CD46 靶点

… Moesin and CD46 were shown to be colocalized by confocal analysis. Moesin cross-linkages between actin and CD46. In human Mφ, CD46 CYT recruits several PTKs and CD46 …

Graphical abstract

… CD46 (also known as membrane cofactor protein or MCP) is a regulator of complement activation that is expressed in most or all human nucleated cell types. CD46 … , bovine CD46 was …

… /67 as human membrane cofactor protein (CD46) and shown that it … CD46 have identical electrophoretic profiles and Mr heterogeneity. We have further shown that expression of CD46 …

C4b and C3b deposited on host cells undergo limited proteolytic cleavage by regulatory proteins. Membrane cofactor protein (MCP; CD46), factor H, and C4b binding protein mediate this reaction, known as cofactor activity, that also requires the plasma serine protease factor I. To explore the roles of the fluid phase regulators vs those expressed on host cells, a model system was used examining complement fragments deposited on cells transfected with human MCP as assessed by FACS and Western blotting. Following incubation with Ab and complement on MCP+ cells, C4b was progressively cleaved over the first hour to C4d and C4c. There was no detectable cleavage of C4b on MCP− cells, indicating that MCP (and not C4BP in the serum) primarily mediates this cofactor activity. C3b deposition was not blocked on MCP+ cells because classical pathway activation occurred before substantial C4b cleavage. Cleavage, though, of deposited C3b was rapid (<5 min) and iC3b was the dominant fragment on MCP− and MCP+ cells. Studies using a function-blocking mAb further established factor H as the responsible cofactor. If the level of Ab sensitization was reduced 8-fold or if Mg2+-EGTA was used to block the classical pathway, MCP efficiently inhibited C3b deposition mediated by the alternative pathway. Thus, for the classical pathway, MCP is the cofactor for C4b cleavage and factor H for C3b cleavage. However, if the alternative pathway mediates C3b deposition, then MCP’s cofactor activity is sufficient to restrict complement activation.

The tetraspans associate with a large number of surface molecules, including a subset of beta1 integrins and, indirectly through CD19, with the complement receptor CD21. To further characterize the tetraspan complexes we have raised and selected monoclonal antibodies (mAb) for their ability to immunoprecipitate a molecule associated with CD9. A unique mAb was identified which recognizes the complement regulator CD46 (membrane cofactor protein). CD46 associated in part with several tetranspans and with all beta1 integrins that were tested (CD29/CD49a, CD29/CD49b, CD29/CD49c, CD29/CD49e, CD29/CD49f) but not with beta4 integrins. These data, together with cross-linking experiments showing the existence in living cells of CD46/integrin complexes, suggest that CD46 associates directly with beta1 integrins and indirectly with tetraspans. CD46 also acts as a receptor for measles virus; however, mAb to various integrins and tetraspans did not modify the virus fusion entry step.

SUMMARYHuman membrane cofactor protein (MCP; CD46) is a widely distributed complement regulator. In the mouse, expression of MCP is largely restricted to the testis while a related, widely expressed protein (Crry) appears to perform MCP's (CD46) regulatory activity. We have developed two mouse strains transgenic for human MCP (CD46) utilizing an ∼ 400 kb YAC clone carrying the complete gene. A third mouse strain was generated using an overlapping YAC clone isolated from a second library. The expression of human MCP (CD46) in these mouse strains was characterized by immunohistochemistry, FACS, Western blotting and RT-PCR. No differences were detected in the isoform pattern or distribution among the three strains, although the expression level varied according to how many copies of the gene were integrated. The expression profile closely mimicked that observed in humans, including the same pattern of isoform expression as the donor. In addition, tissue-specific isoform expression in the kidney, salivary gland and brain paralleled that observed in man. The transgenic mice expressed low levels of MCP (CD46) on their E, in contrast to humans but in line with most other primates. These mice should be a useful tool to analyse tissue-specific expression, to establish animal models of infections and to characterize the role of MCP (CD46) in reproduction.

… complement C3b protein on the surface of infected cells (4). CD46, or membrane cofactor protein, … to inhibit deposition of complement proteins C3b and C4b on host cells (5). Recently, …

… proteins that share a homologous structural motif termed the short consensus repeat. Membrane cofactor protein (MCP), or CD46, is a short consensus repeat-containing protein found …

… Mutations in the plasma complement regulators factor H and factor I and the widely expressed membrane cofactor protein (MCP; CD46) have been described recently. This study looked …

Adherence of group A streptococcus (GAS) to keratinocytes is mediated by an interaction between human CD46 (membrane cofactor protein) with streptococcal cell surface M protein. CD46 belongs to a family of proteins that contain structurally related short consensus repeat (SCR) domains and regulate the activation of the complement components C3b and/or C4b. CD46 possesses four SCR domains and the aim of this study was to characterize their interaction with M protein. Following confirmation of the M6 protein-dependent interaction between GAS and human keratinocytes, we demonstrated that M6 protein binds soluble recombinant CD46 protein and to a CD46 construct containing only SCRs 3 and 4. M6 protein did not bind to soluble recombinant CD46 chimeric proteins that had the third and/or fourth SCR domains replaced with the corresponding domains from another complement regulator, CD55 (decay-accelerating factor). Homology-based molecular modeling of CD46 SCRs 3 and 4 revealed a cluster of positively charged residues between the interface of these SCR domains similar to the verified M protein binding sites on the plasma complement regulators factor H and C4b-binding protein. The presence of excess M6 protein did not inhibit the cofactor activity of CD46 and the presence of excess C3b did not inhibit the ability of CD46 to bind M6 protein by ELISA. In conclusion, 1) adherence of M6 GAS to keratinocytes is M protein dependent and 2) a major M protein binding site is located within SCRs 3 and 4, probably at the interface of these two domains, at a site distinct from the C3b-binding and cofactor site of CD46.

