⁸¹ Additionally, GPR37 exhibits increased expression in the superior olivary complex in mice capable of processing auditory signals, suggesting a physiological role in the auditory system.⁸²

Fetal alcohol spectrum disorders (FASD) represent the leading cause of non-heritable, preventable mental disability, affecting nearly 5% of births in the United States.⁸³ These disorders result in physical, behavioral, and cognitive impairments, including deficits associated with the cerebellum. Despite their significant impact, there is currently no known cure for FASD, and the underlying mechanisms behind these disorders remain poorly understood. Studies have demonstrated that microglia and Purkinje cells are affected in FASD models, displaying increased cell death, immune activation in microglia, and altered firing patterns in Purkinje cells. Alcohol-induced neuroinflammation near the Purkinje cell layer is crucial for cerebellar immune responses. Higher expression of GPR37 has been observed in rat lines with a propensity for high ethanol consumption.⁸⁴ Ethanol suppresses GPR37 expression, which may be expected to increase inflammation in the developing cerebellum.⁸³ This neuroinflammation contributes to the neuropathological consequences of FASD. Despite substantial personal and societal loss caused by FASD, no effective treatment has been approved by the FDA. Therapeutics that modulate GPR37 expression may be a new horizon for developing potential FASD therapies.

The gene for GPR37 was found within the first Autism Spectrum Disorder (ASD) locus identified, localized on chromosome 7q31-33, called the AUTS1 region.^{85, 86} The inheritance patterns of the mutated GPR37 in autism suggest a predisposing variant may be passed from an unaffected parent to affected offspring in a dominant manner, with variable penetrance that is higher in males and lower in females. It may indicate a distinct pattern of autism inheritance, possibly involving additional genes or factors, meaning autism is a multigenetic trait, and a single variant alone is unlikely to reach the threshold for expressing autism pathology. Identifying these additional predisposing variants is an important goal for future research. GPR37 co-localizes with multiple PDZ domain protein 1 (MUPP1) and contactin-associated protein-like 2 (CASPR2) at the synapse, while the ASD-related mutation of GPR37 may disrupt the CASPR2-MUPP1-GPR37 complex on dendrites, contributing to ASD pathogenesis. Overall, these hypotheses highlight the complexity of autism genetics, involving interactions between various genes and coupling factors. Understanding the role of GPR37 and its mutations in autism will contribute to unraveling the underlying mechanisms of the disorder and may provide insights for future diagnostic and therapeutic approaches.

GPR37 and GPR37L1 have been identified as significant players in cerebellar development through their involvement in the Sonic hedgehog (Shh) signaling pathway.^{30, 87} Shh is a critical morphogen that regulates various aspects of brain development, including cell proliferation, differentiation, and patterning.⁸⁸ During postnatal cerebellar development, Purkinje cells release Shh, which is pivotal in stimulating the proliferation of granule cell progenitor cells and Bergmann glia.^26–28 This process involves the activation of downstream signaling cascades triggered by Shh binding to its receptor protein Patched-1 (Ptch1), relieving Ptch1-mediated inhibition on Smoothened (Smo), a transmembrane protein. The activation of Smo leads to the initiation of intracellular signaling events that ultimately influence gene expression and cellular responses. GPR37 and GPR37L1 contribute to this intricate process by participating in Shh signaling.^{89, 90} These receptors are expressed in cerebellar astrocytes and granule cell progenitors, and Shh induces their expression. The activation of these receptors by Shh binding appears to be a crucial step in amplifying the Shh signal within these cell populations. Consequently, the expression of key components of the Shh signaling pathway, such as Smo and Ptch1, is upregulated in response to GPR37 and GPR37L1 activation. This amplification of Shh signaling enhances the cellular responses to Shh and contributes to the proper development of cerebellar structures. A significant aspect of Shh signaling in the cerebellum is its role in oligodendrocyte (OL) development and myelination. GPR37-deficient mice exhibit abnormalities in OL differentiation, proliferation, and myelination.^{32, 36, 91, 92} Shh signaling is critically involved in regulating OL development, and GPR37’s interaction with the Shh pathway is vital for ensuring proper OL differentiation and function. The absence of GPR37 appears to disrupt the balance of Shh signaling, leading to premature OL differentiation and hypermyelination. Interestingly, Shh signaling also influences the timing of neuron-glia fate switching, a process that determines whether a neural precursor cell will differentiate into a neuron or a glial cell. Altered expression of Shh and its signaling partners, such as GPR37, could potentially underlie the observed precocious OL differentiation seen in GPR37-deficient mice. This suggests that the delicate interplay between Shh and its signaling components, including GPR37 and GPR37L1, plays a critical role in coordinating the intricate cellular processes that drive cerebellar development and myelination. Overall, GPR37 and GPR37L1 are important modulators of Shh signaling in the cerebellum, influencing various aspects of cerebellar development, including granule cell proliferation, Bergmann glia activation, and proper OL differentiation and myelination. Their role in amplifying Shh signaling and coordinating complex cellular responses highlights their significance in orchestrating the precise developmental processes that shape the cerebellum. Such neuroprotective functions could explain GPR37’s role in megalencephalic leukoencephalopathy with subcortical cysts (MLC), an inherited disorder characterized by seizures and developmental delay in infants and children, leading to a progressive decline in motor skills and cognitive functions during later stages of life.⁹³ The pathophysiological mechanisms leading to MLC are still unclear, but the negative regulation GPR37 on myelin homeostasis and astrocyte maturation may grant important insights into modulating seizure responses, and understanding type or location of seizures.^93–95

