Pharmacology Cambridge RSS Paper feed

Front Immunol. 2025 Mar 11;16:1547330. doi: 10.3389/fimmu.2025.1547330. eCollection 2025.

ABSTRACT

INTRODUCTION: The nuanced roles of neuropilin (NRP) isoforms, NRP1 and NRP2, have attracted considerable scientific interest regarding cancer progression. Their differential expressions across various cancer types are specific to NRP isoforms which are shown in a cancer type-dependent manner. It accounts for the different mechanisms involved, driven by a co-expression of gene-sets associated with overexpressed NRP1 or NRP2. Their different expressions on tumour-associated macrophages (TAMs) with disparate markers are associated with the heterogenous tumour microenvironment (TME) through their plasticity and pro-tumorigenic activities.

METHODS: Single-cell RNA sequencing (scRNA-seq) analyses were performed on tumours from clear cell Renal Cell Carcinoma (ccRCC) and skin cutaneous melanoma (SKCM) which exhibit the highest expressions of NRP1 and NRP2, respectively. Datasets were processed using established bioinformatics pipelines, including clustering algorithms, to determine cellular heterogeneity and quantify NRP isoform expression within distinct macrophage populations. Using differential gene expression analysis (DEGA) alongside co-enrichment studies, we explored gene-sets associated with NRP1 or NRP2 overexpression in TAMs.

RESULTS: Our analysis revealed a marked upregulation of NRP1 in TAMs isolated from ccRCC and elevated NRP2 expression in SKCM-derived TAMs. Both NRP1+ and NRP2+ macrophages showed an M2-like polarisation characterised by immune suppression and extracellular matrix degradation. Coupled with the previously uncharacterised NRP isoform specific- subpopulations within these cancers identified by DEGA, co-enrichment analyses demonstrated that the upregulation of gene-sets associated with NRP1 is associated with angiogenesis and tumour progression through VEGF signalling, while gene-sets with NRP2 showed dual functionality in the TME-dependent manner. Their distinct roles in regulating macrophage plasticity, tumour invasion, and metastasis were highlighted.

DISCUSSION: These findings underscore distinct isoform-specific mechanisms by which NRP1 and NRP2 contribute to TAM-mediated cancer progression. This study aims to establish a foundation for future research, leading to biological experiments with focused gene-sets derived from our findings. This approach can contribute to the development of immunomodulatory strategies targeting specific NRP isoforms in macrophages, tailored to individual cancer types and abnormal expressions of those gene markers, potentially offering a more effective therapeutic approach compared to broad-spectrum NRP inhibition strategies.

PMID:40134439 | PMC:PMC11933088 | DOI:10.3389/fimmu.2025.1547330

Neuropharmacology. 2025 Feb 28:110391. doi: 10.1016/j.neuropharm.2025.110391. Online ahead of print.

ABSTRACT

Pro-inflammatory mediators can directly activate pain-sensing neurons, known as nociceptors. Additionally, these mediators can sensitise ion channels and receptors expressed by these cells through transcriptional and post-translational modulation, leading to nociceptor hypersensitivity. A well-characterised group of ion channels that subserve nociceptor sensitisation is the transient receptor potential (TRP) superfamily of cation channels. For example, the roles of TRP channels vanilloid 1 (TRPV1) and ankyrin 1 (TRPA1) in nociceptor sensitisation and inflammatory pain have been extensively documented. In the case of TRP melastatin 3 (TRPM3), however, despite the increasing recognition of this channel's role in inflammatory pain, the mediators driving its sensitisation during inflammation remain poorly characterised. Here, using Ca2+ imaging, we found that an inflammatory soup of bradykinin, interleukin 1β (IL-1β) and tumour necrosis factor α (TNFα) sensitised TRPM3 function in isolated mouse sensory neurons; IL-1β and TNFα, but not bradykinin, independently potentiated TRPM3 function. TRPM3 expression and translocation to the membrane remained unchanged upon individual or combined exposure to these inflammatory mediators, which suggests that post-translational modification might occur. Finally, using the complete Freund's adjuvant-induced model of knee inflammation, we found that systemic pharmacological blockade of TRPM3 does not alleviate inflammatory pain (as assessed through evaluation of digging behaviour and dynamic weight bearing), which contrasts with previous reports using different pain models. We propose that the nuances of the immune response may determine the relative contribution of TRPM3 to nociceptive signalling in different neuro-immune contexts. Collectively, our findings improve insight into the role of TRPM3 sensitisation in inflammatory pain.