Membrane cofactor protein (MCP; CD46) is a widely expressed type 1 transmembrane glycoprotein that inhibits complement activation on host cells. It also is a receptor for several pathogens including measles virus, Streptococcus pyogenes, Neisseria gonorrhea, and Neisseria meningitidis. That MCP may have signaling capability was suggested by its microbial interactions. That is, binding of MCP on human monocytes by measles virus hemagglutinin or cross-linking by an anti-MCP Ab resulted in IL-12 down-regulation, while binding to MCP by Neisseria on epithelial cells produced a calcium flux. Through alternative splicing, MCP is expressed on most cells with two distinct cytoplasmic tails of 16 (CYT-1) or 23 (CYT-2) amino acids. These play pivotal roles in intracellular precursor processing and basolateral localization. We investigated the putative signal transduction pathway mediated by MCP and demonstrate that CYT-2, but not CYT-1, is phosphorylated on tyrosine. We examined MCP tail peptides and performed Ab cross-linking experiments on several human cell lines and MCP isoform transfectants. We found an MCP peptide of CYT-2 was phosphorylated by a src kinase system. Western blots of the cells lines demonstrated that cells bearing CYT-2 were also phosphorylated on tyrosine. Additionally, we provide genetic and biochemical evidence that the src family of kinases is responsible for the latter phosphorylation events. In particular, the src kinase, Lck, is required for phosphorylation of MCP in the Jurkat T cell line. Taken together, these studies suggest a src family-dependent pathway for signaling through MCP.

… are produced by alternative splicing of the CD46 … CD46 becomes proteolytically modified on cell membranes. We observed that tumor cells liberated intact 60–65 kDa forms of CD46 …

… Membrane cofactor protein (MCP, CD46) is an integral protein that serves as a cofactor for factor I in inactivating C3b/C4b deposited on the same cell membrane as C3bi/C4c+C4d.This …

The cleavage of C3 is a critical step for complement (C) activation in the classical and alternative pathways. This reaction is controlled by the regulators of C activation protein family. Membrane cofactor protein (MCP) is a cofactor for the factor I-mediated inactivation of C3b and C4b. As a widely distributed membrane protein, MCP may protect host cells from inadvertent C activation. Human MCP has recently been shown to protect transfected rodent cells from human C-mediated lysis. In this report the relationship of MCP expression to C3b deposition and cytoprotection was examined using NIH/3T3 cells transfected with human MCP and exposed to human serum as a source of C and naturally occurring anti-mouse antibody. MCP inhibited C3b deposition in a dose-dependent fashion and inhibited lysis of the mouse cells expressing it. MCP did not inhibit lysis on bystander cells. These results demonstrate the protective role of MCP, at the cellular level, by an intrinsic mechanism.

Membrane cofactor protein (MCP) (CD46) of the C system binds to C3b and C4b, functions as a cofactor for their cleavage, and protects autologous cells from C-mediated injury. The predominant structural motif of MCP is the short consensus repeat (SCR), a repeating domain involved in ligand binding of other related C regulatory proteins. SCR deletion mutants were constructed to determine which of the four SCR of MCP contribute to ligand binding and cofactor activity. ELISA were developed to evaluate binding efficiency of mutants to ligand. Analysis of the deletion mutants indicated that the third and fourth SCR were important for both ligand binding and cofactor activity of C3b (iC3) and C4b. In addition, the same SCR were required for efficient binding of an mAb known to inhibit MCP function. The mutant deleted of SCR-2 bound but lacked cofactor activity for iC3. It did not bind or possess cofactor activity for C4b. Deletion of the first (amino-terminal) SCR had a minimal effect on iC3 binding and cofactor activity but reduced the efficiency of C4b binding. The results identify the SCR of MCP that contribute to ligand binding and cofactor activity. The data also suggest the presence of distinguishable iC3 and C4b binding sites and provide evidence that iC3 binding is not always sufficient for cofactor activity.

Membrane cofactor protein (MCP; CD46) is a widely distributed C3b/C4b-binding glycoprotein that inhibits complement activation on host cells. MCP is expressed primarily as four isoforms that arise by alternative splicing of a single gene. The differences reside in the domains for O-glycosylation and cytoplasmic tails. Tissue-specific expression of isoforms and the differential processing of precursors mediated by the cytoplasmic tails suggest that isoform variations are biologically significant. The goal of these experiments was to characterize the complement inhibitory profile of the four commonly expressed isoforms. The MCP isoforms (BC) with a larger O-glycosylation domain bound C4b more efficiently than the C isoforms, which are smaller and less glycosylated in this region. Additionally, cytoprotection assays of individual clones of transfected isoforms bearing equivalent copy numbers demonstrated that the BC isoforms also provided enhanced protection in a classical pathway-mediated system and cleaved cell-bound C4b more efficiently than the C isoforms. Taken together, these data demonstrate that BC isoforms preferentially protect against the classical pathway of complement. Such findings indicate a physiologic role for isoform variation and have therapeutic implications for use of MCP isoforms as complement inhibitors in such areas as xenotransplantation.

… membrane cofactor protein (CD46) gene of the human complement system and comparison to a membrane cofactor protein-like genetic element. J Immunol 151: 4137-4146, 1993. 62. …

… CD46 is involved in more than complement regulation. In experiments designed to block fertilization using mAbs to CD46… however, for the mechanism by which CD46 is connected to an …

Membrane cofactor protein (MCP; CD46) is an ubiquitously expressed complement regulatory protein that protects host cells from injury by complement. This type-I membrane glycoprotein serves as a cofactor for the serine protease factor I to mediate inactivation of C3b and C4b deposited on host cells. More than 60 disease-associated mutations in MCP have now been identified. The majority of the mutations are linked to a rare thrombotic microangiopathic-based disease, atypical hemolytic uremic syndrome (aHUS), but new putative links to systemic lupus erythematosus, glomerulonephritis, and pregnancy-related disorders among others have also been identified. This review summarizes our current knowledge of disease-associated mutations in this complement inhibitor.

