Neuronal Pentraxin Receptor, a Novel Putative Integral Membrane Pentraxin That Interacts with Neuronal Pentraxin 1 and 2 and Taipoxin-associated Calcium-binding Protein 49

Neuronal pentraxins (NPs) function in the extracellular matrix to bind alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. Three NPs have been described, neuronal activity-regulated pentraxin (Narp), which is regulated as an immediate early gene, NP1, and neuronal pentraxin receptor (NPR). Narp and NP1 enhance synaptogenesis and glutamate signaling by clustering AMPA receptors, whereas NPR contributes to removing AMPA receptors during group I metabotropic glutamate receptor-dependent long-term depression. Here, we examine mice with genetic deletions [knockout (KO)] of each NP to assess their contributions to cocaine-induced neuroplasticity. Consistent with a shared AMPA receptor clustering function for Narp and NP1, deletion of either NP caused similar behavioral alterations. Thus, although both Narp and NP1 deletion promoted cocaine-induced place preference, NPR deletion was without effect. In addition, although Narp and NP1 KO showed reduced time in the center of a novel environment, NPR KO mice spent more time in the center. Finally, although Narp and NP1 KO mice showed blunted locomotion after AMPA microinjection into the accumbens 3 weeks after discontinuing repeated cocaine injections, the AMPA response was augmented in NPR KO. Likewise, endogenous glutamate release elicited less motor activity in Narp KO mice. Consistent with reduced AMPA responsiveness after chronic cocaine in Narp KO mice, glutamate receptor 1 was reduced in the PSD fraction of Narp KO mice withdrawn from cocaine. These data indicate that NPs differentially contribute to cocaine-induced plasticity in a manner that parallels their actions in synaptic plasticity.

Neuronal pentraxin 1 (NP1), neuronal pentraxin 2 (NP2), and neuronal pentraxin receptor (NPR) are members of a new family of proteins identified through interaction with a presynaptic snake venom toxin taipoxin. We have proposed that these three neuronal pentraxins represent a novel neuronal uptake pathway that may function during synapse formation and remodeling. We have investigated the mutual interactions of these proteins by characterizing their enrichment on taipoxin affinity columns; by expressing NP1, NP2, and NPR singly and together in Chinese hamster ovary cells; and by generating mice that fail to express NP1. NP1 and NP2 are secreted, exist as higher order multimers (probably pentamers), and interact with taipoxin and taipoxin-associated calcium-binding protein 49 (TCBP49). NPR is expressed on the cell membrane and does not bind taipoxin or TCBP49 by itself, but it can form heteropentamers with NP1 and NP2 that can be released from cell membranes. This is the first demonstration of heteromultimerization of pentraxins and release of a pentraxin complex by proteolysis. These processes are likely to directly effect the localization and function of neuronal pentraxins in neuronal uptake or synapse formation and remodeling.

e15022 Background: The human neuronal pentraxin receptor (NPTXR) gene encodes a type II transmembrane protein that functions as a trans-synaptic organizer and anchors neuronal pentraxin complexes to plasma membranes. Beside a subset of neuronal cells (Hippocampus and Cerebral Cortex), expression of NPTXR is not detected in all other tissues [ The Human Protein Atlas database]. In recent studies [Kanda et al. (2020) Mol. Cancer. doi: 10.1186/s12943-020-01251-0], transcriptome and bioinformatics analysis of gastric cancer (GC) tissues from patients with or without metastasis have revealed that NPTXR expression was associated with disease progression. NPTXR expression was also correlated negatively with survival in GC patients. It appears that NPTXR does not have a major physiological function; e.g., NPTXR−/− mice showed no abnormalities in reproduction, development, metabolism, or motor function. NPTXR is expressed in a number of GI cancers, i.e., gastric, esophageal, colorectal, pancreatic, bladder, liver, as well as breast and thyroid cancer. YB-800 is a fully humanized monoclonal antibody against NPTXR (identified by in silico analysis). Methods: The present studies were conducted to provide a rationale for the design of a YB-800-based antibody drug conjugate (ADC) for therapeutic intervention in oncology. Hence, YB-800 binding to and internalization in selected tumor cells, as well as immunohistochemistry (IHC) in tumor biopsies, were evaluated. Results: Taken together experimental data confirm that: (i) YB-800 binds to a new target (NPTXR) on selected tumor cells; confirmed by FACS analysis; (ii) IHC analysis in tumor samples demonstrates specific immunoreaction in bladder, gastric, prostate, hepatocellular carcinoma, colorectal cancer and cervical cancer, but not in adjacent healthy tissues; and (iii) once YB-800 binds to NPTXR, the conjugate is internalized in selected tumor cells in a time-dependent manner. Conclusions: In conclusion, YB-800 represents a first in class ADC candidate that could be used as novel treatment for gastro-intestinal cancers. Hence, as initial proof of concept, tesirine (DNA intercalating agent) and deruxtecan (topoisomerase I inhibitor) containing protease-sensitive linkers were coupled to YB-800 by stochastic maleimide conjugations to interchain cysteines. Initial results with YB-800-based ADCs will be discussed.

