Neural Cell Adhesion Molecule Research Articles

The existence of cells exhibiting characteristics of both Type II and Type III cells in rat taste buds. An immunohistochemical and electron-microscopic study.

Taste bud cells are classified into four types by their ultrastructural features. Immunohistochemical detection of taste-signaling molecules is used to distinguish cell types of taste bud cells; however, the characteristics of taste cell types such as the immunoreactivity for taste-signaling molecules have long remained unclear. We investigated the detailed characteristics of taste cells in rat vallate taste buds by electron microscopy and immunohistochemistry for gustducin, neural cell adhesion molecule (NCAM) and vesicle-associated membrane protein 2 (VAMP2), which are known as markers of Type II cells, Type III cells and both cell types, respectively. Triple immunostaining for these molecules discriminated seven kinds of cell, including the totally immunopositive cell. Electron microscopy revealed Type III cells with a typical synaptic structure and subsurface cisterna as a specialized contact between a nerve and a Type II cell. The present study clarified the existence of cells with features of both Type II and TypeIII cells as a subtype of taste bud cells in the rat taste bud.

Neuropilin2 in Mesenchymal Stromal Cells as a Potential Novel Therapeutic Target in Myelofibrosis.

Bone marrow fibrosis in myeloproliferative neoplasm (MPN), myelodysplastic syndromes (MDS), MPN/MDS overlap syndromes and acute myeloid leukemia (AML) is associated with poor prognosis and early treatment failure. Myelofibrosis (MF) is accompanied by reprogramming of multipotent bone marrow mesenchymal stromal cells (MSC) into osteoid and fiber-producing stromal cells. We demonstrate NRP2 and osteolineage marker NCAM1 (neural cell adhesion molecule 1) expression within the endosteal niche in normal bone marrow and aberrantly in MPN, MDS MPN/MDS overlap syndromes and AML (n = 99), as assessed by immunohistochemistry. Increased and diffuse expression in mesenchymal stromal cells and osteoblasts correlates with high MF grade in MPN (p < 0.05 for NRP2 and NCAM1). Single cell RNA sequencing (scRNAseq) re-analysis demonstrated NRP2 expression in endothelial cells and partial co-expression of NRP2 and NCAM1 in normal MSC and osteoblasts. Potential ligands included transforming growth factor β1 (TGFB1) from osteoblasts and megakaryocytes. Murine ThPO and JAK2V617F myelofibrosis models showed co-expression of Nrp2 and Ncam1 in osteolineage cells, while fibrosis-promoting MSC only express Nrp2. In vitro experiments with MC3T3-E1 pre-osteoblasts and analysis of Nrp2-/- mouse femurs suggest that Nrp2 is functionally involved in osteogenesis. In summary, NRP2 represents a potential novel druggable target in patients with myelofibrosis.

Extremely low-frequency electromagnetic field (ELF-EMF) induced alterations in gene expression and cytokine secretion in clear cell renal carcinoma cells.

The study aimed to investigate the influence of extremely low-frequency electromagnetic fields (ELF-EMF) on clear cell renal cell carcinoma (ccRCC) by assessing alterations in gene expression and the secretion of cytokines and chemokines. Three ccRCC cell lines (786-O, 769-P, and CAKI-1) and a healthy HEK293 cell line were subjected to ELF-EMF exposure (frequency 50 Hz, magnetic field strength 4.5 mT) for 30 min daily for 5 days. The study examined the expression of ADAM28, NCAM1, and VEGFC genes, along with the secretion of 30 cytokines and chemokines. Notably, primary tumor-derived cell lines, but not those from metastatic sites, exhibited ADAM28 gene expression, which increased following ELF-EMF exposure. A statistically significant reduction in VEGFC gene expression was observed in 769-P cells after ELF-EMF exposure. Additionally, NCAM1 gene expression was upregulated in HEK293, 769-P, and 786-O cells, representing normal embryonic kidney cells and primary tumor cells, but not in CAKI-1 cells, which model metastatic sites. After EMF exposure, there was a statistically significant decrease in transforming growth factor β1 (TGF-β1) concentration in the cell culture supernatants of HEK293 and CAKI-1 cell lines, with no other significant changes in the secretion of tested cytokines. Given the study's findings and available research, caution is warranted when drawing conclusions about the potential inhibitory effect of ELF-EMF on ccRCC progression. Standardization of experimental models is imperative when assessing the effects of EMF in a human context. Med Pr Work Health Saf. 2024;75(2):133-141.