T he classic pathogen-fighting functions of our microbe sensing systems, such as the TLRs, the Nod-like receptors, and several proteins of the complement system, had been defined relatively rapidly after their discovery (1, 2). A more recent trend, however, clearly indicates that some of these evolutionarily ancient systems also serve additional, noncanonical roles in normal cell physiology. In this article, we highlight how the functions of the complement component CD46 have expanded dramatically beyond its initial discovery as a regulator of complement activation. We now recognize this ancient molecule as a biological focal point for our continuously evolving understanding of the diverse roles of complement as a key orchestrator of (immunological) health. Although identified in the early 1900s, complement traces its origins to more than a billion years ago, when primitive proteins evolved to protect cells from pathogens and to engage in other intracellular processes (reviewed in Refs. 3–5). The contemporary complement system consists of three independently triggered activation pathways (classical, alternative, lectin) and a terminal cytolytic pathway common to all. It engages both innate and adaptive immunity. Complement component C3, the most abundant of its proteins, is the nexus where all three activation pathways converge. The proteolytic cleavage of C3 generates C3a (an anaphylatoxin) and C3b (an opsonin and critical component of the convertase complexes). Unbridled complement activation, however, would just as powerfully attack self-tissue as it does pathogens. Thus, activation must be strictly controlled to maintain appropriate homeostasis while avoiding damage to self. In this Pillars of Immunology article, Cole et al. (6) focused on the regulatory side of complement: in particular, on proteins that bind C3. During the 1980s, a family of structurally, functionally, and genetically related regulators was being elucidated as inhibitors of complement activation via their interactions with fragments of C3 (and/or C4). The regulators of complement activation (RCA) family, consisting of serum and cell-anchored proteins, employed two key processes: decay accelerating activity and cofactor activity. The former refers to the permanent dissociation of activating complexes (i.e., convertases), whereas the latter refers to a role as cofactor for the proteolytic cleavage of C3 (or C4) fragments in association with the serine protease, factor I. The Atkinson laboratory had been studying polymorphisms of two such regulators, complement receptors (CR) CR1 and CR2 using C3 (i.e., C3b or iC3) affinity chromatography of surface-labeled peripheral blood cells (7). However, they routinely observed a third group of molecules when studying human leukocytes. The study by Cole et al. (6) delved into this phenomenon by examining the cell distribution, relative mobility, and antigenic specificity of “. . . a heretofore unrecognized group of 45,000–70,000 Mr C3-binding molecules.” The authors found that every tested cell population possessed this new factor as a broad band or doublet pattern. They dubbed the new protein gp45–70. A year later, Tsukasa Seya, of the same group, developed a purification scheme characterizing two distinct species, “upper” and “lower,” each of which possessed cofactor activity for C3b cleavage (8). Interestingly, the C3b cofactor activity was unique when compared with CR1. To reflect the growing structure/function information, the Atkinson group renamed the molecule as “membrane cofactor protein (MCP),” later designated as CD46 (reviewed in Ref. 9). Since these pioneering studies, we have learned that CD46 is ubiquitously expressed on all cells, except erythrocytes, and is a cofactor for C3b and C4b cleavage. Its cloning and characterization revealed that CD46 is a type 1 transmembrane glycoprotein coexpressed on most cells as four isoforms, which arise by alternative splicing of a single gene that lies within the RCA cluster on chromosome 1q32.2 (10–13). The structural heterogeneity is in part accounted for by alternative splicing in an extracellular region for O-glycosylation (BC region) and by having one of two intracellular cytoplasmic tails (tail 1 or tail 2; *Division of Rheumatology, Department of Medicine, Washington University School of Medicine, Saint Louis, MO 63110; Immunology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892; School of Immunology and Microbial Sciences, Faculty of Life Sciences and Medicine, King’s College London, London SE1 9RT, United Kingdom; and xInstitute for Systemic Inflammation Research, University of L€ubeck, 23562 L€ubeck, Germany

… mediated cytotoxicity is one mechanism of immune suppression by CD3/CD46-generated T-… through cytotoxic mechanisms. The data just discussed indicate that CD46induced Tr1-like …

… has not yet been matched with a comprehensive understanding of the molecular mechanisms by which CD46 affects immune function. This review summarizes the evidence that points …

… of the workings of complement regulatory protein, CD46 (membrane … complement inhibitor. Referred to as a “pathogen magnet,” … events to complement-dependent effector mechanisms. …

… cascade as an effector of humoral immune mechanisms to a network involved actively in … Mice express crry/p65, which shares some of the complement regulatory properties of CD46 …

The human membrane cofactor protein (MCP, CD46) is a central component of the innate immune system. CD46 protects autologous cells from complement attack by binding to complement proteins C3b and C4b and serving as a cofactor for their cleavage. Recent data show that CD46 also plays a role in mediating acquired immune responses, and in triggering autophagy. In addition to these physiologic functions, a significant number of pathogens, including select adenoviruses, measles virus, human herpes virus 6 (HHV-6), Streptococci, and Neisseria, use CD46 as a cell attachment receptor. We have determined the crystal structure of the extracellular region of CD46 in complex with the human adenovirus type 11 fiber knob. Extracellular CD46 comprises four short consensus repeats (SCR1-SCR4) that form an elongated structure resembling a hockey stick, with a long shaft and a short blade. Domains SCR1, SCR2 and SCR3 are arranged in a nearly linear fashion. Unexpectedly, however, the structure reveals a profound bend between domains SCR3 and SCR4, which has implications for the interactions with ligands as well as the orientation of the protein at the cell surface. This bend can be attributed to an insertion of five hydrophobic residues in a SCR3 surface loop. Residues in this loop have been implicated in interactions with complement, indicating that the bend participates in binding to C3b and C4b. The structure provides an accurate framework for mapping all known ligand binding sites onto the surface of CD46, thereby advancing an understanding of how CD46 acts as a receptor for pathogens and physiologic ligands of the immune system.