Neuronal pentraxins comprise three neuronal proteins, neuronal pentraxin receptor (NPR) which is a type-II transmembrane protein on the neuronal surface, and secreted neuronal pentraxin-1 and NARP. The general functions of neuronal pentraxins at synapses have not been explored, except for their basic AMPAR binding properties. Here, we examined the functional role of NPR at synapses because it is the only neuronal pentraxin that is anchored to the neuronal cell-surface membrane. We find that NPR is a potent inducer of both excitatory and inhibitory heterologous synapses, and that knockdown of NPR in cultured neurons decreases the density of both excitatory and inhibitory synapses. Our data suggest that NPR performs a general, previously unrecognized function as a universal organizer of synapses.

Neurons connect to each other via synapses to form neural circuits. Recent research has shown that neuropsychiatric disorders and neurological disorders, such as autism spectrum disorders and Alzheimer's disease, are synaptic diseases caused by abnormalities of synapses. Synaptic organizers are molecules responsible for synapse formation. Neuronal pentraxin 2 (NP2) is a synaptic organizer and a secreted protein that is expressed mainly in the hippocampus and cerebellum, and it contributes to synaptic plasticity. NP2 forms clusters with its family proteins, NP1 and neuronal pentraxin receptor, and binds to postsynaptic amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid–type receptors. In recent years, research has revealed the disease relevance of NP2. For example, it can be a biomarker of Alzheimer's disease, and its overexpression in the peripheral nervous system has been reported to cause chronic itch. However, the mechanism of NP2 function has not been well described at the molecular level. In this study, we developed a variable domain of a heavy-chain antibody (VHH) against NP2 to elucidate its molecular mechanism of action and to regulate its function of NP2. The obtained VHH N1 showed high specificity and affinity to NP2, and its binding mechanism was elucidated by X-ray crystallography. Furthermore, VHH N1 inhibited the binding of NP2 to amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid–type receptors, and this inhibitory activity was confirmed in cells. These results provide useful insights into the molecular mechanism of NP2 function and highlight the potential application of VHH N1 as a detection agent for NP2 or as a therapeutic agent for chronic itch.

Neuronal pentraxin 1 expression is regulated by hypoxia inducible factor-1α

Specific sensitivity of small cell lung cancer cell lines to the snake venom toxin taipoxin