While most studies on GPR37 focus on its role in the brain, it is also highly expressed in Sertoli cells (SCs) in the testis, which support spermatogenesis.^{96, 97} Proper maturation timing of SCs is essential for determining the total number of SCs and sperm production. Desert hedgehog (Dhh) is an important factor produced by SCs and essential for proper testicular development and spermatogenesis. It serves as a key regulator of germ cell proliferation, differentiation, and survival. By binding to its receptor patched-1 on adjacent cells, Dhh triggers a signaling cascade that influences the expression of various genes involved in spermatogenesis. This signaling pathway helps maintain the integrity of the seminiferous epithelium, regulates the production of testosterone, and influences the overall development and function of the male reproductive system. GPR37-deficient mice exhibit premature SC maturation, leading to a reduction in the final number of mature SCs.⁹⁸ Furthermore, these mice display altered expression levels of SC maturation markers and premature elevation of Dhh, patched-1, and Gli2 expression.⁹⁹ Overall, the findings suggest that GPR37 plays diverse roles in various developmental processes and physiological systems, including brain function, reproduction, and the regulation of metabolic and auditory processes.

2.3. RELEVANCE OF GPR37 HETEROMERIC RECEPTOR AND TRANSPORTER INTERACTIONS

The primary hallmark of PD is the progressive degeneration of dopamine-producing neurons in a brain region called the substantia nigra.⁵⁶ The loss of dopamine-producing neurons disrupts the delicate balance of dopamine in the brain, leading to the motor symptoms characteristic of PD, such as tremors, rigidity, and bradykinesia.⁵⁶ Studies have revealed the physical interaction between GPR37 and GPR37L1 with the dopamine transporter (DAT) as well as dopaminergic receptors D₁R and D₂R.^{38, 100, 101} Both GPR37, the D₂R autoreceptor and DAT are all co-expressed presynaptically in dopaminergic neurons that project to the mouse striatum and GPR37 negatively regulates DAT through endocytosis and trafficking. In cells, co-expression of GPR37 with D₂R increased GPR37 expression and enhanced D₂R ligand affinity, as determined by ³H-spiperone competition binding assays.³⁸ Mice lacking GPR37 also showed reduced affinity of D₂R ligands.¹⁰⁰ These mice exhibited elevated surface expression of DAT and increased dopamine uptake in the striatum.^{100, 102} The impact of altered DAT expression in GPR37-deficient mice was assessed through their response to psychostimulants like cocaine, which blocks DAT and reduces dopamine reuptake. GPR37-deficient mice displayed increased sensitivity to amphetamine but reduced cocaine-induced locomotor activity, both acting on DAT but through different mechanisms.¹⁰² In conditional place preference behavioral tests, mice lacking the GPR37 gene failed to respond to either stimulus when used as an incentive.¹⁰³ Evaluation of basal synaptic transmission and paired-pulse stimulation in cocaine-treated mouse slices showed no differences in the cocaine-induced modification of basal synaptic transmission. GPR37 is involved in this modification without affecting cocaine’s effects on short-term plasticity.¹⁰⁴ The loss of these receptors in mouse models suggests that GPR37 plays a role in the proper development and normal signaling of the dopaminergic system.¹⁰³ Fluorescence cross-correlation spectroscopy was employed in live N2a cells to study interactions between GPR37 and two splice variants of D₂R. GPR37 exhibited an interaction with both splice forms of D₂R. Additionally, 4-phenylbutyrate, a neuroprotective chemical chaperone, increased GPR37 expression at the cell surface and the number of interacting proteins.¹⁰⁵ Pramipexole, a D₂R agonist used in PD treatment, also enhanced this interaction, suggesting potential clinical relevance.¹⁰⁵ GPR37 is dysregulated in various neurological disorders also associated with dopaminergic systems including bipolar disorder, anxiety, major depressive disorder, and schizophrenia, despite the exact role of GPR37 in these diseases is not fully understood.^{13, 106–109} In summary, these findings suggest that pharmacological modulation of the heteromeric complex formed by GPR37 and D₂R may be relevant to the treatment of PD and other CNS disorders.

The deletion of GPR37 leads to the upregulation of the striatal Adenosine A(2A) receptor (A_2AR), which correlates with agonist-induced catalepsy and cAMP accumulation.¹¹⁰ Furthermore, GPR37 exhibits chaperone-like activity by repressing A_2AR cell surface targeting and function. In GPR37-deficient mice, treatment with an A_2AR antagonist, SCH58261, increased locomotor sensitization.^{110, 111} Chronic A_2AR blockade also enhanced long-term striatal depression in corticostriatal synapses of GPR37-deficient mice.¹¹⁰ The control exerted by A_2AR over parkinsonian tremor, specifically pilocarpine-induced tremulous jaw movements, is lost in the absence of GPR37.¹¹² The ability of striatal GPR37 to form oligomers with A_2AR and D₂R may allow for fine-tuning multiple receptor signaling pathways and harmonizing neurotransmission, potentially impacting PD management.^{113, 114}

Advanced biophysical approaches, including Fluorescence Cross-Correlation Spectroscopy, Förster Resonance Energy Transfer, and Fluorescence Lifetime Imaging Microscopy, have demonstrated that GPR37 and GPR37L1 form hetero/homodimers in the plasma membrane of live N2a cells, with only GPR37 aggregating in the cytoplasm.¹¹⁵ Parkin has been identified as a crucial regulator of GPR37 aggregation, effectively preventing the accumulation of GPR37 aggregates in the live cell cytoplasm.^{18, 19, 57, 62, 63} This data further supports the involvement of GPR37 in cytosolic aggregation processes associated with PD pathology.