The dynamic TRPV2 ion channel proximity proteome reveals functional links of calcium flux with cellular adhesion factors NCAM and L1CAM in neurite outgrowth

TRPV2 voltage-insensitive, calcium-permeable ion channels play important roles in cancer progression, immune response, and neuronal development. Despite TRPV2’s physiological impact, underlying endogenous proteins mediating TRPV2 responses and affected signaling pathways remain elusive. Using quantitative peroxidase-catalyzed (APEX2) proximity proteomics we uncover dynamic changes in the TRPV2-proximal proteome and identify calcium signaling and cell adhesion factors recruited to the molecular channel neighborhood in response to activation.Quantitative TRPV2 proximity proteomics further revealed activation-induced enrichment of protein clusters with biological functions in neural and cellular projection. We demonstrate a functional connection between TRPV2 and the neural immunoglobulin cell adhesion molecules NCAM and L1CAM. NCAM and L1CAM stimulation robustly induces TRPV2 [Ca2+]I flux in neuronal PC12 cells and this TRPV2-specific [Ca2+]I flux requires activation of the protein kinase PKCα. TRPV2 expression directly impacts neurite lengths that are modulated by NCAM or L1CAM stimulation. Hence, TRPV2’s calcium signaling plays a previously undescribed, yet vital role in cell adhesion, and TRPV2 calcium flux and neurite development are intricately linked via NCAM and L1CAM cell adhesion proteins.

Abstract PS02-05: Trajectories and predictors of peripheral neuropathy after neoadjuvant chemotherapy in a prospective cohort of 11,014 patients with early breast cancer

Abstract Background Peripheral neuropathy (PN) is a debilitating adverse event in patients with early breast cancer (BC) receiving (neo)adjuvant chemotherapy (CT). We harnessed the CANTO cohort study to detail clinical trajectories and explore clinical and genetic predictors of PN. Methods CANTO (CANcer TOxicities - NCT01993498) prospectively enrolled invasive stage I-III BC patients (pts) of 26 French comprehensive cancer centers. Pts were assessed at diagnosis, 3-6 (M6), 12 (M12), 36 (M36), and 60 (M60) months after treatment, defined as completion of surgery, CT or radiotherapy, whichever comes last. At each time point, PN events including paresthesia, sensory and motor neuropathy were collected according to NCI-CTC v4.0 criteria. A genome-wide association study (GWAS) was conducted to identify genetic predictors of PN, using Illumina GSA BeadChips. Minimac4/1000G was used to impute additional single nucleotide polymorphisms (SNPs). After stringent quality control measures, 1,894,475 SNPs with a minor allele frequency (MAF) &amp;gt;0.05 were evaluated. Longitudinal trajectories of PN events were descriptively examined. Statistical associations between each SNP and PN events measured at different time points, and between SNPs and trajectories, were performed with logistic regression assuming a log-additive genetic model. All analyses were adjusted for key clinical parameters and the first ten axes of principal component analysis of the genetic data to control for population stratification. Results Of 12,012 included pts (data lock Aug. 2022), 11,014 (91.7%) were analyzed. Age was &amp;lt; 50 and &amp;gt;65 in 3407 and 2759 pts (31% and 25%, respectively). Overweight/obesity and diabetes were recorded in 5328 (48%) and 458 pts (4%). A neurologic history was observed in 1360 pts (13%). Stage 0-I and II-III were observed in 5356 (49%) and 5512 pts (51%), and 4011 pts had an axillary dissection (37%). CT was administered in 5790 pts (53%), including a taxane (tax) in 5542 pts (96%). Shortly, all grade and grade 2+ PN was observed in 29% and 12%; 27% and 10%; 20% and 10%; 13% and 10% of pts at M6, M12, M36 and M60, respectively. We derived 5 trajectories of overall PN, deemed “never”, “always”, “appears”, “transient” and “disappears”, in respectively 54%, 10%, 9%, 7% and 19% of the pts. Similar trajectories were built for the 3 categories of PN events. We built 9 predictive nomograms including key clinical parameters and time of analysis. E.g., grade 2+ PN at M6 was independently predicted by a medical history of carpal tunnel syndrome, past neurologic history and exposure to tax. Grade 2+ PN at M36 was predicted by past neurologic history and exposure to tax, and largely by previous PN at M6 or M12. The GWAS analysis included 7633 pts (84%). Four independent SNPs adding predictive value to the clinical nomograms were identified at a suggestive level of association (p &amp;lt; 1e-6), including one SNP in the NCAM1 gene (involved in neurogenesis) for M6 PN (OR=1.40, 95%CI 1.23-1.60, p=4.8e-7, see table) and one SNP in the CLDN11 gene (regulates oligodendrocytes) for M12 sensory PN (OR=1.32; 95%CI 1.17-1.49, p=7.5e-6). For rare ( &amp;lt; 5%) toxicities, 10 independent suggestive SNPs were identified, including one SNP in the NELL1 gene (neural cell growth regulation and differentiation) for the “always” trajectory (OR=1.61, 95%CI 1.31-1.99, p=7.6e-6) and one SNP in the KCNIP1 gene (neuronal sensor) for M36 motor PN (OR=1.5, 95%CI 1.7-3.5, p=3.1e-07). Conclusions Risk of early peripheral neuropathy is associated with previous neurological history and taxane exposure. Patients with PN at M6 or M12 are highly exposed to long-term PN. Some key SNPs may add independent predictive value for specific PN endpoints. Exploratory results of interest were produced for rarer toxicities and typical clinical trajectories. Detailed results will be presented at the meeting. All PN, M6 Multivariate analyses of the risk of peripheral neuropathy, all grades, at M6, without and with GWAS input. Citation Format: Youenn Drouet, Emilie Thomas, Florence Lerebours, Barbara Pistilli, Olivier Trédan, Christelle Jouannaud, Marion Fournier, Philippe Rouanet, Laurence Vanlemmens, CHARLES COUTANT, Asma Dhaini Merimeche, Baptiste Sauterey, Christelle Levy, Mario Campone, Carole Tarpin, Marie-Ange Mouret-Reynier, Olivier Rigal, Thierry Petit, Sophie Guillermet, Antoine Arnaud, Mahmoud Ibrahim, Sylvie Giacchetti, Florence Dalenc, Johanna Wassermann, Olivier Arsène, Ariane Darut-Jouve, Sibille Everhard, Ines Vaz Luis, Anne-Laure Martin, Fabrice André, Jean-François Deleuze, Alain Viari, Matthieu Carton, Paul Cottu. Trajectories and predictors of peripheral neuropathy after neoadjuvant chemotherapy in a prospective cohort of 11,014 patients with early breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS02-05.

