Neuregulin Research Articles

3D genomics identifies previously unreported yet clinically actionable gene fusions and complex rearrangements in a cohort of driver-negative colorectal carcinoma tumors.

e15608 Background: Molecular characterization by next-generation sequencing (NGS) has been unable to detect pathogenic driver mutations in a significant number of tumors. For example, 40% of colorectal carcinomas (CRC) are RAS and BRAF wild-type and do not have therapeutic targets after comprehensive molecular characterization using FISH, IHC and NGS. Analysis of 3D genomics data produced by DNA-based chromosome conformation capture (Hi-C) assay has been shown to detect gene fusions and other rearrangements in tumor biopsies. We applied Hi-C in a cohort of CRC tumors previously characterized using FISH, IHC, whole exome sequencing (WES) and whole transcriptome sequencing (WTS) by a large commercial reference laboratory, where no targets had been reported, to evaluate the potential for Hi-C to identify novel therapeutic targets. Methods: FFPE tissue from 14 CRC tumors were subjected to Hi-C using the Arima HiC+ for FFPE kit. Hi-C libraries were sequencing to ~100M raw read-pairs on an Illumina NovaSeq, and fusions were called using the Arima-SV pipeline. To attribute clinical significance, we characterized fusions based on whether the fusion event involved a Tier 1 therapeutic level biomarker or a Tier 3 diagnostic or prognostic biomarker according to NCCN and OncoKb, or a Tier 2 biomarker if the biomarker was the target of a therapy in an ongoing clinical trial. Results: Hi-C detected fusions implicating a Tier 1 biomarker in 50% (7/14) of tumors, a Tier 2 level biomarker in 14% (2/14), and a Tier 3 level biomarker in 14% (2/14). Homologous recombination repair (HRR) gene rearrangements were detected in 29% (n = 4/14) of CRC tumors, disrupting HRR genes ( BRIP1, BRCA1, BRCA2, RAD51D). NRG1 gene fusions were detected in 14% (2/14), and amplified complex rearrangement carrying KRAS was detected in one CRC case. A complex rearrangement involving extrachromosomal DNA (ecDNA) amplification carrying EGFR was detected in two cases that were both previously found to have EGFR amplifications, but the ecDNA nature was not previously known and was confirmed by FISH. Lastly, Hi-C detected gene rearrangements involving POLE or APC in two CRC cases, alterations in which are known to increase risk for colonic polyposis. Conclusions: Fusion detection by Hi-C in CRC may uncover numerous therapeutic biomarkers not currently detected by molecular profiling. Our data suggests that homologous recombination deficiency (HRD) may frequently result from HRR gene rearrangements readily detectable by Hi-C, and this alternate mechanism for HRD may serve as a therapeutic biomarker for HRD directed therapies. Our data also suggests that therapeutically targetable gene fusions and complex rearrangements involving therapeutic level biomarkers are also readily detectable by Hi-C, detection of which may increase the number of CRC patients treatable with targeted therapies.

Association of a haplotype in the NRG1 gene with schizophrenia: a case-control study

Abstract Objectives Schizophrenia (SZ) is a severe multifactorial disease. NRG1 is a gene acting in the development of SZ. A number of NRG1 single nucleotide polymorphisms (SNPs) and their haplotypes are associated with SZ. In the present study, we investigated the association of a NRG1 haplotype (G-C in rs6988339-rs3757930 frame) which was reported to be associated with SZ, and two other SNPs in the same gene (rs74942016, rs80127039) whose rare missense alleles were found in SZ patients. Also, we analyzed disease associations of potential new haplotypes constructed by the variants of these SNPs. Methods We genotyped 4 SNPs in a sample consisting of 302 SZ patients and 333 controls from a local Turkish population. We tested the disease associations of these variants at single SNP, haplotype and diplotype levels in case-control design. Results At single SNP level, the CC genotype of rs3757930 was associated with SZ (p=0.038). The previously reported association of G-C haplotype in rs6988339-rs3757930 frame was absent (p=0.416), but we found another haplotype (C-G in rs3757930-rs74942016, p=0.018) and three diplotypes (A-C/G-C diplotype of rs6988339-rs3757930 frame, C-G/C-G diplotype of rs3757930-rs74942016 frame, and A-C-G/G-C-G diplotype of rs6988339-rs3757930-rs74942016 frame) associated with schizophrenia in our sample. Conclusions Our study indicated the associations of a SNP, a haplotype, and a diplotype of NRG1 with schizophrenia and supported the involvement of NRG1 gene in the development of the disease. Since our sample was collected from a limited geographic area, the associations we have reported need to be supported by further studies in different populations.