The interactions between GPR37 and dopamine-related receptors and their functional consequences, such as altered ligand affinity and dopamine uptake, suggest a novel avenue for targeting and modulating dopaminergic signaling. The dysregulation of GPR37 in bipolar disorder, anxiety, major depressive disorder, and schizophrenia, as well as its involvement in PD pathogenesis, highlights GPR37 as a promising therapeutic target for the CNS disorders. Additionally, the chaperone-like activity of GPR37 in repressing the A_2AR function suggests that targeting GPR37-A_2AR interactions may also be relevant for PD treatment. Large-scale interactome mapping for human GPCRs has provided a valuable resource for analyzing signaling pathways involving druggable families of integral membrane proteins.¹¹⁶ Although 5-hydroxytryptamine receptor 4 (5-HT₄) has been identified as a potential coupling partner of GPR37 through this technique, its biological relevance remains to be demonstrated.¹¹⁶ Understanding the function and heteromeric coupling of GPR37 can offer valuable insights into the development of pharmacological strategies to address different pathologies that impact dopaminergic, serotonergic, or adenosine function.

2.4. CANCER STUDIES AND PROGNOSTIC SIGNIFICANCE OF GPR37

GPR37, known for its high expression in the central nervous system (CNS), has been linked to medulloblastoma (MB), a malignant brain tumor that originates in the lower posterior region of the brain responsible for muscle coordination, balance, and movement.¹¹⁷ A multi-omics pilot study using mass spectrometry on cerebrospinal fluid (CSF) from recurrent MB patients revealed the upregulation of GPR37, as well as ADAMTS1 and GAP43 indicating the presence of hypoxic conditions, in the CSF of these patients.¹¹⁷ Functional studies have demonstrated that elevated levels of GPR37 promote tumor cell proliferation, migration, and invasion while suppressing GPR37 through knockdown techniques inhibits malignant behaviors.^{118, 119} GPR37 is crucial to comprehend the role of GPR37 in peripheral cancers, as protein-protein interaction (PPI) analysis has revealed its involvement in a functional pathway that facilitates breast cancer metastasis to the brain and lungs.^{120, 121}

Furthermore, analysis of the Cancer Genome Atlas and Oncomine databases has shown high expression of GPR37 in lung adenocarcinoma (LUAD), with upregulation associated with unfavorable outcomes.^{122, 123} Gain- or LOF assays have demonstrated that increased GPR37 expression enhances the activation of TGF-β1, Smad2, and Smad3 phosphorylation, leading to improved proliferation, migration, and invasion of carcinoma cells in vitro.¹²⁴ Conversely, knocking down GPR37 impedes malignant behaviors. In LUAD, GPR37 plays a significant role in constructing a prognostic prediction model for competitive endogenous RNA (ceRNA) and tumor-infiltrating immune cells (TIICs). Among the three types of TIICs (Monocytes, Macrophages M1, activated mast cells) found to be significantly associated with LUAD prognosis, GPR37, and Macrophages M1 exhibit a close relationship.¹²⁵ Additionally, a PPI network suggests that GPR37 genes are upregulated in samples with TP53/EGFR co-mutations, making it a potential novel prognostic marker and therapeutic target for patients with dual TP53/EGFR mutation LUAD.¹²⁶

Other studies have indicated lower GPR37 expression in human hepatocellular (HuH7) carcinoma compared to adjacent non-tumorous tissues.¹²⁷ Transient knockdown of GPR37 using siRNA in HuH7 cells has shown a significant decrease in hepatoma cell apoptosis by activating the phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway.¹²⁸ AKT plays a crucial role in promoting cell survival and growth. In multiple myeloma cells, GPR37 is implicated in regulating cell proliferation through the modulation of cell adhesion ability and AKT and ERK activity.¹²⁸ GPR37 gene expression is also upregulated in ovarian cancer samples and Schwann cell tumors.^{129, 130} The upregulation of GPR37 in human glioma U251 cells leads to increased proliferation, a decrease in G1/G0 phase cells, an increase in S and G2 phase cells, and enhanced phosphorylation of p-AKT (Ser473).¹³¹ Increased phosphorylation of p-AKT suggests the activation of signaling pathways associated with cell survival and proliferation. This finding further supports the notion that GPR37 upregulation may contribute to cancer cell growth and survival. Additionally, the alteration of cell cycle distribution is noteworthy. The decrease in G1/G0 phase cells and the increase in S and G2 phase cells indicate a disruption in the normal cell cycle regulation, potentially promoting uncontrolled cell division. GPR37 is found in the same complex as REG4, which mediates signal transduction and promotes peritoneal metastasis of gastric cancer cells.¹³² High expression of REG4 is associated with advanced stage and poor survival prognosis in gastric cancer patients. By expressing GPR37, REG4 contributes to peritoneal metastasis and establishes a positive feedback loop in gastric cancer.¹³³ These findings indicate that targeting GPR37 holds promise as a potential therapeutic approach for the development of novel treatments for various types of cancers and their metastasis.