CD46 is a ubiquitously expressed type I transmembrane protein, first identified as a regulator of complement activation, and later as an entry receptor for a variety of pathogens. The last decade has also revealed the role of CD46 in regulating the adaptive immune response, acting as an additional costimulatory molecule for human T cells and inducing their differentiation into Tr1 cells, a subset of regulatory T cells. Interestingly, CD46 regulatory pathways are defective in T cells from patients with multiple sclerosis, asthma and rheumatoid arthritis, illustrating its importance in regulating T cell homeostasis. Indeed, CD46 expression at the cell surface is tightly regulated in many different cell types, highlighting its importance in several biological processes. Notably, CD46 is the target of enzymatic processing, being cleaved by metalloproteinases and by the presenilin/gamma secretase. This processing is required for its functions, at least in T cells. This review will summarize the latest updates on the regulation of CD46 expression and on its effects on T cell activation.

The cell surface complement regulatory (CReg) proteins CD46, CD55 and CD59 are widely expressed on human lymphoid and non‐lymphoid cells. This study aimed to compare …

In this study we demonstrate a new form of immunoregulation: engagement on CD4+ T cells of the complement regulator CD46 promoted the effector potential of T helper type 1 cells (TH1 cells), but as interleukin 2 (IL-2) accumulated, it switched cells toward a regulatory phenotype, attenuating IL-2 production via the transcriptional regulator ICER/CREM and upregulating IL-10 after interaction of the CD46 tail with the serine-threonine kinase SPAK. Activated CD4+ T cells produced CD46 ligands, and blocking CD46 inhibited IL-10 production. Furthermore, CD4+ T cells in rheumatoid arthritis failed to switch, consequently producing excessive interferon-γ (IFN-γ). Finally, γδ T cells, which rarely produce IL-10, expressed an alternative CD46 isoform and were unable to switch. Nonetheless, coengagement of T cell antigen receptor (TCR) γδ and CD46 suppressed effector cytokine production, establishing that CD46 uses distinct mechanisms to regulate different T cell subsets during an immune response.

… , CD46, and CD55 in human blood cells, and the differences in these complement regulatory … We have studied the mechanism of CD46 up-regulation in our laboratory in leukemia cell …

BACKGROUND Complement is thought to play a role in immunoglobulin A nephropathy (IgAN), though the activating mechanisms are unknown. This study focused on the gene expression of CD46 and CD55, two key molecules for regulating C3 convertase activity of lectin and alternative complement pathways at a cellular level. METHODS The transcriptional expression in peripheral white blood cells (WBCs) of CD46 and CD55 was investigated in 157 patients enrolled by the Validation of the Oxford Classification of IgAN group, looking for correlations with clinical and pathology features and estimated glomerular filtration rate (eGFR) modifications from renal biopsy to sampling. Patients had a previous median follow-up of 6.4 (interquartile range 2.8-10.7) years and were divided into progressors and non-progressors according to the median value of their velocity of loss of renal function per year (-0.41 mL/min/1.73 m2/year). RESULTS CD46 and CD55 messenger RNA (mRNA) expression in WBCs was not correlated with eGFR values or proteinuria at sampling. CD46 mRNA was significantly correlated with eGFR decline rate as a continuous outcome variable (P = 0.014). A significant difference was found in CD46 gene expression between progressors and non-progressors (P = 0.013). CD46 and CD55 mRNA levels were significantly correlated (P < 0.01), although no difference between progressors and non-progressors was found for CD55 mRNA values. The prediction of progression was increased when CD46 and CD55 mRNA expressions were added to clinical data at renal biopsy (eGFR, proteinuria and mean arterial blood pressure) and Oxford MEST-C (mesangial hypercellularity, endocapillary hypercellularity, segmental glomerulosclerosis, tubular atrophy/interstitial fibrosis, presence of any crescents) score. CONCLUSIONS Patients with progressive IgAN showed lower expression of mRNA encoding for the complement inhibitory protein CD46, which may implicate a defective regulation of C3 convertase with uncontrolled complement activation.

In mammals, CD46 is involved in the inactivation of complement by factor I (FI). In teleost, study on the function of CD46 is very limited. In this study, we examined the immunological property of a CD46 molecule (CsCD46) from tongue sole, a teleost species with important economic value. We found that recombinant CsCD46 (rCsCD46) interacted with FI and inhibited complement activation in an FI-dependent manner. rCsCD46 also interacted with bacterial pathogens via a different mechanism to that responsible for the FI interaction, involving different rCsCD46 sites. Cellular study showed that CsCD46 was expressed on peripheral blood leukocytes (PBL) and protected the cells against the killing effect of complement. When the CsCD46 on PBL was blocked by antibody before incubation of the cells with bacterial pathogens, cellular infection was significantly reduced. Consistently, when tongue sole were infected with bacterial pathogens in the presence of rCsCD46, tissue dissemination and survival of the pathogens were significantly inhibited. These results provide the first evidence to indicate that CD46 in teleosts negatively regulates complement activation via FI and protects host cells from complement-induced damage, and that CD46 is required for optimal bacterial infection probably by serving as a receptor for the bacteria.

Background CD46 has been revealed to be a key factor in malignant transformation and cancer treatment. However, the clinical significance of CD46 in cervical cancer remains unclear, and this study aimed to evaluate its role in cervical cancer diagnosis and prognosis evaluation. Methods A total of 180 patients with an initial diagnosis of cervical cancer were enrolled at Taizhou Hospital of Zhejiang Province, China. The plasma levels of soluble CD46 (sCD46) and the expression of membrane-bound CD46 (mCD46) were detected by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry (IHC), respectively. Results CD46 was found to be significantly upregulated in cervical cancer tissues vs. normal tissues, while no CD46 staining was detected in paired adjacent noncancerous tissues. CD46 staining was more pronounced in cancer cells than in stromal cells in situ (in tissues). Moreover, the plasma levels of sCD46 were able to some extent discriminate between cancer patients and healthy women (AUC=0.6847, 95% CI:0.6152–0.7541). Analysis of Kaplan–Meier survival curves revealed that patients with low CD46 expression had slightly longer overall survival (OS) than patients with high CD46 expression in the tumor microenvironment, but no significant difference. Univariate Cox regression analysis revealed that CD46 (P=0.034) is an independent risk factor for OS in cervical cancer patients. Conclusion The present study demonstrated that cervical cancer patients exhibit aberrant expression of CD46, which is closely associated with a poor prognosis, suggesting that CD46 plays a key role in promoting cervical carcinogenesis and that CD46 could serve as a promising potential target for precision therapy for cervical cancer.