Scalp acupuncture improves disordered behavior in the valproic acid-induced rat model of autism: 头针改善丙戊酸钠诱导的孤独症大鼠行为学及其机制研究

ObjectivesTo investigate the underlying mechanisms of scalp acupuncture treatment (SAT) on autism spectrum disorder (ASD). MethodsThirty male Wistar rat pups that had been prenatally exposed to valproic acid sodium (VPA) were randomly divided into the VPA, VPA+acupoint, and VPA+ non-acupoint groups using the random number table method, with 10 rats in each group. Ten pups who had been prenatally exposed to saline were assigned to the control group (CG). There was no intervention in either the control or VPA groups. In the acupoint group, “Shenting (GV24),” bilateral “Benshen (GB13)” were manipulated. In the non-acupoint group, the area below the costal space was stimulated. Acupuncture stimulation lasted for 40 min, with manual twisting of the needles every 10 min, 5 days/week, with 2 days of rest per week, for a total duration of 4 weeks. After the corresponding treatments, behavioral tests (including the open field, social interaction, and Morris water maze tests) were performed to evaluate the therapeutic effects. RT-PCR and Western blotting were performed to detect the expression of neuregulin 1 (NRG1)/ErbB4. ResultsIn the open field test, the activity time spent in the central area in the VPA+acupoint group was significantly longer than that in the VPA group and VPA+ non-acupoint group (both P<0.05). The total length in the VPA+acupoint group was significantly longer than that in the VPA group (P<0.05). The number of bouts in the central area of the VPA+acupoint group was significantly higher than that of the VPA group (P<0.05). In session I of social interaction test, all experimental rats spent more time interacting with stranger 1 (all P<0.05). In session II, the CG and VPA+acupoint groups rats showed more interest in searching for new strangers, but the VPA+non-acupoint group spent more time interacting with stranger 1 than with stranger 2(all P<0.05). In the Morris water maze test, compared with the VPA group, the latency of the VPA+acupoint group was shorter (day 2, 3, 4, 5, P<0.05); compared with VPA+acupoint group, the latency of the VPA+non-acupoint group was longer (day 2, 4, P<0.05). The mean distance in the VPA+acupoint group was shorter than that in the VPA group (day 3, 5, P<0.05). The platform quadrant time of the VPA+non-acupoint group was significantly shorter than that of the VPA+acupoint group (P<0.05) (day 6). The VPA+acupoint group had more platform crossings than the VPA group (P<0.05), and the VPA+ non-acupoint group had fewer platform crossings than the VPA+acupoint group (P<0.05) (day 6). After SAT, the expression levels of NRG1 and ErbB4 proteins in the VPA+acupoint group were significantly increased than those in the VPA group (both P<0.05) in the medial prefrontal cortex (mPFC). The expression levels of NRG1 and ErbB4 proteins in VPA+non-acupoint group were significantly less than those in the VPA+acupoint group (both P<0.05) in the mPFC. After SAT, the expression levels of NRG1 and ErbB4 proteins in the VPA+acupoint group were significantly higher than those in the VPA group (both P<0.05) in the hippocampus. The expression levels of NRG1 and ErbB4 proteins in the VPA+non-acupoint group were significantly lower than those in the VPA+acupoint group (both P<0.05) in the hippocampus. After SAT, the expression levels of mRNA levels of NRG1-ErbB4 in the mPFC and hippocampus in VPA+acupoint group were significantly higher than those in the VPA group (all P<0.05). The mRNA expression levels of NRG1-ErbB4 in the mPFC and hippocampus in VPA+ non-acupoint group were significantly lower than those in the VPA+acupoint group (all P<0.05). ConclusionsOur results suggest that SAT can improve ASD-like behaviors in young rats in a VPA model of ASD. This may be related to its function in upregulating the expression of protein and gene of NRG1 and its receptor ErbB4 in the mPFC.

HER3- A key survival pathway and an emerging therapeutic target in metastatic colorectal cancer and pancreatic ductal adenocarcinoma.

Colorectal cancer (CRC) and pancreatic ductal adenocarcinoma (PDAC) are highly metastatic cancers with poor survival rates. The tumor microenvironment has been shown to play a critical role in cancer progression and response to therapies. Endothelial cells (ECs) are a key component of the tumor microenvironment and promote cancer cell survival by secreting soluble factors that activate cancer-promoting signaling pathways. Studies from us and others identified HER3 as a key mediator of liver EC-induced chemoresistance and cancer cell growth in metastatic CRC and PDAC. In this article, we discuss that HER3-targeted therapies may be effective in treating patients with HER3-expressing CRC and PDAC, and highlight the importance of applying HER3 expression as a predictive biomarker for patient response to HER3-targeted therapies. We also discuss the challenges encountered in past clinical trials of HER3-targeted therapies, including the role of NRG1 gene fusions, alternative HER3 activation mechanisms, and adaptive resistance mechanisms. Finally, we conclude by suggesting the future directions of HER3-targeted therapies, including novel approaches to overcome chemoresistance and promote cancer cell death.