BackgroundAlzheimer's dementia (AD) is a neurodegenerative condition characterized by progressive cognitive decline. Synaptopathy—defined as loss and dysfunction of existing synapses—is a hallmark pathological feature of AD and can directly contribute to underlying cognitive deficits. In this study, we meta‐analyzed several cerebrospinal fluid (CSF) and blood exosomal biomarkers associated with synaptopathy in AD and healthy controls (HCs).MethodOriginal peer‐reviewed articles that reported synaptic biomarker concentrations in CSF or blood exosomes were reviewed. Specifically, synaptosome associated protein‐25 (SNAP‐25), growth associated protein‐43 (GAP‐43), neuronal pentraxin receptor (NPTXR), neuronal pentraxin‐1 (NPTX‐1), neuronal pentraxin‐2 (NPTX‐2), complexin‐2, syntaxin‐1B, syntaxin‐7, and vesicle‐associated membrane protein‐2 (VAMP‐2) in AD and HCs were included for meta‐analysis. A random‐effects model was used to determine standardized mean differences (SMDs) and 95% confidence intervals (CIs). Heterogeneity was quantified using I2.ResultThe meta‐analysis included 43 study cohorts. In CSF, concentrations of SNAP‐25 (NAD/NHC = 394/539, SMD [95% CI] = 1.08 [0.73, 1.42], p < 0.001; I2 = 81.43%), GAP‐43 (NAD/NHC = 851/557, SMD [95% C] = 1.02 [0.69, 1.34], p < 0.001; I2 = 83.97%), and VAMP‐2 (NAD/NHC = 398/490, SMD [95% CI] = 0.32 [0.05, 0.60], p = 0.02; I2 = 64.91%) were elevated, and NPTXR (NAD/NHC = 575/470, SMD [95% CI] = ‐0.68 [‐0.96, ‐0.40], p < 0.001; I2 = 75.59%), NPTX‐1 (NAD/NHC = 344/333, SMD [95% CI] = ‐0.48 [‐0.64, ‐0.31], p < 0.001; I2 = 9.05%), and NPTX‐2 (NAD/NHC = 462/496, SMD [95% CI] = ‐0.78 [‐1.02, ‐0.54], p < 0.001; I2 = 66.43%) were decreased in AD compared to HCs. In blood exosomes, SNAP‐25 (NAD/NHC = 161/159, SMD [95% CI] = ‐1.05 [‐1.28, ‐0.82], p < 0.001; I2 = 0%) and GAP‐43 (NAD/NHC = 110/110, SMD [95% CI] = ‐1.66 [‐2.43, ‐0.88], p < 0.001; I2 = 77%) concentrations were decreased in AD.ConclusionThis study found that several synaptic biomarkers were significantly altered in AD in CSF and blood exosomes. There was significant heterogeneity for most comparisons (with the exception of CSF NPTX‐1 and blood exosome SNAP‐25) that remains to be explored. Nonetheless, further review of the identified biomarkers may provide fundamental insight into AD pathophysiology and disease trajectory.

BackgroundApproximately a third of frontotemporal dementia (FTD) is genetic with mutations in three genes accounting for the majority of the inheritance: C9orf72, GRN and MAPT. Synaptic dysfunction is a common mechanism in all of them and the use of fluid biomarkers could be helpful to improve the diagnostic accuracy and useful as a readout of cellular dysfunction within therapeutic trials.MethodA total of 193 cerebrospinal fluid samples from the GENetic FTD Initiative including 77 presymptomatic (31 C9orf72, 23 GRN, 23 MAPT), 55 symptomatic mutation carriers (26 C9orf72, 17 GRN, 12 MAPT) and 61 mutation‐negative controls were measured using a microflow LC PRM‐MS set‐up targeting 15 synaptic proteins: 14‐3‐3 proteins (eta, zeta/delta and epsilon), AP‐2 complex subunit beta, beta‐synuclein, gamma‐synuclein, complexin‐2, neurogranin, neuronal pentraxin receptor (NPTXR), neuronal pentraxin 1 (NPTX1), neuronal pentraxin 2 (NPTX2), phosphatidylethanolamine‐binding protein 1 (PEBP‐1), rab GDP dissociation inhibitor α (rab GDIα), syntaxin‐1B and syntaxin‐7. Mutation carrier groups were compared to each other and to controls using a bootstrapped linear regression model, adjusting for age and sex.ResultEight proteins were increased only in symptomatic MAPT mutation carriers (compared with controls) and not in symptomatic C9orf72 or GRN mutation carriers: 14‐3‐3‐eta, beta‐synuclein, gamma‐synuclein, neurogranin, PEBP‐1, rab GDIα, syntaxin‐1B and syntaxin‐7. In contrast, NPTX1 and NPTX2 were affected in all three genetic groups (decreased compared to controls), with NPTXR being affected in C9orf72 and GRN mutation carriers only (decreased compared to controls). No changes were seen in presymptomatic mutation carriers in these proteins. Figure 1 contains p‐values for all significant changes.ConclusionDifferential involvement of synaptic markers is seen in the genetic forms of FTD, with impairment particularly in those with MAPT mutations, with only the neuronal pentraxins affected in GRN and C9orf72 mutation carriers. Further work is needed to explore correlations with clinical and imaging biomarkers, whether there are changes in the late presymptomatic period, and how these markers change over time.