… complement regulators (CRegs), CD46, CD55 and CD59 in human mesothelial cells, and correlated with clinical background and level of complement … CD55 but not CD46 and CD59 in …

… that a CD46-targeted immunoPET radiotracer could enable a whole-body assessment of CD46 … burden, and aid patient selection and treatment monitoring for CD46-targeted therapies. …

… Thus, genomic amplification of CD46 may serve as a surrogate for target amplification that … CD46-targeted therapy. Overall, these findings indicate that CD46 is a promising target for …

… In light of these findings, CD46 may serve as a potential target not only for adenovirus-… gene therapy but also for any targeted therapy that depends on CD46 for cellular entry, target …

… of CD46-CAR T cells by suppressing the proliferation and liver metastasis in CD46 high-expressed CRC PDX mice. Collectively, our findings highlight that CD46-CAR T cell therapy …

Although initially responsive to androgen signaling inhibitors (ASIs), metastatic castration-resistant prostate cancer (mCRPC) inevitably develops and is incurable. In addition to adenocarcinoma (adeno), neuroendocrine prostate cancer (NEPC) emerges to confer ASI resistance. We have previously combined laser capture microdissection and phage antibody display library selection on human cancer specimens and identified novel internalizing antibodies binding to tumor cells residing in their tissue microenvironment. We identified the target antigen for one of these antibodies as CD46, a multifunctional protein that is best known for negatively regulating the innate immune system. CD46 is overexpressed in primary tumor tissue and CRPC (localized and metastatic; adeno and NEPC), but expressed at low levels on normal tissues except for placental trophoblasts and prostate epithelium. Abiraterone- and enzalutamide-treated mCRPC cells upregulate cell surface CD46 expression. Genomic analysis showed that the CD46 gene is gained in 45% abiraterone-resistant mCRPC patients. We conjugated a tubulin inhibitor to our macropinocytosing anti-CD46 antibody and showed that the resulting antibody-drug conjugate (ADC) potently and selectively kills both adeno and NEPC cell lines in vitro (sub-nM EC50) but not normal cells. CD46 ADC regressed and eliminated an mCRPC cell line xenograft in vivo in both subcutaneous and intrafemoral models. Exploratory toxicology studies of the CD46 ADC in non-human primates demonstrated an acceptable safety profile. Thus, CD46 is an excellent target for antibody-based therapy development, which has potential to be applicable to both adenocarcinoma and neuroendocrine types of mCRPC that are resistant to current treatment.

… of CD46 including the CD46 domains that engage with endogenous and/or pathogenic ligands and key disease states associated with perturbations in CD46 … in combination therapy. …

Abstract Purpose: Multiple myeloma is a plasma cell malignancy with an unmet clinical need for improved imaging methods and therapeutics. Recently, we identified CD46 as an overexpressed therapeutic target in multiple myeloma and developed the antibody YS5, which targets a cancer-specific epitope on this protein. We further developed the CD46-targeting PET probe [89Zr]Zr-DFO-YS5 for imaging and [225Ac]Ac-DOTA-YS5 for radiopharmaceutical therapy of prostate cancer. These prior studies suggested the feasibility of the CD46 antigen as a theranostic target in multiple myeloma. Herein, we validate [89Zr]Zr-DFO-YS5 for immunoPET imaging and [225Ac]Ac-DOTA-YS5 for radiopharmaceutical therapy of multiple myeloma in murine models. Experimental Design: In vitro saturation binding was performed using the CD46 expressing MM.1S multiple myeloma cell line. ImmunoPET imaging using [89Zr]Zr-DFO-YS5 was performed in immunodeficient (NSG) mice bearing subcutaneous and systemic multiple myeloma xenografts. For radioligand therapy, [225Ac]Ac-DOTA-YS5 was prepared, and both dose escalation and fractionated dose treatment studies were performed in mice bearing MM1.S-Luc systemic xenografts. Tumor burden was analyzed using BLI, and body weight and overall survival were recorded to assess antitumor effect and toxicity. Results: [89Zr]Zr-DFO-YS5 demonstrated high affinity for CD46 expressing MM.1S multiple myeloma cells (Kd = 16.3 nmol/L). In vitro assays in multiple myeloma cell lines demonstrated high binding, and bioinformatics analysis of human multiple myeloma samples revealed high CD46 expression. [89Zr]Zr-DFO-YS5 PET/CT specifically detected multiple myeloma lesions in a variety of models, with low uptake in controls, including CD46 knockout (KO) mice or multiple myeloma mice using a nontargeted antibody. In the MM.1S systemic model, localization of uptake on PET imaging correlated well with the luciferase expression from tumor cells. A treatment study using [225Ac]Ac-DOTA-YS5 in the MM.1S systemic model demonstrated a clear tumor volume and survival benefit in the treated groups. Conclusions: Our study showed that the CD46-targeted probe [89Zr]Zr-DFO-YS5 can successfully image CD46-expressing multiple myeloma xenografts in murine models, and [225Ac]Ac-DOTA-YS5 can effectively inhibit the growth of multiple myeloma. These results demonstrate that CD46 is a promising theranostic target for multiple myeloma, with the potential for clinical translation.