RNA Sequencing Identifies Novel NRG1 Fusions in Solid Tumors that Lack Co-Occurring Oncogenic Drivers

NRG1 gene fusions are rare, therapeutically relevant, oncogenic drivers that occur across solid tumor types. To understand the landscape of NRG1 gene fusions, 4397 solid tumor formalin-fixed, paraffin-embedded samples consecutively tested by comprehensive genomic and immune profiling during standard care were analyzed. Nineteen NRG1 fusions were found in 17 unique patients, across multiple tumor types, including non-small-cell lung (n=7), breast (n=2), colorectal (n=3), esophageal (n=2), ovarian (n=1), pancreatic (n=1), and unknown primary (n=1) carcinomas, with a cumulative incidence of 0.38%. Fusions were identified with breakpoints across four NRG1 introns spanning 1.4 megabases, with a mixture of known (n=8) and previously unreported (n=11) fusion partners. Co-occurring driver alterations in tumors with NRG1 fusions were uncommon, except colorectal carcinoma, where concurrent alterations in APC, BRAF, and ERBB2 were present in a subset of cases. The overall lack of co-occurring drivers highlights the importance of identifying NRG1 gene fusions, as these patients are unlikely to harbor other targetable alterations. In addition, RNA sequencing is important to identify NRG1 gene fusions given the variety of fusion partners and large genomic areas where breakpoints can occur.

Genetic labeling reveals spatial and cellular expression pattern of neuregulin 1 in mouse brain

BackgroundWhere the gene is expressed determines the function of the gene. Neuregulin 1 (Nrg1) encodes a tropic factor and is genetically linked with several neuropsychiatry diseases such as schizophrenia, bipolar disorder and depression. Nrg1 has broad functions ranging from regulating neurodevelopment to neurotransmission in the nervous system. However, the expression pattern of Nrg1 at the cellular and circuit levels in rodent brain is not full addressed.MethodsHere we used CRISPR/Cas9 techniques to generate a knockin mouse line (Nrg1Cre/+) that expresses a P2A-Cre cassette right before the stop codon of Nrg1 gene. Since Cre recombinase and Nrg1 are expressed in the same types of cells in Nrg1Cre/+ mice, the Nrg1 expression pattern can be revealed through the Cre-reporting mice or adeno-associated virus (AAV) that express fluorescent proteins in a Cre-dependent way. Using unbiased stereology and fluorescence imaging, the cellular expression pattern of Nrg1 and axon projections of Nrg1-positive neurons were investigated.ResultsIn the olfactory bulb (OB), Nrg1 is expressed in GABAergic interneurons including periglomerular (PG) and granule cells. In the cerebral cortex, Nrg1 is mainly expressed in the pyramidal neurons of superficial layers that mediate intercortical communications. In the striatum, Nrg1 is highly expressed in the Drd1-positive medium spiny neurons (MSNs) in the shell of nucleus accumbens (NAc) that project to substantia nigra pars reticulata (SNr). In the hippocampus, Nrg1 is mainly expressed in granule neurons in the dentate gyrus and pyramidal neurons in the subiculum. The Nrg1-expressing neurons in the subiculum project to retrosplenial granular cortex (RSG) and mammillary nucleus (MM). Nrg1 is highly expressed in the median eminence (ME) of hypothalamus and Purkinje cells in the cerebellum.ConclusionsNrg1 is broadly expressed in mouse brain, mainly in neurons, but has unique expression patterns in different brain regions.

Neuregulin-1 reduces vascular leak and airway epithelium permeability caused by respiratory viral infection

Abstract Respiratory viruses, such as Respiratory Syncytial Virus (RSV), are associated with significant morbidity and mortality. Mice with pre-existing atopy are protected from mortality to Sendai virus (SeV), a rodent virus related to RSV. We found that lungs of atopic mice had increased levels of neuregulin-1(NRG1), and hypothesized that SeV induces death via lung vascular leak, and that NRG1 might inhibit vascular leakage. Non-atopic C57Bl/6 mice were treated with NRG1 (500ng/30 mcl, i.n) or PBS for five days before infection with SeV (2×10 5pfu) or UV-inactivated SeV (UV-SeV) and 8 days post viral inoculation, Evan’s Blue Dye (EBD) was injected i.v with EBD concentration determined spectrophotometrically one hour later in homogenized lungs. Vascular leak in SeV-infected mice was 65.25% and 46.62% more than in UV-SeV-treated or SeV-infected mice pre-treated with NRG1, respectively. We also found that RSV infection of well-differentiated human bronchial epithelial cells (hBEC) grown at the air-liquid interface caused 70KDa FITC-Dextran leakage 8 days post-inoculation. NRG1 pretreatment for 5 days before RSV inoculation reduced epithelial leakage 8 days post-inoculation by 2.5 fold (p&amp;lt;0.001). These results suggest NRG1 has the potential to improve epithelial barrier function in rodent and human systems and may explain the increased survival following a severe respiratory viral infection. Abigail Wexner Research Institute at Nationwide Children’s Hospital, Start-up fund to MHG

Influence of functional-assembled-FGL polypeptide timbering material on expression of glial cell line-derived neurotropic growth factor mRNA in damaged rat’s spinal cord

Spinal cord injury is one of most serious complications when there is spine injury in patients. Spinal cord injury clinically involves anesthesia and mobility loss in leg in most of patients. Rats were herein molded and divided into groups, and Basso-Beattie-Bresnahan (BBB) method was adopted to assess the motor function of rats. The quantities of cassyl aspartic acid-specified protease (Caspase-3), neurofilament protein-200 (NF-200) and Glial Fibrillary Acidic Protein (GFAP)-positive cell were detected by immunohistochemical staining method. The expression of Glial cell line-derived neurotropic growth factor (GDNF) mRNA in spinal cord tissue was detected by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) method. The quantity of Caspase-3 positive cell and area of glial scar in the fiber grating laser-National Security (FGL-NS) group were significantly reduced compared with those from the other two groups. The quantities of NF-200 positive cell, graded result of Basso-Beattie-Bresnahan method and content of GDNF mRNA in the FGL-NS group were increased compared to those from the other two groups. The FGL-NS improved the expression of GDNF mRNA so as to repair the damaged neuronal cells.