3. GPR37 PHARMACOLOGY AND POTENTIAL LIGANDS

The GPR37 amino acid sequence exhibits significant homology to peptide-specific GPCRs, and the invertebrate-derived neuropeptide “head activator” (HA, Figure 3) has been shown to activate Ca²⁺ signaling via GPR37 using the G_α16/aequorin assay with an EC₅₀ value of 3.3 nM.^{37, 134} In addition, HA has been reported to modulate NFAT signaling and inhibit Forskolin-mediated cAMP production through GPR37.¹³⁵ To overcome challenges in validating ligand-GPR37 pairings using recombinant GPR37, and GPR37L1 expressed in HEK293 cells, recent studies have turned to primary cell cultures for successful ligand identification.⁴⁶ These efforts have revealed the involvement of osteocalcin (OCN), a bone-derived protein crucial for brain development and neural cognitive function, in interacting with GPR37 to regulate OL differentiation, myelination, and myelin production and remyelination after demyelinating injury.³⁶ Dose–response studies revealed that OCN activates GPR37 with an apparent EC₅₀ of 10.2 nM. OCN treatment has shown protective effects against LPS-induced inflammation, which are absent in GPR37-deficient mice.¹³⁶ Furthermore, OCN activates intracellular responses via GPR37 in macrophages, regulating inflammatory factor release and phagocytic function. These findings highlight the protective role of OCN in acute inflammation and suggest the therapeutic potential of activating the OCN/GPR37 regulatory axis for treating inflammatory diseases.

Figure 3:

Potential ligands of GPR37 and their reported potencies in the relevant assays: proteins (not shown in figure) PSAP (EC₅₀ of 7.0 nM), and OCN (EC₅₀ of 10.2 nM); protein fragments HA (EC₅₀ of 3.3 nM), and TX14A (EC₅₀ of 19.4 nM); small molecules ARU (K_D of 17.0 μM), NPD1 (EC₅₀ of 26 nM), and CGA (activity at 100 μM). Shared backbone motifs between HA and TX14A are highlighted in red.

Another suspected peptide ligand with an EC₅₀ of 7.0 nM for GPR37 is prosaposin (PSAP), a multifunctional protein precursor to Saposin A-D, that internally regulates lysosomal enzyme function and externally acts as a neuroprotective and glio-protective secretory factor.^{48, 137, 138} It has been found that extracellular PSAP is necessary for the partitioning of GM1 ganglioside-containing lipid rafts in the plasma membrane of live cells expressing turbo green fluorescent protein-tagged (tGFP) GPR37.⁶⁶ Lipid rafts are specialized structures on the plasma membrane enriched in cholesterol, certain gangliosides, and lipids, and these membrane domains can coordinate the signaling of membrane receptors, including GPCRs.¹³⁹ Lipid raft disturbance with methyl-β-cyclodextrin or cholesterol oxidase results in reduced GPR37(tGFP) surface density and decreased cell viability in N2a cells.⁶⁶ Furthermore, GPR37(tGFP) was observed to form complexes with GM1 ganglioside when labeled with cholera toxin on the plasma membrane, which also suggested a protective mechanism of GPR37, GM1, and lipid rafts against toxic GPR37 aggregates observed in PD.¹⁴⁰

In studies involving rats injected with kainic acid (KA), the expression of PSAP and GPR37 was found to alleviate the nerve damage caused by KA.¹⁴¹ Moreover, in facial nerve transection, PSAP and GPR37 were shown to produce neurotrophic factors involved in neuronal repair in microglial and astrocytes.^{142, 143} Saposin C and its neuroactive fragment TX14A (Figure 3) were specifically associated with the neuroprotective effects of GPR37-expressing astrocytes on neurons subjected to oxidative stress.⁴⁸ The administration of TX14A in GPR37-transfected cells induced a dose-dependent increase of luciferase activity with an EC₅₀ of 19.4 nM.³⁶ Biotinylated TX14A demonstrated the ability to reduce GPR37 levels, and ERK phosphorylation was sensitive to pertussis toxin in cells transfected with GPR37. Additionally, when co-expressed with G_αi1, GPR37, and GLR37L1 exhibited increased accumulation of ³⁵S-GTPγS, indicating that the receptors likely signal via G_αi/o family proteins. The interaction of GPR37 with TX14A has also been demonstrated by fluorescence-correlated spectroscopy, which indicates that GPR37 is coordinated with TX14A in lipid rafts of N2a cells expressing the tGFP version of GPR37.⁶⁶ However, studies investigating the functional interactions between GPR37/GPR37L1 and PSAP/TX14A have yielded mixed results.⁴⁶ There are some similarities between the backbones of Saposin C, its fragment TX14A, and HA, which may grant insights into possible binding interactions and the rational design of GPR37 small molecule ligands.

Recent findings indicate that macrophage GPR37 plays a role in reversing inflammatory pain by enhancing phagocytosis and shifting cytokines toward an anti-inflammatory profile.^{47, 144–146} Pain is a characteristic feature of inflammation and can have both protective and detrimental effects in acute or chronic stages. The resolution of inflammation is a critical process facilitated by specialized pro-resolving mediators (SPMs). However, therapeutic targeting of these receptors has proven challenging, with only a few ligands advancing to preclinical or clinical development. To control inflammation and pain, macrophage-nociceptor interactions can be targeted.¹⁴⁷ Notably, Artesunate (ARU, Figure 3), an antimalarial drug, has concentration-dependent binding with a K_D of 17.0 μM to GPR37 in macrophages and promotes the phagocytosis and clearance of pathogens.¹⁴⁸ Reduced expression of the GPR37 gene in goats seropositive for small ruminant lentiviruses (SRLV) indicates the presence of an ongoing inflammatory process in the early stages of infection, compared to SRLV-seronegative goats.¹⁴⁹ Mice lacking GPR37 showed increased mortality and delayed resolution of hypothermia when challenged with lipopolysaccharide (LPS), Listeria bacteria, or the Plasmodium berghei malaria parasite.¹⁴⁸ The specialized proresolving mediator Neuroprotectin D1 (NPD1, Figure 3) with an EC₅₀ of 26 nM for GPR37 and ARU protected wild-type mice from LPS- and Listeria-induced sepsis, but these protective effects were lost in GPR37-deficient mice.^{47, 148} Moreover, the elimination of macrophages worsened infection severity, sepsis, and associated effects, while the transfer of NPD1- or ARU-primed macrophages mitigated infection, sepsis, and pain-related behaviors. Recently, Chlorogenic acid (CGA, Figure 3), at a concentration of 100 μM, has been shown to reduce inflammatory cytokine and enhance phagocytosis in RAW264.7 cells stimulated with LPS.¹⁵⁰ CGA significantly alleviated lung inflammation, reduced bacterial lung load, improved phagocytosis by alveolar macrophages, and increased survival rates in mice with acute respiratory distress syndrome (ARDS) induced by cecal ligation and perforation. The absence of protective effects of CGA against ARDS in GPR37 knockout mice indicates the essential role of GPR37 in mediating these effects. These findings highlight the potential of targeting GPR37 as a therapeutic strategy for ARDS treatment and emphasize the importance of further investigating the molecular mechanisms underlying this interaction to develop effective treatments.