CNTN4 modulates neural elongation through interplay with APP.

The neuronal cell adhesion molecule contactin-4 (CNTN4) is genetically associated with autism spectrum disorder (ASD) and other psychiatric disorders. Cntn4-deficient mouse models have previously shown that CNTN4 plays important roles in axon guidance and synaptic plasticity in the hippocampus. However, the pathogenesis and functional role of CNTN4 in the cortex has not yet been investigated. Our study found a reduction in cortical thickness in the motor cortex of Cntn4 -/- mice, but cortical cell migration and differentiation were unaffected. Significant morphological changes were observed in neurons in the M1 region of the motor cortex, indicating that CNTN4 is also involved in the morphology and spine density of neurons in the motor cortex. Furthermore, mass spectrometry analysis identified an interaction partner for CNTN4, confirming an interaction between CNTN4 and amyloid-precursor protein (APP). Knockout human cells for CNTN4 and/or APP revealed a relationship between CNTN4 and APP. This study demonstrates that CNTN4 contributes to cortical development and that binding and interplay with APP controls neural elongation. This is an important finding for understanding the physiological function of APP, a key protein for Alzheimer's disease. The binding between CNTN4 and APP, which is involved in neurodevelopment, is essential for healthy nerve outgrowth.

Exploring the role of neuronal-enriched extracellular vesicle miR-93 and interoception in major depressive disorder

Major depressive disorder (MDD) is associated with interoceptive processing dysfunctions, but the molecular mechanisms underlying this dysfunction are poorly understood. This study combined brain neuronal-enriched extracellular vesicle (NEEV) technology and serum markers of inflammation and metabolism with Functional Magnetic Resonance Imaging (fMRI) to identify the contribution of gene regulatory pathways, in particular micro-RNA (miR) 93, to interoceptive dysfunction in MDD. Individuals with MDD (n = 41) and healthy comparisons (HC; n = 35) provided blood samples and completed an interoceptive attention task during fMRI. EVs were separated from plasma using a precipitation method. NEEVs were enriched by magnetic streptavidin bead immunocapture utilizing a neural adhesion marker (L1CAM/CD171) biotinylated antibody. The origin of NEEVs was validated with two other neuronal markers - neuronal cell adhesion molecule (NCAM) and ATPase Na+/K+ transporting subunit alpha 3 (ATP1A3). NEEV specificities were confirmed by flow cytometry, western blot, particle size analyzer, and transmission electron microscopy. NEEV small RNAs were purified and sequenced. Results showed that: (1) MDD exhibited lower NEEV miR-93 expression than HC; (2) within MDD but not HC, those individuals with the lowest NEEV miR-93 expression had the highest serum concentrations of interleukin (IL)-1 receptor antagonist, IL-6, tumor necrosis factor, and leptin; and (3) within HC but not MDD, those participants with the highest miR-93 expression showed the strongest bilateral dorsal mid-insula activation during interoceptive versus exteroceptive attention. Since miR-93 is regulated by stress and affects epigenetic modulation by chromatin re-organization, these results suggest that healthy individuals but not MDD participants show an adaptive epigenetic regulation of insular function during interoceptive processing. Future investigations will need to delineate how specific internal and external environmental conditions contribute to miR-93 expression in MDD and what molecular mechanisms alter brain responsivity to body-relevant signals.