ErbB3 Control of Pmp22 Regulates Intestinal Barrier Function

Background: ErbB3 is a member of the ErbB/EGFR family of receptor tyrosine kinases and is the constitutively-expressed neuregulin (NRG) receptor in the intestinal epithelium. Pathways downstream of ErbB3 have been implicated in regulating tight junctions in other systems, but a role for ErbB3 in barrier function in the intestines has not been described. Here we tested the hypothesis that ErbB3 is required for maintenance of normal barrier function in the intestine. Methods: We generated Villin-Cre;ErbB3 flox/flox mice (ErbB3-IEKO) with ErbB3 deletion in the intestinal epithelium. All comparisons are to ErbB3 flox/flox littermate controls. Mice were given a FITC-dextran 4 kDa (FD4) gavage to assess intestinal permeability. Isolated epithelial ileal samples were collected for RT-qPCR, immunohistochemistry, and bulk RNA-seq analysis. Enteroids were generated and treated with the ErbB3 ligand NRG-1β as well as PD153035 and CP-724714 to inhibit EGFR and ErbB2, respectively. Enteroids were also fixed and stained for whole mount immunofluorescence staining. Caco-2BBe cells were grown on Transwell filters, treated with NRG-1β, and assessed for transepithelial electrical resistance (TEER) and FD4 flux. Results: ErbB3-IEKO mice displayed increased intestinal barrier permeability to FD4 compared to controls (p &lt; 0.01). RNA-seq analysis showed a decrease in the expression of the putative tight junction protein Pmp22 in ErbB3-IEKO mice, while other tight junctional components were unchanged. RT-qPCR analysis of ileal samples showed an 80% Pmp22 downregulation (p &lt; 0.01) with ErbB3 loss compared to controls, with similar results in ErbB3-deficient enteroids. Immunohistochemical analysis confirmed near complete loss of PMP22 in ErbB3-IEKO intestinal epithelium. In vitro, treatment with NRG-1β induced Pmp22 expression (p &lt; 0.001) in control, but not ErbB3-deficient, enteroids. Treatment with PD153035 blocked NRG-1β-driven expression of Pmp22 while treatment with CP-724714 did not, suggesting that EGFR is a critical ErbB3 heterodimer partner in regulating Pmp22. Whole mount staining of enteroids confirmed co-localization of PMP22 with the tight junction protein ZO-1 at the apical junctions. Furthermore, treatment of Caco-2BBe monolayers with NRG-1β increased TEER and reduced FD4 flux, demonstrating active regulation of permeability. Conclusions: We demonstrate that loss of ErbB3 in the intestinal epithelium leads to reduced expression of the tight junction protein PMP22 and impaired barrier function in mice. Co-localization of PMP22 with ZO-1 suggests a role for PMP22 in forming the barrier and may represent a mechanism for regulation of intestinal permeability through the ErbB3-PMP22 axis. Furthermore, increased TEER in Caco-2BBe monolayers following NRG-1β treatment suggests induction of PMP22 may represent a means to enhance barrier function and a potential target for future therapeutic interventions targeting the barrier. Supported by NIH awards R01DK095004 and F31DK131856 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Gestational 1-nitropyrene exposure causes anxiety-like behavior partially by altering hippocampal epigenetic reprogramming of synaptic plasticity in male adult offspring

1-Nitropyrene (1-NP), a typical nitro-polycyclic aromatic hydrocarbon, is a developmental toxicant. This study was to evaluate gestational 1-NP-induced anxiety-like behavior in male adult offspring. Pregnant mice were orally administered to 1-NP daily throughout pregnancy. Anxiety-like behaviors, as determined by Elevated Plus-Maze (EPM) and Open-Field Test (OFT), were showed in male adult offspring whose mothers were exposed to 1-NP. Gestational 1-NP exposure reduced dendritic arborization, dendritic length and dendritic spine density in ventral hippocampus of male adult offspring. Additional experiments showed that gephyrin, an inhibitory synaptic marker, was reduced in fetal forebrain and hippocampus in male adult offspring. Nrg1 and Erbb4, two gephyrin-related genes, were reduced in 1-NP-exposed fetuses. Accordingly, 5hmC contents in two CpG sites (32008909 and 32009239) of Nrg1 gene and three CpG sites (69107743, 69107866 and 69107899) of Erbb4 gene were decreased in 1-NP-exposed fetuses. Mechanistically, ten-eleven translocation (TET) activity and alpha-ketoglutarate (α-KG) content were decreased in 1-NP-exposed fetal forebrain. Supplementation with α-KG alleviated 1-NP-induced downregulation of gephyrin-related genes, prevented hippocampal synaptic damage, and improved anxiety-like behavior in male adult offspring. These results indicate that early-life 1-NP exposure causes anxiety-like behavior in male adulthood partially by altering hippocampal epigenetic reprogramming of synaptic plasticity.