CD46 is a complement inhibitor membrane cofactor which also acts as a receptor for various microbes, including species B adenoviruses (Ads). While most Ad gene therapy vectors are derived from species C and infect cells through coxsackie-adenovirus receptor (CAR), CAR expression is downregulated in many cancer cells, resulting inefficient Ad-based therapeutics. Despite a limited knowledge on the expression status of many cancer cells, an increasing number of cancer gene therapy studies include fiber-modified Ad vectors redirected to the more ubiquitously expressed CD46. Since our finding from tumor microarray indicate that CD46 was overexpressed in cancers of the prostate and colon, fiber chimeric Ad5/35 vectors that have infection tropism for CD46 were employed to demonstrate its efficacy in colorectal cancers (CRC). CD46-overexpressed cells showed a significantly higher response to Ad5/35-GFP and to Ad5/35-tk/GCV. While CRC cells express variable levels of CD46, CD46 expression was positively correlated with Ad5/35-mediated GFP fluorescence and accordingly its cell killing. Injection of Ad5/35-tk/GCV caused much greater tumor-suppression in mice bearing CD46-overexpressed cancer xenograft compared to mock group. Analysis of CRC samples revealed that patients with positive CD46 expression had a higher survival rate (p=0.031), carried tumors that were well-differentiated, but less invasive and metastatic, and with a low T stage (all p<0.05). Taken together, our study demonstrated that species B-based adenoviral gene therapy is a suitable approach for generally CD46-overexpressed CRC but would require careful consideration preceding CD46 analysis and categorizing CRC patients.

CD46 is generally overexpressed in many human cancers, representing a prime target for CD46-binding adenoviruses (Ads). This could help to overcome low anti-tumoral activity by coxsackie-adenoviral receptor (CAR)-targeting cancer gene therapy viruses. However, because of scarce side-by-side information about CAR and CD46 expression levels in cancer cells, mixed observations of cancer therapeutic efficacy have been observed. This study evaluated Ad-mediated therapeutic efficacy using either CAR-targeting Ad5 or CD46-targeting Ad5/35 fiber chimera in bladder cancer cell lines. Compared with normal urothelia, bladder cancer tissue generally overexpressed both CAR and CD46. While CAR expression was not correlated with disease progression, CD46 expression was inversely correlated with tumor grade, stage, and risk grade. In bladder cancer cell lines, expression levels of CD46 and CAR were highly correlated with Ad5/35- and Ad5-mediated gene transduction and cytotoxicity, respectively. In a human EJ bladder cancer xenograft mouse model, with either overexpressed or suppressed CD46 expression levels, Ad5/35-tk followed by ganciclovir (GCV) treatment significantly affected tumor growth, whereas Ad5-tk/GCV had only minimal effects. Overall, our findings suggest that bladder cancer cells overexpress both CAR and CD46, and that adenoviral cancer gene therapy targeting CD46 represents a more suitable therapy option than a CAR-targeting therapy, especially in patients with low risk bladder cancers.

One of the most challenging aspects of cancer therapeutics is target selection. Recently, CD46 (membrane cofactor protein; MCP) has emerged as a key player in both malignant transformation as well as in cancer treatments. Normally a regulator of complement activation, CD46 is co-expressed as four predominant isoforms on almost all cell types. CD46 is highly overexpressed on a variety of human tumor cells. Clinical and experimental data support an association between increased CD46 expression and malignant transformation and metastasizing potential. Further, CD46 is a newly discovered driver of metabolic processes and plays a role in the intracellular complement system (complosome). CD46 is also known as a pathogen magnet due to its role as a receptor for numerous microbes, including several species of measles virus and adenoviruses. Strains of these two viruses have been exploited as vectors for the therapeutic development of oncolytic agents targeting CD46. In addition, monoclonal antibody-drug conjugates against CD46 also are being clinically evaluated. As a result, there are multiple early-phase clinical trials targeting CD46 to treat a variety of cancers. Here, we review CD46 relative to these oncologic connections.

… levels of expression of CD46 on EOC cells, we evaluated the targeting potential of the Ad5T*… transduce cells expressing CD46. We demonstrate a significant, CD46-dependent increase …

Initially identified as a regulator of complement activation on host cells, the known roles of CD46 (membrane cofactor protein [MCP]) have expanded. We now know that this ancient molecule is expressed on almost all nucleated cells as a family of four predominant isoforms. It also is involved in human reproduction, modulation of T cell activation and immunoinflammatory effector functions, autophagy, and the newly identified intracellular complement system (complosome). CD46 is also known as a “pathogen” magnet, being a port of entry for at least seven bacteria and five viruses. Moreover, CD46 has recently emerged as a key player in cancer biology. Numerous studies provide evidence of the association among elevated CD46 expression, malignant transformation, and metastasizing potential. These features, along with its roles as pathogen receptor, have made CD46 a target for cancer therapeutics. Thus, modified viral vectors (such as strains of adenovirus and measles virus) targeting CD46 currently are being exploited against a wide range of cancers. Another oncologic treatment utilizes a CD46-targeting human mAb as an antibody-drug conjugate. Herein, we review CD46 and its “multiverse” of cancer interactions.

PURPOSE FOR46, a fully human antibody conjugated to monomethyl auristatin E, targets a tumor-selective epitope of CD46, which is overexpressed in metastatic castration-resistant prostate cancer (mCRPC). FOR46 demonstrates potent nonclinical activity in enzalutamide-resistant CRPC models. PATIENTS AND METHODS This was a phase I, first-in-human, dose escalation/expansion study in patients with progressive mCRPC after treatment with ≥one androgen signaling inhibitors (ClinicalTrials.gov identifier: NCT03575819). The starting dose of FOR46 was 0.1 mg/kg given intravenously every 3 weeks. The primary objective was to determine the maximally tolerated dose (MTD). Whole-blood mass cytometry (cytometry by time of flight) was used to characterize peripheral immune response and CD46 expression in CRPC tissue that underwent central pathology review. RESULTS Fifty-six patients were enrolled. Dose-limiting toxicities included neutropenia (n = 4), febrile neutropenia (n = 1), and fatigue (n = 1). The MTD was 2.7 mg/kg using adjusted body weight. The most common grade ≥3 adverse events across all dose levels were neutropenia (59%), leukopenia (27%), lymphopenia (7%), anemia (7%), and fatigue (5%). One grade 3 febrile neutropenia event was observed. There were no treatment-related deaths. In the efficacy evaluable subset (patients with adenocarcinoma treated with a starting dose ≥1.2 mg/kg, n = 40), the median radiographic progression-free survival was 8.7 months (range, 0.1-33.9). Fourteen of 39 evaluable patients (36%) achieved a PSA50 response. The confirmed objective response rate was 20% (5 of 25 RECIST-evaluable patients). The median duration of response was 7.5 months. Responders had a significantly higher on-treatment frequency of circulating effector CD8+ T cells. CONCLUSION FOR46 demonstrated encouraging preliminary clinical activity with a manageable safety profile. Targeting CD46 elicited an immune priming effect that was associated with clinical outcomes.