Information about ligands that functionally interact with GPR37 helps guide drug discovery by providing a starting point to identify and design novel small molecule ligands with better drug-like properties. Overall, the reported GPR37 ligands, as shown in Figure 3, have been associated with various therapeutic effects, including neuroprotection, resolution of inflammation, and pain relief. Due to the unselective function of these ligands, further research is needed to identify and develop more specific and potent GPR37 ligands that can provide targeted therapeutic benefits while minimizing potential off-target effects.²² Understanding the binding interactions and effects of these ligands on GPR37 can aid in the development of novel therapeutics targeting GPR37.

4. HOMOLOGY MODEL AND STRUCTURE OF GPR37

In GPCR research, phylogenetic relationships are crucial for understanding pharmacological similarities based on both primary sequence homology and three-dimensional structures to establish receptor-ligand pairings.¹⁵¹ Homology modeling has witnessed significant opportunities and challenges driven by advancements in experimental techniques and computational methods. With the increasing availability of high-quality protein 3D structures, particularly through methods like cryo-electron microscopy (Cryo-EM), the role of homology modeling in predicting 3D structures of protein sequences within families has grown. However, high-resolution structures for oGPCRs are largely lacking, including for GPR37, posing challenges for modeling and rational drug design. Numerous methods and algorithms exist for homology modeling, each with its own strengths and limitations. The choice of method depends on the specific protein and intended applications of the model, emphasizing the importance of selecting the appropriate approach. Traditional homology modeling relies on sequence similarity, often neglecting ligand information present in experimental structures. Ligand-sensitive methods have been introduced to address this limitation, but their manual interventions require time and expertise. A significant goal is to develop fully automated homology modeling tools capable of handling ligand-related challenges. Homology modeling initially developed for GPCRs can generally be effectively applied to oGPCRs, providing a helpful starting point for ligand development.⁴⁰ Homology modeling also aids in comprehending the pharmacological similarity of oGPCRs, which is of utmost importance for predicting off-target effects of ligands, repurposing drugs, and discovering new ligands.¹⁵² Homology modeling may still leave questions unanswered in computational models. Improved methods for model refinement and co-coordinate adjustments parallel to the native state contribute to better coverage and accuracy. Modeling loops and inserts remains challenging without template data, and optimizing these regions and side chains is crucial for accurate models. Molecular dynamics simulations play a role in refining generated models. In cases of low sequence similarity, using multiple templates can enhance accuracy, although careful template selection is necessary to avoid alignment aberrations. Building multiple models during homology modeling presents an opportunity, but selecting the best model requires rigorous assessment. Various parameters like DOPE score, TM score, and RMSD value are used for model comparison, with the choice of determinant parameter based on the modeling purpose.

GPR37 is a membrane-embedded protein with 613 amino acids and seven transmembrane domains with an unusually long N-terminal chain (Figure 4).⁴³ GPR37, and GPR37L1 exhibit the highest sequence similarity and identity to the ET_BR. Even within the same GPCR subfamily, individual receptors can exhibit substantial structural variability. This variability can arise from differences in the length and composition of extracellular loops, ligand-binding pocket variations, and unique domains. For instance, TM2 contains important and conserved interactions for peptide-binding receptors.¹⁵³ Homology modeling of GPR37 has been performed using crystal structures of δ opioid receptors and M2-muscarinic acetylcholine as templates.¹⁴⁸ The predicted binding pocket of GPR37 and GPR37L1 shows similarity to the bombesin, orexin, and neuropeptide S receptors.⁴⁰ Two potential binding sites have been identified using a homology model: one for small molecules like ARU, NPD1, and CGA (Figure 5A) and another for peptides like TX14A (Figure 5B). However, these binding sites have not yet led to the discovery of effective ligands for GPR37, highlighting the challenges in ligand discovery and the need for further research and innovative approaches. Ultimately, a solved high-resolution structure of GPR37 in complex with a ligand or an unoccupied form would provide a key next step to elucidate the ligand binding pocket, crucial ligand and amino acid interactions, and the overall structure of GPR37 with atomic detail. Recent successes solving oGPCR structures using CryoEM suggest a good probability that a similar approach could be used to solve the GPR37 structure in the future.^154–157

Figure 4:

Alignment of GPR37 with its closest homologs GPR37L1 and ET_BR was performed using pairwise alignment BLOSUM45 in Schrodinger’s Multiple Sequence Viewer. Each residue is displayed by single letter amino acid code, and colors are based on the similarity of their physiochemical properties (red is acidic hydrophilic; blue is basic hydrophilic; green is neutral hydrophobic aliphatic; orange is neutral hydrophobic aromatic; cyan is neutral hydrophilic; yellow is primary thiol; gray is proline).