Abstract CT229: CRESTONE: A Phase 2 study of seribantumab in adult patients with neuregulin-1 (NRG1) fusion positive locally advanced or metastatic solid tumors

Abstract Background: NRG1 fusions represent a rare and potentially actionable oncogenic alteration found in ~0.2% of solid tumors. Seribantumab is a fully human anti-HER3 IgG2 monoclonal antibody that blocks aberrant NRG1 fusion protein binding to HER3 thereby preventing ligand-dependent activation and dimerization resulting in complete inhibition of the PI3K/AKT and MAPK downstream signaling pathways. Seribantumab has shown significant anti-tumor activity in PDX models and in patients (pts) with solid tumors harboring NRG1 fusions. Methods: CRESTONE (NCT04383210) is a phase 2 study of seribantumab in adult pts with solid tumors harboring NRG1 fusions who received at least one prior therapy and were naïve to ERBB-targeted therapy (Cohort 1); pts previously treated with ERBB-targeted therapies (exploratory Cohort 2) and pts with tumors harboring additional molecular alterations (exploratory Cohort 3). Here, we present updated efficacy results for pts in Cohort 1 dosed at 3 g IV QW. Safety data are evaluated for pts enrolled in Cohort 1 and exploratory Cohorts 2 and 3. Results: As of Dec 2, 2022, 51 pts were enrolled across all cohorts with 30 pts in Cohort 1 dosed at 3 g IV QW. Median age was 62 years (range 19-84); median prior lines of therapy was 2 (range 1-7); tumor types included non-small cell lung cancer (NSCLC, n=30), pancreatic adenocarcinoma (PDAC, n=6), biliary tract/cholangiocarcinoma (CCA, n=6), breast (n=4), and others (n=5); 15 different NRG1 fusion partners were identified with CD74 (22%) and SLC3A2 (16%) as the most frequently reported. In the overall safety population, 41 pts (80%) reported at least one treatment-related adverse event (TRAE). The most common TRAEs (occurring in ≥20% of pts) were diarrhea (41%), fatigue (29%), rash (24%), and nausea (22%). Four pts (8%) experienced Gr 3/4 TRAEs; no Gr 5 TRAEs. The safety profile for Cohort 1 was similar to the overall safety population. Among the 30 pts in Cohort 1, 22 were evaluable for investigator assessed (INV) response per RECIST v1.1; 2 pts (9%) had confirmed complete response (CR), 6 pts (27%) had confirmed partial response (PR), and 13 pts (59%) had stable disease (SD) as their best overall response (BOR). The INV objective response rate (ORR, confirmed PR + CR) was 36% and disease control rate (DCR, confirmed PR+CR+SD) was 95%. The overall duration of response ranged from 1.4 to 17.2 months. In pts with NSCLC, the INV-ORR was 39% and DCR was 94%. Conclusions: Seribantumab has an acceptable safety and tolerability profile as a single agent in pts with solid tumors harboring NRG1 fusions. Updated efficacy data indicate seribantumab has robust and durable clinical activity, including CRs, across different tumor types harboring an NRG1 fusion and is a promising treatment option. Citation Format: Tejas Patil, Daniel R. Carrizosa, Mark E. Burkard, Karen L. Reckamp, Jayesh Desai, Young Kwang Chae, Stephen V. Liu, Kartik Konduri, Shirish M. Gadgeel, Jessica J. Lin, Parneet K. Cheema, David R. Spigel, Alison M. Schram, Valerie M. Jansen, Yasir Y. Elamin. CRESTONE: A Phase 2 study of seribantumab in adult patients with neuregulin-1 (NRG1) fusion positive locally advanced or metastatic solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT229.

Abstract 3636: Liver endothelium secreted LRG1 promotes metastatic colorectal cancer growth through the HER3/RSK/EIF4B AXIS