PMID:40024472 | DOI:10.1016/j.neuropharm.2025.110391

Nat Commun. 2025 Feb 27;16(1):2020. doi: 10.1038/s41467-025-57034-y.

ABSTRACT

G protein-coupled receptors (GPCRs) constitute a functionally diverse protein family and are targets for a broad spectrum of pharmaceuticals. Technological progress in X-ray crystallography and cryogenic electron microscopy has enabled extensive, high-resolution structural characterisation of GPCRs in different conformational states. However, as highly dynamic events underlie GPCR signalling, a complete understanding of GPCR functionality requires insights into their conformational dynamics. Here, we present a large dataset of molecular dynamics simulations covering 60% of currently available GPCR structures. Our analysis reveals extensive local "breathing" motions of the receptor on a nano- to microsecond timescale and provides access to numerous previously unexplored receptor conformational states. Furthermore, we reveal that receptor flexibility impacts the shape of allosteric drug binding sites, which frequently adopt partially or completely closed states in the absence of a molecular modulator. We demonstrate that exploring membrane lipid dynamics and their interaction with GPCRs is an efficient approach to expose such hidden allosteric sites and even lateral ligand entrance gateways. The obtained insights and generated dataset on conformations, allosteric sites and lateral entrance gates in GPCRs allows us to better understand the functionality of these receptors and opens new therapeutic avenues for drug-targeting strategies.

PMID:40016203 | DOI:10.1038/s41467-025-57034-y

Curr Opin Biotechnol. 2025 Feb 5;92:103267. doi: 10.1016/j.copbio.2025.103267. Online ahead of print.

ABSTRACT

Intermolecular relationships at the cell surface dictate the behavior and regulatory network of cells. Such interactions often require precise spatial control for optimal response. By binding simultaneously to two different target sites, bispecific binders can bridge molecules of interest. Despite decades of bispecific development, only recently have bispecifics been engineered with programmable, tuneable geometries to replicate natural interaction geometries or achieve new responses from unnatural arrangements. This review highlights emerging methods of protein engineering and modular bioconjugation to control pairing and orientation of binders in bispecific scaffolds. We also describe novel biophysical and phenotypic assays, which reveal how bispecific geometries change cell fate. These approaches are informing design of next-generation precision therapeutics, as well as uncovering fundamental features of signal integration.

PMID:39914134 | DOI:10.1016/j.copbio.2025.103267

NPJ Biosens. 2025;2(1):2. doi: 10.1038/s44328-024-00018-7. Epub 2025 Jan 16.

ABSTRACT

The reactivation of heterotrimeric protein phosphatase 2A (PP2A) through small molecule activators is of interest to therapeutic intervention due to its dysregulation, which is linked to chronic conditions. This study focuses on the PP2A scaffold subunit PR65 and a small molecule activator, ATUX-8385, designed to bind directly to this subunit. Using a label-free single-molecule approach with nanoaperture optical tweezers (NOT), we quantify its binding, obtaining a dissociation constant of 13.6 ± 2.5 μM, consistent with ensemble fluorescence anisotropy results but challenging to achieve with other methods due to low affinity. Single-molecule NOT measurements reveal that binding increases optical scattering, indicating PR65 elongation. This interpretation is supported by all-atom molecular dynamics simulations showing PR65 adopts more extended conformations upon binding. This work highlights NOT's utility in quantifying binding kinetics and structural impact, offering insights valuable for drug discovery.

PMID:39830999 | PMC:PMC11738983 | DOI:10.1038/s44328-024-00018-7

J Cell Sci. 2025 Jan 1;138(1):JCS263434. doi: 10.1242/jcs.263434. Epub 2025 Jan 15.