mGluR1/5-Dependent Long-Term Depression Requires the Regulated Ectodomain Cleavage of Neuronal Pentraxin NPR by TACE

Parallel age-associated changes in brain and plasma neuronal pentraxin receptor levels in a transgenic APP/PS1 rat model of Alzheimer's disease

BackgroundApproximately a third of frontotemporal dementia (FTD) is genetic with mutations in three genes accounting for most of the inheritance: C9orf72, GRN, and MAPT. Impaired synaptic health is a common mechanism in all three genetic variants, so developing fluid biomarkers of this process could be useful as a readout of cellular dysfunction within therapeutic trials.MethodsA total of 193 cerebrospinal fluid (CSF) samples from the GENetic FTD Initiative including 77 presymptomatic (31 C9orf72, 23 GRN, 23 MAPT) and 55 symptomatic (26 C9orf72, 17 GRN, 12 MAPT) mutation carriers as well as 61 mutation-negative controls were measured using a microflow LC PRM-MS set-up targeting 15 synaptic proteins: AP-2 complex subunit beta, complexin-2, beta-synuclein, gamma-synuclein, 14–3-3 proteins (eta, epsilon, zeta/delta), neurogranin, Rab GDP dissociation inhibitor alpha (Rab GDI alpha), syntaxin-1B, syntaxin-7, phosphatidylethanolamine-binding protein 1 (PEBP-1), neuronal pentraxin receptor (NPTXR), neuronal pentraxin 1 (NPTX1), and neuronal pentraxin 2 (NPTX2). Mutation carrier groups were compared to each other and to controls using a bootstrapped linear regression model, adjusting for age and sex.ResultsCSF levels of eight proteins were increased only in symptomatic MAPT mutation carriers (compared with controls) and not in symptomatic C9orf72 or GRN mutation carriers: beta-synuclein, gamma-synuclein, 14–3-3-eta, neurogranin, Rab GDI alpha, syntaxin-1B, syntaxin-7, and PEBP-1, with three other proteins increased in MAPT mutation carriers compared with the other genetic groups (AP-2 complex subunit beta, complexin-2, and 14–3-3 zeta/delta). In contrast, CSF NPTX1 and NPTX2 levels were affected in all three genetic groups (decreased compared with controls), with NPTXR concentrations being affected in C9orf72 and GRN mutation carriers only (decreased compared with controls). No changes were seen in the CSF levels of these proteins in presymptomatic mutation carriers. Concentrations of the neuronal pentraxins were correlated with brain volumes in the presymptomatic period for the C9orf72 and GRN groups, suggesting that they become abnormal in proximity to symptom onset.ConclusionsDifferential synaptic impairment is seen in the genetic forms of FTD, with abnormalities in multiple measures in those with MAPT mutations, but only changes in neuronal pentraxins within the GRN and C9orf72 mutation groups. Such markers may be useful in future trials as measures of synaptic dysfunction, but further work is needed to understand how these markers change throughout the course of the disease.