Abstract Background: 25% of patients diagnosed with colorectal cancer (CRC) have liver metastasis at presentation, and ~80% of all metastatic CRC are developed in the liver. We previously reported that liver endothelial cells (ECs), a key component of the liver microenvironment, secrete LRG1 to promote CRC growth via activating human epidermal growth factor receptor (ERbB3, also known as HER3). However, we found that LRG1-induced HER3 activation is distinct from the canonical neuregulin 1 (NRG1)-induced HER3 pathway. The present study further validated LRG1 as a new HER3 ligand for promoting mCRC growth and elucidated the novel downstream signaling pathway induced by LRG1-HER3. Methods: We first measured the binding affinity between HER3 and LRG1 by Biolayer interferometry (BLI). We then used in vitro and in vivo xenograft approaches to determine the effect of LRG1 monoclonal antibody (15C4) on HER3 activation and CRC growth. To further determine the role of LRG1 in promoting CRC growth in the liver, we used murine CRC cells in a syngeneic orthotropic liver injection model to establish CRC allografts in the liver of LRG1−/− mice with systemic LRG1 knockout and wild-type siblings (LRG1+/+). We also performed unbiased phospho-MS analysis and subsequent validations to determine the downstream signaling pathway activated by LRG1-HER3. Results: We identified that LRG1 binds to HER3 with the affinity at ~100nM. The LRG1 antibody 15C4 completely attenuated LRG1-induced HER3 activation and in vitro and xenograft growth in vivo. Moreover, LRG1−/− mice with CRC allografts in the liver had 2 times longer overall survival than tumor-bearing LRG1+/+ mice. Lastly, unbiased -omics analysis identified eIF4-protein synthesis is significantly activated by LRG1. With target-specific inhibitors, we further determined that LRG1-HER3 activates the PI3K-PDK1-RSK1/3-eIF4 axis independent of AKT. Conclusions: We identified LRG1 as a novel HER3 ligand and demonstrated that the liver microenvironment-derived LRG1 plays a key oncogenic role in mCRC, by activating a novel RSK-eIF4 survival pathway. Our findings highlighted the potential of blocking LRG1-HER3 and involved downstream pathways for treating patients with mCRC. Citation Format: Moeez Ghani Rathore, Michelle Wright, Wei Huang, Derek Taylor, Yamu Li, Jordan Winter, Zhenghe Wang, John Greenwood, Stephen Moss, Rui Wang. Liver endothelium secreted LRG1 promotes metastatic colorectal cancer growth through the HER3/RSK/EIF4B AXIS. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3636.

STAT5b is a key effector of NRG‐1/ERBB4‐mediated myocardial growth

The growth factor Neuregulin‐1 (NRG‐1) regulates myocardial growth and is currently under clinical investigation as a treatment for heart failure. Here, we demonstrate in several in vitro and in vivo models that STAT5b mediates NRG‐1/EBBB4‐stimulated cardiomyocyte growth. Genetic and chemical disruption of the NRG‐1/ERBB4 pathway reduces STAT5b activation and transcription of STAT5b target genes Igf1, Myc, and Cdkn1a in murine cardiomyocytes. Loss of Stat5b also ablates NRG‐1‐induced cardiomyocyte hypertrophy. Dynamin‐2 is shown to control the cell surface localization of ERBB4 and chemical inhibition of Dynamin‐2 downregulates STAT5b activation and cardiomyocyte hypertrophy. In zebrafish embryos, Stat5 is activated during NRG‐1‐induced hyperplastic myocardial growth, and chemical inhibition of the Nrg‐1/Erbb4 pathway or Dynamin‐2 leads to loss of myocardial growth and Stat5 activation. Moreover, CRISPR/Cas9‐mediated knockdown of stat5b results in reduced myocardial growth and cardiac function. Finally, the NRG‐1/ERBB4/STAT5b signaling pathway is differentially regulated at mRNA and protein levels in the myocardium of patients with pathological cardiac hypertrophy as compared to control human subjects, consistent with a role of the NRG‐1/ERBB4/STAT5b pathway in myocardial growth.

Fibroblast-derived EGF ligand neuregulin 1 induces fetal-like reprogramming of the intestinal epithelium without supporting tumorigenic growth.

Growth factors secreted by stromal fibroblasts regulate the intestinal epithelium. Stroma-derived epidermal growth factor (EGF) family ligands are implicated in epithelial regeneration and tumorigenesis, but their specific contributions and associated mechanisms remain unclear. Here, we use primary intestinal organoids modeling homeostatic, injured and tumorigenic epithelia to assess how the fibroblast-derived EGF family ligands neuregulin 1 (NRG1) and epiregulin (EREG) regulate the intestinal epithelium. NRG1 was expressed exclusively in the stroma, robustly increased crypt budding and protected intestinal epithelial organoids from radiation-induced damage. NRG1 also induced regenerative features in the epithelium, including a fetal-like transcriptome, suppression of the Lgr5+ stem cell pool and remodeling of the epithelial actin cytoskeleton. Intriguingly, unlike EGF and EREG, NRG1 failed to support the growth of pre-tumorigenic intestinal organoids lacking the tumor suppressor Apc, commonly mutated in human colorectal cancer (CRC). Interestingly, high expression of stromal NRG1 was associated with improved survival in CRC cohorts, suggesting a tumor-suppressive function. Our results highlight the power of stromal NRG1 in transcriptional reprogramming and protection of the intestinal epithelium from radiation injury without promoting tumorigenesis.