The Bifunctional Effects of Lactoferrin (LFcinB11) in Inhibiting Neural Cell Adhesive Molecule (NCAM) Polysialylation and the Release of Neutrophil Extracellular Traps (NETs).

The expression of polysialic acid (polySia) on the neuronal cell adhesion molecule (NCAM) is called NCAM-polysialylation, which is strongly related to the migration and invasion of tumor cells and aggressive clinical status. Thus, it is important to select a proper drug to block tumor cell migration during clinical treatment. In this study, we proposed that lactoferrin (LFcinB11) may be a better candidate for inhibiting NCAM polysialylation when compared with CMP and low-molecular-weight heparin (LMWH), which were determined based on our NMR studies. Furthermore, neutrophil extracellular traps (NETs) represent the most dramatic stage in the cell death process, and the release of NETs is related to the pathogenesis of autoimmune and inflammatory disorders, with proposed involvement in glomerulonephritis, chronic lung disease, sepsis, and vascular disorders. In this study, the molecular mechanisms involved in the inhibition of NET release using LFcinB11 as an inhibitor were also determined. Based on these results, LFcinB11 is proposed as being a bifunctional inhibitor for inhibiting both NCAM polysialylation and the release of NETs.

Diagnostic Challenges and Emerging Pathogeneses of Selected Glomerulopathies.

Recent progress in glomerular immune complex and complement-mediated diseases have refined diagnostic categories and informed mechanistic understanding of disease development in pediatric patients. Herein, we discuss selected advances in 3 categories. First, membranous nephropathy antigens are increasingly utilized to characterize disease in pediatric patients and include phospholipase A2 receptor (PLA2R), Semaphorin 3B (Sema3B), neural epidermal growth factor-like 1 (NELL1), and protocadherin FAT1, as well as the lupus membranous-associated antigens exostosin 1/2 (EXT1/2), neural cell adhesion molecule 1 (NCAM1), and transforming growth factor beta receptor 3 (TGFBR3). Second, we examine advances in techniques for paraffin and light chain immunofluorescence (IF), including the former's function as a salvage technique and their necessity for diagnosis in adolescent cases of membranous-like glomerulopathy with masked IgG kappa deposits (MGMID) and proliferative glomerulonephritis with monotypic Ig deposits (PGNMID), respectively. Finally, progress in understanding the roles of complement in pediatric glomerular disease is reviewed, with specific attention to overlapping clinical, histologic, and genetic or functional alternative complement pathway (AP) abnormalities among C3 glomerulopathy (C3G), infection-related and post-infectious GN, "atypical" post-infectious GN, immune complex mediated membranoproliferative glomerulonephritis (IC-MPGN), and atypical hemolytic uremic syndrome (aHUS).

NrCAM-deficient mice exposed to chronic stress exhibit disrupted latent inhibition, a hallmark of schizophrenia.

The neuronal cell adhesion molecule (NrCAM) is widely expressed and has important physiological functions in the nervous system across the lifespan, from axonal growth and guidance to spine and synaptic pruning, to organization of proteins at the nodes of Ranvier. NrCAM lies at the core of a functional protein network where multiple targets (including NrCAM itself) have been associated with schizophrenia. Here we investigated the effects of chronic unpredictable stress on latent inhibition, a measure of selective attention and learning which shows alterations in schizophrenia, in NrCAM knockout (KO) mice and their wild-type littermate controls (WT). Under baseline experimental conditions both NrCAM KO and WT mice expressed robust latent inhibition (p = 0.001). However, following chronic unpredictable stress, WT mice (p = 0.002), but not NrCAM KO mice (F < 1), expressed latent inhibition. Analyses of neuronal activation (c-Fos positive counts) in key brain regions relevant to latent inhibition indicated four types of effects: a single hit by genotype in IL cortex (p = 0.0001), a single hit by stress in Acb-shell (p = 0.031), a dual hit stress x genotype in mOFC (p = 0.008), vOFC (p = 0.020), and Acb-core (p = 0.032), and no effect in PrL cortex (p > 0.141). These results indicating a pattern of differential effects of genotype and stress support a complex stress × genotype interaction model and a role for NrCAM in stress-induced pathological behaviors relevant to schizophrenia and other psychiatric disorders.