Figure 5:

GPR37 is shown as a blue ribbon using a homology model. Potential binding site based on GPR37 homology modeling was determined using the human M2-Muscarinic Acetylcholine receptor (PDB ID: 4MQS) and predicted by molecular dynamics simulation (MDS) in the Schrodinger software suite.¹⁴⁸ A) Key residues at the binding site are in stick representation. NPD1 (magenta) is in the CPK presentation. B) Distinct binding site on GPR37 for peptide agonist TX14A (Orange ribbon) predicted by MDS.

Understanding the interactions between GPR37 and various proteins is crucial for comprehending its structural, functional regulation, and physiological significance. Several studies have described interactions between the C-terminal postsynaptic density protein-95/Discs large/ZO-1 (PDZ) domain of C-kinase (PICK1) PDZ and GPR37 and/or GPR37L1.^{38, 158, 159} PDZ domain-containing proteins are cytoplasmic scaffolding proteins that assemble various multiprotein signaling complexes, influencing the signaling, trafficking, and function of targeted receptors.^{159, 160} In-silico modeling suggests a novel interaction between GPR37 and γ-aminobutyrate type A receptor-associated protein-like 2 (GABARAPL2), an autophagosome-specific Ub-like protein involved in vesicle trafficking and autophagy.¹⁶¹ The cysteine-rich region at the C-terminal of GPR37 is proposed to interact with the GABA(A) binding site of GABARAPL2. GPR37 expression is likely regulated by several proteins, including parkin, PICK1, and GABARAPL2, through mechanisms such as ubiquitination, proteasomal degradation, and autophagy.

While N-terminal cleavage is rarely described for class A GPCRs, both GPR37 and GPR37L1 undergo this process.^{38, 162–164} A Disintegrin and Metalloprotease 10 (ADAM10) is suggested to cleave GPR37 at Glu167↓Gln168 into mature protein lacking the first 167 amino acids in the native N-terminus.¹⁶⁵ Expression of GPR37 modified with dual epitope tags at the N and C termini has revealed the existence of different-sized versions of the receptor.¹⁶² Studies using pulse-chase labeling and immunofluorescence microscopy have shown that full-length receptors are present in the secretory pathway until a rapid and efficient proteolytic N-glycan modification occurs constitutively in the Golgi.¹⁶⁵ Broad-range metalloproteinase inhibitors, such as GM6001, TAPI-1, and marimastat, prevent GPR37 cleavage and increase the number of full-length receptors on the cell surface.¹⁶² Moreover, this unconventional N-terminal processing may have significant implications for PD and neuronal function pathophysiology, as cleavage enhances plasma membrane insertion and increases receptor surface expression.^{38, 166} The functional relevance of these cleavage events for GPR37 and GPR37L1 requires further investigation.

5. GAPS IN BASIC AND TRANSLATIONAL GPR37 RESEARCH

To date, significant knowledge gaps remain in understanding how the activation of GPR37 and/or GPR37L1 and their downstream signaling results in efficient resolution of inflammation, ubiquitination, proteasomal degradation, autophagy, and macrophages. Non-selective ligands make pinpointing the exact function of GPR37 difficult. With the numerous functions of GPR37 in the CNS and cancers (summarized in Table 1), it is crucial to identify selective ligands to probe receptor function. Despite the growing interest in GPR37 and its potential as a therapeutic target for CNS disorders, there are still major gaps in both basic and translational research that need to be addressed. Such gaps hinder our comprehensive understanding of the receptor functions and limit the development of effective therapeutic strategies.

Table 1:

GPR37-associated diseases and relevant findings.

Disease	Relevant Findings
Parkinson’s Disease (PD)	GPR37 misfolding is found within Lewy bodies in PD.57–60 Overexpression of GPR37 reduces β-amyloid aggregation and ER stress that exacerbate the death of dopaminergic neurons.18, 62–64 Mutations in GPR37 are implicated in PD pathogenesis.53 Mitochondrial complex I activity has also been shown to be reduced in parkin knockout mice.70
Pain	Macrophage GPR37 may reverse inflammatory pain by enhancing phagocytosis and shifting cytokines toward an anti-inflammatory profile.47, 144–146
Stroke	GPR37 regulates inflammation, cell death pathways, and progenitor cell responses after ischemic stroke.72, 73
Megalencephalic Leukoencephalopathy with Subcortical Cysts (MLC)	Mechanisms leading to MLC are still unclear, but the negative regulation of GPR37 on myelin homeostasis may play a role.93
Seizure Susceptibility	GPR37 and GPR37L1’s impact on susceptibility to seizures might stem from their involvement in astrocyte maturation.94, 95
Multiple Sclerosis	GPR37 influences OL differentiation and myelin formation. GPR37 deletion leads to increased susceptibility to demyelination.75
Autism Spectrum Disorder (ASD)	The GPR37 gene is within an autism-related locus.85 Interactions with CASPR2-MUPP1-GPR37 complex suggest a role in ASD pathogenesis.86
Fetal Alcohol Spectrum Disorder (FASD)	Ethanol suppresses GPR37 expression, potentially contributing to neuroinflammation in the cerebellum, a key feature of FASD.83, 84
Bipolar Disorder, Anxiety, Major Depressive Disorder, and Schizophrenia	GPR37 dysregulation is observed in other CNS disorders, but the exact mechanisms and potential contributions through dopaminergic systems are not wholly elucidated.13, 106–109
Medulloblastoma (MB)	Elevated levels of GPR37 promote tumor cell proliferation, migration, and invasion.117
Lung Adenocarcinoma (LUAD)	GPR37 is highly expressed in LUAD, associated with unfavorable outcomes.123, 126 It enhances TGF-β1 activation and carcinoma cell behaviors.167
Hepatocellular Carcinoma	GPR37 knockdown decreases apoptosis in hepatoma cells by activating the PI3K-AKT pathway.127
Ovarian Cancer	GPR37 is upregulated in ovarian cancer and implicated in cell proliferation and AKT/ERK activity.130
Glioma	GPR37 upregulation in glioma cells leads to increased proliferation and activation of p-AKT signaling pathway.131
Gastric Cancer	GPR37 is in the same complex as REG4, which promotes signal transduction and peritoneal metastasis.132