Rats, Neuregulins and Radical Prostatectomy: A Conceptual Overview

In the contemporary era of early detection, with mostly curative initial treatment for prostate cancer (PC), mortality rates have significantly diminished. In addition, mean age at initial PC diagnosis has decreased. Despite technical advancements, the probability of erectile function (EF) recovery post radical prostatectomy (RP) has not significantly changed throughout the last decade. Due to virtually unavoidable intraoperative cavernous nerve (CN) lesions and operations with younger patients, post-RP erectile dysfunction (ED) has now begun affecting these younger patients. To address this pervasive limitation, a plethora of CN lesion animal model investigations have analyzed the use of systemic/local treatments for EF recovery post-RP. Most promisingly, neuregulins (NRGs) have demonstrated neurotrophic effects in both neurodegenerative disease and peripheral nerve injury models. Recently, glial growth factor 2 (GGF2) has demonstrated far superior, dose-dependent, neuroprotective/restorative effects in the CN injury rat model, as compared to previous therapeutic counterparts. Although potentially impactful, these initial findings remain limited and under-investigated. In an effort to aid clinicians, our paper reviews post-RP ED pathogenesis and currently available therapeutic tools. To stimulate further experimentation, a standardized preparation protocol and in-depth analysis of applications for the CN injury rat model is provided. Lastly, we report on NRGs, such as GGF2, and their potentially revolutionary clinical applications, in hopes of identifying relevant future research directions.

Correlation of rs35753505 polymorphism in Neuregulin 1 gene with psychopathology and intelligence of people with schizophrenia

Background and aimSchizophrenia is one of the most severe psychiatric disorders. About 0.5 to 1% of the world's population suffers from this non-Mendelian disorder. Environmental and genetic factors seem to be involved in this disorder. In this article, we investigate the alleles and genotypic correlation of mononucleotide rs35753505 polymorphism of Neuregulin 1 (NRG1), one of the selected genes of schizophrenia, with psychopathology and intelligence. Materials and methods102 independent and 98 healthy patients participated in this study. DNA was extracted by the salting out method and the polymorphism (rs35753505) were amplified by polymerase chain reaction (PCR). Sanger sequencing was performed on PCR products. Allele frequency analysis was performed using COCAPHASE software, and genotype analysis was performed using Clump22 software. ResultsAccording to our study's statistical findings, all case samples from the three categories of men, women, and overall participants significantly differed from the control group in terms of the prevalence of allele C and the CC risk genotype. The rs35753505 polymorphism significantly raised Positive and Negative Syndrome Scale (PANSS) test results, according to a correlation analysis between the two variables. However, this polymorphism led to a significant decrease in overall intelligence in case samples compared to control samples. ConclusionIn this study, it seems that the rs35753505 polymorphism of NRG1 gene has a significant role in the sample of patients with schizophrenia in Iran and also in psychopathology and intelligence disorders.

Study on inhibitory effect of GC-miR-143 inhibitor nanoparticles on D2 receptor-mediated neurological behavioral damage in lacunar infarction

Blocking the dopamine D2 receptor is a key link in anti-psychiatric disease. Knockdown of miR-143 improved neurological behavioral damage by regulating the D2 receptor signaling pathway. The risk of neurological behavior damage can be increased 20 times by 1–2 lacunar infarction lesions. Based on the fact that miR-143 can target and regulate the D2 receptor signaling pathway, this study focused on analyzing the inhibitory effect of GC-miR-143 inhibitor nanoparticles on neurological behavior damage in lacunar infarction. 60 SD rats were separated into sham operation, model, vehicle, vehicle+inhibitor, inhibitor, or D2 agonist group (n = 10). The vascular dementia (VD) model was made by ligating bilateral common carotid arteries. Morris water maze assessed rats behavioral changes. Quantitative Real-Time Reverse Transcription PCR (QRT-PCR) was applied to detect levels of miRNA-143. Proteins were extracted from the prefrontal cortex, and immunoblotting was performed to measure the downstream of signal proteins of Neuregulin 1 (NRG1) and D2 receptor signaling pathways, p-protein kinase B (AKT) and p-Glycogen Synthase Kinase 3β (GSK3β). On days 1 to 5 of the water maze experiment, the escape latency for the five groups of rats were longer than in sham group; while escape latency for vehicle+inhibitor group and inhibitor group was shorter. Compared to VD model rats, vehicle+inhibitor group and inhibitor group had significantly reduced escape latency. Cross-platform time number for five groups was decreased and it was increased in vehicle+inhibitor group and inhibitor group with decreased number for D2 agonist group w. Compared with D2 agonist group, the vehicle+inhibitor group and inhibitor group showed more cross-platform times. Compared to sham operation group, levels of NRG1, p-AKT/GSK3β in remaining five groups were significantly decreased. Their levels in vector+inhibition group and inhibitor group were dramatically up-regulated, and levels of the above-mentioned proteins in D2 agonist group were significantly decreased. Their levels in vector+inhibitor group and inhibitor group were up-regulated relative to D2 agonist group. The GC-miR-143 inhibitor nanoparticles can up-regulate the expression of NRG1 and antagonize the D2 receptor to weaken the inhibitory effect of D2 receptor-mediated signaling pathways and inhibit neurological behavioral damage in lacunar cerebral infarction.