Background Alzheimer's disease (AD) is the most prevalent form of dementia. Currently, the most studied biomarkers of AD are cerebrospinal fluid (CSF) amyloid β 1-42, total tau and phosphorylated tau. However, misdiagnosis can exceed 20%. Recently, we found that CSF amyloid β precursor-like protein-1 (APLP1) and neuronal pentraxin receptor (NPTXR) are promising biomarkers of AD. The aim of the present study is to validate CSF APLP1 and NPTXR as biomarkers of AD severity. Methods APLP1 and NPTXR concentrations were measured in the CSF of patients with mild cognitive impairment (MCI) (n = 14), mild AD (n = 21), moderate AD (n = 43) and severe AD (n = 30) using enzyme-linked immunosorbent assays (ELISAs). Results CSF APLP1 and NPTXR were not associated with age or sex. CSF APLP1 was not different between any of the AD severity groups (p = 0.31). CSF NPTXR was significantly different between MCI and mild AD (p = 0.006), mild and moderate AD (p = 0.016), but not between moderate and severe AD (p = 0.36). NPTXR concentration progressively declined from MCI to mild, to moderate and to severe AD patients (p < 0.0001, Kruskal-Wallis test). CSF NPTXR positively correlated with the Mini-Mental Status Examination (MMSE) score (p < 0.001). Conclusions NPTXR concentration in CSF is a promising biomarker of AD severity and could inform treatment success and disease progression in clinical settings.

Stroke is a growing burden for healthcare systems globally, with 1 in 4 people over the age of 25 expected to have a stroke in their lifetime. Ischemic stroke – the most common form of stroke – is characterized by rapid and propagating cell death and increased levels of the excitatory neurotransmitter glutamate in the affected tissue. However, the mechanisms underlying cell death and increased cell vulnerability are not fully understood. Neuronal activity-regulated pentraxin (aka Narp) is a calcium-dependent lectin and immediate early gene that is implicated in the aggregation of ionotropic glutamate receptor α-amino-3-hydroxy-5-methyl-4- isoxazolepropionic acid (AMPA) at excitatory synapses – which has implications in a cell’s sensitivity to glutamate. Using the photothrombosis model of stroke, Narp was significantly upregulated in ipsilateral and contralateral hemispheres in a time-dependent manner. In mice treated with Narp via a contralateral intracerebroventricular injection, there was a significant decrease in the expression of active and inactive Tumor Necrosis Factor Converting Enzyme (TACE) as well as its cleavage product Neuronal Pentraxin Receptor (NPR) – a pentraxin that interacts with Narp and has been implicated in AMPA receptor internalization. Additionally, Narp treatment in vitro decreased NPR expression in a model of glutamate excitotoxicity. These data suggest that Narp may participate in cell death mechanisms in the context of photothrombosis by modulating the expression of TACE and consequential cleavage of NPR.

BackgroundEarly deficits of cognitive symptoms have molecular background closely related to Alzheimer’s Disease (AD). Before developing full‐blown AD, mild cognitive impairment (MCI) develops. It has been suggested that synaptic pathology is closely associated with memory impairment in the early phase of the disease and may be monitored by assessment of the synaptic proteins in cerebrospinal fluid (CSF), like neuronal pentraxin receptor (NPTXR). The highest expression of NPTXR and involvement in neuronal processes have been observed in the hippocampus and cortex. This candidate biomarker of synaptic dysfunction may have a crucial role in synaptic transmission and modulation of memory processes with other synaptic proteins. The purpose of the our investigation was to assessed the neuronal pentraxin receptor (NPTXR) level in cerebrospinal fluid (CSF) in MCI patients.MethodThe study included 17 patients with MCI and 17 non‐demented controls. The CSF levels of NPTXR and classical AD biomarkers, such as Aβ‐42, Aβ‐42/Aβ‐40, Tau, and pTau181, were assessed by commercially available immunoenzyme assays.ResultThe CSF concentration of NPTXR was significantly lower in MCI patients compared to non‐demented controls. Moreover, NPTXR level was positively correlated with Tau181, Aβ‐42, and total Tau proteins in MCI patients.ConclusionOur results suggest that Neuronal Pentraxin Receptor may be one of the biomarker reflecting synaptic dysfunction in MCI patients. These preliminary results seem very promising, particularly for the early diagnosis of Alzheimer’s disease. Future research concerning NPTXR is necessary to understand better the role of the protein in early pathological processes of the disease.