Abstract 6996: A role for the cell adhesion molecule NCAM1 in regulating activity of the receptor tyrosine kinase RET in cancer

Abstract RET (REarranged during Transfection) is a receptor tyrosine kinase with essential roles in early development; however, genetic alterations lead to aberrant RET activity that contributes to oncogenic processes in several cancers. While some of the mechanisms by which RET mediates these processes are known, the network of proteins facilitating its activity remains largely undiscovered. To address this, we performed a genome-wide synthetic dosage lethal screen which identified NCAM1 (Neural Cell Adhesion Molecule 1) as one potential candidate modulating RET activity. Using an NCAM1 knockdown (KD) in SH-SY5Y neuroblastoma cells, we show that loss of NCAM1 reduces total protein levels of RET with no effect on RET transcription. Despite reduced abundance of RET, we show that the mature form of RET has a prolonged half-life in the absence of NCAM1. Assessing the functional outcomes of reduced RET protein levels, we observe reduced activation of RET by its extracellular ligand GDNF (Glial cell line-derived neurotrophic factor) and a subsequent reduction in activation of proliferative signaling pathways downstream of RET. As a result, RET-mediated proliferation is diminished in the absence of NCAM1, with reduced viability of NCAM1 KD SH-SY5Y observed in response to GDNF. Together, our data suggest that the synthetic lethal relationship between RET and NCAM1 may arise from a role for NCAM1 in regulating total RET protein available for activation, thereby reducing RET-mediated cell viability and proliferation. Our work has uncovered a role for NCAM1 in modulating RET activity in cancer and validates the use of our synthetic dosage lethal approach in uncovering proteins facilitating oncogenic RET activity. Citation Format: Montdher H. Hussain, Brandy D. Hyndman, Frederick S. Vizeacoumar, Mathieu J. Crupi, Franco J. Vizeacoumar, Lois M. Mulligan. A role for the cell adhesion molecule NCAM1 in regulating activity of the receptor tyrosine kinase RET in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6996.

Abstract 1884: DXC006, Anti-CD56-CPT113 ADC displays favorable anti-tumor efficacy, pharmacokinetic and safety profiles in preclinical studies

Abstract CD56, also known as neural cell adhesion molecule 1 (NCAM1), is a type I plasma membrane glycoprotein involved in cell-cell and cell-matrix adhesion. It is overexpressed in almost all neuroblastomas, 95% of small cell lung cancers and 78% of multiple myeloma patients[1], whereas CD56 is less frequently or less abundantly expressed in normal tissues, making it a potential target for tumor therapy. So far, no CD56-targeting ADCs have entered late-stage clinical trials.DXC006 is an antibody-drug conjugate of a humanized anti-CD56 antibody (DXA006) linked to the topoisomerase I inhibitor CPT113 through a cleavable peptidyl linker and CROSSCONJU™ technology. The pharmacologic activity and mechanism of action of DXC006 were investigated in several human cancer cell lines and xenograft mouse models. Pharmacokinetic and safety profiles were also assessed in rats and cynomolgus monkeys.DXC006 binds specifically to CD56 and internalizes into tumor cells, after intracellular trafficking to lysosome and CPT113 releasing, induces DNA damage. In vitro cell based assays showed that DXC006 remarkably inhibited the cell growth of CD56-positive cell lines of IMR-32 (neuroblastoma), NCI-H526 (small cell lung cancer), and NCI-H929 (multiple myeloma), with IC50 of 0.0046~0.061 nmol/L. Although DXC006 also binds to CD56 expressed NK cells, we found that the inhibition of DXC006 to human NK cells was much less effective, with IC50 higher than 675 nmol/L, which is ten thousand-folds less potent than that of targeted abnormal cells. In vivo, DXC006 demonstrated good durable antitumor activities in three CD56-positive xenograft models of IMR-32, NCI-H526 and NCI-H929 cell lines, with a single dose of 2 mg/Kg, 3 mg/Kg, and 5 mg/Kg, respectively. No animal death or moribund outcome was observed in two months during preclinical safety studies in 150 mg/Kg Q2W dosed rats. For cynomolgus monkeys, the highest non-severely toxic dose (HNSTD) of DXC006 was 20 mg/Kg, in Q2W, four times of administration. PK profiles of both DXC006 the naked antibody DXA006 were comparable in both rats and monkeys. In summary, DXC006 demonstrated strong antitumor activity in various types of cancer cells both in vitro and in vivo, and good pharmacokinetic and safety profiles in rat and monkeys, a potential to be explored in clinical applications. [1] Yang Feng, Yanping Wang, Zhongyu Zhu, et al. Differential killing of CD56-expressing cells by drug-conjugated human antibodies targeting membrane-distal and membrane-proximal nonoverlapping epitopes. mAbs 2016; 799-810. Citation Format: Xiangfei Kong, Huihui Guo, Yong Du, Zhicang Ye, Yongxiang Chen, Zhixiang Guo, Lingli Zhang, Lu Bai, Junxiang Jia, Yunxia Zheng, Wei Zheng, Jun Zheng, Wenjun Li, Yuanyuan Huang, Zhongliang Fan, Mengmeng Liu, Binbin Chen, Meng Dai, Juan Wang, Fang Du, Miaomiao Chen, Qingliang Yang, Robert Y. Zhao. DXC006, Anti-CD56-CPT113 ADC displays favorable anti-tumor efficacy, pharmacokinetic and safety profiles in preclinical studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1884.