Dual-targeting therapy against HER3/MET in human colorectal cancers.

Colorectal cancer (CRC) is the most common malignancy in the world, and novel molecular targeted therapies for CRC have been vigorously pursued. We searched for novel combination therapies based on the expression patterns of membrane proteins in CRC cell lines. A positive correlation was observed between the expression of human pidermal growth factor receptor (HER) 3 and mesenchymal-to-epithelial transition factor (MET) on the cell surface of CRC cell lines. The brief stimulation of HER3/MET-high SW1116 CRC cells with both neuregulin-1 (NRG1) and hepatocyte growth factor enhanced ERK phosphorylation and cell proliferation more than each stimulation alone. In addition, a prolonged NRG1 stimulation resulted in the tyrosine phosphorylation of MET. In this context, the Forkhead Box protein M1 (FOXM1)-regulated tyrosine phosphorylation of MET by NRG1 was demonstrated, suggesting the existence of a signaling pathway mediated by FOXM1 upon the NRG1 stimulation. Since the co-expression of HER3 and MET was also demonstrated in in vivo CRC tissues by immunohistochemistry, we investigated whether the co-inhibition of HER3 and MET could be an effective therapy for CRC. We established HER3-and/or MET-KO SW1116 cell lines, and HER3/MET-double KO resulted in the inhibition of in vitro cell proliferation and in vivo tumor growth in nude mice by SW1116 cells. Furthermore, the combination of patritumab, an anti-HER3 fully human mAb, and PHA665752, a MET inhibitor, markedly inhibited in vitro cell proliferation, 3D-colony formation, and in vivo tumor growth in nude mice by SW1116 cells CONCLUSION: The dual targeting of HER3/MET has potential as CRC therapy.

ER-PM Junctions on GABAergic Interneurons Are Organized by Neuregulin 2/VAP Interactions and Regulated by NMDA Receptors.

Neuregulins (NRGs) signal via ErbB receptors to regulate neural development, excitability, synaptic and network activity, and behaviors relevant to psychiatric disorders. Bidirectional signaling between NRG2/ErbB4 and NMDA receptors is thought to homeostatically regulate GABAergic interneurons in response to increased excitatory neurotransmission or elevated extracellular glutamate levels. Unprocessed proNRG2 forms discrete clusters on cell bodies and proximal dendrites that colocalize with the potassium channel Kv2.1 at specialized endoplasmic reticulum-plasma membrane (ER-PM) junctions, and NMDA receptor activation triggers rapid dissociation from ER-PM junctions and ectodomain shedding by ADAM10. Here, we elucidate the mechanistic basis of proNRG2 clustering at ER-PM junctions and its regulation by NMDA receptors. Importantly, we demonstrate that proNRG2 promotes the formation of ER-PM junctions by directly binding the ER-resident membrane tether VAP, like Kv2.1. The proNRG2 intracellular domain harbors two non-canonical, low-affinity sites that cooperatively mediate VAP binding. One of these is a cryptic and phosphorylation-dependent VAP binding motif that is dephosphorylated following NMDA receptor activation, thus revealing how excitatory neurotransmission promotes the dissociation of proNRG2 from ER-PM junctions. Therefore, proNRG2 and Kv2.1 can independently function as VAP-dependent organizers of neuronal ER-PM junctions. Based on these and prior studies, we propose that proNRG2 and Kv2.1 serve as co-regulated downstream effectors of NMDA receptors to homeostatically regulate GABAergic interneurons.

Neuregulin-1/PI3K signaling effects on oligodendrocyte proliferation, remyelination and behaviors deficit in a male mouse model of ischemic stroke

In this study, we investigated the effect of neuregulin-1 (NRG1) on demyelination and neurological function in an ischemic stroke model, and further explored its neuroprotective mechanisms. Adult male ICR mice underwent photothrombotic ischemia surgery and were injected with NRG1 beginning 30 min after ischemia. Cylinder and grid walking tests were performed to evaluate the forepaw function. In addition, the effect of NRG1 on neuronal damage/death (Cresyl violet, CV), neuronal nuclei (NeuN), nestin, doublecortin (DCX), myelin basic protein (MBP), non-phosphorylated neurofilaments (SMI-32), adenomatous polyposis coli (APC), erythroblastic leukemia viral oncogene homolog (ErbB) 2, 4 and serine-threonine protein kinase (Akt) in cortex were evaluated using immunohistochemistry, immunofluorescence and western blot. The cylinder and grid walking tests exposed that treatment of NRG1 observably regained the forepaw function. NRG1 treatment reduced cerebral infarction, restored forepaw function, promoted proliferation and differentiation of neuron and increased oligodendrogliogenesis. The neuroprotective effect of NRG1 is involved in its activation of PI3K/Akt signaling pathway via ErbB2, as shown by the suppression of the effect of NRG1 by the PI3K inhibitor LY294002. Our results demonstrate that NRG1 is effective in ameliorating the both acute phase neuroprotection and long-term neurological functions via resumption of neuronal proliferation and differentiation and oligodendrogliogenesis in a male mouse model of ischemic stroke.