One of the major gaps in GPR37 research is the lack of knowledge regarding its physiological functions and endogenous ligands. Although several peptides, such as HA, saposin C, and TX14A, and small molecules, like ARU and NPD1, have been identified as potential ligands, their exact interactions and functional significance, as well as target specificity, remain unclear.^{37, 148} Subsequent optimizations of hit compounds are needed to fine-tune their properties and enhance potency, selectivity, and pharmacokinetic profiles. The synthesis of derivatives for small molecule leads, like ARU, NPD1, and CGA, would impose serious challenges owing to the multiple stereocenters and labile function groups. Developing peptidomimetics, a class of compounds designed to mimic the structural and functional attributes of peptides, offers a promising avenue for addressing challenges related to the stability and bioavailability of peptides, ultimately leading to improved pharmacokinetic properties and prolonged systemic exposure.¹⁶⁸ Additionally, the strategic design of peptidomimetics enables fine-tuning of receptor binding affinity, facilitating enhanced target engagement and cellular response. This precise modulation of binding interactions holds immense therapeutic potential, as it allows for the creation of high-affinity ligands with improved efficacy. Moreover, the versatility of peptidomimetic design provides a platform for generating highly selective agents, minimizing off-target effects, and promoting safer therapeutic interventions.¹⁶⁸ Through rational design and innovative synthetic approaches, peptidomimetics empower researchers to unlock the full potential of peptide-based therapeutics, offering a robust toolkit to develop more potent, target specific, and effective treatments across a spectrum of medical conditions. Further studies are required to validate these ligand-receptor interactions and elucidate the downstream signaling pathways activated by GPR37. Additionally, the identification of endogenous ligands will provide valuable insights into the physiological roles of GPR37 and its potential involvement in CNS disorders. The expression and localization of GPR37 in different cell types within the CNS require further investigation.¹⁶⁰ Although GPR37 has been found to be highly expressed in myelinating OLs, astrocytes, and specific neuronal subtypes, its precise distribution and functional implications in these cells are not fully understood.³¹ Comprehensive studies using advanced techniques such as single-cell RNA sequencing and immunohistochemistry can help elucidate the cell-specific expression patterns and subcellular localization of GPR37, providing valuable information about its roles in various CNS cell types.

Understanding the molecular interactions of GPR37 and downstream signaling pathways is crucial for unraveling its functional mechanisms. While interactions with PDZ domain-containing proteins, such as PICK1 and GABARAPL2, as well as serial truncation of the N-terminus, have been reported, the specific consequences of these interactions and their impact on GPR37 signaling remain unclear.^{161, 165} Further molecular and biochemical studies can shed light on the PPI involving GPR37 and unravel the signaling cascades activated upon ligand binding.¹¹⁶

Another critical gap in GPR37 research is the limited availability of suitable preclinical models that recapitulate the pathophysiological aspects of CNS disorders associated with GPR37 dysfunction. While animal studies have provided valuable insights into the potential roles of GPR37 in neurodegenerative diseases, more comprehensive models are needed to fully understand its involvement in disease progression and evaluate the therapeutic potential of GPR37 modulation.³⁰ The development of genetically modified animal models, including conditional knockout or transgenic mice targeting GPR37, can provide valuable tools to investigate the receptor’s functions in vivo.

Despite the promising potential of GPR37 as a therapeutic target, developing potent, specific, and effective ligands or modulators remains a challenge. Identifying small molecules or peptides that can selectively activate or inhibit GPR37 signaling pathways is crucial for developing targeted therapies. High-throughput screening and structure-based drug design approaches can aid in the discovery of novel ligands with high affinity and selectivity for GPR37. Homology modeling benefits from increasing experimental structural data and computational advancements, offering insights into protein 3D structures. However, challenges persist in ligand docking, modeling loops, and selecting optimal models. To address this challenge, the binding pockets, conformational dynamics, and signaling pathways associated with GPR37 must be thoroughly investigated to gain a deeper understanding of its unique properties. This knowledge is essential for designing ligands that precisely target GPR37 while minimizing interactions with other receptors. Moreover, advancements in computational modeling, virtual screening, and high-throughput screening technologies offer promising avenues for rationalizing and screening novel ligands with enhanced specificity and affinity for GPR37. A concerted effort to uncover the molecular underpinnings of GPR37’s function and its interactions within complex cellular networks will pave the way for developing tailored therapeutics that unlock the full potential of GPR37 modulation, providing targeted benefits and reducing the risk of off-target effects. Continued efforts in method development, automation, and refinement are necessary to harness the full potential of homology modeling in structural biology and rational drug design.