Abstract 2364: Anti-CADM1 chimeric antigen receptor (CAR) is a novel therapeutic strategy against small cell lung cancer

Abstract Small cell lung cancer (SCLC) is a highly aggressive form accounting for 15% of all lung cancers. SCLC is characterized by high proliferative rates, early metastasis, and poor prognosis. The median survival is less than 2 years even in patients with early-stage disease and less than a year in patients with metastatic disease which highlights the need for new therapeutic strategies. Cell Adhesion Molecule (CADM) 1 encodes a glycoprotein which belongs to an immunoglobulin superfamily of cell adhesion molecules responsible for epithelial cell adhesion through its homophilic trans-interaction between the adjacent cells. CADM1 is highly expressed on the cell surface of 80% of the SCLC cells, associated with its neuroendocrine features, along with the expression of Neural Cell Adhesion Molecule (NCAM). CADM1 is known to act as tumor promoter through its interactions with actin binding proteins. Consistently, we observed CADM1 expression in H69, H82, and H510 SCLC cell lines as assessed by western immuno blot. More importantly they were all susceptible to NK mediated cytotoxicity when assessed against NK cell line, NK92. We previously demonstrated that CADM1 mediates NK-mediated immune surveillance in non-small cell lung cancer, suggesting that CADM1 could be a potential immunotherapeutic target for SCLC. Here we report the development of an anti-CADM1 chimeric antigen receptor (CAR) specific for T and NK cells engineered with T and NK cell specific adaptor domains. Assessed the specificity of CARs by expressing them in NK92 cells and testing them against CADM1 expressing and CADM1-null A549 cells as targets. Finally, we evaluated the efficacy of anti-CADM1 CAR-T and CAR-NK cells against SCLC targets and observed a robust killing, largely proportionate to the levels of CADM1 expression in them. We are further assessing the efficacy of anti-CADM1 CAR-T and CAR-NK products in inhibiting tumor growth in NSG mice against SCLC xenografts. Together, these observations demonstrate that targeting CADM1 with CAR-T and CAR-NK cells is a potential new therapeutic strategy for SCLC. Citation Format: Shiva Krishna Katkam, Sergei Zolov, Delaney Shield, Sergei Chuikov, Venkateshwar G Keshamouni. Anti-CADM1 chimeric antigen receptor (CAR) is a novel therapeutic strategy against small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2364.

CD56/NCAM mediates cell migration of human NK cells by promoting integrin-mediated adhesion turnover.

Natural killer (NK) cells patrol tissue to mediate lysis of virally infected and tumorigenic cells. Human NK cells are typically identified by their expression of neural cell adhesion molecule (NCAM, CD56), yet despite its ubiquitous expression on NK cells, CD56 remains a poorly understood protein on immune cells. CD56 has been previously demonstrated to play roles in NK cell cytotoxic function and cell migration. Specifically, CD56-deficient NK cells have impaired cell migration on stromal cells and CD56 is localized to the uropod of NK cells migrating on stroma. Here, we show that CD56 is required for NK cell migration on ICAM-1 and is required for the establishment of persistent cell polarity and unidirectional actin flow. The intracellular domain of CD56 (NCAM-140) is required for its function and the loss of CD56 leads to enlarged actin foci and sequestration of phosphorylated Pyk2 accompanied by increased size and frequency of activated LFA-1 clusters. Together, these data identify a role for CD56 in regulating human NK cell migration through modulation of actin dynamics and integrin turnover.

CircNRCAM up-regulates NRCAM to promote papillary thyroid carcinoma progression.

Papillary Thyroid Carcinoma (PTC) is the most prevalent subtype of Thyroid Carcinoma (THCA), a type of malignancy in the endocrine system. According to prior studies, Neural Cell Adhesion Molecule (NRCAM) has been found to be up-regulated in PTC and stimulates the proliferation and migration of PTC cells. However, the specific mechanism of NRCAM in PTC cells is not yet fully understood. Consequently, this study aimed to investigate the underlying mechanism of NRCAM in PTC cells, the findings of which could provide new insights for the development of potential treatment targets for PTC. Bioinformatics tools were utilized and a series of experiments were conducted, including Western blot, colony formation, and dual-luciferase reporter assays. The data collected indicated that NRCAM was overexpressed in THCA tissues and PTC cells. Circular RNA NRCAM (circNRCAM) was found to be highly expressed in PTC cells and to positively regulate NRCAM expression. Through loss-of-function assays, both circNRCAM and NRCAM were shown to promote the proliferation, invasion, and migration of PTC cells. Mechanistically, this study confirmed that precursor microRNA-506 (pre-miR-506) could bind with m6A demethylase AlkB Homolog 5 (ALKBH5), leading to its m6A demethylation. It was also discovered that circNRCAM could competitively bind to ALKBH5, which restrained miR-506-3p expression and promoted NRCAM expression. In summary, circNRCAM could up-regulate NRCAM by down-regulating miR-506-3p, thereby enhancing the biological behaviors of PTC cells.