ABSTRACT

G protein-coupled receptor (GPCR) signalling pathways underlie numerous physiological processes, are implicated in many diseases and are major targets for therapeutics. There are more than 800 GPCRs, which together transduce a vast array of extracellular stimuli into a variety of intracellular signals via heterotrimeric G protein activation and multiple downstream effectors. A key challenge in cell biology research and the pharmaceutical industry is developing tools that enable the quantitative investigation of GPCR signalling pathways to gain mechanistic insights into the varied cellular functions and pharmacology of GPCRs. Recent progress in this area has been rapid and extensive. In this Review, we provide a critical overview of these new, state-of-the-art approaches to investigate GPCR signalling pathways. These include novel sensors, Förster or bioluminescence resonance energy transfer assays, libraries of tagged G proteins and transcriptional reporters. These approaches enable improved quantitative studies of different stages of GPCR signalling, including GPCR activation, G protein activation, second messenger (cAMP and Ca2+) signalling, β-arrestin recruitment and the internalisation and intracellular trafficking of GPCRs.

PMID:39810711 | DOI:10.1242/jcs.263434

Cells. 2024 Dec 21;13(24):2121. doi: 10.3390/cells13242121.

ABSTRACT

The adenosine A1 receptor (A1R) is a promising target for pain treatment. However, the development of therapeutic agonists is hampered by adverse effects, mainly including sedation, bradycardia, hypotension, or respiratory depression. Recently discovered molecules able to overcome this impediment are the positive allosteric modulator MIPS521 and the A1R-selective agonist BnOCPA, which are both potent and powerful analgesics with fewer side effects. While BnOCPA directly activates the A1R from the canonical orthosteric site, MIPS521 binds to an allosteric site, acting in concert with orthosteric adenosine and tuning its pharmacology. Given their overlapping profile in pain models but distinct mechanisms of action, we combined pharmacology and microsecond molecular dynamics simulations to address MIPS521 and BnOCPA activity and their reciprocal influence when bound to the A1R. We show that MIPS521 changes adenosine and BnOCPA G protein selectivity in opposite ways and propose a structural model where TM7 dynamics are differently affected and involved in the G protein preferences of adenosine and BnOCPA.

PMID:39768211 | DOI:10.3390/cells13242121

ACS Omega. 2024 Dec 2;9(50):49422-49431. doi: 10.1021/acsomega.4c06868. eCollection 2024 Dec 17.

ABSTRACT

Latent membrane protein 1 (LMP1) plays a crucial role in Epstein-Barr virus (EBV)'s ability to establish latency and is involved in developing and progressing EBV-associated cancers. Additionally, EBV-infected cells affect the immune responses, making it challenging for the immune system to eliminate them. Due to the aforementioned reasons, it is crucial to understand the structural features of LMP1, which are essential for the development of novel cancer therapies that target its signaling pathways. To date, there is yet to be a complete LMP1 protein structure; therefore, in our work, we modeled the full-length LMP1 containing the short cytoplasmic N-terminus, six transmembrane domains (TMDs), and a long-simulated C-terminus. Our model showed good stability and protein compactness evaluated through accelerated-molecular dynamics, where the conformational ensemble exhibited compact folds, particularly in the TMDs. Our results suggest that specific domains or motifs, predominantly in the C-terminal domain of LMP1, show promise as potential drug targets. As a whole, our work provides insights into key structural features of LMP1 that will allow the development of novel LMP1 therapies.

PMID:39713625 | PMC:PMC11656244 | DOI:10.1021/acsomega.4c06868

Proc Natl Acad Sci U S A. 2024 Dec 17;121(51):e2410653121. doi: 10.1073/pnas.2410653121. Epub 2024 Dec 11.

ABSTRACT

Inflammation is associated with localized acidosis, however, attributing physiological and pathological roles to proton-sensitive receptors is challenging due to their diversity and widespread expression. Here, agonists of the proton-sensing GPCR, GPR65, were systematically characterized. The synthetic agonist BTB09089 (BTB) recapitulated many proton-induced signaling events and demonstrated selectivity for GPR65. BTB was used to show that GPR65 activation on fibroblast-like synoviocytes (FLS), cells that line synovial joints, results in the secretion of proinflammatory mediators capable of recruiting immune cells and sensitizing sensory neurons. Intra-articular injection of BTB resulted in GPR65-dependent sensitization of knee-innervating neurons and nocifensive behaviors in mice. Stimulation of GPR65 on human FLS also triggered the release of inflammatory mediators and synovial fluid samples from human osteoarthritis patients were shown to activate GPR65. These results suggest a role of GPR65 in mediating cell-cell interactions that drive inflammatory joint pain in both mice and humans.

PMID:39661058 | DOI:10.1073/pnas.2410653121