We recently identified CD46 as a novel prostate cancer cell surface antigen that shows lineage independent expression in both adenocarcinoma and small cell neuroendocrine subtypes of metastatic castration resistant prostate cancer (mCRPC), discovered an internalizing human monoclonal antibody YS5 that binds to a tumor selective CD46 epitope, and developed a microtubule inhibitor-based antibody drug conjugate that is in a multi-center phase I trial for mCRPC (NCT03575819). Here we report the development of a novel CD46-targeted alpha therapy based on YS5. We conjugated ^212Pb, an in vivo generator of alpha-emitting ^212Bi and ^212Po, to YS5 through the chelator TCMC to create the radioimmunoconjugate, ^212Pb-TCMC-YS5. We characterized ^212Pb-TCMC-YS5 in vitro and established a safe dose in vivo. We next studied therapeutic efficacy of a single dose of ^212Pb-TCMC-YS5 using three prostate cancer small animal models: a subcutaneous mCRPC cell line-derived xenograft (CDX) model (subcu-CDX), an orthotopically grafted mCRPC CDX model (ortho-CDX), and a prostate cancer patient-derived xenograft model (PDX). In all three models, a single dose of 0.74 MBq (20 µCi) ^212Pb-TCMC-YS5 was well tolerated and caused potent and sustained inhibition of established tumors, with significant increases of survival in treated animals. A lower dose (0.37 MBq or 10 µCi ^212Pb-TCMC-YS5) was also studied on the PDX model, which also showed a significant effect on tumor growth inhibition and prolongation of animal survival. These results demonstrate that ^212Pb-TCMC-YS5 has an excellent therapeutic window in preclinical models including PDXs, opening a direct path for clinical translation of this novel CD46-targeted alpha radioimmunotherapy for mCRPC treatment.

Radiopharmaceutical therapy is changing the standard of care in prostate cancer (PCa) and other malignancies. We previously reported high CD46 expression in PCa and developed an antibody-drug conjugate and immunoPET agent based on the YS5 antibody, which targets a tumor-selective CD46 epitope. Here, we present the preparation, preclinical efficacy, and toxicity evaluation of [225Ac]DOTA-YS5, a radioimmunotherapy agent based on the YS5 antibody. Our radiolabeled antibody retains binding efficacy and shows a high tumor to background ratio in PCa xenografts. Furthermore, we show that radiolabeled antibody was able to suppress the growth of cell-derived and patient-derived xenografts, including PSMA-positive and deficient models. Nephrotoxicity, not seen at low radioactive doses, is evident at higher radioactivity dose levels, likely due to redistribution of daughter isotope 213Bi. Overall, this preclinical study confirms that [225Ac]DOTA-YS5 is a highly effective treatment and suggests feasibility for clinical translation of CD46 targeted radioligand therapy in PCa.

OBJECTIVE Multiple myeloma (MM) is an incurable B cell malignancy and novel therapeutics are urgently needed. Live attenuated measles virus (MV) has potent oncolytic activity against MM tumor xenografts. The virus is tumor selective and preferentially targets cells that express high levels of CD46 receptors. However, CD46 levels on MM have not previously been evaluated. In this study, we investigated the potential of CD46 as a target for MM therapy and correlated surface levels of CD46 on MM cells with their susceptibility to MV-induced cytopathic effects. MATERIALS AND METHODS CD46 expression on neoplastic plasma cells (PCs) and nonplasma cells (NPCs) from 38 MM patients was analyzed by flow cytometry and receptor numbers were quantitated using BD QuantiBRITE PE beads. RESULTS Results showed that malignant PCs expressed significantly higher levels of CD46 receptors compared to NPCs (p < 0.0001). The mean CD46 receptor numbers on PCs and NPCs were 49,130/cell and 7,340/cell, respectively. Potent cytopathic effects of extensive intercellular fusion were observed in measles-infected PCs but not in NPCs. The extent of MV-induced cytopathic effects of cell fusion correlated with CD46 expression levels on the MM cells. Normal plasma cells do not overexpress CD46 and colony-forming assays demonstrated that MV was not cytotoxic to normal bone marrow progenitor cells. CONCLUSION The present study establishes CD46 as a surface antigen that is expressed more abundantly on primary MM cells compared to normal hematopoietic cells of various lineages in the bone marrow, making CD46 a promising surface marker for targeted cytoreductive therapy of MM.