K-Ras(V12) differentially affects the three Akt isoforms in lung and pancreatic carcinoma cells and upregulates E-cadherin and NCAM via Akt3

K-Ras is the most frequently mutated Ras variant in pancreatic, colon and non-small cell lung adenocarcinoma. Activating mutations in K-Ras result in increased amounts of active Ras-GTP and subsequently a hyperactivation of effector proteins and downstream signaling pathways. Here, we demonstrate that oncogenic K-Ras(V12) regulates tumor cell migration by activating the phosphatidylinositol 3-kinases (PI3-K)/Akt pathway and induces the expression of E-cadherin and neural cell adhesion molecule (NCAM) by upregulation of Akt3. In vitro interaction and co-precipitation assays identified PI3-Kα as a bona fide effector of active K-Ras4B but not of H-Ras or N-Ras, resulting in enhanced Akt phosphorylation. Moreover, K-Ras(V12)-induced PI3-K/Akt activation enhanced migration in all analyzed cell lines. Interestingly, Western blot analyses with Akt isoform-specific antibodies as well as qPCR studies revealed, that the amount and the activity of Akt3 was markedly increased whereas the amount of Akt1 and Akt2 was downregulated in EGFP-K-Ras(V12)-expressing cell clones. To investigate the functional role of each Akt isoform and a possible crosstalk of the isoforms in more detail, each isoform was stably depleted in PANC-1 pancreatic and H23 lung carcinoma cells. Akt3, the least expressed Akt isoform in most cell lines, is especially upregulated and active in Akt2-depleted cells. Since expression of EGFP-K-Ras(V12) reduced E-cadherin-mediated cell-cell adhesion by induction of polysialylated NCAM, Akt3 was analyzed as regulator of E-cadherin and NCAM. Western blot analyses revealed pronounced reduction of E-cadherin and NCAM in the Akt3-kd cells, whereas Akt1 and Akt2 depletion upregulated E-cadherin, especially in H23 lung carcinoma cells. In summary, we identified oncogenic K-Ras4B as a key regulator of PI3-Kα-Akt signaling and Akt3 as a crucial regulator of K-Ras4B-induced modulation of E-cadherin and NCAM expression and localization.

Relationship between patterns of immunohistochemical conventional neuroendocrine markers and efficacy of immune check point inhibitors in patients with extensive disease small cell lung cancer.

Which patients benefit from the addition of immune checkpoint inhibitors (ICIs) to chemotherapy for small cell lung cancer (SCLC) remains unclear. There have been few reports on the efficacy of ICIs based on conventional immunohistochemical neuroendocrine (NE) markers (synaptophysin, chromogranin A, and neural cell adhesion molecule [NCAM]). In the present study, we aimed to analyze the relationship between the expression of immunohistochemical NE markers and the efficacy of ICIs in patients with extensive disease (ED)-SCLC, to assess whether conventional NE markers are predictive of ICIs. Patients with untreated ED-SCLC who received first-line therapy at the Shizuoka Cancer Center between November 2002 and July 2021 were retrospectively reviewed. We evaluated the efficacy of first-line chemotherapy according to the expression status of each immunohistochemical NE marker in patients treated with ICI plus chemotherapy (ICI-chemo group) and with chemotherapy alone (chemo group). A total of 227 patients were included in the ICI-chemo and chemo groups, respectively. The progression-free survival (PFS) tended to be better in patients in the ICI-chemo group than those treated with chemotherapy alone in patients with NE marker-positive SCLC. In particular, it was statistically significant in patients with chromogranin A-positive SCLC (p = 0.036). In patients with NE marker-negative SCLC, no significant differences were observed in PFS between the two groups. There were no significant differences in overall survival (OS), regardless of the expression of any conventional NE marker. Our study suggests that the efficacy of ICIs in addition to chemotherapy may be poor in patients with NE marker-negative SCLC.

NCAM mimetic peptide P2 synergizes with bone marrow mesenchymal stem cells in promoting functional recovery after stroke.