Addressing these gaps in basic and translational GPR37 research will significantly advance our understanding of its biological functions, role in various human diseases, and its potential as a therapeutic target. Such endeavors will facilitate the development of innovative treatment strategies for neurodegenerative diseases and other indications, to improve patient outcomes. In addition, identifying allosteric binding sites in GPR37 would not only shed light on its ability to bind multiple ligands simultaneously but also provide a deeper understanding of its functional versatility.^{4, 169} Allosteric binding sites offer the potential for modulating GPR37 activity through the binding of allosteric modulators, which can fine-tune receptor signaling and provide new opportunities for drug development.¹⁶⁹ Exploring the allosteric regulation of GPR37 could uncover novel therapeutic strategies and expand our knowledge of the complex mechanisms underlying GPR37’s biological functions.

6. CONCLUSIONS AND FUTURE DIRECTIONS

GPR37 has significant potential as a novel therapeutic target for CNS disorders and other conditions. Although our understanding of GPR37 has advanced in recent years, there are several key findings and future directions that emerge from the review of existing literature.¹⁷⁰ Firstly, the physiological functions of GPR37 and its endogenous ligands remain largely unknown. Further research is needed to validate and characterize the proposed ligand-receptor interactions and to elucidate the downstream signaling pathways activated by GPR37. The identification of endogenous ligands will be crucial in understanding its physiological roles and involvement in human diseases. Secondly, investigating the cell-specific expression and localization of GPR37 in different CNS cell types is essential for unraveling its functional implications. Advanced techniques such as single-cell RNA sequencing and immunohistochemistry can provide valuable insights into the expression patterns and subcellular localization of GPR37, shedding light on its roles in various cell types, including myelinating OLs and astrocytes. Moreover, further research is needed to solve the crystal structures and elucidate the molecular interactions and signaling pathways of GPR37. Understanding the consequences of its interactions with PDZ domain-containing proteins and other binding partners will provide insights into the downstream signaling cascades activated by GPR37. Utilizing molecular and biochemical approaches can help unravel the complex PPI and signaling mechanisms involving GPR37. The development of appropriate preclinical models that accurately recapitulate the pathophysiological aspects of human diseases, particularly CNS disorders, associated with GPR37 dysfunction is also crucial. These models will enable a better understanding of the role of the receptor in disease progression and facilitate the evaluation of potential therapeutic interventions. Genetically modified animal models, such as conditional knockout or transgenic mice targeting GPR37, can serve as valuable tools for in vivo studies. Finally, the therapeutic targeting of GPR37 requires the discovery and development of potent and specific ligands, including small molecule agonists, antagonists, and allosteric modulators. High-throughput screening and structure-based drug design approaches are anticipated to aid in the identification of novel compounds that selectively modulate GPR37 signaling. Such ligands could serve as chemical leads for further optimizations and valuable pharmacological tools for elucidating the receptor functions and could potentially lead to the development of novel target-based therapeutics.

In summary, GPR37 represents a promising but still less explored therapeutic target in the field of neuro- and psycho-pharmacology. Addressing the identified gaps in basic and translational research on GPR37, such as understanding its physiological functions, elucidating endogenous ligands, characterizing cell-specific expression, unraveling crystal and CryoEM structures and molecular interactions, developing appropriate preclinical models, and discovering potent and specific ligands, will contribute to a more comprehensive understanding of this emerging drug target and pave the way for the development of novel therapeutic strategies with potential clinical impact.

ABBREVIATIONS

5-HT₄

5-hydroxytryptamine receptor 4

ADAM10

A Disintegrin and Metalloprotease 10

A_2AR

Adenosine A(2A) receptor

ARDS

acute respiratory distress syndrome

ARU

Artesunate

ASD

Autism Spectrum Disorder

CNS

central nervous system

CSF

cerebrospinal fluid

ChR2

channel rhodopsin 2

ceRNA

competitive endogenous RNA

CASPR2

contactin-associated protein-like 2

Dhh

desert hedgehog

DAT

dopamine transporter

D₁R

dopaminergic D1 receptor

D₂R

dopaminergic D2 receptor

EPAC

exchange protein activated by cAMP

ER

endoplasmic reticulum

ERK

extracellular signal-regulated kinases

ET_BR

endothelin-type B

FASD

Fetal Alcohol Spectrum Disorders

FDA

Food and Drug Administration

GPR37

G protein-coupled receptor 37

GPR37L1

G protein-coupled receptor 37 Like 1

GPCR

G protein-coupled receptor

GTEx

Genotype-Tissue Expression

GnRH

Gonadotropin-releasing hormone

HuH7

human hepatocellular

IPA

indole-3-propionic acid

KA

kainic acid

LPS

lipopolysaccharide

LUAD

lung adenocarcinoma

MAPK

mitogen-activated protein kinase

MB

medulloblastoma

MLC

megalencephalic leukoencephalopathy with subcortical cysts

mTOR

mechanistic target of rapamycin

MUPP1

multiple PDZ domain protein 1

NPD1

Neuroprotectin D1

OLs

oligodendrocytes

oGPCR

orphan G protein-coupled receptor

OCN

osteocalcin

PaelR

parkin-associated endothelin receptor-like receptor

PDZ

protein-95/Discs large/ZO-1

PPI

protein-protein interaction

PSAP

prosaposin

Ptch1

Patched-1

SCs

Sertoli cells

Shh

sonic hedgehog

Smo

Smoothened

SRLV

small ruminant lentiviruses

TIICs

tumor-infiltrating immune cells

tGFP

turbo green fluorescent protein tagged