Mutations in CD46 predispose to atypical hemolytic uremic syndrome (aHUS) with low penetrance. Factors driving immune-dysregulatory disease in individual mutation carriers have remained ill-understood. In addition to its role as a negative regulator of the complement system, CD46 modifies T cell-intrinsic metabolic adaptation and cytokine production. Comparative immunologic analysis of diseased vs . healthy CD46 mutation carriers has not been performed in detail yet. In this study, we comprehensively analyzed clinical, molecular, immune-phenotypic, cytokine secretion, immune-metabolic, and genetic profiles in healthy vs . diseased individuals carrying a rare, heterozygous CD46 mutation identified within a large single family. Five out of six studied individuals carried a CD46 gene splice-site mutation causing an in-frame deletion of 21 base pairs. One child suffered from aHUS and his paternal uncle manifested with adult-onset systemic lupus erythematosus (SLE). Three mutation carriers had no clinical evidence of CD46-related disease to date. CD4^+ T cell-intrinsic CD46 expression was uniformly 50%-reduced but was comparable in diseased vs . healthy mutation carriers. Reconstitution experiments defined the 21-base pair-deleted CD46 variant as intracellularly—but not surface-expressed and haploinsufficient. Both healthy and diseased mutation carriers displayed reduced CD46-dependent T cell mitochondrial adaptation. Diseased mutation carriers had lower peripheral regulatory T cell (Treg) frequencies and carried potentially epistatic, private rare variants in other inborn errors of immunity (IEI)-associated proinflammatory genes, not found in healthy mutation carriers. In conclusion, low Treg and rare non- CD46 immune-gene variants may contribute to clinically manifest CD46 haploinsufficiency-associated immune-dysregulation.

Complement is the canonical innate immune system involved in host defense and tissue repair with the clearance of cell debris. In contrast to the robust armory mounted against microbial nonself-pathogens, complement is selectively activated on altered self (i.e. apoptotic and necrotic cells) to instruct the safe demise by poorly characterized mechanisms. Our data shed new light on the role of complement C1q in sensing nucleic acids (NA) rapidly exposed on apoptotic Jurkat T cell membranes and in driving C3 opsonization but without the lytic membrane attack complex. DNA/RNase-treated apoptotic cells failed to activate complement. We found that several other apoptotic cell models, including senescent keratinocytes, ionophore-treated sperm cells, and CMK-derived platelets, stained for cleaved caspase 3 were rapidly losing the key complement regulator CD46. CD46 from nuclear and membrane stores was found to cluster into blebs and shed into microparticles together with NA, phosphatidylserine, C1q, and factor H. Classical and alternative pathways of complement were involved in the recognition of H2O2-treated necrotic cells. Membrane attack complex was detected on necrotic cells possibly as a result of CD46 and CD59 shedding into soluble forms. Our data highlight a novel and universal paradigm whereby the complement innate immune system is using two synergistic strategies with the recognition of altered self-NA and missing self-CD46 signals to instruct and tailor the efficient removal of apoptotic and necrotic cells in immunoprivileged sites.

… and immunoblotting of breast tissue for CD46. CD46 was consistently and strongly expressed in … Immunoblotting of breast tissue for CD46 showed the same size of the molecule as for …

… that CD46 and CD59 are immunogenic in macaques was found when anti-cell antibodies in plasmas from macaques immunized with human cell-grown SIV blocked anti-CD46 and anti-…

The role of membrane-bound complement regulatory proteins (mCRP) in the protection of tumor cells in vivo against elimination by the immune system is still unknown. In this study the …

CD46 is a complement regulator with important roles related to the immune response. CD46 functions as a pathogen receptor and is a potent costimulator for the induction of interferon-γ (IFN-γ)-secreting effector T helper type 1 (TH1) cells and their subsequent switch into interleukin 10 (IL-10)-producing regulatory T cells. Here we identified the Notch family member Jagged1 as a physiological ligand for CD46. Furthermore, we found that CD46 regulated the expression of Notch receptors and ligands during T cell activation and that disturbance of the CD46-Notch crosstalk impeded induction of IFN-γ and switching to IL-10. Notably, CD4+ T cells from CD46-deficient patients and patients with hypomorphic mutations in the gene encoding Jagged1 (Alagille syndrome) failed to mount appropriate TH1 responses in vitro and in vivo, which suggested that CD46-Jagged1 crosstalk is responsible for the recurrent infections in subpopulations of these patients.

Summary Background The increasing prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) incurs substantial morbidity, mortality and healthcare costs. Detection and clinical intervention at early stages of disease improves prognosis; however, we are currently limited by a lack of reliable diagnostic tests for population screening and monitoring responses to therapy. To address this unmet need, we investigated human invariant Natural Killer T cell (iNKT) activation by fat-loaded hepatocytes, leading to the discovery that circulating soluble CD46 (sCD46) levels accurately predict hepatic steatosis. Methods sCD46 in plasma was measured using a newly developed immuno-competition assay in two independent cohorts: Prospective living liver donors (n = 156; male = 66, female = 90) and patients with liver tumours (n = 91; male = 58, female = 33). sCD46 levels were statistically evaluated as a predictor of hepatic steatosis. Findings Interleukin-4-secreting (IL-4+) iNKT cells were over-represented amongst intrahepatic lymphocytes isolated from resected human liver samples. IL-4+ iNKT cells preferentially developed in cocultures with a fat-loaded, hepatocyte-like cell line, HepaRG. This was attributed to induction of matrix metalloproteases (MMP) in fat-loaded HepaRG cells and primary human liver organoids, which led to indiscriminate cleavage of immune receptors. Loss of cell-surface CD46 resulted in unrepressed differentiation of IL-4+ iNKT cells. sCD46 levels were elevated in patients with hepatic steatosis. Discriminatory cut-off values for plasma sCD46 were found that accurately classified patients according to histological steatosis grade. Interpretation sCD46 is a reliable clinical marker of hepatic steatosis, which can be conveniently and non-invasively measured in serum and plasma samples, raising the possibility of using sCD46 levels as a diagnostic method for detecting or grading hepatic steatosis. Funding F.B. was supported by the Else Kröner Foundation (Award 2016_kolleg.14). G.G. was supported by the Bristol Myers Squibb Foundation for Immuno-Oncology (Award FA-19-009). N.S. was supported by a 10.13039/100010269Wellcome Trust Fellowship (211113/A/18/Z). J.A.H. received funding from the 10.13039/501100007601European Union’s Horizon 2020 research and innovation programme (Award 860003). J.M.W. received funding from the Else Kröner Foundation (Award 2015_A10).

… CHS developed equivalently in wild-type mice and all CD46-transgenic lines; this excluded the possibility that the transgene affected the ability of the mice to mount an inflammatory …