The neural cell adhesion molecule (NCAM) promotes neural development and regeneration. Whether NCAM mimetic peptides could synergize with bone marrow mesenchymal stem cells (BMSCs) in stroke treatment deserves investigation. We found that the NCAM mimetic peptide P2 promoted BMSC proliferation, migration, and neurotrophic factor expression, protected neurons from oxygen-glucose deprivation through ERK and PI3K/AKT activation and anti-apoptotic mechanisms in vitro. Following middle cerebral artery occlusion (MCAO) in rats, P2 alone or in combination with BMSCs inhibited neuronal apoptosis and induced the phosphorylation of ERK and AKT. P2 combined with BMSCs enhanced neurotrophic factor expression and BMSC proliferation in the ischemic boundary zone. Moreover, combined P2 and BMSC therapy induced translocation of nuclear factor erythroid 2-related factor, upregulated heme oxygenase-1 expression, reduced infarct volume, and increased functional recovery as compared to monotreatments. Treatment with LY294002 (PI3K inhibitor) and PD98059 (ERK inhibitor) decreased the neuroprotective effects of combined P2 and BMSC therapy in MCAO rats. Collectively, P2 is neuroprotective while P2 and BMSCs work synergistically to improve functional outcomes after ischemic stroke, which may be attributed to mechanisms involving enhanced BMSC proliferation and neurotrophic factor release, anti-apoptosis, and PI3K/AKT and ERK pathways activation.

The effects of induced type I diabetes on developmental regulation of GDNF, NRTN, and NCAM proteins in the dentate gyrus of male rat offspring

BackgroundMaternal diabetes during pregnancy can affect the neurological development of offspring. Glial cell-derived neurotrophic factor (GDNF), neurturin (NRTN), and neural cell adhesion molecules (NCAM) are three important proteins for brain development. Therefore, this study aimed to investigate the impacts of the mentioned neurotrophic factors in the hippocampal dentate gyrus (DG) of rat offspring born to diabetic mothers. MethodsWistar female rats were randomly allocated into diabetic (STZ-D) [(45 mg/kg BW, STZ (Streptozotocin), i.p)], diabetic + NPH insulin (STZ-INS) [(4–6 unit/kg/day SC)], and control groups. The animals in all groups were mated by non-diabetic male rats. Two weeks after birth, male pups from each group were sacrificed and then protein contents of GDNF, NRTN, and NCAM were evaluated using immunohistochemistry. ResultsThe study found that the expression of GDNF and NRTN in the hippocampus of diabetic rat offspring was significantly higher compared to the diabetic+ insulin and control groups, respectively (P < 0.01, P < 0.001). Additionally, the expression of NCAM was significantly higher in the diabetic group the diabetic+ insulin and control groups (P < 0.01, P < 0.001). ConclusionsThe results of the study revealed that diabetes during pregnancy significantly impacts the distribution pattern of GDNF, NRTN, and NCAM in the hippocampus of rat neonates.

Anisotropic microtopography surface of chitosan scaffold regulating skin precursor-derived Schwann cells towards repair phenotype promotes neural regeneration.

For repairing peripheral nerve and spinal cord defects, biomaterial scaffold-based cell-therapy was emerged as an effective strategy, requiring the positive response of seed cells to biomaterial substrate and environment signals. Previous work highlighted that the imposed surface properties of scaffold could provide important guidance cues to adhered cells for polarization. However, the insufficiency of native Schwann cells and unclear cellular response mechanisms remained to be addressed. Given that, this study aimed to illuminate the micropatterned chitosan-film action on the rat skin precursor-derived Schwann cells (SKP-SCs). Chitosan-film with different ridge/groove size was fabricated and applied for the SKP-SCs induction. Results indicated that SKP-SCs cultured on 30 μm size microgroove surface showed better oriented alignment phenotype. Induced SKP-SCs presented similar genic phenotype as repair Schwann cells, increasing expression of c-Jun, neural cell adhesion molecule, and neurotrophic receptor p75. Moreover, SKP-SC-secretome was subjected to cytokine array GS67 assay, data indicated the regulation of paracrine phenotype, a panel of cytokines was verified up-regulated at secreted level and gene expression level in induced SKP-SCs. These up-regulated cytokines exhibit a series of promotive neural regeneration functions, including cell survival, cell migration, cell proliferation, angiogenesis, axon growth, and cellular organization etc. through bioinformatics analysis. Furthermore, the effectively polarized SKP-SCs-sourced secretome, promoted the proliferation and migration capacity of the primarily cultured native rat Schwann cells, and augmented neurites growth of the cultured motoneurons, as well as boosted axonal regrowth of the axotomy-injured motoneurons. Taken together, SKP-SCs obtained pro-neuroregeneration phenotype in adaptive response to the anisotropic topography surface of chitosan-film, displayed the oriented parallel growth, the transition towards repair Schwann cell genic phenotype, and the enhanced paracrine effect on neural regeneration. This study provided novel insights into the potency of anisotropic microtopography surface to Schwann-like cells phenotype regulation, that facilitating to provide promising engineered cell-scaffold in neural injury therapies.